RxPG News : Genetics

Genetic study of bedbugs may help identify pesticide resistance genes

Wed, 19 Jan 2011 17:08:01 PST

( from http://www.rxpgnews.com ) OhioState University entomologists have conducted the first genetic study of bedbugs, paving the road to the identification of potential genes associated with pesticide resistance and possible new control methods for the troublesome insect, whose sudden resurgence in the United States has led to a public health scare.
The discovery was reported Jan. 19 in the online journal PLoS ONE.
"While bedbugs are poised to become one of the major household pests across the United States in the coming years, we know very little about their genetic makeup and their mechanisms of resistance to insecticides," said Omprakash Mittapalli, corresponding author of the study and an assistant professor of entomology with the university's Ohio Agricultural Research and Development Center in Wooster.
"This is the first study to elucidate the genetic make up of the insect and to obtain fundamental molecular knowledge regarding potential defense pathways and genes that may be involved in metabolic resistance to commonly used pesticides."
A minor nuisance since World War II as a result of the widespread use of DDT and other long-lasting residual insecticides, bedbug (Cimex lectularius) numbers have increased in the past decade as much as 500 percent in North America and other parts of the world — costing billions of dollars to homeowners and businesses annually and requiring the use of large quantities of pesticides, many of them ineffective.
Reasons behind the spike in bedbug infestations include a boom in international travel, increased exchange of used furniture, a shift from powerful but dangerous insecticides such as DDT to more selective control tactics, and the development of resistance among bedbug populations to currently used pesticides — pyrethroids in particular.
"The common assumption today is that pesticide resistance in bedbugs results from point mutations in certain genes," Mittapalli explained. "However, the role of detoxification and antioxidant enzymes in pesticide resistance of bedbugs is poorly understood.
Enzymes such as Cytochrome P450s and glutathione S-transferases (GSTs) have been shown in other insects to act as detoxification agents, allowing the insects to get rid of toxic compounds such as insecticides and not be killed by them. Our study looked closely at those groups of enzymes in bedbugs."
For the study, Mittapalli and his team employed 454 sequencing technology, which has recently enabled the application of functional genomics to a broad range of insect species previously unexplored at the molecular level. They analyzed both laboratory-reared bedbugs susceptible to insecticides (the Harlan strain) and pesticide-exposed bedbugs collected from a Columbus, Ohio apartment in 2009 and 2010.
This analysis led to the identification of 35,646 expressed sequence tags, or ESTs, which are instrumental in gene discovery and sequencing work. Before this study was conducted, less than 2,000 ESTs for C. lectularius had been filed in the National Center for Biotechnology Information (NCBI) databases. This information alone is expected to advance additional genetic studies of bedbugs and comparative molecular analyses of blood-feeding insects.
"From the database we created, we profiled the transcript level for a cytochrome P450 (CYP9) and a GST (Delta-epsilon) in different developmental stages (early-stage nymphs, late-stage nymphs and adults) of pesticide-susceptible and pesticide-exposed bedbugs," Mittapalli said. "We found higher transcript levels for CYP9 in all developmental stages in pesticide-exposed populations compared to pesticide-susceptible populations. We also found higher transcript levels of Delta-epsilon in the late-instar nymphs of pesticide-exposed bedbug populations."
Further studies — including gene silencing, or "knocking down," the CYP9 and Delta-epsilon candidate genes to confirm that they are indeed involved in pesticide resistance — are still needed.
"The insecticides being used right now are based on the idea that resistance in bedbugs is caused by point mutations in genes," Mittapalli pointed out. "But we are finding out that the mode of resistance could be attributed to a combination of changes in the bug's genetic makeup (such as mutations) as well as transcriptomic adjustments leading to differential gene expression. Pinpointing such defense mechanisms and the associated genes could lead to the development of novel methods of control that are more effective."

Novel method of database analysis to help identify responsible genes and diagnostic markers

Tue, 11 Jan 2011 17:53:27 PST

( from http://www.rxpgnews.com ) Researchers at the University of Gothenburg, Sweden, have developed new methods for analysing medical databases that can be used to identify diagnostic markers more quickly and to personalise medication for allergic disorders. They could also reduce the need for animal trials in clinical studies.

Published in the journal PLoS Computational Biology, the study builds on data analyses of freely available medical databases representing studies of countless numbers of patients in the PubMed database, and microarray data in another major database. The use of microarrays is a method that allows scientists to study all 20,000 human genes at the same time for various disorders.

Groups of researchers in Gothenburg, Oslo and Rome have developed computational methods to simulate how a change in the interaction between several different genes in the lymphocytes (a kind of white blood cell) controls the immune system. They identified the genes by reviewing abstracts of all 18 million articles included in PubMed, and then constructed a network model of how these genes interact.

"The model can be compared to a printed circuit card in the lymphocyte which the cell uses to make decisions about whether to activate or suppress the immune system," says Mikael Benson, a researcher at the Sahlgrenska Academy's Unit for Clinical Systems Biology and consultant at the Queen Silvia Children's Hospital. "These decisions are made constantly as the lymphocytes are constantly exposed to different particles, just through breathing for example. Some of the particles could be dangerous and need to trigger a decision to mobilise the immune system. However, sometimes wrong decisions are made, which can lead to various disorders such as allergy or diabetes."

The researchers then carried out data simulations of how the network model reacted to repeated exposure to particles, which resulted in four reaction patterns, one of which was to suppress the immune system, while the other three were to trigger it in various ways.

"We found that the genes in the model reacted in lymphocytes from patients with various immunological disorders. We'll be using the model to identify diagnostic markers so that we can personalise medication that we're testing in clinical studies of allergy patients."

Benson believes that these methods will become increasingly important in the future, as the huge amount of information in medical databases is growing all the time. This information could serve as an important resource for researchers in their endeavours to investigate and verify medical hypotheses.

"These methods could reduce the need for animal trials and lead to major savings in both time and money," says Benson. "They could also mean quicker and better-designed experiments and their results could generate new knowledge about diagnostic markers or new medicines."

Environmental influences can be passed down to the next generation

Tue, 28 Dec 2010 09:48:13 PST

( from http://www.rxpgnews.com ) Scientists at the University of Massachusetts Medical School and the University of Texas at Austin have uncovered evidence that environmental influences experienced by a father can be passed down to the next generation, "reprogramming" how genes function in offspring. A new study published this week in Cell shows that environmental cues—in this case, diet—influence genes in mammals from one generation to the next, evidence that until now has been sparse. These insights, coupled with previous human epidemiological studies, suggest that paternal environmental effects may play a more important role in complex diseases such as diabetes and heart disease than previously believed.

"Knowing what your parents were doing before you were conceived is turning out to be important in determining what disease risk factors you may be carrying," said Oliver J. Rando, MD, PhD, associate professor of biochemistry & molecular pharmacology at UMMS and principal investigator for the study, which details how paternal diet can increase production of cholesterol synthesis genes in first-generation offspring.

The human genome is often described as the set of instructions that govern the development and functioning of life. It's not surprising, then, that most contemporary genetic research focuses on understanding and cataloging how mutations and changes to our DNA—the basis of those "instructions"—cause disease and impact health. A number of recent studies, however, have begun to draw attention to the role epigenetic inheritance – inherited changes in gene expression caused by mechanisms other than changes in the underlying DNA sequence – may play in a host of illnesses. "A major and underappreciated aspect of what is transmitted from parent to child is ancestral environment," said Dr. Rando. "Our findings suggest there are many ways that parents can 'tell' their children things."

To test their hypothesis that environmental influences experienced by the father can be passed down to the next generation in the form of changed epigenetic information, Rando and colleagues fed different diets to two groups of male mice. The first group received a standard diet, while the second received a low-protein diet. To control for maternal influences, all females were fed the same, standard diet. Rando and colleagues observed that offspring of the mice fed the low-protein diet exhibited a marked increase in the genes responsible for lipid and cholesterol synthesis in comparison to offspring of the control group fed the standard diet.

These observations are consistent with epidemiological data from two well-known human studies suggesting that parental diet has an effect on the health of offspring. One of these studies, called the Överkalix Cohort Study, conducted among residents of an isolated community in the far northeast of Sweden, found that poor diet during the paternal grandfather's adolescence increased the risk of diabetes, obesity and cardiovascular disease in second-generation offspring. However, because these studies are retrospective and involve dynamic populations, they are unable to completely account for all social and economic variables. "Our study begins to rule out the possibility that social and economic factors, or differences in the DNA sequence, may be contributing to what we're seeing," said Rando. "It strongly implicates epigenetic inheritance as a contributing factor to changes in gene function."

The results also have implications for our understanding of evolutionary processes, says Hans A. Hofmann, PhD, associate professor of integrative biology at the University of Texas at Austin and a co-author of the study. "It has increasingly become clear in recent years that mothers can endow their offspring with information about the environment, for instance via early experience and maternal factors, and thus make them possibly better adapted to environmental change. Our results show that offspring can inherit such acquired characters even from a parent they have never directly interacted with, which provides a novel mechanism through which natural selection could act in the course of evolution." Such a process was first proposed by the early evolutionist Jean-Baptiste Lamarck, but then dismissed by 20th century biologists when genetic evidence seemed to provide a sufficient explanation.

Taken together, these studies suggest that a better understanding of the environment experienced by our parents, such as diet, may be a useful clinical tool for assessing disease risk for illnesses, such as diabetes or heart disease. "We often look at a patient's behavior and their genes to assess risk," said Rando. "If the patient smokes, they are going to be at an increased risk for cancer. If the family has a long history of heart disease, they might carry a gene that makes them more susceptible to heart disease. But we're more than just our genes and our behavior. Knowing what environmental factors your parents experienced is also important."

The next step for Rando and colleagues is to explore how and why this genetic reprogramming is being transmitted from generation to generation. "We don't know why these genes are being reprogrammed or how, precisely, that information is being passed down to the next generation," said Rando. "It's consistent with the idea that when parents go hungry, it's best for offspring to hoard calories, however, it's not clear if these changes are advantageous in the context of a low-protein diet."

Gene found to be key in etiology of cleft palate

Tue, 02 Feb 2010 13:54:58 PST

( from http://www.rxpgnews.com ) Cleft palate has been linked to dozens of genes. During their investigation of one of these genes, researchers at Washington University School of Medicine in St. Louis were surprised to find that cleft palate occurs both when the gene is more active and when it is less active than normal.

They say the finding suggests this gene and processes closely associated with it are central to palate development and could become important targets for investigators seeking nonsurgical treatments to prevent cleft palate before birth. Their report will appear in an upcoming issue of the Proceedings of the National Academy of Sciences.

"Palate formation in the embryo is a complex process, and many things can go wrong," says senior author David M. Ornitz, M.D., Ph.D., Alumni Endowed Professor of Developmental Biology. "A cleft palate is often diagnosed late in pregnancy and treated surgically after birth. But if we understood the genetic causes of this common birth defect, we might be able to diagnose it much earlier. That would potentially allow intervention with prenatal surgery or with drugs or other agents designed to counteract the genetic abnormalities."

Clefts of the lip and palate affect about one in 700 newborns worldwide. Children with cleft lip and palate can have difficulty feeding as infants and can have speech, dental and hearing problems as they grow older. Depending on severity, surgical repair can require several operations over many years, and the estimated average lifetime cost of treatment in the United States is about $100,000 per patient.

"We believe the more information we have on the causes of cleft palate, the better hope we have for possibly preventing and more effectively treating the condition," says lead author Alison K. Snyder-Warwick, M.D., a plastic surgery resident at Barnes-Jewish Hospital.

Although some cases of cleft lip and palate are linked to environmental factors such as maternal smoking, viral infections or certain medications, genetic variations play a significant role in many cases. The Washington University researchers studied the fibroblast growth factor receptor 2 (FGFR2) gene, which earlier research has implicated in cleft palate.

They focused on mice with Crouzon syndrome, a developmental disorder caused by a mutation in FGFR2. The mutation activates the receptor and results in a syndrome that is characterized by abnormal development of the skull, face and mouth and is associated with an increased incidence of cleft palate.

In effect, the FGFR2 mutation prevents specific growth signals from being switched off. Normally, the signals would be turned on and off in a carefully orchestrated manner to ensure proper patterns of growth and development of embryonic tissues. However, the Crouzon syndrome mutation locks the receptor in a permanently on position.

As mouse embryos with the mutation grew, cells destined to become the palate initially grew faster than normal cells, as anticipated. But just before palate formation, the growth of these cells lagged behind their normal pace of proliferation. That was unexpected because the signals created by mutant FGFR2 should logically have maintained an increased rate of cell proliferation in the palate, Ornitz indicates.

In a normal mouse embryo, groups of cells called the palatal shelf on either side of the mouth grow outward, elevate to meet in the middle and fuse to form the palate. But in the mutant mice embryos, the stunted growth of this tissue prevented the palatal shelves from properly elevating, meeting and fusing. In addition, the researchers detected a decrease in some key components of the supporting matrix between cells of the palate.

Another of the study's coauthors, Kai Yu, Ph.D., a scientist in the Ornitz lab, created genetically engineered mice in which FGF receptors were inactivated in tissue that gives rise to the palate. These mice also developed cleft palate.

In palate cells grown in the lab, the researchers looked at the FGF cell-signaling network, in which FGFR2 participates. They compared the effect of increased activity of FGF signaling with decreased activity of the same network, and interestingly, both led to cleft palate.

"We found that overactivation of an important signaling pathway resulted in loss of function," Snyder-Warwick says. "Our results suggest a different way of thinking about mutations in the FGF signaling network. This study clearly showed that this FGF signaling pathway is a critical regulator of palate development."

These findings strengthen the evidence that the developmental processes in which FGFR2 is involved could be targeted with drugs in order to ensure normal palate growth.

"It's possible that someday doctors might be able to administer drugs that would either slightly activate or slightly inhibit FGFR2 function," Ornitz says. "That might be enough to tip the balance from a cleft palate to a normal palate during embryonic development."

History, geography also seem to shape our genome

Thu, 18 Jun 2009 18:00:21 PST

( from http://www.rxpgnews.com ) History and geography shape our genome, according to a new study.

The movements of humans within and among continents, expansions and contractions of populations and vagaries of genetic chance, have influenced the distribution of genetic variations.

In recent years, geneticists have identified a handful of genes that have helped human populations adapt to new environments within just a few thousand years - a strikingly short time scale in evolutionary terms.

However, a team from the Universities of Chicago, California and Stanford, which jointly conducted the study, found that for most genes, it can take at least 50,000-100,000 years for natural selection to spread favourable traits through a human population.

They found that gene variants tend to be distributed throughout the world in patterns that reflect ancient population movements and other aspects of population history.

'We don't think that selection has been strong enough to completely fine-tune the adaptation of individual human populations to their local environments,' says study co-author Jonathan Pritchard, professor in human genetics and Howard Hughes Medical Institute investigator.

'In addition to selection, demographic history -- how populations have moved around -- has exerted a strong effect on the distribution of variants,' he added.

Selection may still be occurring in many regions of the genome, said Pritchard. But if so, it is exerting a moderate effect on many genes that together influence a biological characteristic, according to a Howard Hughes release.

'We don't know enough yet about the genetics of most human traits to be able to pick out all of the relevant variation,' said Pritchard.

'As functional studies go forward, people will start figuring out the phenotypes - associated with selective signals,' said lead study author Graham Coop.

'That will be very important, because then we can figure out what selection pressures underlie these episodes of natural selection.'

The study was published in the Friday edition of the open-access journal PLoS Genetics.

Induced pluripotent stem cell lines from pigs

Sun, 07 Jun 2009 03:44:15 PST

( from http://www.rxpgnews.com ) The discovery that adult skin cells can be 'reprogrammed' to behave like stem cells has been a major scientific boon, providing a way to tap the potential of embryonic stem cells without the associated ethical quandaries. Now, in a study appearing online in JBC, researchers have created a line of such reprogrammed stem cells from adult pigs. As pigs are large animals with a physiology very similar to humans, this work provides a valuable model to study the therapeutic potential of this new "induced pluripotent stem cell" (iPS) technology.

iPS cells have already been developed from both mice and humans. Both systems will help researchers answer many biological and genetic questions about these cells, but still leave a gap before clinical applications can begin. These iPS cells cannot be tested on humans before thorough safety and efficacy trials in animal models, but the size, physiology and short lifespan of mice makes them less than ideal for these trials.

Duanqing Pei and colleagues turned to a better pre-clinical model: pigs. These large animals share a remarkably similar biology to humans, as evidenced by their already extensive contributions to medicine, such as using pig insulin to treat diabetes or pig heart valves in transplant surgery. The research group modified the current iPS protocols to successfully generate a line of stem cells from a miniature Tibetan pig (whose smaller size would make breeding and maintenance easier). A biochemical analysis revealed these cells expressed the key proteins that would classify them as 'stem cells' and had the ability to differentiate into many other types of cells.

Importantly, these pig iPS cells more closely resembled human stem cells than other animals, confirming their value in pre-clinical studies. The researchers believe porcine iPS technology is an emerging and exciting field that should progress quickly and lead to many applications

Egg cells help extend life of sperms

Wed, 25 Mar 2009 15:59:20 PST

( from http://www.rxpgnews.com ) In contrast to women, men are fertile throughout life, but research at the Sahlgrenska Academy, University of Gothenburg, Sweden, has now shown that a fertilising sperm can get help from the egg to rejuvenate. The result is an important step towards future stem cell therapy.

The risk of chromosomal abnormalities in the foetus is highly correlated to the age of the mother, but is nearly independent of the age of the father. One possible explanation is that egg cells have a unique ability to reset the age of a sperm.

"We are the first to show that egg cells have the ability to rejuvenate other cells, and this is an important result for future stem cell research", says Associate Professor Tomas Simonsson, who leads the research group at the Sahlgrenska Academy that has made this discovery.

Each time a cell divides, the genetic material at the ends of the chromosomes becomes shorter. The ends of the chromosomes, known as "telomeres", are important for the genetic stability of the cell and they act as a DNA clock that measures the age of the cell. The cell stops dividing and dies when the telomeres become too short.

The discovery that the egg cell can extend the telomeres of a fertilising sperm cell is important in the development of stem cell therapy. Stem cell therapy involves replacing the cell nucleus in unfertilised egg with a nucleus from a somatic cell that has come from a patient who needs a stem cell transplantation. As soon as the cell has divided a few times, it is possible to harvest stem cells that are then allowed to mature to the cell type that the recipient needs.

"The genetic stability of the transplanted cells has been a serious concern up until now, and it was feared that the lifetime of these cells would depend on the age of the cell nucleus that was transferred. Our results suggest that this is not the case", says Tomas Simonsson.

Family of genes known as KRAB-ZFP regulate genes dealing with stress

Fri, 12 Dec 2008 13:33:47 PST

( from http://www.rxpgnews.com ) Research conducted by a team in Switzerland suggests that a family of genes involved in regulating the expression of other genes in the brain is responsible for helping us deal with external inputs such as stress. Their results, appearing in the December 11 advance online version of the journal Neuron, may also give a clue to why some people are more susceptible to anxiety or depression than others.

The researchers from EPFL and the National Competence Center "Frontiers in Genetics" studied the role of a family of genes known as KRAB-ZFP, which acts like a group of genetic censors, selectively silencing the expression of other genes. These repressors make up about 2% of our genetic material, but little is known about how this "epigenetic" silencing process works, what the long-term consequences are, and even which genes are targeted. (Epigenetics refers to a change in gene expression that is caused by something other than a change in the underlying DNA sequence.)

The researchers bred a strain of mice that lacked in the hippocampus, a part of the forebrain involved in short-term memory and inhibition, a key cofactor used by the KRAB family. The genetically altered mice appeared completely normal until they were placed in a stressful situation. Then they became extremely anxious. Although the normal mice quickly adapted, the altered mice never managed to overcome their stress, and remained anxious and unable to complete simple cognitive tasks. The disruption of the KRAB-mediated regulatory process thus altered the mice's normal behavioral response to stress.

"The KRAB regulators appeared fairly recently on an evolutionary scale," notes EPFL professor Didier Trono, lead author on the study, " and it's very likely that there is a fair degree of polymorphism between individuals. We postulate that variability in these genes is one factor that may participate in predisposing people to anxiety syndromes or depression. "

Because epigenetic alterations are often long-lasting and sometimes permanent, one could also interpret them as a way in which an individual's personal history can have a lasting impact on his or her genetic expression. "It's a way for a cell to have a sort of memory," explains Trono.

This work opens promising leads for further exploration, because evidence of epigenetic modification has been observed in animal models of depression, addiction, schizophrenia and neuro-developmental disorders. Some psychoactive drugs like cocaine or anti-psychotics also cause changes in some of the co-factors involved in this genetic regulatory system. With an understanding of the molecular mechanisms involved in epigenetic modulation, it might be possible to develop targeted therapies for those individuals in whom it malfunctions.

New screening strategy increases Down's syndrome detection before birth

Sat, 29 Nov 2008 03:34:26 PST

( from http://www.rxpgnews.com ) A new national screening strategy in Denmark has halved the number of infants born with Down's syndrome and increased the number of infants diagnosed before birth by 30%, according to a study published on bmj.com.

Many countries, including England, Australia and New Zealand, are trying to introduce national screening strategies for Down's syndrome, but are facing a variety of problems because of a lack of consensus about the screening policy and logistical challenges.

In 2004, the Danish National Board of Health issued new guidelines for prenatal screening and diagnosis. These included the offer of a combined test for Down's syndrome (based on combination of maternal age, plus serum and nuchal screening) in the first trimester. This test gave women a risk assessment for Down's syndrome at an early stage in the pregnancy. Women whose risk was higher than a defined cut off were referred for invasive diagnostic tests (chorionic villus sampling or amniocentesis).

In the previous guidelines screening for Down's syndrome was based on maternal age and a diagnostic test was mainly offered to women above 35 years.

Professor Ann Tabor and colleagues from Denmark, evaluated the impact of the new national screening strategy on the number of infants born with Down's syndrome and the number of referrals for invasive procedures. They analysed data from the 19 Danish departments of gynaecology and obstetrics and the national cytogenetic registry, for an average of 65,000 births each year, between 2000 and 2007.

Uptake was good, by June 2006 all 15 Danish counties followed the guidelines from 2004 and offered the new screening strategy. In 2006 approximately 84% of pregnant women had a risk assessment for Down's syndrome.

The researchers found that the new strategy was associated with improved earlier detection of Down's syndrome, low false positive rates, and more than a 50% decrease in the number of invasive tests carried out each year.

They report that the number of infants born with Down's syndrome decreased from 55󈞭 per year during 2000𔃂, to 31 in 2005 and 32 in 2006. The total number of invasive tests fell sharply from 7524 in 2000 to 3510 in 2006.

The detection rate in the screened population was 86% in 2005 and 93% in 2006. With 3.9% (17) of women receiving a false positive result in 2005 and 3.3% (7) in 2006.

The authors point out that the value of this new screening strategy is that all women can be assessed early in pregnancy (in the first trimester). The national guidelines emphasise that risk assessment should only be done if women choose the test on the basis of informed choice, therefore despite the programme being available to all pregnant in women in Denmark, some will still choose not to be screened.

The authors conclude by emphasising Denmark's success at building a strong national organisation for fetal medicine and a national quality database that allows follow-up of all screened women at a national level and quality control of the new national screening programme.

Can genetic research spur racist attitudes?

Wed, 19 Nov 2008 11:59:33 PST

( from http://www.rxpgnews.com ) Toronto, Nov 18 - People might be different in many ways but genetically they are quite similar. However, is it possible that genetic research may evoke racist attitudes, asks University of Alberta's Tim Caulfield. He organised a seminar to examine the issue.

Last year, Nobel Prize winning geneticist James Watson claimed there are genes responsible for creating differences in human intelligence. These comments made international headlines and Watson later apologised.

Caulfield knows that studying racial groups is important. For example, if a researcher is studying health disparities in the US, they want to know why African Americans have poorer outcomes than those of European descent.

'In that case you're not saying that there's a biological difference because you're incorporating social and economic factors to that definition,' said Caulfield.

Accordingly, Caulfield brought together an interdisciplinary group to discuss the concerns of the scientific community and come up with ways to avoid it. This group included professionals in anthropology, bioethics, clinical medicine and law.

'It was a very interesting group of individuals that haven't always agreed in the past,' said Caulfield. They managed to come together and agree on this topic, though, detailing a number of steps to ensure biomedical research doesn't stir up racism.

'For example, scientists must justify in the study why they're studying that certain group. When a discovery is made, researchers are to ensure the evidence is defined properly in the hard copy of the study and to the media,' according to an Alberta release.

Besides, Caulfied's group will continue to track the ways published studies reference ethnic groups. 'We're trying to trace how race-based studies are described in various stages.'

'We're continuing to study the issue in how race is represented,' said Caulfield, whose study will appear in January edition of Genome Medicine.

Atlas of kidney genome created

Thu, 13 Nov 2008 10:43:26 PST

( from http://www.rxpgnews.com ) A comprehensive genome-based atlas, created by researchers, would help shed light on healthy and abnormal kidney development and disease. The atlas shows how the entire genome is regulated to produce thousands of specific genes that are mixed and re-mixed to form genetic teams.

It is the joint outcome of the work by Cincinnati Children's Hospital Medical Centre, Institute for Molecular Bioscience - University of Queensland - and Harvard University researchers.

The teams jointly directed formation of 15 embryonic sections in developing kidney - from the earliest phases when stem cells are told how to differentiate into specific kidney cells, to the development of nephrons, the kidney's primary functioning unit.

Given that about one in every 500 births results in a kidney development abnormality, the study is a beginning for providing new insight into genes and genetic programmes that are critical to determining how kidney stem cells develop into structures in the adult kidney.

'Researchers can refer to the atlas to see the gene expression patterns in a normal developing kidney,' said Melissa Little, a professor who led the Australian team.

'It will provide a basis of comparison for scientists studying abnormal kidney development, so they can see where gene interactions have gone awry to produce the abnormality.'

Researchers created the atlas by analysing mouse embryonic kidneys aged 15.5 days, according to a University of Queensland release.

One of the more unexpected discoveries was the observation of new domains of gene expression that marked clusters of cells not previously known to be distinct.

The data has been released as an open-access resource for researchers around the world as part of the GenitoUrinary Development Molecular Anatomy Project.

Customised DNA-based prescriptions to avert drug reactions

Fri, 24 Oct 2008 16:42:27 PST

( from http://www.rxpgnews.com ) Washington, Oct 24 - Customised DNA-based prescriptions could help avert adverse drug reactions, a new research has found.

Warfarin, the most widely prescribed anti-coagulant, heads the list of problem drugs.

Evgeny Krynetskiy, associate professor and director of Jayne Haines Centre for Pharmacogenomics and Drug Safety of Temple University, has focused his research on that drug.

'Prescribing this medicine is like trial and error in finding the right dosage that works best for you,' says Krynetskiy. 'Five milligrams is a typical dose, but a little less or a little more could have dramatic consequences or no benefit at all.'

Accordingly, the medical community is learning how to use genetic information to tailor drug regimens for patients, and so are medical students, by genotyping themselves. All 153 medicos will extract their own DNA through collected saliva samples to see how they would react to the anti-tuberculosis drug Isoniazid.

Doctors call this optimal dosage the therapeutic window, and Krynetskiy is trying to find it through pharmacogenomics, the study of a person's response to drugs based on their genetic makeup, said a Temple University release.

It's a collaboration that crosses campuses and includes Krynetskiy and fellow clinical faculty at the School of Pharmacy, clinicians at Temple University Hospital and Jeannes Hospital.

Researchers are studying why people process the same drug differently. In this case, they are trying to find the correlation between genotypes, or a person's inner code of DNA, and the correct dosage of Warfarin.

By collecting saliva samples and extracting DNA from 77 participants already on the drug, the researchers can look for variances, genetic clues, which make people metabolise the same drug in very different ways.

That would allow doctors to prescribe the correct dosage of Warfarin and decrease the risk of adverse drug reactions: Too low a dose can increase the risk of dangerous blood clots, while too large can cause life-threatening bleeding.

XXYY syndrome- features and treatment options elucidated by researchers

Sun, 24 Aug 2008 01:16:40 PST

( from http://www.rxpgnews.com ) Researchers at the UC Davis M.I.N.D. Institute and The Children's Hospital in Denver have conducted the largest study to date describing the medical and psychological characteristics of a rare genetic disorder in which males have two "X" and two "Y" chromosomes, rather than the normal one of each. The study, published in the June 15, 2008, issue of the American Journal of Medical Genetics Part A, also offers treatment recommendations for men and boys with the disorder.

"We found that there are a variety of behaviors, learning disabilities and emotional problems that are unique to patients with XXYY syndrome that may be better addressed with more targeted therapies," said Randi Hagerman, medical director of the M.I.N.D. Institute and senior author of the study. "Our research is important because it provides an accurate picture of what patients are experiencing that can help physicians who treat patients with the disorder."

XXYY syndrome is a sex chromosome anomaly that is thought to occur in about one in 18,000 males in the general population. Boys with XXYY syndrome usually come to the attention of physicians because of unique facial features, developmental delays, late puberty and behavioral problems. It was once thought to be a variant of Klinefelter syndrome, in which males have one extra X chromosome. While the two disorders are similar in some ways, clinicians have become increasingly aware that they are distinct in some significant ways. The current study set out to identify the unique features of patients with XXYY for the purposes of informing the medical community and improving treatment approaches.

"Until now, physicians have had to search the medical literature to patch together a treatment plan mostly based on information on Klinefelter syndrome," said Nicole Tartaglia, an assistant professor of pediatrics at the University of Colorado Denver School of Medicine who was a fellow at the M.I.N.D. Institute when the study was conducted. "As a result, people with XXYY weren't being screened for the specific medical problems associated with their disorder. They weren't receiving therapies or medications for the behavioral and neurodevelopmental issues that are more profound for them. And they weren't receiving the types of community services that can help them live independent lives. Our research is an important resource for families and practitioners."

For the current study, Tartaglia and Hagerman examined 95 males with XXYY syndrome between the ages of one and 55 years of age. Among their medical findings were that 19.4 percent had cardiac abnormalities such as congenital heart defects and mitral valve prolapse, 87.6 percent had dental problems such as severe dental caries and malocclusion, 15 percent had seizures and 59.8 percent had asthma or other respiratory issues. Intention tremor became more common with age and was present in 71 percent of study participants over 20 years old. 45.7 percent who underwent brain MRIs showed abnormal white matter that may explain some learning difficulties.

Psychologically, the researchers found that 72.2 percent had attention-deficit/hyperactivity disorder and up to 28.3 percent had autism spectrum disorders. In the previous literature, mental retardation was the norm. This study, however, found that only 29.1 percent had IQ scores within the mental retardation range. Learning disabilities were the more common cognitive impairments, affecting 70.9 percent of study participants.

"Life skills are more of a struggle for these males, and they may need different medications, a broader array of behavioral therapies and more intensive community support than those with Klinefelter syndrome," Tartaglia said.

Lack of comprehensive information about the syndrome is what drove the current study. For years, parents of boys with XXYY syndrome supported each other over the Internet, sharing stories of heartbreak and frustration. While their sons suffered everything from heart defects to learning disabilities, they could only point doctors and teachers to a 1960s scientific paper that first identified the condition along with a few outdated notes on its outcomes.

"We knew we needed a more complete description," said Renee Beauregard, of Aurora, Col., whose 26-year-old son, Kyle, was diagnosed with XXYY syndrome at age 10. "We were tired of having our families running around the country looking for answers from people who didn't have them," said Beauregard, who is also a co-author on the study.

In 2003, Beauregard and other parents turned their frustration into advocacy and established the XXYY Project to support families.

"The more we talked, the more we realized our boys had things in common that were not addressed in the literature," said Beauregard, the project's director. "We had to do something."

The parents had their children take part in the study, and they flew Tartaglia to the United Kingdom so that she could include XXYY boys living there in the research as well.

Now, with more concrete answers, parents like Beauregard and children like Kyle can find some peace of mind.

"Kyle knows that people don't understand XXYY and therefore don't understand him as a person, she said. "The study helps the world know why he is like he is. It validates what he knows about himself and what we know about him. When he can't follow directions, it's not because he's stupid."

UC Davis M.I.N.D institute to start screening for fragile X

Tue, 19 Aug 2008 23:49:03 PST

( from http://www.rxpgnews.com ) Researchers at the UC Davis M.I.N.D. Institute will launch the first widespread newborn screening for the genetic mutation that results in fragile X syndrome, the single most common inherited cause of mental retardation.

Using a test they developed, the researchers will screen as many as 30,000 infants during the next five years as part of a $2.3 million pilot study that lays the groundwork for universal newborn screening of all infants in the United States for the fragile X mutation.

"This is a very important advance for the understanding and treatment of fragile X syndrome and paves the way for early identification and early intervention for these children," said M.I.N.D. Institute medical director and study senior investigator Randi Hagerman.

The study will for the first time allow families of infants with fragile X to learn shortly after birth whether their child will have the disorder, which is associated with physical anomalies, intellectual deficits, learning disabilities and behavioral and psychiatric problems.

"Identification of the condition in early infancy will allow families to seek crucial early intervention services for their children that we hope will mitigate the disabling affects of the disorder," said lead study investigator Flora Tassone, associate research biochemist in the department of biochemistry and molecular medicine.

Though fragile X syndrome is rare — occurring in approximately 1 in 3,600 males and 1 in 4,000 females — researchers believe widespread genetic testing will reveal that a much higher incidence of the condition exists than is currently believed.

Fragile X syndrome is also the most common known genetic cause of autism, which affects 1 in 150 children in the United States. Approximately one-third of all children with fragile X syndrome have autistic-like symptoms. Of children diagnosed with an autism spectrum disorder, approximately 5 percent have fragile X syndrome.

The blood spot study will allow researchers to offer screening for fragile X to the parents of all newborns at UC Davis Medical Center in Sacramento, Calif., and to all parents of infants born at Rush University Medical Center in Chicago. Study participants in Chicago will be followed by co-principal investigator Liz Berry-Kravis.

The significance of widespread newborn screening is amplified by the fact that adults related to children with fragile X syndrome often suffer from associated but frequently misdiagnosed ailments.

One such ailment, first identified by a team led by Hagerman, is fragile X-associated tremor/ataxia syndrome or FXTAS (pronounced fax-tass), a condition affecting older males and some older females that is routinely mistaken for Parkinson's disease or other movement disorders.

Males tend to be more severely affected by fragile X than females because women have two X chromosomes. If one of a woman's chromosomes has the fragile X mutation, the second X chromosome can compensate.

Hagerman, one of the world's leading experts on fragile X and related conditions, said that, while the newborn screening study is not specifically designed to offer treatment, the diagnosis will open the door to new therapies for both infants and adult family members who are subsequently diagnosed with related conditions.

"Once we have identified the affected infants, we will propose treatment options for them and also will assess the gene's impact on other family members. In many instances, families may not even know that the gene exists," Hagerman said.

"This will allow us to also look at a larger group of extended family members and the effects that the mutation may cause," she said. "That includes emotional problems, aging disorders, FXTAS and premature ovarian failure — all of that."

In addition, genetic counseling for parents of children identified as having fragile X syndrome will be crucial, said Hagerman, because such families are at an increased risk of having additional children with the condition.

The blood spot test was developed by Tassone and Paul Hagerman, professor of biochemistry and molecular medicine. They worked on optimization of the methodology for the test for the past two years.

Fragile X syndrome and its associated disorders are the result of a mistake in the number of repeats of three nucleotides on the FMR1 gene on the X chromosome. A normal X chromosome generally has between five and 55 repeats of these nucleotides. Repeats above 200 result in fragile X syndrome. Individuals with between 55 and 200 repeats, called carriers, are susceptible to a wide range of ailments, like FXTAS and premature ovarian failure.

The test uses small drops of blood drawn from infants shortly after birth to test for the fragile X mutation. It employs a polymerase chain reaction (PCR) technique that amplifies the expression of the FMR1 gene, allowing researchers to detect the number of nucleotide repeats, from the normal number of repeats to the full fragile X mutation of 200 repeats or more.

Existing tests for fragile X have had a variety of limitations, Tassone said, including not being able to detect the full range of genetic mutation in both males and females.

"For effective newborn screening, a test has to be quick, cheap, it has to work on a very small amount of DNA and it has to detect everyone," Tassone said. "This test responds to all of those requirements."

A tall story? Genes do manipulate height

Mon, 07 Apr 2008 09:41:15 PST

( from http://www.rxpgnews.com ) New York, April 7 - Scientists now have a far clearer picture of what makes some people tall - and others not so.

Researchers who last year identified the first gene influencing height have now identified a further 20 regions of the genome which, together, can make a difference of up to six centimetres in a person's height.

The results, based on DNA samples of 30,000 people, and published Sunday with two independent studies in the journal Nature Genetics, implies that scientists now know of dozens of genetic regions that influence our height.

This provides scientists with a fascinating insight into how the body grows and develops normally and may shed light on diseases such as osteoarthritis and cancer.

Unlike a number of other body size characteristics such as obesity, caused by a mix of genetic and environmental factors, 90 percent of normal variation in human height is due to genetic factors rather than, for example, diet.

Last year, a team including Tim Frayling from the Peninsula Medical School, Exeter, and Mark McCarthy of Oxford, identified the first common gene variant to affect height, though it made a difference of only 0.5 cm.

'The number and variety of genetic regions that we have found show that height is not just caused by a few genes operating in the long bones,' said Frayling.

On the basis of DNA samples, researchers have identified 20 regions of genetic code, common variations of which influence adult height.

Half of these regions contain genes whose functions are well documented. Other genes have a role in cell-to-cell signalling, an important process in the early development of embryos in the womb.

One region in particular is also linked to osteoarthritis, the most common form of arthritis.

Large gap between Genomic Medicine and Clinical practice

Tue, 18 Mar 2008 09:31:45 PST

( from http://www.rxpgnews.com ) A large gap exists between what knowledge is available about genomic medicine and incorporating it into clinical practice for assessing the risk of and treating common chronic diseases, such as cardiovascular disease, diabetes mellitus, and cancer, according to a systematic review in the March 19 issue of JAMA, a theme issue on Genetics and Genomics.

Maren T. Scheuner, M.D., M.P.H., of the RAND Corporation, Santa Monica, Calif., presented the findings of the study at a JAMA media briefing at the National Press Club in Washington, D.C.

“The greatest public health benefit of advances in understanding the human genome will likely occur as genomic medicine expands from its focus from rare genetic disorders to inclusion of more common chronic diseases, such as coronary heart disease, stroke, diabetes mellitus, and cancer,” the authors provide as background information in the article. “With genomics discoveries relating to common chronic diseases, numerous genetic tests may emerge that hold promise for significant changes in the delivery of health care, particularly in preventive medicine and in tailoring drug treatment.”

Dr. Scheuner and colleagues analyzed the medical literature for research articles and systematic reviews published between Jan. 2000 and Feb. 2008 dealing with common chronic adult-onset conditions. The authors included 68 articles in the analysis and assessed four key areas: outcomes of genomic medicine, consumer information needs, delivery of genomic medicine, and challenges and barriers to integration of genomic medicine.

“Generally there were modest positive effects on psychological outcomes such as worry and anxiety, behavioral outcomes have shown mixed results, and clinical outcomes were less well studied,” the authors report. “The most important and consistent finding from our literature review is that the primary care workforce, which will be required to be on the front lines of the integration of genomics into the regular practice of medicine, feels woefully underprepared to do so.”

The authors note that consumers are unsure about the value of genetic testing and have concerns about privacy issues and discrimination in health insurance and employment. However, the consumers were interested in the technology to help better identify diseases for which they and their family members are at increased risk.

The analysis identified the need to better understand the outcomes of genomic medicine interventions. “More research describing clinical outcomes is needed: do patients who receive counseling and testing have better clinical outcomes in terms of mortality, decreases in incidence of disease, and better clinical responses to pharmaceuticals? And at what cost?”

Other barriers to the clinical integration of genomic medicine for common chronic diseases were identified by these authors in addition to the perceived inadequacy of the primary care workforce. “The most prominent of these include health professionals’ lack of basic knowledge about genetics and their lack of confidence in interpreting familial patterns of disease, which limits their ability to appropriately counsel their patients, order and accurately interpret genetic tests, and refer their patients for genetics consultation.”

In conclusion, the authors write: “It will be a lost opportunity if the health services research components of genomic medicine fail to keep pace with the rapid basic science advances and clinical discoveries.”

Moving an active gene from the interior of the nucleus can silence genes , preventing their transcription . scientists report .

Thu, 14 Feb 2008 04:00:00 PST

( from http://www.rxpgnews.com ) Moving an active gene from the interior of the nucleus to its periphery can inactivate that gene report scientists from the University of Chicago Medical Center .

Attachment to the inner nuclear membrane, they show, can silence genes, preventing their transcription--a novel form of gene regulation.

Several years ago, we and others described the correlation between nuclear positioning and gene activation, said study author Harinder Singh, Louis Block Professor of Molecular Genetics and Cell Biology and an Investigator in the Howard Hughes Medical Institute at the University of Chicago.

With that in mind, we wanted to take the next step, to design an experiment that could test causality. Could we move a gene from the center of the nucleus to the periphery, we asked, and then measure the consequences of such repositioning?

In mammalian nuclei, chromatin--a complex of DNA and associated proteins--is organized into structural domains through interactions with distinct nuclear compartments. In this study, the authors developed the molecular tools to take specific genes from these interior compartments, move them to the periphery and attach them to the nuclear membrane--which turned those genes off.

Not only were selected test genes that served as markers turned off after being attached to the inner nuclear membrane, but also nearby real genes the scientists quoted.

Europe's most common genetic disease is a liver disorder

Wed, 06 Feb 2008 20:55:00 PST

( from http://www.rxpgnews.com ) Much less widely known than the dangerous consequences of iron deficiencies is the fact that too much iron can also cause problems. The exact origin of the genetic iron overload disorder hereditary hemochromatosis (HH) has remained elusive. In a joint effort, researchers from the European Molecular Biology Laboratory (EMBL) and the University of Heidelberg, Germany, have now discovered that HH is a liver disease. They report in the current issue of Cell Metabolism that the disorder develops when a crucial gene is lacking in liver cells.

Iron is essential for our body, because it is a central component of red blood cells. Too little iron can lead to dangerous anemias, but also too much iron can be detrimental as it promotes the formation of toxic radicals that lead to tissue damage. Hereditary hemochromatosis is an iron overload disorder that, affecting about one in 300 people, is probably the most common genetic disorder in Europe. Scientists have identified a gene, called HFE, that when mutated causes hemochromatosis in mice and humans. But as yet it is unknown in which tissue or organ the gene is acting to prevent iron overload.

A group of researchers around Matthias Hentze at EMBL and Martina Muckenthaler and Wolfgang Stremmel at the University of Heidelberg have now found that mice that are genetically engineered to lack HFE only in liver cells show all central features of the disease.

“For a long time scientists thought of HH as a disease of the intestine, because this is where iron uptake actually takes place,” says Matthias Hentze, Associate Director of EMBL. “Our research now reveals the crucial point is actually the liver and explains why HH patients suffer from increased iron absorption.”

HFE encodes a protein that is likely involved in transmitting signals about the current iron contents of the body to liver cells. In response to these signals, the liver cells make a special iron hormone, hepcidin that is released into the blood stream and reduces iron uptake in the intestine.

“HFE influences hepcidin expression through a series of intermediate molecules, but when the HFE gene is mutated the result is that less hepcidin is produced. This in turn means iron uptake in the intestine cannot be limited as effectively and an overload develops,” says Martina Muckenthaler, professor at the University of Heidelberg.

Genetic variations may predispose some men to suicidal thoughts during treatment for depression

Thu, 07 Jun 2007 16:00:00 PST

( from http://www.rxpgnews.com ) Genetic variations may help explain why some men with depression develop suicidal thoughts and behaviors after they begin taking antidepressant medications, while most do not, according to a report in the June issue of Archives of General Psychiatry, one of the JAMA/Archives journals.

Although most patients with depression respond favorably to antidepressant medications, a very small subgroup may experience worse symptoms after beginning treatment, according to background information in the article. "Regardless of treatment specificity, nearly all antidepressant medication studies find that some patients experience suicidality [suicidal thoughts and behaviors] after treatment initiation," the authors write. "Identification of this subpopulation before treatment would have tremendous clinical utility."

Roy H. Perlis, M.D., of Massachusetts General Hospital and Harvard Medical School, Boston, and colleagues studied 1,447 individuals with depression who were part of the Sequenced Treatment Alternatives to Relieve Depression (STAR*D) study, which was conducted from July 2001 to September 2006, and who did not express suicidal thoughts at the beginning of the study. The participants were men and women ages 18 to 75 years who had been diagnosed with non-psychotic major depressive disorder. They took the antidepressant citalopram hydrobromide for up to 12 weeks, following a protocol that advised follow-up treatment visits at two, four, six, nine and 12 weeks, with an optional visit at 14 weeks if needed. The patients' DNA was analyzed for common types of mutations nearby or within the CREB1 gene, which codes for a protein previously suggested to be involved in both antidepressant effects and suicide.

Of the 1,447 patients, 123 (8.5 percent) reported suicidal thoughts or behaviors during at least one follow-up visit, including 54 (10 percent) of the 539 men. Two of five single nucleotide polymorphisms (SNPs) - variations that occur when a single building block of DNA is altered - were significantly and strongly associated with the onset of suicidality in men, but not in women.

The researchers performed additional analyses suggesting these variations are not linked to suicidal thoughts and behaviors in men before treatment. "No statistically significant association was noted between any SNP and the presence or absence of baseline suicidality," the authors write. "Likewise, no evidence of association was noted between any SNP and history of lifetime suicide attempt."

Studies that link genes to illnesses are most compelling when there is additional evidence of that gene's function, the authors note. "We recently observed an association between the same CREB1 polymorphisms and a measure of anger expression among males but not females in a sample of 94 patients with major depressive disorder; hostility and anger expression have also been associated with suicide," they write.

"If replicated, this finding would suggest that pharmacogenetic testing could facilitate the identification of the small subset of individuals at greater risk during short-term antidepressant treatment," the authors conclude.

Switching genes to overdrive improves muscular dystrophy symptoms in mice

Sun, 01 Apr 2007 11:56:50 PST

( from http://www.rxpgnews.com ) Scientists at Dana-Farber Cancer Institute have shown in a laboratory study that revving up a crucial set of muscle genes counteracts the damage caused by a form of muscular dystrophy.

Reporting in the April 1 issue of Genes and Development, the researchers demonstrated that manipulating a genetic molecular switch increased the genesâ activity in the muscles of mice with Duchenne muscular dystrophy, slowing the disease-associated muscle wasting. The authors caution that they have not yet found a way to tweak the switch, known as PGC-1alpha, in humans.

âI think that if we could elevate the levels of PGC-1alpha in the muscles of patients with Duchenne muscular dystrophy, it is likely that we could slow or reduce the course of the disease,â said Bruce Spiegelman, PhD, the Dana-Farber researcher who led the team along with Christoph Handschin, PhD, formerly of Dana-Farber and now at the University of Zurich. Other authors are from the University of Iowa College of Medicine.

Duchenne muscular dystrophy (DMD) is the most common type of muscular dystrophy in children, occurring once in about every 5,000 live births of boys, and is ultimately fatal. The average age of death is the mid-teens, and most patients die by their 30s. In the United States, about 400 to 600 boys are born each year with DMD or Becker Muscular Dystrophy, a milder form of the disease. The cause is a mutation, either inherited or occurring spontaneously, that affects a muscle protein called dystrophin.

Spiegelman, whose laboratory discovered PGC-1alpha in 1998, led the new study which was aimed at determining whether increasing levels of PGC-1alpha in the muscles of mice could increase the activity of genes that are known to behave abnormally in muscular dystrophy.

PGC-1alpha is known as a âtranscriptional coactivatorâ that functions as a switch, or perhaps more accurately, like a light dimmer that increases or decreases the activity of genes under its control. Exercising a muscle raises PGC-1alpha levels, causing the formation of more mitochondria, the chemical power plants that create energy in cells.

PGC-1alpha is also required for the normal operation of genes that control the development of neuromuscular junctions (NMJ) â sites on muscle fibers where nerves attach and signal the fibers to contract. Part of the reason that exercise builds stronger muscles is that it increases PGC-1alpha activity. Conversely, disease or lack of exercise reduces PGC-1alpha activity, causing a loss of NMJ function and weakening, or atrophying, of muscles.

Spiegelmanâs team had previously bred a strain of mice with higher-than-normal levels of PGC-1alpha in their muscles. Also available for the research was a mouse model of Duchenne muscular dystrophy, the MDX mouse. In the new experiment, the scientists bred male high-PGC-1alpha mice with female MDX mice (the muscular dystrophy gene is carried by females in mouse and in humans.) As a result, the offspring of these matings had muscular dystrophy but also had elevated PGC-1alpha. Using exercise and chemical tests, the researchers compared muscle function in the offspring with MDX mice having no additional PGC-1alpha.

Both sets of rodents were run on a treadmill for one hour, then again 24 hours later. Normal mice completed the runs easily on both days, while untreated MDX rodents were exhausted halfway through each run. The MDX mice with increased PGC-1alpha activity performed almost as well as normal mice on the first day; their performances decreased on the second day, but they still did better than the untreated MDX mice on both runs.

The exercise tests and microscopic and chemical examinations of the muscles showed that boosting PGC-1alpha caused âa clear and substantial improvement in the structure and function of skeletal muscle in this disease model,â the scientists wrote.

Gene mutation associated with X-linked mental retardation revealed

Mon, 19 Mar 2007 22:53:10 PST

( from http://www.rxpgnews.com ) Researchers have identified a novel gene mutation that causes X-linked mental retardation for which there was no previously known molecular diagnosis, according to an article to be published electronically on Tuesday, March 20, 2007 in The American Journal of Human Genetics.

Investigators F. Lucy Raymond (Cambridge Institute of Medical Research, University of Cambridge, Cambridge, UK) and Patrick S. Tarpey (Wellcome Trust Sanger Institute, Hixton, UK) describe the ZDHHC9 gene found in those with severe retardation as being mutated to the point of entirely losing function.

"ZDHHC9 is a novel gene," explains Dr. Raymond. "This gene would not have been predicted to play a role in mental retardation based on the previous genetics work. It was found only because we were systematically looking at all the genes on the X chromosome irrespective of what they do."

X-linked mental retardation is severe. Some patients require total care and may not have language ability. The condition runs in families and only affects the male offspring. So far only a few of these genes have been identified.

Working through a large, international collaboration, the researchers collected genetic samples from 250 families in which at least two boys have mental retardation to help identify novel genes that cause X-linked mental retardation. The investigators systematically analyzed the X chromosome for gene mutations.

Dr. Raymond says that the families are receiving information from the study and using it to make decisions in their lives. "We cannot currently make their children better, but knowing that we found a genetic abnormality gives them an explanation for what has happened," she explains. "We had one family that said this knowledge was the best news they had ever been given."

"We have identified the cause of problems in certain families and are able to tell whether or not women are carriers of the condition," Dr. Raymond comments. "Consequently, the families that had previously chosen to forego having children because there was no method of testing can now be tested. We have been able to test a substantial number of people to identify whether are not they are carriers, and we can offer prenatal testing to the carriers who want it."

In the broader picture, this research is not only benefiting families with X-linked mental retardation, but it is also defining the genes involved in intellectual development. "If you find genes that are abnormal, it is a reasonable assumption that the identified genes are involved in the formation of normal intellectual processing as well," concludes Dr. Raymond.

Now that a posttranslational modification enzyme has been found to be mutated in X-linked mental retardation, the researchers expect to find similar genes related to other mental retardation syndromes.

RNA Silencing Sheds Light on the RNA World

Tue, 05 Dec 2006 07:51:55 PST

( from http://www.rxpgnews.com ) RNA silencing — also known as RNA interference — is an intriguing phenomenon in which short, double-stranded RNA “triggers” can prevent the expression of specific genes. First discovered in plants, RNA interference is now recognized as a widespread, if not ubiquitous, phenomenon, and it is causing great excitement as an experimental technique for selectively blocking gene expression.

The mechanisms of RNA silencing have been intensively studied. One important step is the formation of single-stranded RNA pieces (called siRNAs) from the double-stranded triggers. In “lower” organisms—including plants, protozoa, fungi, and nematode worms—it also involves an enzyme—called RNA-dependent RNA polymerase—that can generate a strand of RNA using existing RNA as a template. This means that it can create double-stranded RNA from single-stranded pieces of RNA. By doing this, it generates more triggers and so amplifies the effect of RNA silencing. Paula Salgado and her colleagues have studied the structure of one such polymerase, called QDE-1, and found that it provides clues to the earliest stages of evolution.

When a gene is transcribed and translated to generate a protein, the process begins with a DNA-dependent RNA polymerase. Like QDE-1, DNA-dependent RNA polymerases generate strands of RNA—the difference is that they use a DNA template to do it. The RNA they generate is called messenger RNA and is in turn used as the template for building a protein out of amino acids. The structures of DNA-dependent RNA polymerases have been described previously, so that the authors of this study could compare them with their new structure of QDE-1.

Schematic showing the fold of the QDE-1 RNA interference polymerase. The dimeric molecule is shown with the polypeptide chains colored from blue at the N termini to red at the C termini.

What they found was a remarkable similarity. Both DNA-dependent RNA polymerases and QDE-1 have an active catalytic site—the working core of the enzyme—that is formed by two distinctive structural domains called double-psi β-barrels. This strong structural resemblance between QDE-1 and the DNA-dependent RNA polymerases points towards an evolutionary link between the two types of RNA polymerase.

An influential theory on the origin of life proposes that RNA molecules were the first self-replicating molecules, forming a kind of precellular life in an “RNA world.” Initially, RNA molecules would have had to act as enzymes as well as genetic information so that they could replicate, but it is likely that an RNA-dependent RNA polymerase would have been one of the earliest protein-based enzymes to evolve.

The similarity between QDE-1 and the DNA-dependent RNA polymerases suggests that both evolved from one ancestor, because otherwise, the resemblance between their active sites would be highly unlikely. This common ancestor might have been a primordial RNA polymerase in an RNA world. The authors suggest that this ancestor would have had just one double-psi β-barrel, and that gene duplication led to an enzyme with two barrels. From this early polymerase evolved both QDE-1–like RNA-dependent RNA polymerases and a diverse array of specialized DNA-dependent RNA polymerases. The diversification of DNA-dependent RNA polymerases would have been facilitated by the splitting of the two double-psi β-barrels onto separate subunits, rather than being borne on the same subunit as in QDE-1, so that different subunits could combine to create specialized polymerases.

These findings provide a link between RNA silencing and the earliest mechanisms of RNA transcription—perhaps shedding light on both the origins of RNA replication (and therefore life) and the evolution of RNA silencing. They also provide insights into the mechanism of action of QDE-1 that might apply across the board to RNA-dependent RNA polymerases, and that will be built upon by further work.

Link between Huntington's and abnormal cholesterol levels in brain discovered

Sun, 03 Dec 2006 15:10:30 PST

( from http://www.rxpgnews.com ) Mayo Clinic researchers have discovered a protein interaction that may explain how the deadly Huntington's disease affects the brain. The findings, published in and featured on the cover of the current issue of Human Molecular Genetics, show how the mutated Huntington's protein interacts with another protein to cause dramatic accumulation of cholesterol in the brain.

"Cholesterol is essential for promoting the connection network among brain cells and in maintaining their membrane integrity. Both the level of cholesterol and its delivery to the proper locations in the cell are essential for the survival of neurons," explains Mayo Clinic molecular biologist Cynthia McMurrary, Ph.D.

"Our discovery that the mutant Huntington's disease protein derails the cholesterol delivery system and causes cholesterol accumulation in neurons provides us with key results and solid clues to the mechanism of this disease," says Dr. McMurray. "Fully understanding the mechanism of toxicity is the key to developing treatments."

Huntington's disease -- sometimes called Huntington's chorea or St. Vitus' dance -- is a progressive, degenerative condition that causes nerve cells in the brain to waste away. Symptoms include uncontrolled movements, emotional disturbances and mental deterioration.

The mutant protein of Huntington's attacks the railroad system of brain cells and impairs transport of essential materials required for neurons to function. When this transportation system goes awry in the parts of the brain affected in Huntington's disease, motor skills, cognitive skills and even speech can be affected. A person cannot move without shaking, and physical control gradually deteriorates, often with accompanying personality changes, depression and increased risk of suicide. Those who have Huntington's commonly die from complications of the disease, such as falls or infections.

Mouse control neuron (left) and neuron showing cholesterol accumulation in Huntington's disease.

Approximately 30,000 Americans have Huntington's disease. Another 150,000 carry the gene and have a 50 percent risk of passing it on to their children. The disease is easily diagnosed by a blood test, but symptoms usually don't appear until middle age.

Their findings, say the researchers, provide the first direct link between the Huntington's protein and the protein that controls capture and trafficking inside the cell. Their research suggests a possible means by which Huntington's disease functions.

Because no one knows how the disease is incurred or spreads, this new information is critical and establishes a clear path for investigations to move forward.

The Mayo researchers observed the abnormal accumulation of cholesterol in cultured neuronal cells in the laboratory and in the brains of animal models. They found that this happens only when the mutant Huntington's protein is expressed together with the molecule, caveolin-1. Caveolin-1 is the major structural protein of small vesicles called caveolae, which capture cholesterol and move it in and out of the neuronal membranes. When the researchers "knocked out" expression of caveolin, the neurons expressing mutant Huntington's protein stopped accumulating cholesterol.

Reinventing Human genome

Thu, 23 Nov 2006 16:27:05 PST

( from http://www.rxpgnews.com ) Nearly six years after the sequence of the human genome was sketched out, one might assume that researchers had worked out what all that DNA means. But a new investigation has left them wondering just how similar one person's genome is to another's.

Geneticists have generally assumed that your string of DNA 'letters' is 99.9% identical to that of your neighbour's, with differences in the odd individual letter. These differences make each person genetically unique — influencing everything from appearance and personality to susceptibility to disease.

But hold on, say the authors of a new study published in Nature1. They have identified surprisingly large chunks of the genome that can differ dramatically from one person to the next. "Everyone has a unique pattern," says one of the lead authors, Matthew Hurles at the Wellcome Trust Sanger Institute in Cambridge, UK.

The differences in question - made up of stretches of DNA that span tens to hundreds of thousands of chemical letters — are called 'copy-number variants', or CNVs. Within a given stretch of DNA, one person may carry one copy of a DNA segment, another may have two, three or more. The region might be completely absent from a third person's genome. And sometimes the segments are shuffled up in different ways.

These variable regions received short shrift for many years. When the human genome sequence was pieced together, they were largely glossed over, because researchers were focused on finding one overarching reference sequence — and because the repetitive nature of the segments makes them hard to sequence. "It was swept under the rug," says Michael Wigler who is also mapping CNVs at Cold Spring Harbor Laboratory, New York.

The new study, led by Hurles and Stephen Scherer of the Hospital for Sick Children in Toronto, Canada, and their colleagues is the most detailed attempt to find how CNVs are scattered across the whole human genome. To do this, they compared genome chunks from 270 people of European, African or Asian ancestry.
They found nearly 1,500 such regions, taking up some 12% of the human genome. That doesn't mean that your DNA is 12% different from mine (or 88% similar), because any two people's DNA will differ at only a handful of these spots.

According to the team's back-of-the-envelope calculations, one person's DNA is probably 99.5% similar to their neighbour's. Or a bit less. "I've tried to do the calculation and it's very complicated," says Hurles. "It all depends on how you do the accounting."

The answer is also unclear because researchers think that there are many more variable blocks of sequence that are 10,000 or 1,000 letters long and were excluded from the current study. Because of limits with their methods, the new map mainly identified variable chunks larger than 50,000 letters long.

Many of these CNVs are thought to be important in our biology. The team found that 10% of human genes are spanned by these regions, meaning that they might be doubled, deleted or otherwise jumbled in a way that could help to determine whether and when we develop diseases.

CNVs have already been linked with susceptibility to Alzheimer's disease, kidney disease and HIV, among others, and the new map will help researchers to make connections to other conditions. "There's a general expectation that these things are quite influential," Wigler says.

The new map adds to a whole library of genetic cartography that already points out other landmarks in the human genome. A lot of attention has focused on mapping the places where single letters vary between individuals (single-nucleotide polymorphisms, or SNPs). Other researchers are identifying hard-to-spot regions where a segment can be flipped around so it runs backwards.

But there is plenty more for geneticists to navigate and undoubtedly more maps to come. Some will reveal the smaller regions of variation excluded from Hurle's map. Other projects are attempting to mark every single sequence that does something biologically useful, such as making proteins or packaging up DNA into chromosomes.

The precise degree to which each person's DNA differs from another may not become clear until geneticists devise a way to read through the entire genome of many different people and compare them all in detail, something that is far too expensive and time consuming today but may become possible with the advent of faster, cheaper sequencing machines.

Scherer and his team have already lined up the only two complete human genome sequences produced by the publicly funded Human Genome Project and the private company Celera. They identified both single-letter changes and small and large regions of variation and report their results in Nature Genetics

Wnt reactivates dormant limb regeneration program

Sat, 18 Nov 2006 13:52:57 PST

( from http://www.rxpgnews.com ) Chop off a salamander's leg and a brand new one will sprout in no time. But most animals have lost the ability to replace missing limbs. Now, a research team at the Salk Institute for Biological Studies has been able to regenerate a wing in a chick embryo – a species not known to be able to regrow limbs - suggesting that the potential for such regeneration exists innately in all vertebrates, including humans.

Their study, published in the advance online edition of Genes and Development on Nov. 17, demonstrates that vertebrate regeneration is under the control of the powerful Wnt signaling system: Activating it overcomes the mysterious barrier to regeneration in animals like chicks that can't normally replace missing limbs while inactivating it in animals known to be able to regenerate their limbs (frogs, zebrafish, and salamanders) shuts down their ability to replace missing legs and tails.

"In this simple experiment, we removed part of the chick embryo's wing, activated Wnt signaling, and got the whole limb back - a beautiful and perfect wing," said the lead author, Juan Carlos Izpisúa Belmonte, Ph.D., a professor in the Gene Expression Laboratory. "By changing the expression of a few genes, you can change the ability of a vertebrate to regenerate their limbs, rebuilding blood vessels, bone, muscles, and skin - everything that is needed."

This new discovery "opens up an entirely new area of research," Belmonte says. "Even though certain animals have lost their ability to regenerate limbs during evolution, conserved genetic machinery may still be present, and can be put to work again," he said. Previously, scientists believed that once stem cells turned into muscles, bone or any other type of cells, that was their fate for life – and if those cells were injured, they didn't regenerate, but grew scar tissue.

Manipulating Wnt signaling in humans is, of course, not possible at this point, Belmonte says, but hopes that these findings may eventually offer insights into current research examining the ability of stem cells to build new human body tissues and parts. For example, he said Wnt signaling may push mature cells go back in time and "dedifferentiate" into stem-like cells, in order to be able to then differentiate once more, producing all of the different tissues needed to build a limb.

"This is the reverse of how we currently are thinking of using stem cells therapeutically, so understanding this process could be very illuminating," he says. "It could be that we could use the Wnt signaling pathway to dedifferentiate cells inside a body at the site of a limb injury, and have them carry out the job of building a new structure."

Members of the Wnt gene family (for "wingless," originally discovered in fruit flies) are known to play a role in cell proliferative processes, like fetal growth and cancer development, and Belmonte's lab has characterized the crucial role of Wnt signaling in limb growth. In 1995, the Salk researchers were first to demonstrate that they could induce the growth of extra limbs in embryonic chicks, and in 2001, they found that the Wnt signaling system played a critical role in triggering both normal and abnormal limb growth.

The current study was designed to see if Wnt signaling also was involved in the regeneration of limbs and included three groups of vertebrates: zebrafish and salamanders, which can regenerate limbs throughout their lives; frogs, which can only regenerate new limbs during a limited period during their fetal development; and chicks, which cannot regenerate limbs.

To manipulate animals' regeneration ability, the Salk researchers used inhibitory and excitatory factors for Wnt signaling, which they delivered directly to the remaining bulge after they cut a limb from the experimental embryos.

In adult zebrafish and salamanders, they found that blocking Wnt signaling with the inhibitory factors, prevented normal regeneration. And, conversely, when they treated mutant adult zebrafish that cannot regenerate with the excitatory agent, the ability to regenerate their fins was rescued, Belmonte says.

Using an inhibitory agent on frogs before the regeneration-enabled developmental window closed resulted in loss of that ability, but treating them with the excitatory agent after they had lost their regenerative capacity induced new limb growth.

They then performed the key experiment, successfully testing the ability of an excitatory factor to produce limb regeneration in chick embryos. "The signal restarted the process, and genes that were involved in the initial development of the limb were turned back on," Belmonte says. "It is simply amazing."

The procedure was tricky, however. Belmonte noted that if Wnt signaling is activated for too long of a period in these animals, cancer results. "This has to be done in a controlled way, with just a few cells for a specific amount of time," he says. "The fact is that this pathway is involved in cell proliferation, whether it is to generate or regenerate limbs, control stem cells, or produce cancer."

New research into csd genes could help designing strategies for breeding honey bees

Fri, 27 Oct 2006 16:46:00 PST

( from http://www.rxpgnews.com ) Three years ago, scientists pinpointed a gene called csd that determines gender in honey bees, and now a research team led by University of Michigan evolutionary biologist Jianzhi "George" Zhang has unraveled details of how the gene evolved. The new insights could prove useful in designing strategies for breeding honey bees, which are major pollinators of economically important cropsand notoriously tricky to breed.

The findings of Zhang and collaborators appear in a special issue of Genome Research devoted to the biology of the honey bee. The issue will be published online and in print Oct. 26, coinciding with the publication of the honey bee genome sequence in the journal Nature.

Scientists have long known that in beesas well as wasps, ants, ticks, mites and some 20 percent of all animalsunfertilized eggs develop into males, while females typically result from fertilized eggs. But that's not the whole story, and the discovery in 2003 of csd (the complementary sex determination gene) helped fill in the blanks. The gene has many versions, or alleles. Males inherit a single copy of the gene; bees that inherit two copies, each a different version, become female. Bees that have the misfortune of inheriting two identical copies of csd develop into sterile males but are quickly eaten at the larval stage by female worker bees.

The system works fine in nature, where it prevents the colony from wasting precious energy and resources on abnormal males incapable of carrying out the all-important role of mating. But in bees raised for honey or for pollinating crops, the sex-determination system can cause problems. Beekeepers inbreed bees to select desirable traits, but inbreeding raises the odds of producing fertilized eggs with two copies of the same csd allele. If too many sterile males result, the colony may die out.

"If we know more details about how many alleles there are and what their frequencies are, bee breeders can design better strategies to avoid producing sterile males," Zhang said. "Our work aids in this effort by providing a direct tool to examine alleles from different populations."

In the research, Zhang and coworkers from U-M, Michigan State University and the University of Kansas sequenced csd genes from individuals in three closely related species of honey bee: the familiar backyard denizen Apis mellifera and the Asian honey bees Apis dorsata and Apis cerana. The group also sequenced six so-called neutral regions of the genome which, unlike genes, do not carry codes telling cells how to make proteins. Then, the researchers constructed gene genealogiesfamily trees for both the csd gene and the neutral regions.

Their results showed that csd is about seven times more variable than neutral regions of the honey bee genome. In addition, many csd variants are shared among the three species, evidence that the many different alleles have been preserved in these lineages for a very long time.

Such a pattern supports the idea that an evolutionary mechanism known as balancing selection has been at work. Evolution works through the process of natural selection, in which genetic mutations that offer some advantage are favored, and those that have harmful effects are weeded out. Typically, this results in one version of a gene becoming very common and other versions becoming rare or disappearing altogether. When balancing selection operates, however, natural selection favors a diverse mix of alleles, as seen with csd in honey bees.

The research also showed just how long the csd alleles have been around.

"We estimated the age of the alleles at about 14 million years," said Zhang. "We don't know for sure when the species formed, but it's thought to be about six to eight million years ago, so the alleles are even older than the species."

Williams Syndrome, the brain and music

Thu, 05 Oct 2006 00:58:00 PST

( from http://www.rxpgnews.com ) Children with Williams syndrome, a rare genetic disorder, just love music and will spend hours listening to or making music. Despite averaging an IQ score of 60, many possess a great memory for songs, an uncanny sense of rhythm, and the kind of auditory acuity, than can discern differences between different vacuum cleaner brands.

A study by a multi-institutional collaboration of scientists, published in a forthcoming issue of NeuroImage, identified structural abnormalities in a certain brain area of people afflicted with Williams syndrome. This might explain their heightened interest in music and, in some cases, savant-like musical skill.

Professor Ursula Bellugi, director of the Laboratory for Cognitive Neuroscience at the Salk Institute for Biological Studies the central hub of this unique scientific alliance explains, "Understanding the connections between missing genes, the resulting changes in brain structure and function, and ultimately behavior may help us to reveal how the brain works."

The current study is just the latest chapter in a story that's been unfolding for quite some time gaining increasing momentum in recent years. It all started when Bellugi reached out across disciplines and assembled a team of experts under the umbrella of a Program Project from the National Institutes of Child Health and Human Development to help her trace the influence of individual genes on the development and functioning of the brain.

Along with co-author Albert Galaburda, a professor at the Harvard Medical School's Department of Neurology, Professor Allan L. Reiss, Director of the Center for Interdisciplinary Brain Sciences Research at Stanford University and senior author of the current study, focuses on the overall morphology of the brain, zooming in on the cellular architecture of the brain. Molecular geneticist Julie R. Korenberg, a professor in the Department of Pediatrics at UCLA, digs even deeper and studies the genes missing in people with Williams syndrome, whereas Debra Mills, an associate professor in the Department of Psychology at Emory University, concentrates on the neurophysiology, the electrical activity of behaving neural networks. Says Bellugi, who studies the cognitive aspects of the disorder: "Things are really starting to come together now."

Identified more than 40 years ago, Williams syndrome arises from a faulty recombination event during the development of sperm or egg cells. As a result, almost invariably the same set of about 20 genes is deleted from one copy of chromosome seven, catapulting the carrier of the deletion into a world where people make much more sense than objects do.

"Williams syndrome is a perfect example where a genetic predisposition interacts with the environment to sculpt the brain in unique ways," says Reiss. "It provides a unique window of understanding on how the brain develops under typical and atypical conditions," he adds.

People with Williams syndrome are irresistibly drawn to strangers, remember names and faces with ease, show strong empathy and have fluent and exceptionally expressive language. Yet, they are confounded by the visual world around them: While they can't scribble more than a few rudimentary lines to illustrate an elephant, they can verbally describe one in almost poetic detail.

"The discrepancy between their engaging social use of language and their poor visual-spatial skills is startling," says Bellugi. "I am confident that once all the evidence is in, we will have identified genes and pathways in the Williams syndrome deletion that underlie these drastic differences in modalities," she adds.

Despite whole brain volumes that are about 15 percent smaller than normal, the temporal lobe, which lies above the ear canal and, among other things, is involved in processing sounds and interpreting music and language, is of approximately normal volume in people with Williams syndrome. In their study, the researchers tried to answer the question of whether an atypical development of the planum temporale, which is part of the temporal lobe and thought to be involved many auditory tasks, including perfect pitch, may underlie the unusual musical and language skills.

First author Mark Eckert, formerly at Stanford and now an assistant professor at the Medical University of South Carolina, and his colleagues used data from brain scans of 42 individuals with Williams syndrome and 40 control participants to compare the surface folds of the planum temporale. In most people, the structure, a slender inch-long piece of tissue, is larger on the left side of the brain than the right.

In people with Williams syndrome, however, both sides tended toward symmetry. "There are different possible explanations: Either the left side didn't grow enough or the right side grew larger than usual," says Galaburda. The folding pattern, in particular one groove called the Sylvian fissure, pointed to an increase size of the right planum temporale.

But size alone might not explain the unusual auditory strengths of people with Williams syndrome. A more general explanation includes variations in the connectivity of certain brain regions that might contribute to the specific strengths and weaknesses in Williams syndrome."

In recent studies, Galaburda had found that cells in the primary visual cortex of carriers of the Williams deletion are smaller and more densely packed -- allowing for fewer connections between cells. Neurons in the primary auditory cortex, on the other hand, were larger and loosely packed, denoting increased "connectedness."

"These differences in cell size and density may underlie the strengths in auditory phonology, language and possibly music, and the difficulties in visual spatial construction for primary visual areas," says Bellugi, adding, "This is really just part of the overall effect of the genes' deletion on brain development."

"Relatively subtle developmental defects can have a significant impact on neurological function," says Dennis O'Leary, a Salk neurobiologist who studies the development of the visual system. "This work opens the door to explaining how genes works through the brain and make us who we are," he adds.

Genetic mutation identified as cause of cranio-lenticulo-sutural dysplasia

Fri, 29 Sep 2006 16:08:00 PST

( from http://www.rxpgnews.com ) A research team led by a UC Davis Childrens Hospital scientist has identified a genetic mutation as the cause of a congenital craniofacial birth defect called cranio-lenticulo-sutural dysplasia. The mutation closes off a pathway that is vital to the transport of cellular proteins and, in doing so, significantly alters normal growth patterns of skeletal and connective tissue. The research finding, which appears in the October issue of Nature Genetics (http://press.nature.com), is independently confirmed in a Vanderbilt University Medical Center study published in the same issue.

We know defects in this cellular pathway cause genetic diseases that lead to bleeding problems or skeletal defects in humans, said Simeon Boyadjiev Boyd, chief of the Section of Genetics with UC Davis Childrens Hospital and lead author of the UC Davis study. But this is the first time these defects have been linked to craniofacial abnormalities. Although cranio-lenticulo-sutural dysplasia is rare, this new finding opens up the intriguing possibility that more skeletal and connective tissue disorders are also caused by mutations in the pathway.

Congenital birth defects of the face and skull are relatively common and are estimated to affect one in every 500 to 1,000 babies born in the United States. Boyd and colleagues first identified cranio-lenticulo-sutural dysplasia in a consanguinous Saudi Arabian family in 2003. Affected children inherit the defective gene from both parents and can have skull bones that take an abnormally long time sometimes well into teenage years to completely close. As a result, the childrens heads are very wide at the top and almost pointed in the chin region. They can also have scoliosis, eye cataracts, unusually wide set eyes and developmental delays.

After identifying the disorder, Boyd, who was then at the Johns Hopkins University School of Medicine, set about finding the cause. Using genetic linkage analysis, he and his research team were able to map the defect to a particular region of human chromosome 14 and find the exact mutation in gene SEC23A. They learned that the mutation inactivated the SEC23A protein, an integral component of a critical intracellular pathway called the COPII-mediated secretory pathway that moves proteins from one part of a cell to another. To create healthy tissue and bones, certain proteins must move through this secretory pathway. For individuals with cranio-lenticulo-sutural dysplasia, this transition is impossible. As a result, the proteins accumulate and enlarge one part of the cell called the endoplasmic reticulum, which was confirmed for this study using electron microscopy and immunofluorescence.

In the Vanderbilt study, Lang, et. al., describes a similar mutation in the SEC23A gene that causes defects in zebrafish, including a malformed craniofacial skeleton, kinked pectoral fins and a short body length. Other studies have found that SEC23A mutations cause cellular defects in lower organisms. Boyd predicts that other human diseases resulting from mutations in the transport system are likely to be discovered.

People with these defects live well into adulthood but, because of the redundancy of systems in the body, they may have a series of medical problems involving multiple organs, the face, the brain and the skeletal system. They bounce from doctor to doctor, each of whom treats a particular concern such as cataracts, skeletal defect or mental retardation, but nobody knows why those patients have breakage of multiple systems, he said. Our goals now are to identify these disorders using a reverse genetic approach and knock-out gene studies and to explore if the specific cellular changes can be used to screen for other disorders in the secretory pathway.

Boyds research may also have implications in the areas of aging, neurodegeneration and endocrinology, since cell function decline and damage of the endoplasmic reticulum occur with age and could play a role in Alzheimers and Parkinsons diseases.

We also expect that the disorders in this particular part of the secretory pathway will manifest with endocrine problems, such as diabetes. Most hormones, including insulin, are secretory proteins that must be processed in the endoplasmic reticulum and exported elsewhere, he said.

Boyds research team included Lelio Orci from the University of Geneva, Switzerland, a recognized leader on cell morphology; Randy Schekman and Chris Fromme from the University of California, Berkeley, who used biochemical characterization of the mutant cells to precisely pinpoint the mechanism leading to cranio-lenticulo-sutural dysplasia; and Samuel Chong from University of Singapore, who inactivated the corresponding gene in zebrafish and was able to produce very similar skeletal defects.

This project is especially rewarding as it demonstrates how collaboration between scientists with similar interests but different expertise can rapidly advance the field, said Boyd.

Chance Fluctuations in mRNA Output in Mammalian Cells

Wed, 13 Sep 2006 03:44:00 PST

( from http://www.rxpgnews.com ) In the drama of cell biology, both genetics and environment write the script, and chance throws in the twists of plot. In general, most cells live relatively predictable lives: divide, differentiate, and die. Yet chance leaves its imprint even in ordinary cells. For instance, bacterial or yeast cells in culture are known to produce widely different amounts of certain proteins, even when they are genetically identical. Scientists attribute such cell-to-cell variations to chance fluctuations in the cellsâ ability to make these proteins. They also speculate that such fluctuations may benefit the cells in their struggle to adapt and survive.

But opinions vary as to which step in protein production is subject to random fluctuations. Proteins result from the translation of messenger RNAs (mRNAs), which come from the transcription of genes. Fluctuations in protein output could reflect a whimsy intrinsic to the expression of the coding gene, or random swings in global, or extrinsic, regulators of the geneâs expression. In yeast, extrinsic causes, such as variations in cellular volume, seem to predominate. But in a new study, Arjun Raj and his colleagues show that in mammalian cells, intrinsic causes, specifically the genesâ ability to transition randomly between active and inactive transcriptional states, can be crucial.

Individual reporter mRNA molecules are shown in a field of clonal CHO cells. Though genetically identical, the cells display extreme variations in their expression levels.

The researchers developed a sensitive technology to detect mRNAs in single cells and examined the output of a genetically engineered reporter gene they called M1, which they had introduced into Chinese hamster ovary (CHO) cells. After staining cultures of identical cells with a fluorescent probe specific to M1, they counted fluorescent dots corresponding to single M1 mRNA molecules in individual cells. At any given time, a few cells displayed a large, bright cluster of these spots, indicative of recently transcribed mRNAs still densely packed around the M1 gene. These cells also had the largest number of M1 mRNAs (over 150). In these cells, the M1 gene was therefore actively churning out mRNAs. But the majority of cells displayed fewer than 50 M1 mRNAs that were dispersed over their entire volume. In these cells, the M1 gene had become silent. These results showed that the M1 gene is expressed via infrequent bursts of transcriptional activity in the CHO cells.

If these bursts reflected the uneven distribution of transcription factors among CHO cells, then multiple reporter genes in a single cell should burst at the same time. The researchers generated cell lines that contained two versions of the reporter gene (M1 and M2) that were controlled by the same transcriptional activators but whose mRNAs could be distinguished with probes of different colors. In some lines, the two genes had landed in separate genomic locations. The researchers found that they burst independently of each other. They concluded that bursts are not induced by extrinsic factors such as M1- or M2-specific transcriptional activators. Rather, they reflect the intrinsically random ability of both the M1 and M2 genes to switch between an inactive and an active state.

Nevertheless, when the two versions of the reporter gene landed next to each other, they did burst in synchrony, as if switching spread locally among contiguous genes. This observation suggests that switching follows the waves of condensation and decondensation that randomly breathe through the coils of genomic DNA (chromatin). Indeed, genes are mostly silent when packed into dense chromatin. But when they decondense, the transcription machinery can latch on to their DNA and begin making RNA. Raj and his colleagues propose that genes switch to an active state as a consequence of randomly initiated decondensation, and that transcription factors merely stabilize their active state. Consistent with this proposal, they find that reducing the availability of a transcriptional factor, or increasing M1âs affinity for this factor, does not significantly increase the frequency of bursts. Only the size of the burstsâthat is, the amount of RNA produced in each burstâincreases.

Random bursting is not restricted to artificial genes such as M1. The researchers document the same behavior in the gene encoding RNA polymerase, the lead actor of transcription. That cells would tolerate chance fluctuations in an mRNA so fundamental to their survival comes as a surprise. The researchers show that in the case of the M1 gene, protein stability buffers the consequences of erratic mRNA production. This is because stable proteins are only âtopped upâ by the occasional bursts of transcription, whereas unstable proteins follow the variations in mRNA levels more closely. This finding indicates that protein stability may be a critical factor in the cellâs ability to tolerate variations in transcription. Whether chance fluctuations in RNA production sometimes produce beneficial twists of fate for mammalian cells remains to be shown.

Transposon Silencing Keeps Jumping Genes in Their Place

Wed, 13 Sep 2006 03:40:00 PST

( from http://www.rxpgnews.com ) Nearly a century ago, two geneticists described ârogueâ pea plants with an unorthodox pattern of inheritance. William Bateson and Caroline Pellew found that crossing inferior rogues with normal plants always produced rogue offspring, suggesting that the rogue appearance was a dominant trait. The real surprise came when rogue progeny were crossed back to normal plants. Following the principles of Mendelian inheritance, these crosses should have produced a mix of normal and rogue plants, but they produced only rogue plants. The phenomenon, later dubbed âparamutation,â allowed the rogues to break the rules by acting âepigeneticallyââinducing heritable changes in gene expression without DNA mutations. In one-sided interactions between gene pairs, or alleles, only âparamutagenicâ alleles can attenuate, and eventually silence, the expression of âparamutableâ alleles.

Epigenetic silencing involves chemical modifications to DNA and the histone proteins that remodel the chromatin surrounding DNA, rendering genes inaccessible to transcription-related proteins. Epigenetic silencing also targets âtransposons,â genetic elements that can jump around the genome. Both paramutagenic alleles and transposons contain tandem or inverted repetitive DNA sequences. Recent work in a variety of species suggests that such repeats can induce heritable silencing when they trigger the production of double-stranded RNAs, which are then processed into small interfering RNAs that can inactivate genes through DNA methylation and other mechanisms.

Heavy spotting on corn kernels reveals the activity of the Mutator system. (Photo: Damon Lisch)

A new study by Margaret Roth Woodhouse, Michael Freeling, and Damon Lisch sheds light on these somewhat mystifying processes by identifying a gene that keeps both transposons and paramutated color genes silenced in maize, confirming results recently published in Nature. The gene, Mediator of paramutation1 (Mop1), encodes an RNA-processing enzyme called RNA-dependent RNA polymerase 2 (RDR2) that is required to make the small RNAs needed to maintain silencing of a transposon (called MuDR). While Mop1 maintains MuDR silencing, the authors show that a second gene is required to establish heritability of silencing.

MuDR can be heritably silenced by a paramutagenic gene called Mu killer (Muk)âan inverted repeat variant of the MuDR transposon. MuDR encodes two genesâmudrA causes excision of the element and, with mudrA, mudrB helps reinsert it. Muk produces hairpin double-stranded RNAs that trigger rapid processing of full-length mudrA into small RNAs, leading to the destruction of mudrA transcripts and the methylation and silencing of MuDR. Muk-induced MuDR silencing begins with mudrA, then spreads to the adjacent mudrB gene.

Mutations in Mop1 interfere with both Mutator transposon methylation and paramutation of several maize color genes. Woodhouse et al. found that mop1 is an evolutionary cousin of the RDR2 found in Arabidopsis, where it effects transposon silencing. Maize plants carrying two mutant copies of mop1 failed to produce small RNAs corresponding to mudrA and mudrBâconfirming Mop1âs role as an RNA-processing enzyme.

To test the mop1 mutantâs effects on Muk-induced MuDR silencing, the researchers bred plants that carried both Muk and MuDR in the presence or absence of a functional copy of the Mop1 gene. Mutant plants showed clear evidence of MuDR silencing, suggesting that Mop1 is not required to initiate silencing. These plants also continued to produce small RNAs specifically associated with the initiation of silencing. Thus, although Mop1 is required to make the small RNAs associated with the maintenance of mudrA and mudrB silencing, it is not required to make the small RNAs associated with the initiation of Muk-induced MuDR silencing. Further, the progeny of these mutant plants carried only inactive MuDR elements, indicating that Mop1 is not required for heritable silencing of MuDRâthough there appear to be other factors that are required for this process. Offspring of plants that had both MuDR and Muk but lacked functional nucleosome assembly protein 1 (NAP1, a chromatin-building protein) gave rise to heavily spotted kernelsâthe sign that heritable MuDR silencing had been disrupted. Yet the loss of NAP1 did not block the initiation of silencing, because these plants had methylated Mutator elements whether or not they expressed NAP1, which suggests that mudrA activity had been lost in both cases. Thus, while losing NAP1 didnât prevent Muk from initiating MuDR silencing, it did prevent Muk from establishing a stable, heritably silenced state.

Altogether, these results show that distinct factors initiate, establish, and maintain MuDR silencing. Muk initiates silencing by targeting mudrA with its hairpin RNAs, leading to the destruction of mudrA transcripts and methylation of the transposonâs terminal inverted repeats. NAP1 is required to establish heritable silencing, likely by changing chromatin into a transcription-unfriendly state. Mop1/RDR2 then maintains silencing by using RNA processing to mediate continued DNA methylation.

Given the damage that transposons can cause by inserting themselves into essential genes, itâs not surprising that organisms have evolved enduring mechanisms to keep jumping genes in their place. This study contributes a valuable framework for identifying the factors that regulate the enigmatic epigenetic processes that defend the genome against invasive elementsâand helps explain how these changes can persist and be transmitted to the next generation.

GATA2 - predicting susceptibility to coronary artery disease

Sat, 26 Aug 2006 02:15:00 PST

( from http://www.rxpgnews.com ) Variations in a gene that acts as a switch to turn
on other genes may predispose individuals to heart disease, an international team of researchers led by Duke University Medical Center scientists has discovered. Their report was published in the Open Access journal PLoS Genetics

Further study of this master switch -- a gene called GATA2 -- and the genes it controls may uncover a regulatory network that influences whether a person inherits coronary artery disease, the most common form of heart disease in the Western world, according to
the researchers. The discovery also may lead to development of genetic tests to predict an individuals risk of developing coronary artery disease, the scientists said.

We hope that one day it will be possible to use these gene variations to predict who is susceptible to cardiovascular disease," said Jessica J. Connelly, a postdoctoral fellow at the Duke Center for Human Genetics and lead author on the study. "This finding is the
first step before we can develop such a test for use in patients.

People who know they are at higher risk may be encouraged to take early steps to modify behaviors, such as smoking or consuming foods high in saturated fats, that are known to play a role in promoting heart disease, the scientists said.

The team reports its findings in the August 2006 issue of Public Library of Science (PLoS) Genetics. The research was sponsored by the National Institutes of Health.

Coronary artery disease affects more than 13 million Americans and is one of the nation's leading causes of death. The disease occurs when the arteries supplying blood to the heart become narrowed or clogged by plaque deposits. Left untreated, the disease can completely block the blood flow to the heart, leading to a heart attack.

Coronary artery disease is what scientists call a complex genetic
disease that is, it results from the accumulation of a number of
small genetic changes that influence an individuals ability to cope
with environmental and biological effects. While risk factors such as
smoking, high blood pressure and high cholesterol are known to
contribute to coronary artery disease, little is known about genes
that render an individual susceptible to developing the disease,
according to the Duke researchers.

It is extremely difficult to pin down genes associated with a
complex disease such as heart disease, said Simon G. Gregory, Ph.D.,
assistant professor of medicine at the Duke Center for Human Genetics
and senior investigator on the study.

Two pieces of evidence pointed the researchers' attention to GATA2
as a likely candidate, Gregory said.

In a previous study, the researchers had scanned the entire genome
-- the body's genetic blueprint -- of a group of families with at
least two siblings with early onset coronary artery disease, looking
for regions of linkage where DNA variations appeared to be
inherited along with the disease. They found just such a region: a
small section of the long arm of chromosome 3 where GATA2 is located.
Chromosome 3 is one of the 23 pairs of chromosomes that comprise the
human genome.

In a separate study led by David Seo, M.D., assistant professor of
medicine, another group of Duke researchers found that GATA2 was
turned on, or expressed, differently in diseased areas of aorta, the
primary artery supplying the heart, in people with damaged hearts.
This finding suggested that the gene could be involved in
susceptibility to coronary artery disease, the researcher said.

In the current study, the researchers focused on specific gene
variants, called single nucleotide polymorphisms (SNPs), which occur
when a single nucleotide building block in the long strand of DNA is
altered. The researchers sought SNPs that occurred more or less often
in individuals with coronary artery disease than in individuals
without it, as such a link would indicate that these gene variants
were associated with the disease.

The researchers obtained DNA from 3,000 individuals from 1,000
families affected by coronary artery disease, through a collaborative
study, called GENECARD, led by William E. Kraus, M.D., associate
professor of medicine, under way at the Duke Center for Human
Genetics. Using these DNA samples, the researchers scanned the GATA2
gene for SNPs that differed in sequence between individuals with and
without coronary heart disease.

We identified five SNPs that were significantly associated with
early onset coronary artery disease, Gregory said.

The researchers then looked for the same SNPs in a separate group of
600 patients with early onset coronary artery disease who had
volunteered to be studied while being examined at the cardiac
catheterization laboratories at Duke University Hospital. The team
identified significant association of two of the same SNPs in this
independent group of patients. This finding, according to the
researchers, validated the suspected link between GATA2 and coronary
artery disease.

A huge strength of our study is that we used two separate
populations, finding the association between GATA2 and coronary
artery disease in one population and then validating the finding in
another, Connelly said.

GATA2 is a transcription factor, a master switch that controls when
and where other genes are expressed. According the researchers, the
SNPs identified in this study may change the ability of this
transcription factor to influence the activity of many other genes,
demonstrating how small genetic changes can influence multiple
genetic outcomes.

The researchers said they now can use molecular techniques to look
at where GATA2 acts within the genome to see what other genes also
contribute to cause cardiovascular disease.

As science progresses in this field, we are compiling a portfolio
of genes that contribute to cardiovascular disease, Gregory said.

The hope, according to the Duke team, is that this discovery will be
followed by others that eventually will enable scientists to identify
people who are predisposed to developing coronary artery disease long
before they develop any symptoms of the disease.

What we have found is that changes in GATA2 affect susceptibility
to developing coronary artery disease, Connelly said. Eventually,
we hope to create a diagnostic test containing all the genes affected
in cardiovascular disease and use it to identify which SNPs are
present in an individual. This approach will enable us to generate a
profile of risk for developing cardiovascular disease.

The profile would not say conclusively whether or not a person would
develop coronary heart disease, but could tell individuals whether
they are more or less at risk for developing the disease, Gregory
said.

Exploring genetics of congenital malformations

Sat, 19 Aug 2006 21:42:00 PST

( from http://www.rxpgnews.com ) New research published in the August issue of the Journal of Cell Biology explains for the first time why congenital heart defects so often occur with limb deformities.

In their research into the molecular mechanisms that control embryonic limb and heart development, Northwestern University researcher Hans-Georg Simon and his laboratory group recently identified a new protein, LMP4, which binds and regulates activity of the Tbx4 and Tbx5 transcription factors. Tbx5 and Tbx4 proteins play a key role in limb and heart formation in virtually all vertebrates, from fish to birds to mice to humans.

Mutations in the respective Tbx5 and Tbx4 genes can cause severe birth defects characterized by upper limb and heart defects (Holt-Oram syndrome) or patella, hip and foot malformations (small patella syndrome), respectively, Simon explained.

"Despite the importance in embryogenesis and disease, the mechanisms by which the transcription factors encoded by these genes exert their functions are not well understood," Simon said.

In studies using chicken and zebrafish model systems, Simon and his lab members are trying to gain a complete picture of how the Tbx and LMP4 proteins interact in order to control the growth and particular shaping of the limbs and heart.

LMP4 apparently regulates Tbx protein activity in the cell by keeping the Tbx transcription factors bound to the actin cytoskeleton or releasing them to the nucleus. Actin is a contractile protein of muscle and is a major component of the cytoskeleton the "scaffolding" of the cell.

As the researchers wrote in their paper, this is the first demonstration of a Tbx transcription factor to be localized outside the cell nucleus by a specific protein. In addition, they demonstrate that removal of Tbx5 from the nucleus represses the transcription factor's ability to activate target genes in the limbs and heart.

"We are just beginning to understand the multitude of different cellular process that the Tbx proteins are involved in," concluded Simon. "The next step will be to identify the signals that regulate the dynamic interaction between LMP4 and Tbx5."

Genome insertions and deletions (INDELs) provide expanded view of human genetic differences

Fri, 11 Aug 2006 14:19:00 PST

( from http://www.rxpgnews.com ) Emory University scientists have identified and created a map of more than 400,000 insertions and deletions (INDELs) in the human genome that signal a little-explored type of genetic difference among individuals. INDELS are an alternative form of natural genetic variation that differs from the much-studied single nucleotide polymorphisms (SNPs). Both types of variation are likely to have a major impact on humans, including their health and susceptibility to disease.

The INDEL research, led by Scott Devine, PhD, assistant professor of biochemistry at Emory University School of Medicine, has been posted online and will be published in the September issue of the journal Genome Research.

The human genome sequence in our DNA contains three billion base pairs of four chemical building blocks Ð adenine, thymine, cytosine, and guanine (A, T, C, G), strung together in different combinations in long chains within 23 pairs of chromosomes. When the first human genome was being sequenced, it became apparent that additional human genomes would have to be sequenced to identify the places in the genetic code that account for human variation. Scientists now know that humans share about 97-99 percent of the genetic code, and the remaining 1-3 percent dictates individual differences. These naturally occurring differences, called polymorphisms, help explain differences in appearance, susceptibility to diseases, and responses to the environment.

SNPs are differences in single chemical bases in the genome sequence, and INDELs result from the insertion and deletion of small pieces of DNA of varying sizes and types. If the human genome is viewed as a genetic instruction book, then SNPs are analogous to single letter changes in the book, whereas INDELs are equivalent to inserting and deleting words or paragraphs.

Most polymorphism discovery projects have focused on SNPs, resulting in the International HapMap Project Ð a catalog and map of more than 10 million SNPs derived from diverse individuals throughout the globe. Dr. Devine and postdoctoral researcher Ryan Mills, PhD, focused instead on INDELs, using a computational approach to examine DNA re-sequences that originally were generated for SNP discovery projects. Thus far they have identified and mapped 415,436 unique INDELs, but they expect to expand the map to between 1 and 2 million by continuing their efforts with additional human sequences.

Dr. Devine says INDELs can be grouped into five major categories, depending on their effect on the genome: (1) insertions or deletions of single base pairs; (2) expansions by only one base pair (monomeric base pair expansions); (3) multi-base pair expansions of 2 to 15 repeats; (4) transposon insertions (insertions of mobile elements); (5) and random DNA sequence insertions or deletions. INDELs already are known to cause human diseases. For example, cystic fibrosis is frequently caused by a three-base-pair deletion in the CFTR gene, and DNA insertions called triplet repeat expansions are implicated in fragile X syndrome and Huntington's disease. Transposon insertions have been identified in hemophilia, muscular dystrophy and cancer.

"We are entering an exciting new era of predictive health where an individuals personal genetic code will provide guidance on healthcare decisions", says Dr. Devine. "Our maps of insertions and deletions will be used together with SNP maps to create one big unified map of variation that can identify specific patterns of genetic variation to help us predict the future health of an individual. The next phase of this work is to figure out which changes correspond to changes in human health and develop personalized health treatments. This could include specific drugs tailored to each individual, given their specific genetic code.

Ultimately, each person's genome could be re-sequenced in a doctor's office and his or her genetic code analyzed to make predictions about their future health. Dr. Devine believes the technology holds the promise of predicting whether a person will develop diabetes, mental disorders, cancer, heart disease and a range of other conditions.

BRIT1 gene identified as protector of DNA

Sun, 06 Aug 2006 11:17:00 PST

( from http://www.rxpgnews.com ) A single gene plays a pivotal role launching two DNA damage detection and repair pathways in the human genome, suggesting that it functions as a previously unidentified tumor suppressor gene, researchers at The University of Texas M. D. Anderson Cancer Center report in Cancer Cell.

The advance online publication also reports that the gene - called BRIT1 - is under-expressed in human ovarian, breast and prostate cancer cell lines.

Defects in BRIT1 seem to be a key pathological alteration in cancer initiation and progression, the authors note, and further understanding of its function may contribute to novel, therapeutic approaches to cancer.

"Disruption of BRIT1 function abolishes DNA damage responses and leads to genomic instability," said senior author Shiaw-Yih Lin, Ph.D., assistant professor in the Department of Molecular Therapeutics at M. D. Anderson. Genomic instability fuels the initiation, growth and spread of cancer.

A signaling network of molecular checkpoint pathways protects the human genome by detecting DNA damage, initiating repair and halting division of the damaged cell so that it does not replicate.

In a series of laboratory experiments, Lin and colleagues show that BRIT1 activates two of these checkpoint pathways. The ATM pathway springs into action in response to damage caused by ionizing radiation. The ATR pathway responds to DNA damage caused by ultraviolet radiation.

By using small interfering RNA (siRNA) to silence the BRIT1 gene, the scientists shut down both checkpoint pathways in cells exposed to either type of radiation.

Researchers then used siRNA to silence the gene in normal human mammary epithelial cells (HMEC). The result: Inactivation of the gene caused chromosomal aberrations in 21.2 to 25.6 percent of cells. Control group HMEC had no cells with chromosomal aberrations. In cells with the gene silenced that were then exposed to ionizing radiation, 80 percent of cells had chromosomal aberrations.

"We also found that BRIT1 expression is aberrant in several forms of human cancer," Lin said. The team found reduced expression of the gene in 35 of 87 cases of advanced epithelial ovarian cancer. They also found reduced expression in breast and prostate cancer tissue compared with non-cancerous cells.

Genetic analysis of breast cancer specimens revealed a truncated, dysfunctional version of the BRIT1 protein in one sample.

Loss of the DNA damage checkpoint function and the ability to proliferate indefinitely are two cellular changes required for the development of cancer. Lin and colleagues have now tied the gene to both factors. They previously identified BRIT1 as a repressor of hTERT, a protein that when reactivated immortalizes cells, allowing them to multiply indefinitely.

FDA Approves Idursulfase As First Treatment for Hunter Syndrome

Wed, 02 Aug 2006 12:30:00 PST

( from http://www.rxpgnews.com ) The Food and Drug Administration (FDA) approved Elaprase (idursulfase), the first product for the treatment of Hunter syndrome (Mucopolysaccharidosis II, or MPS II), a rare inherited disease which can lead to premature death. Elaprase is a new molecular entity, which is an active ingredient never before marketed in the United States.

Hunter Syndrome, which usually becomes apparent in children one to three years of age, is a disease in which the person's body is defective in producing the chemical iduronate-2-sulfatase, which is needed to adequately breakdown complex sugars produced in the body. Symptoms include growth delay, joint stiffness, and coarsening of facial features. In severe cases, patients experience respiratory and cardiac problems, enlargement of the liver and spleen, neurological deficits, and death.

Elaprase was designated as an orphan product by FDA. Orphan products, such as Elaprase, are generally developed to treat rare diseases or conditions that affect fewer than 200,000 people in the U.S. The Orphan Drug Act provides a seven-year period of exclusive marketing to the first sponsor who obtains marketing approval for a designated orphan product. Hunter syndrome is diagnosed in approximately one out of 65,000 to 132,000 births.

"This is the first product that brings help to a very small group of seriously ill patients who have no other treatment option," said Dr. Steven Galson, Director, Center for Drug Evaluation and Research. "This approval is a good example of how the Orphan products program can benefit the public health with urgently needed products that would otherwise not be commercially available."

Elaprase was approved after a randomized, double-blind, placebo-controlled study of 96 patients with Hunter syndrome showed that the treated participants had an improved capacity to walk. At the end of the 53week trial, patients who received Elaprase infusions experienced on average a 38-yard greater increase in the distance walked in six minutes compared to the patients on placebo.

The most serious adverse events reported during the trial were hypersensitivity reactions to Elaprase that could be life-threatening. They included respiratory distress, drop in blood pressure, and seizure. Other frequent, but less serious adverse events included fever, headache and joint pain.

Because of the potential for severe hypersensitivity reactions, appropriate medical support should be readily available when Elaprase is administered. Patients and their physicians are encouraged to participate in a voluntary Hunter Outcome Survey which has been established to monitor and evaluate the safety and effects of long-term treatment with Elaprase.

PARP1 inhibitors can protect Huntington's disease affected cells from damage

Sun, 30 Jul 2006 02:41:00 PST

( from http://www.rxpgnews.com ) An enzyme known to be critical for the repair of damaged cells and the maintenance of cellular energy may be a useful target for new strategies to treat Huntington's disease (HD) and other disorders characterized by low cellular energy levels. In the August issue of Chemistry & Biology, a research team from the MassGeneral Institute for Neurodegenerative Disease (MIND) describes their discovery of a novel inhibitor of Poly (ADP-ribose) polymerase (PARP1) and their findings that PARP1 inhibitors can protect HD-affected cells from damage in laboratory assays.

"While PARP1 is essential for the repair of damaged DNA, we also know that, if overactivated, it can cause cell death by excessive energy depletion," says Aleksey Kazantsev, PhD, director of the MIND High Throughput Drug Screening Laboratory, who led the current study. "It has recently been shown that neurons from patients with Huntington's appear to be energy-deficient, so we hypothesized that modest stresses that would be tolerated by healthy cells could send HD cells below a viable energy threshold and that blocking PARP1 activation could be protective."

To test this hypothesis the MIND researchers first ran a computer search of their small-molecule library for potential novel inhibitors of PARP1, searching for those with structural similarities to known inhibitors. "Safety and efficacy of human drugs depends on many factors, so it's hard to predict which inhibitor would be most effective against a specific disorder. The more diverse novel inhibitors can be identified, the more chances there are of developing safe and effective drugs," Kazantsev explains.

Two candidate molecules were identified as potential PARP1 inhibitors based on their structure, and both of them were confirmed to inhibit the enzyme's activity in an in vitro assay. However, when tested using cultured human and rat cells, only one of the candidate molecules, K245-14, successfully prevented the death of cells in which PARP1 had been overactivated.

The next assays examined whether blocking PARP1 activity with K245-14 could reduce energy depletion in cells with the HD genetic mutation. Using cells from human HD patients and from a mouse model of the disorder, the MIND researchers compared the reactions of HD cells to oxidative stress caused by the application of hydrogen peroxide with the reactions of normal cells. Although all of the cells reacted with a loss of ATP, a key source of cellular energy, the HD cells which had much lower ATP levels to begin with were much more vulnerable to stress-induced energy loss. Inhibiting PARP1 by means of K245-14 reduced ATP loss in all tested cells and significantly protected against both energy loss and cell death in the HD cells.

"While we were pleased to observe these predicted protective effects in our experiments, validation of PARP1 as a useful HD drug target will require the testing of inhibitors in animal trials," Kazantsev explains. "The process of identifying the best candidates for trials will be very complex, since any drug treating a central nervous system disorder needs to penetrate the blood-brain barrier. We will be working with our collaborators at the Scripps Research Institute world leaders in computational chemistry to conduct a more comprehensive virtual screen and select additional promising candidates for drug development.

"Inhibition of PARP1 activity is thought to be potentially beneficial for treatment of cancer, neurodegenerative conditions such as Parkinson's disease, and over twenty other human disorders," he adds. "We envision broad therapeutic applications for small molecule inhibitors of PARP1." Kazantsev is an assistant professor of Neurology at Harvard Medical School.

Genetic Gender Gap in Disease Risk, Drug Response

Mon, 10 Jul 2006 06:29:00 PST

( from http://www.rxpgnews.com ) UCLA researchers report that thousands of genes behave differently in the same organs of males and females something never detected to this degree. Published in the August issue of Genome Research, the study sheds light on why the same disease often strikes males and females differently, and why the genders may respond differently to the same drug.

"We previously had no good understanding of why the sexes vary in their relationship to different diseases," explained Xia Yang, Ph.D., first author and postdoctoral fellow in cardiology at the David Geffen School of Medicine at UCLA. "Our study discovered a genetic disparity that may explain why males and females diverge in terms of disease risk, rate and severity."

"This research holds important implications for understanding disorders such as diabetes, heart disease and obesity, and identifies targets for the development of gender-specific therapies," said Jake Lusis, Ph.D., co-investigator and UCLA professor of human genetics.

The UCLA team examined brain, liver, fat and muscle tissue from mice with the goal of finding genetic clues related to mental illnesses, diabetes, obesity and atherosclerosis. Humans and mice share 99 percent of their genes.

The scientists focused on gene expression -- the process by which a gene's DNA sequence is converted into cellular proteins. With the help of Rosetta Informatics, the team scrutinized more than 23,000 genes to measure their expression level in male and female tissue.

What they found surprised them. While each gene functioned the same in both sexes, the scientists found a direct correlation between gender and the amount of gene expressed.

"We saw striking and measurable differences in more than half of the genes' expression patterns between males and females," said Dr. Thomas Drake, co-investigator and UCLA professor of pathology. "We didn't expect that. No one has previously demonstrated this genetic gender gap at such high levels."

UCLA is the first to uncover a gender difference in gene expression in fat and muscle tissue. Earlier studies have identified roughly 1,000 sex-biased genes in the liver, and other research has found a combined total of 60 gender-influenced genes in the brain about one-tenth of what the UCLA team discovered in these organs.

Even in the same organ, the researchers identified scores of genes that varied in expression levels between the sexes. Gender consistently influenced the expression levels of thousands of genes in the liver, fat and muscle tissue. This effect was slightly more limited in the brain, where hundreds, not thousands, of genes showed different expression patterns.

"Males and females share the same genetic code, but our findings imply that gender regulates how quickly the body can convert DNA to proteins," said Yang. "This suggests that gender influences how disease develops."

The gender differences in gene expression also varied by tissue. Affected genes were typically those most involved in the organ's function, suggesting that gender influences important genes with specialized roles, not the rank-and-file.

In the liver, for example, the expression of genes involved in drug metabolism differed by sex. The findings imply that male and female livers function the same, but work at different rates.

"Our findings in the liver may explain why men and women respond differently to the same drug," noted Lusis. "Studies show that aspirin is more effective at preventing heart attack in men than women. One gender may metabolize the drug faster, leaving too little of the medication in the system to produce an effect."

"At the genetic level, the only difference between the genders is the sex chromosomes," said Drake. "Out of the more than 30,000 genes that make up the human genome, the X and Y chromosomes account for less than 2 percent of the body's genes. But when we looked at the gene expression in these four tissues, more than half of the genes differed significantly between the sexes. The differences were not related to reproductive systems they were visible across the board and related to primary functions of a wide variety of organs."

The UCLA findings support the importance of gender-specific clinical trials. Most medication dosages for women have been based on clinical trials primarily conducted on men.

"This research represents a significant step forward in deepening our understanding of gender-based differences in medicine," said Dr. Janet Pregler, director of the Iris Cantor-UCLA Women's Health Center. The center's executive advisory board, a group of businesswomen interested in advancing women's health, helped fund the study.

"Many of the genes we identified relate to processes that influence common diseases," said Yang. "This is crucial, because once we understand the gender gap in these disease mechanisms, we can create new strategies for designing and testing new sex-specific drugs."

'Molecular assassin' targets disease gene

Wed, 05 Jul 2006 15:16:00 PST

( from http://www.rxpgnews.com ) University of New South Wales (UNSW) researchers have announced they are developing a new class of experimental drug that has the potential to treat a diverse range of health problems, from inflammation and cancer through to eye and heart disease.

Certain types of skin cancers and blindness due to age-related macular degeneration (AMD) and diabetic retinopathy are likely to be among the first uses for the drug. AMD is the most common cause of blindness in Australia (Macular Degeneration Foundation).

The experimental drug has already been shown to be effective on skin cancers in pre-clinical models, in another paper published this month by Professor Khachigian's team in the journal, Oncogene.

"This may be a 'one-size fits all' therapy, because it targets a master regulator gene called c-Jun which appears to be involved in all of these diseases," said UNSW Professor Levon Khachigian, of the Centre for Vascular Research (CVR), who is the senior author of the Nature Biotechnology paper.

"c-Jun is an important disease-causing gene," said Professor Khachigian, a molecular biologist. "It stands out because we don't see much of it in normal tissue but it is highly expressed in diseased blood vessels, eyes, lungs, joints, and in the gut in any number of areas involving inflammation and aggressive vascular growth.

"Our experimental drug, Dz13, is like a secret agent that finds its target, c-Jun, within the cell and destroys it," he said. "It is a specific, pre-programmed 'molecular assassin'."

The paper in Nature Biotechnology shows the potential of c-Jun as a drug target in inflammation. It details tests in a variety of pre-clinical models showing how effective Dz13 is in problems such as eye disease and arthritis.

The next phase in the therapy's development would be a trial, involving up to 10 people with non-melanoma skin cancers. The tumours would be injected with the drug over an eight-week period.

"If such a trial were successful, it would be a significant development given the high rates of skin cancer and because the main treatment currently is surgical excision, which can cause scarring," said Professor Khachigian.

"Conventional anti-inflammatory drugs are associated with a whole host of side-effects. Our therapeutic may potentially avert some of these."

A third paper using the same technology, but focusing on a different master regulator, Egr-1, has also been published this month by Professor Khachigian's group in the Journal of Thrombosis and Haemostasis and shows that heart muscle damage is reduced by the drug after a heart attack.

Scientists Uncover Rules for Gene Amplification

Fri, 30 Jun 2006 13:17:00 PST

( from http://www.rxpgnews.com ) Gene amplification plays an important role in causing cancers via activation of oncogenes. If scientists can determine the rules as to which segments of genetic material become amplified and how, oncologists and drug researchers may be able to interrupt that process and prevent the formation and growth of some tumors. Using yeast as a model organism, researchers at the Georgia Institute of Technology have discovered that the location of a hairpin-capped break relative to the end of the chromosome will determine the fate of the amplification event.

Gene amplification is the increase in copy number of a particular piece of DNA and
is a hallmark of tumor cells. Amplified genomic segments are frequently manifested in one of two cytologically recognizable forms. Double minutes are extrachromosomal segments of amplified DNA. Homogeneously staining regions are amplified intrachromosomal segments forming large genomic regions. Some strategies of pharmaceutical research in cancer prevention and treatment could involve curbing cancer development via restricting gene amplification. The first step towards achieving this is to discover the rules that govern whether an amplification event is a double minute or a homogenously-staining region.

Its known that regions of chromosomes that are prone to amplification have
palindromic sequences of DNA, which are weak places where the chromosome can break. These palindromic sequences can be naturally found in human genome. The distribution of such sequences can vary from one individual to another. Researchers at the Georgia Institute of Technology have discovered that a particular type of DNA break, a hairpin-capped double strand break, induced by these palindromic sequences, is a precursor to amplification.

We have a developed a system in yeast which would mimic the situation in human cancer cells wherein oncogenes might be located next to palindromic sequences. Using this system we have discovered the rules that determine how double minutes or homogeneously staining regions can be generated, said Kirill Lobachev, assistant professor in Georgia Techs School of Biology.

If these rules operating in yeast can be extended to higher eukaryotes then we can propose that if the oncogene is located between the hairpin-capped break and the telomere, then the amplification event will result in a double minute. If the break occurs between the oncogene and the telomere, then the amplification would yield a homogenously-staining region. adds Vidhya Narayanan a Ph.D. student in Kirill Lobachevs lab and first author of the study.

The findings can help researchers understand the cause of cancer in diseased individuals and also to potentially identify individuals who might be prone for cancer.

In addition to Lobachev and Narayanan, the research team consisted of Hyun-Min Kim from Georgia Tech and collaborators Piotr A. Mieczkowski and Thomas D. Petes from Duke University. This work was supported by funds from National Science Foundation and National Institute of Health.

Gene therapy protects neurons in Huntington's disease

Fri, 30 Jun 2006 03:02:00 PST

( from http://www.rxpgnews.com ) Researchers at Rush University Medical Center, Chicago, and Ceregene Inc., San Diego, have successfully used gene therapy to preserve motor function and stop the anatomic, cellular changes that occur in the brains of mice with Huntington's disease (HD). This is the first study to demonstrate that, using this delivery method, symptom onset might be prevented in HD mice with this treatment.

"This could be an important step toward a disease modifying therapy," says co-author Jeffrey H. Kordower, PhD, director of the Research Center for Brain Repair at Rush. "We could potentially be stopping the disease process in its tracks, delaying symptoms from ever showing up."

Huntington's disease is an inherited degenerative disease that progressively robs patients of the ability to think, judge appropriately, control their emotions and perform coordinated tasks. HD typically begins in mid-life, between the ages of 40 and 50. There is no effective treatment or cure for this fatal illness that affects 30,000 Americans and places another 75,000 at risk.

Kordower says this research, if eventually applied to humans, could help those who have HD or, due to the presence of a genetic test, are known to be destined to get HD.

"Each child of an affected parent has a 50 percent risk for inheriting the disease. Genetic testing can identify mutated gene carriers destined to suffer from HD. Unlike other neurodegenerative disorders, identification of the genetic markers provides a unique opportunity to intercede therapeutically before or extremely early in the disease processonly a small fraction of potential carriers get tested. But, if there was a treatment, especially one that altered the natural course of disease, potentially halting it, we would hope every potential patient would get tested so they could avail themselves to the therapy."

Researchers used a defective virus, adenoassociated viral vector, (AAV) to deliver gene therapy (glial-derived neurotrophic factor (GDNF) directly to the brain cells of mice.

GDNF is one of two closely related, naturally-occurring nutrients that strengthen and protect brain cells that would normally die in this disease. The other neural nutrient is called neurturin (NTN). GDNF and NTN also increase production of the chemical neurotransmitter dopamine, which sends signals in the brain that enable people to move smoothly and normally. Ceregene, Inc, whose scientists co-authored this paper, is developing AAV-NTN (called CERE-120) as a potential treatment for several neurodegenerative diseases, while using AAV-GDNF for 'proof of principle' research studies.

The mice in this study were injected with the gene for GDNF encased in a harmless viral coating, which protects the gene and facilitates its delivery to brain cells. The virus coating (AAV vector) that carries the gene is well studied and has been used in several other gene transfer studies to deliver different genes for Parkinson's disease and Alzheimer's disease patients. The vector is no longer a true virus as it cannot replicate on its own and no longer contains any of its own genes. The vector has been engineered to transfer the gene for the brain nutrient selectively to the area of the brain where it is needed to protect the degenerating cells.

Three groups of mice were involved in the 4 month study. All mice were modeled to have the genetics of HD. The HD mice exhibited symptoms of motor deficits including loss of control, gait abnormalities, hypokinesia (abnormally decreased mobility and motor function), hind limb clasping behaviors and muscle weakness. One control group of mice did not receive any gene therapy. A second control group was injected with a placebo gene therapy. The third group received the active GDNF gene therapy.

To measure fine motor coordination, balance and fatigue, researchers evaluated mice walking on a rotating rod. Mice injected with the gene therapy performed significantly better than the other mice. These mice also showed diminished hind limb clasping, (a simulation of motor control behavior in HD patients). Perhaps most importantly, gene delivery of GDNF provided neuroprotection in the brain, with reduced density of brain inclusions and less cell death.

The authors wrote "Although GDNF's exact role in preventing cell death in mice modeled with HD remains to be established, we speculate the increase trophic support and inhibiting apoptosis (programmed cell death) via these two pathways likely played integral roles."

Kordower says the study suggests a new approach to forestall disease progression in newly diagnosed HD patients by delivering potent trophic factors with effects that are long-term and non-toxic." "If these results can be replicated in HD patients, it would represent a significant advance in the treatment of this tragic disease", agreed Dr. Jeffrey Ostrove, President and CEO of Ceregene.

"We are pleased with the results of this 'proof of concept' study with AAV-GDNF in HD mice", stated Raymond T. Bartus, Ph.D., Sr. Vice President, Clinical and Preclinical R&D and COO, Ceregene. "We now look forward to completing ongoing studies with our product, AAV-NTN (CERE-120), in HD mice, also performed in collaboration with Dr. Kordower and Rush University Medical Center", Bartus added.

Ceregene's lead program with CERE-120 is in Parkinson's disease (PD). The company completed enrollment of a Phase I trial with CERE-120 at UCSF and Rush University Medical Center, which was reported to be safe and well tolerated in PD patients at the American Association of Neurology meeting last spring. Initial efficacy results of this Phase I trial are expected to be presented this fall and a double-blinded, controlled Phase II trail in PD patients is planned for later this year.

DNA damage resets the cellular circadian clock

Fri, 30 Jun 2006 02:42:00 PST

( from http://www.rxpgnews.com ) Dartmouth Medical School geneticists have discovered that DNA damage resets the cellular circadian clock, suggesting links among circadian timing, the cycle of cell division, and the propensity for cancer.

Their work, reported June 29 in Science Express, the advance electronic publication of Science, implies a protective dimension for the biological clock in addition to its pacemaker functions that play such a sweeping role in the rhythms and activities of life.

The notion that the clock regulates DNA-damage input and that mutation can affect the clock as well as the cell cycle is novel, says Jay Dunlap, professor and chair of genetics at DMS. It suggests a fundamental connection among circadian timing, cell cycle progress, and potentially the origins of some cancers.

Dunlap is a co-author of the paper with DMS colleagues, Jennifer Loros, professor of biochemistry, graduate student Christopher L. Baker, and former students António M. Pregueiro and Qiuyun Liu.

The team of Loros and Dunlap were among to first to delineate the intricate web of clockwork genes, proteins and feedback loops that drive circadian rhythms, working chiefly in the classic genetic model organism Neurospora, the common bread mold.

One gene (period-4) was identified over 25 years ago by a mutation that affects two clock properties, shortening the circadian period and altering temperature compensation. For this study, the researchers cloned the gene based on its position in the genome, and found it was an important cell cycle regulator. When they eliminated the gene from the genome, the clock was normal, indicating that the mutation interfered in some way with the clock, rather than supplying something that the clock normally needs to run.

Biochemically, the mutation results in a premature modification of the well understood clock protein, frequency (FRQ). The investigators demonstrated that this was a direct result of action by an enzyme, called in mammals checkpoint kinase-2 (CHK2), whose normal role is exclusively in regulating the cell division cycle. CHK2 physically interacts with FRQ; the mutation makes this interaction much stronger. However, a mutant enzyme that has lost its activity has no effect on the clock.

Normally CHK2 is involved in the signal response pathway that begins when DNA is damaged and results in a temporary stoppage of cell division until the damage is fixed. The researchers found that the resetting effect of DNA damage requires the period-4 clock protein, and that period-4 is the homolog, the Neurospora version, of the mammalian checkpoint kinase.

Moreover, the clock regulates expression of the period-4 gene. This closes a loop connecting the clock to period-4 and period-4 to the clock and the cell cycle. The clock normally modulates expression of this gene that encodes an important cell cycle regulator, and that cell cycle regulator in turn affects not only the cell cycle but also the clock.

Recent evidence in mammalian cells shows that other cell cycle regulators physically interact with clock proteins. Loss of at least one clock protein (mammalian period-2) is known to increase cancer susceptibility. The coordination of the clock and cell division through cell cycle checkpoints, supports the clocks integral role in basic cell biology, conclude the researchers. Their work can help advance understanding of cancer origins as well as the timing of anti-cancer treatment.

Human cells use complex system of transcription-factor combinations

Mon, 19 Jun 2006 01:22:00 PST

( from http://www.rxpgnews.com ) Scientists eager to help develop a new generation of pharmaceuticals are studying cellular proteins called transcription factors, which bind to upstream sequences of genes to turn the expression of those genes on or off. Some pharmaceutical companies are also hoping to develop drugs that selectively block the binding of transcription factors as a way to short-circuit the harmful effects of diseases, and researchers at the University of California, San Diego on June 16 reported new findings that could aid that effort.

Bioengineering researchers at UCSD and two research institutes in Germany report in the June 16 issue of PLoS Computational Biology that transcription factors act not only in isolation, but also in pairs, trios, and combinations of up to 13 to regulate distinct sets of genes. The researchers, led by UCSD bioengineering professor Trey Ideker, reported a list with 363 combinations of 91 transcription factors that regulate a large proportion of genes in the yeast genome. The team used rigorous statistical tests to discover active combinations of transcription factors, as if a cell were mixing and matching parts of its regulatory-protein wardrobe to respond to different environmental conditions.

The researchers expect that human cells use a similar system of transcription-factor combinations, but on a larger scale.

âA cellâs surprising ability to mix and match so many different combinations of these factors to achieve a high degree of complexity and specificity in the expression of its genes is impossible for even the most experienced cell biologists to conceptualize,â said Andreas Beyer, a post-doctoral fellow at the UCSD Jacobs School of Engineeringâs Department of Bioengineering. âThatâs why we have computers.â

The researchers combined the results of their laboratory with other large-scale measurements of transcription factor-gene binding, such as those reported earlier by MIT biology professor Richard A. Young and his collaborators.

Idekerâs team was able to identify new transcription factor binding patterns by borrowing a concept from computer science. The team considered the binding of one transcription factor to one gene as analogous to one âhopâ of a data packet from one Internet router to another.

In the case of gene regulation, Idekerâs team identified â2hopâ relationships by first focusing on single transcription factor-gene associations, plus other experimental evidence that indicates that that gene regulates a second gene.

To enlarge the scope of the model further, Idekerâs group also incorporated other previously discovered transcription-factor interactions and related genetic results. They relied on a total of eight types of direct and indirect evidence to create a model. That model predicts 980 as-yet-undiscovered transcription factor-gene binding interactions.

âThis âsystems biologyâ approach, using so many different lines of evidence, has given us a much more revealing and detailed picture of how cells orchestrate gene regulation to cope with different environments,â said Ideker. âWeâre far from understanding the full picture of gene regulation in a cell, but this new information should give scientists who are interested in blocking transcription factors a powerful new tool to narrow their search to the most promising candidates.â

Huntingtin cleavage is caused by caspase-6

Sat, 17 Jun 2006 20:10:00 PST

( from http://www.rxpgnews.com ) Researchers at the University of British Columbia's Centre for Molecular Medicine and Therapeutics (CMMT) have provided ground-breaking evidence for a cure for Huntington disease in a mouse offering hope that this disease can be relieved in humans.

Published today in Cell journal, Dr. Michael Hayden and colleagues discovered that by preventing the cleavage of the mutant huntingtin protein responsible for Huntington disease (HD) in a mouse model, the degenerative symptoms underlying the illness do not appear and the mouse displays normal brain function. This is the first time that a cure for HD in mice has been successfully achieved.

"Ten years ago, we discovered that huntingtin is cleaved by 'molecular scissors' which led to the hypothesis that cleavage of huntingtin may play a key role in causing Huntington disease", said Dr. Michael Hayden, Director and Senior Scientist at the Child and Family Research Institute's Centre for Molecular Medicine and Therapeutics. Dr. Hayden is also a Canada Research Chair in Human Genetics and Molecular Medicine.

Now a decade later, this hypothesis has resulted in a landmark discovery. "This is a monumental effort that provides the most compelling evidence of this hypothesis to date", said Dr. Marian DiFiglia, Professor in Neurology, Massachusetts General Hospital, Harvard Medical School and one of the world's leading experts on Huntington disease. "Dr. Hayden and his team have shown in convincing fashion that many of the changes seen in HD patients can be erased in HD mice simply by engineering a mutation into the disease gene that prevents the protein from getting cleaved at a specific site".

To explore the role of cleavage, Dr. Hayden's team established an animal model of HD that replicated the key disease features seen in patients. A unique aspect of this particular animal model is that it embodied the human HD gene in exactly the same way seen in patients. This replication allowed researchers to examine the progression of HD symptoms including the inevitable cleavage of the mutant huntingtin protein. In the study, researchers confirmed that the deadly cleavage is caused by a key enzyme called caspase-6. By blocking the action of this target, they showed that the mouse did not develop any symptoms of Huntington disease.

Hayden's team is now trying to test this model of prevention in a mouse using drug inhibitors and then ultimately in humans. "Our findings are important because they tell us exactly what we need to do next", said Dr. Rona Graham, Post Doctoral Fellow at the CMMT and lead author in the study.

This work is also pivotal for the individuals and families affected by Huntington disease. "Patients of this disease should know that this is a research milestone for all and that this work brings the field closer to finding effective treatment for a devastating disorder", said Dr. DiFiglia.

The Huntington Society of Canada (HSC), a national network of volunteers and professionals united in the fight against HD, echoed this sentiment. "This ground-breaking research provides great hope for the Huntington community", said Don Lamont, the Society's CEO and Executive Director. "This research brings us closer to treatment and ultimately a cure".

Huntington disease is a degenerative brain disease that affects one in every 10,000 Canadians. One in 1,000 is touched by HD -- for example, as a person with HD, a family member, a person at risk, caregiver or friend. The disease results from degeneration of neurons in certain areas of the brain causing uncontrolled movements, loss of intellectual faculties, and emotional disturbances. Currently, there is no treatment to delay or prevent HD in patients.

The risk of transmission of genetic disorders through donor's sperm

Fri, 19 May 2006 19:25:00 PST

( from http://www.rxpgnews.com ) As medical technology continues to advance, fertility procedures such as in-vitro fertilization and donor insemination are becoming more commonplace. However, a study in the May issue of The Journal of Pediatrics warns that, even after thorough screenings of sperm donors, genetic disorders can be transmitted to the conceived children.

Laurence Boxer, MD, and colleagues from the University of Michigan and the Severe Chronic Neutropenia International Registry investigated the cases of five children conceived by in-vitro fertilization or donor insemination who had severe congenital neutropenia (SCN)--a genetic disorder characterized by abnormally low levels of certain white blood cells in the body. Because these white blood cells help fight bacterial infections by destroying invading bacteria, people with SCN are more susceptible to recurring infections and are at greater risk for developing leukemia.

The study results showed that the same sperm donor was used for all five pregnancies. After conducting advanced genetic testing, the authors established that the donor was the carrier of the gene, not the mothers. The sperm bank was informed of this evidence, and all remaining samples were discarded.

The authors conclude that, because it is presently difficult to screen for all conceivable genetic disorders, it is imperative that potential mothers be properly counseled and informed prior to the procedures. "The mothers need to be prepared that there is always an inherent risk of a genetic disorder being transmitted by the donor's sperm," says Dr. Boxer.

How can DNA be damaged

Thu, 18 May 2006 02:58:00 PST

( from http://www.rxpgnews.com ) Researchers have known for years that damaged DNA can lead to human diseases such as cancer, but how damage occurs--and what causes it--has remained less clear.

Now, computational chemists at the University of Georgia have discovered for the first time that when a proton is knocked off one of the pairs of bases that make up DNA, a chain of damage begins that causes "lesions" in the DNA. These lesions, when replicated in the copying mechanisms of DNA, can lead to serious disorders such as cancer.

The research, just published in the Proceedings of the National Academy of Sciences (PNAS), was led by doctoral student Maria Lind and Henry F. Schaefer III, Graham-Perdue Professor of Chemistry. Other authors on the paper are doctoral student Partha Bera, postdoctoral associate Nancy Richardson and recent doctoral graduate Steven Wheeler.

Call it a "pinball proton." While chemists have shown other causes of DNA damage, the report in PNAS is the first to report how protons, knocked away by such mechanisms as radiation or chemical exposure, can cause lesions in DNA. The work was done entirely on computers in the Center for Computational Chemistry, part of the Franklin College of Arts and Sciences at UGA.

"This kind of damage in DNA subunits is about as basic as you can get," said Schaefer. "This is the simplest kind of lesion possible for such a system."

The double-helix structure of DNA has been known for more than half a century. This basic building block of life can "unzip" itself to create copies, a process at the heart of cell replication and growth. DNA is made of four "bases," Adenine, Guanine, Thymine and Cytosine, and each one pairs with its opposite to form bonds where the "information" of life is stored. Thus, Guanine pairs with Cytosine, and Thymine with Adenine.

The team at the University of Georgia studied how the removal of a proton from the Guanine-Cytosine (G-C) base pair is involved in creating lesions that can lead to replication errors. This pair has 10 protons, meaning there are numerous targets for processes that knock the protons off.

The lesions are breaks in the hydrogen bonds, of which there are two in the G-C base pair. (The Adenine-Thymine pair has three hydrogen bonds.)

"Our real goal is to examine all possible lesions in DNA subunits," said Lind.

The team discovered that the base pair minus its knocked-off proton can either break entirely or change its bonding angle--something that also causes improper replication.

"The C-G subunit is usually totally planar [flat]," said Lind. "If it twists, it could simply pull apart."

Though it has already been suspected that lesions in DNA caused by both high- and low-energy electrons result in cancer cell formation, the new study is the first evidence that protons do the same thing.

The study in PNAS also has other implications. Researchers are beginning to understand how DNA can be used as "molecular wire" in constructing electrical circuits. Such a breakthrough would allow small electronic devices to shrink even further, but how the electrical properties of DNA would work in such a context is not yet understood. The UGA research adds important knowledge about how so-called "deprotonated" DNA base pairs work and could be important in creating "DNA wire."

Could cellular defenses against sunlight be the key to effective gene therapy?

Tue, 25 Apr 2006 19:33:00 PST

( from http://www.rxpgnews.com ) An early study has demonstrated for the first time that laser light can target gene therapy right up to the edge of damaged cartilage, while leaving nearby healthy tissue untouched, according to an article published in the April edition of the Journal of Bone and Joint Surgery. True repair of injuries to articular cartilage would enable millions of patients, currently consigned to worsening arthritis and joint replacement, to return to athletic exercise.

Study authors say that dramatic progress is being made toward a new form of light-activated gene therapy for cartilage repair that will be safe, fast, easy on patients and compatible with techniques used by most surgeons (e.g. arthroscopy). Beyond knee injuries, researchers believe the technology could one day guide precision gene therapy for cancer or heart disease, restore vision by repairing eye tissue and rebuild skin destroyed by burns.

As many as 10 percent of young, active patients with bleeding in the knee joint following injury have damaged articular cartilage, the sponge-like layer that protects joints from the punishing impact of running and jumping. Unlike a broken leg bone that mends itself, damaged cartilage does not. Over time, damaged cartilage erodes to become a leading cause of osteoarthritis, which causes joint inflammation and pain in 40 million Americans. Many eventually require total joint replacement surgery.

"For years researchers have been trying to turn on gene therapy precisely within areas of damaged tissue without harming surrounding healthy tissue," said Edward M. Schwarz, Ph.D., professor of Orthopaedics within the Center for Musculoskeletal Research at the University of Rochester Medical Center. "Our study shows that we can use our cellular defenses against, of all things, sunlight, to finally achieve safe, precise control over tissue repair."

Limits of Current Therapy

Present surgical treatments for damaged cartilage like lavage and debridement relieve pain and inflammation, but leave a hole in the cartilage. Other techniques are too expensive and invasive or apply to few injuries, according to the study authors. Given the limits of surgical repair, researchers have been attempting for years to use gene therapy to truly repair, or re-grow, articular cartilage.

The blueprint for the human body is encoded in genes, which store information that is converted into proteins which carry out bodily functions. Gene therapy works by inserting specially designed genes into cells that can, for instance, direct cells to divide, which makes a tissue grow. Animals like amphibians re-grow amputated limbs, including articular cartilage, but humans grow cartilage only once, during development, without help.

To deliver the genes into cells, researchers need an effective delivery vehicle, or vector. Viruses have evolved for millions of years to invade human cells and insert DNA into their prey. Researchers have harnessed their useful qualities while removing the harmful ones. Unfortunately, no viral vector to date has been able to turn on genes only in the damaged areas because they infect cells in nearby healthy tissue as well.

Sunlight: A Specific Solution

The solution to the problem of how to target some cells for gene therapy, while missing their neighbors, came from a strange source: our cellular defenses against sunlight. The sun gives off ultraviolet (UV) light, which can cause destructive changes (genetic mutations) when exposed to sensitive molecules like DNA. If not defended against, the changes in DNA caused by UV light would cause humans to constantly develop cancer, for instance, in exposed tissue. Thus, an SOS system evolved that calls for genetic repairs when UV light causes too many mutations. Specifically, UV light turns on signaling proteins called stress kinases, which activate DNA polymerase, the enzyme that re-builds DNA chains when damaged.

Current technologies can direct UV light with great precision. That, combined with the ability of UV light to turn on DNA polymerase, has granted researchers the ability to turn on gene therapy in one cell, but not its neighbors. In recent years, researchers have been working to develop a system where UV light pre-treats target tissue, so that only the cells exposed to light gain the ability to copy themselves and grow. What remained was to find the right combination of vector and light to make the therapy safe as well as effective.

Recombinant adeno-associated virus (rAAV) turned out to be the right vector because it has evolved to deliver into the cell only a single strand of deoxyribonucleic acids (DNA), not the usual two strands of molecules. A second strand of DNA must be built by DNA polymerase to form active, double-stranded DNA before genes, or a gene therapy, can take effect. Single-stranded delivery is the key rAAV's usefulness as part of light-activated gene therapy because, of the all the cells infected with a gene therapy, only those struck by UV light will turn on DNA polymerase. Only those cells will activate the therapeutic gene, divide and re-grow tissue.

In addition, rAAV vectors used in human trials appear to be very safe. Other viral vectors used in early attempts at human gene therapy in some cases made permanent changes that caused violent immune reactions, and even cancer in a few cases, along with the therapeutic changes made. With safety more important than ever, it is almost impossible for rAAV to be dangerous because it does not contain any viral genes and is delivered using a harmless virus, Schwarz said. Lastly, rAAV appears to be the kind of virus that makes changes in the DNA of human cells for a few weeks, but then stops. This happens to be the perfect amount of time for a regenerative gene therapy, shutting down before too much re-growth occurs, according to researchers.

The Long and Short of UV Light

When discussing light-activated gene therapy, UV light used is categorized by its wavelength. The higher the intensity of light; the shorter its wavelength. An early attempt at light-activated gene therapy used short wavelength UV light (254 nm) because it can turn on DNA polymerase and direct tissue to re-grow. Unfortunately, it also destroys DNA in nearby healthy tissues. To overcome this obstacle, Schwarz and colleagues have been developing a system based on long-wavelength UV light with the goal of safely achieving site-specific therapy.

Long wavelength UV light is not absorbed by DNA, but still triggers the same natural SOS system. Instead of triggering the system by directly causing dangerous mutations like short wavelength UV light, the longer wavelength creates molecules called free radicals. Free radicals are highly reactive molecules that can themselves create mutations in DNA, but not nearly to the extent of short wavelength light. Free radicals, thus, turn on gene therapy before mutations are formed.

In addition, long wavelength UV light can be captured and transmitted through a cable, allowing for arthroscopic approaches, where short wavelength UV cannot. In arthroscopy, surgeons examine the injury with a fiber optic cable through the small incision, and in many cases make the repair through the same incision. Traditional surgery requires large incisions with long recovery times. This arthroscopic capability gives light-activated gene therapy the potential to become part of most orthopaedic medical practices in the country.

Thirdly, researchers developed a laser that can deliver the effective dose of long wavelength UV light within seconds. Using this laser light source means that the light-activated gene therapy could be completed in moments, not hours, for less pain and trouble on the part of patients.

Study Methods

The current study evaluated the ability of long-wavelength ultraviolet light to stimulate gene expression following infection by rAAV. Researchers evaluated the safety and efficacy of long-wavelength ultraviolet laser light to induce light-activated gene therapy in articular cartilage cells (chondrocytes). The study authors believe this is the first demonstration that site-directed gene delivery can safely and effectively treat articular defects in higher animal cartilage cells.

Given the safety concerns found with short wavelength, researchers were excited to find that the new long wavelength system is an order of magnitude more likely to turn on gene therapy as designed than to cause death by mutation (cytotoxity). Along with previous studies, the current research found rAAV to be highly efficient at turning on gene therapy in articular chondrocytes. Pretreatment with 6000 Joules per meter squared, a standard dose of UV light, led to a tenfold increase in the effect of gene therapy in target cells after one week. In addition, nearly half of cells exposed to the light expressed the inserted, therapeutic gene.

Schwarz is also president and founder of LAGeT, Inc. In 2003, LAGeT Inc. licensed the technology used in the study from the medical center, where he and his partners developed it. LAGeT is initially focusing on the development of therapies for musculoskeletal diseases, including osteoarthritis and spine-related repair, but recognizes its broader potential. One early project will attempt to use rAAV gene therapy to re-grow bone in patients who have lost bone to severe trauma or a bone tumor, and would otherwise lose the limb.

"While LAGeT's current focus is on musculoskeletal-related disorders, this approach could just as readily be used to deliver therapeutic genes to treat cancer and cardiovascular disease," Schwarz said. "We believe this area of research could represents a quantum leap from current treatments if successful because nothing out there yet brings about true regeneration of lost tissue."

Liver transplants provide metabolic cure for maple syrup urine disease

Tue, 11 Apr 2006 22:28:00 PST

( from http://www.rxpgnews.com ) Liver transplants cured the metabolic symptoms of 11 patients with a rare but devastating genetic condition known as Maple Syrup Urine Disease (MSUD), according to a study by researchers from Children's Hospital of Pittsburgh and the Clinic for Special Children.

All patients from the study (ranging in age from 1-20) are alive and well with normal liver function, according to the researchers. Amino acid levels in the study patients stabilized within 6-12 hours of transplant and remained stable since transplant despite unrestricted intake of protein.

MSUD is a metabolic disease which causes amino acids from proteins to accumulate in the body. The disease gets its names from the sweet smell of the urine. The accumulation of amino acids in the blood can cause metabolic crisis at any age, which can lead to brain swelling, stroke and even sudden death. Over a patient's lifetime, chronic instability of blood amino acids can result in serious learning disabilities and mental illness.

Before transplant, the only treatment was strict adherence to a diet almost devoid of protein. Despite adherence to this diet, patients were still at risk of metabolic crisis from something as simple as a common cold, which can disrupt the body's metabolism and cause rapid neurological deterioration.

In 1997, an MSUD patient at another hospital received a liver transplant due to an unrelated medical condition and physicians noticed the symptoms of her MSUD were alleviated.

Based on this serendipitous result, physicians from Children's and the Clinic for Special Children, located in Strasburg, Pa., began working collaboratively to develop a liver transplant protocol for MSUD which optimized patient safety. With a comprehensive, multidisciplinary protocol established, Children's transplant surgeons began performing liver transplants on MSUD patients in May 2004. Children's has performed 18 MSUD liver transplants since then.

The study by Children's and the Clinic for Special Children involved 11 of these MSUD patients, including the original patient. Results of the study are published in the March issue of the American Journal of Transplantation.

"The development of liver transplantation as a treatment for MSUD has dramatically improved our patients' quality of life," said George V. Mazariegos, director of Pediatric Transplantation at Children's and one of the study authors. "Our MSUD patients and their families had lived in fear of everything from a chicken nugget to a common cold. Liver transplantation is not without risks, but for some patients, it is the best option and it has allowed these recipients and their families to live without fear of simple things most people take for granted."

Kevin A. Strauss, MD, a pediatrician at the Clinic for Special Children and a co-author of the study, said that over the past 15-20 years, early diagnosis of MSUD followed by careful nutritional therapy have improved the health and developmental outcome of affected individuals.

"Nevertheless, the risk for metabolic crisis and acute neurological injury is always present, and many older individuals with MSUD suffer from depression, anxiety, and impaired concentration and learning," Dr. Strauss said. "Liver transplantation protects patients from these acute and chronic neurological complications. It is a reasonable alternative to nutritional therapy, particularly for patients with poor access to specialized medical care. However, liver transplantation is not without serious risks, and decisions about the best course of therapy will vary on an individual basis."

Gene therapy helps two Germans in global first

Mon, 03 Apr 2006 14:38:00 PST

( from http://www.rxpgnews.com ) For the first time, gene therapy has been used to alter the cells of sick adults and allow them greater resistance to bacteria and fungi, a Germany-based team of scientists said.

The research was described in the online version of the journal Nature Medicine. The cutting-edge techniques have mainly been used in the past to help sick children.

Dorothee von Laer, gene therapy coordinator at the Georg Speyer laboratory in Frankfurt, said two adult males aged 25 and 26 had been treated for a hereditary autoimmune disease over the past two years at Frankfurt University Hospital.

She said the new technique might prove suitable for other diseases where people are born with a defective gene.

The team of 27 scientists was able to raise above 50 percent the proportion of healthy immune cells in the men's blood, leading to a lessening of infection and avoiding serious new infections.

The rare condition the two men suffer from is termed chronic granulomatous disease - their bodies cannot kill invading bacteria and fungi and they often have internal infections. Currently the disease is treated using bone-marrow transplants.

The scientists took blood stem cells from the men and introduced a healthy gene to the cells. The cells were then re-injected into the men. The key to the technique was a new, more efficient method of inserting the genes in many cells at the same time.

Concern remains that such therapy could have a negative side effect: causing cancer of the blood. Research is continuing on how to make such techniques safer.

Mutations Change the Boolean Logic of Gene Regulation

Wed, 29 Mar 2006 06:39:00 PST

( from http://www.rxpgnews.com ) It is easy to think of a gene acting like a light bulb, switching either on or off, remaining silent, or being transcribed by the RNA-making machinery. The region of DNA that controls the gene's output is called its regulatory region, and in this simple (and too simplistic) scenario, that region would act like a simple onoff switch.

But the regulatory regions of real genes are more complex, and act more like molecular computers, combining the effects of multiple inputs and calibrating the gene's output accordingly. The inputs are the various molecules that affect gene activity by binding to sites in the regulatory region. These molecules combine their effects in complex ways. Sometimes the gene remains silent unless both are present. Sometimes they are additive, such that the output when two factors are present is twice the output when only one is present. Sometimes they cancel each other outin the presence of either, the gene is transcribed, but in the presence of both, it is not.

Thus, the regulatory region acts as a Boolean logic function, whose simple ANDs, ORs, and NOTs combine to determine the output of the gene. In a new study, Avi Mayo, Uri Alon, and colleagues show that mutations in the regulatory region affect this logic function in a simple and well-studied genetic system, the lac operon in Escherichia coli bacteria, whose suite of genes regulate metabolism of lactose.

The authors began by creating multiple strains of bacteria with mutations in the binding sites for the two regulators of the gene, called CRP and LacI, that respond to cyclic AMP and IPTG, an analog of lactose. They analyzed the effect of these mutations on the rate of gene transcription in the presence of varying concentrations of the two inducers. Previously, the authors showed that the function of the unmutated regulatory region was intermediate between a pure AND gate (in Boolean parlance) and a pure OR gate: that is, at certain concentrations the first regulator AND the second were needed, but at others, one OR the other sufficed. In the mutated strains, they found that some mutations replicated this behavior, while others switched the regulatory region to a more purely AND or purely OR gate, independent of concentration. Some mutations left the regulator almost like a simple light switch, whose on-or-off state depended almost entirely on one, but not the other, regulator.

Next, they developed a mathematical model that links the binding strengths of the regulators for each mutation (the inputs of the regulatory function) to the gene output. Based on this model, they propose that point mutations in this system cannot create all of the 16 possible two-input gates described by Boolean logic. For instance, since both regulators stimulate gene activity, no simple mutation is likely to switch the system to an AND NOT gate, in which one input can stimulate only when the other is not present.

The authors suggest that applying this kind of logic analysis to genetic circuits may aid in the design of artificial genetic systems, and in understanding more complex gene regulatory regions. With only 30,000 genes, it is clear that humans and other complex creatures must depend on exquisitely regulated gene expression to develop and adapt to environmental changes. The findings in this study support the growing appreciation that, from bacterium to baleen whale, complexity is highly dependent on fine-tuning gene regulation.

Synthetic biology experiment turns up a previously unrecognized gene-expression phenomenon

Thu, 16 Feb 2006 19:47:00 PST

( from http://www.rxpgnews.com ) An experiment designed to show how a usually innocuous bacterium regulates the expression of an unnecessary gene for green color has turned up a previously unrecognized phenomenon that could partially explain a feature of bacterial pathogenicity.

In a paper published in the Feb. 16 issue of Nature, researchers at Boston University (BU) and the University of California, San Diego (UCSD) reported that computer modeling predicted the new phenomenon before they confirmed it in laboratory experiments. The group led by James J. Collins, a biomedical engineering professor at BU, and Jeff Hasty, a bioengineering professor at UCSD, reported that the rise and fall in the amount of green-fluorescence protein in computer modeling matched the pattern recorded in E. coli cells grown in various laboratory conditions.

The researchers were surprised that cell-to-cell variation in the expression of the synthetic gene increased sharply as growth slowed and then stopped. "We were initially skeptical of our own results because they were so counterintuitive," said Collins. "But our laboratory experiments confirmed this increase in gene-expression variability, or noise, when growth stops. We think there may be some very interesting biology to explore in this situation."

Variability in gene expression could offer distinct survival advantages to a bacterium. Like a cruise ship whose life boats have been stocked with different combinations of food, first-aid kits, rain jackets, and flotation devices, a microscopic version of Survivor could occur in which only those individual bacterial cells with opportune combinations of proteins are able to weather harsh growth conditions in a pond or even inside a human body.

"This phenomenon could be relevant to bacterial 'persisters' - dormant cells that are highly resistant to antibiotics," said Collins. "Many bacterial pathogens can generate these persisters, which over many months can become the source of chronic infections. We don't understand the how persisters arise, but we think this unexpected gene-expression variability in bacterial cells is an interesting phenomenon that should be explored."

The group of researchers came up with the novel finding by using a relatively new research approach that involves the synthesis of simple gene networks, in this case one that produces a green-fluorescence protein. They measured expression of green fluorescence in a laboratory strain of E. coli under different growth conditions where other genes and proteins could potentially complicate the situation. They incorporated that information into a mathematical model.

The authors say their findings demonstrate the value of a so-called "bottom-up" approach to synthetic biology: models of relatively simple cellular processes can be used to predict the behavior of larger, more complex ones.

"We're excited by this study because the model itself led to a counterintuitive prediction that was supported by experimentation," said UCSD's Hasty. "The logical next step is to examine noise in the expression of proteins that would be essential to a bacterium's survival," Hasty said. "We've only begun to get an inkling of how noise in gene expression may be involved in the life of a cell."

Spinocerebellar ataxia type 5 (SCA5) gene pinpointed

Mon, 23 Jan 2006 16:12:00 PST

( from http://www.rxpgnews.com ) Researchers at the University of Minnesota Medical School have discovered the gene responsible for a type of ataxia, an incurable degenerative brain disease affecting movement and coordination.

This is the first neurodegenerative disease shown to be caused by mutations in the protein â-III spectrin which plays an important role in the maintaining the health of nerve cells. The scientific discovery has historical implications as well--the gene was identified in an 11-generation family descended from the grandparents of President Abraham Lincoln, with the President having a 25 percent risk of inheriting the mutation.

"We are excited about this discovery because it provides a genetic test that will lead to improved patient diagnoses and gives us new insight into the causes of ataxia and other neurodegenerative diseases, an important step towards developing an effective treatment," said Laura Ranum, Ph.D., senior investigator of the study and professor of Genetics, Cell Biology and Development at the University of Minnesota.

Understanding the effects of this abnormal protein, which provides internal structure to cells, will clarify how nerve cells die and may provide insight into other diseases, including amyotrophic lateral sclerosis (Lou Gehrig's disease) and Duchenne muscular dystrophy. The research will be published in the February print issue of Nature Genetics, and posted online Jan. 22, 2006.

Ataxia is a hereditary disease that causes loss of coordination resulting in difficulty with everyday tasks such as walking, speech, and writing. About 1 in 17,000 people have a genetic form of ataxia.

Spinocerebellar ataxia type 5 (SCA5) is a dominant gene disorder; if a parent has the disease, each of their children has a 50 percent chance of inheriting the mutation and developing ataxia sometime during their lifetime. The onset of SCA5 usually occurs between the ages of 30 and 50, but can appear earlier or later in life, with reported ages of onset ranging from 4 to more than 70 years of age.

Now that researchers have identified the specific mutation that causes SCA5, testing of patients at risk of developing this disease is possible before any symptoms appear. The availability of predictive testing allows people with a family history of the disease to determine whether they will develop the disease and whether their children are at risk of inheriting the mutation. In addition, the prognoses of the different types of ataxias vary greatly, so identifying the specific type of ataxia provides patients with a more accurate picture of what the future holds.

Ranum added: "Finding the SCA5 mutation in Lincoln's family makes it possible to test Lincoln's DNA if it becomes available to unequivocally determine if he carried the mutation and had or would have developed the disease." Biographical texts of Lincoln include descriptions of his uncoordinated and uneven gait, suggesting the possibility that he showed early features of the disease.

Ranum started this historical and scientific journey more than a decade ago. She and her colleagues John Day, M.D., Ph.D., University of Minnesota, and Larry Schut, M.D., CentraCare Clinic in St. Cloud, Minn., examined and collected DNA samples from more than 300 Lincoln family members who live across the country, tracking descendants from two major branches of the family.

MDC1 protein amplifies DNA injury signals

Sat, 21 Jan 2006 15:39:00 PST

( from http://www.rxpgnews.com ) A Mayo Clinic-led research collaboration has discovered that the protein MDC1 amplifies weak DNA injury signals so genetic repair can begin. Once amplified, even low-level damage signals become strong enough to activate the cell's natural repair processes while the injury is most tractable to repair. How this "distress call" was communicated wasn't clear until this finding, which appears in the January 20 issue of Molecular Cell. The research was conducted in collaboration with colleagues from Harvard University and the University of Texas, Austin.

"It's important that DNA lesions get repaired because then we don't get mutations," says Junjie Chen, Ph.D., Mayo Clinic oncology researcher and leader of the Mayo Clinic team. "This is just one mechanism involved in communicating injury to the repair processes, but it's an important start to understanding how we might one day design new treatments that help this repair system recover from injury or resist injury."

Dysfunction of the DNA damage response pathway makes the gene unstable. Genomic instability is the driving force in tumor formation, which is why cancer researchers around the world are focusing on understanding the DNA damage response pathway. Knowing how the cell communicates DNA injury to alert the repair system is an important first step to designing new therapies for cancers and other diseases.

The damage control process is continual and essential to health. DNA must repair itself so the instructions it gives to operate bodily functions are correct. In earlier work, the Mayo Clinic researchers determined that MDC1 is important to the repair process -- but they didn't know its role.

DNA is easily and often damaged by environmental and chemical sources such as ultraviolet radiation, cigarette smoke, and other natural and artificial toxins. These create injury sites or lesions. In healthy situations, DNA repair signal pathways are competently monitoring for damage and alerting the repair system when DNA lesions are detected.

"Most of the time we don't really encounter severe damage in the cell; most of the damage to DNA is mild injury -- such as low doses of sunlight," notes Dr. Chen. "But it's still injury, and we want to repair it as soon as possible so things don't get worse. That's why our question was: How does the cell detect low-dose damage signals? We believe this amplification process involving MDC1 is the answer to that question, and that it is critical because it's involved in even very subtle injury, such as a single DNA strand break -- which is very small. It is a very sensitive communication pathway."

To investigate the role of the protein MDC1, the researchers disrupted the MDC1 gene in mice and compared them to normal mice. The engineered strain of mice lacking MDC1 was extremely sensitive to DNA damage -- and unable to repair it. The MDC1-deficient mice showed symptoms of growth retardation, male infertility, immune defects and chromosome instability.

Now that they understand MDC1's role in amplifying distress calls from injured DNA to cue the repair process, the Mayo researchers are investigating another system that appears to play a similar role in the cell. "If we can understand all the pathways involved in signaling the DNA repair process, we may be able to develop a comprehensive approach to managing the signals to treat disease," says Dr. Chen.

Breakthrough in master gene mapping

Thu, 19 Jan 2006 12:46:00 PST

( from http://www.rxpgnews.com ) Researchers have broken new ground with their work on the key tumour suppressor gene "p53", raising hopes of better detection and treatment of cancer, a published report said Thursday.

Singapore's Agency for Science, Technology and Research and the Genome Institute developed the technology to map out which genes are affected by p53, the most studied gene in the world.

This will help understand what the gene does, and hopefully lead to better ways of detecting and treating cancer, researchers said.

Using home-grown gene-sequencing methods, they discovered nearly 100 genes controlled by p53, a master gene.

The ground-breaking effort, published in the journal Cell, has effectively doubled the number of genes discovered over the past two decades.

"With this understanding of how human genes are regulated, we can uncover more potentially important pathological and clinical roles of p53," said Ruan Yijun, a principal investigator.

A one-stop online portal is being developed to integrate information on all aspects of p53.

Research points to possible therapy to prevent congenital skull malformation

Sun, 25 Dec 2005 00:56:00 PST

( from http://www.rxpgnews.com ) Craniofacial researchers have developed an animal model that explains how skull malformations occur and how they might be prevented.

Birth defects of the face and skull are relatively common in humans, striking one in 500 to 1,000 babies. Defects can include cleft lip or palate, congenitally missing teeth and severe malformations of the skull.

A group led by Yang Chai, chair of the division of craniofacial sciences and therapeutics in the USC School of Dentistry, has identified the genetic factor leading to malformation of the forehead and frontal part of the skull. The discovery was published online Dec. 20 by the journal Development.

Children with frontal bone defects lack vital protection for their brain. They also may develop bulging, irregularly shaped heads.

Chai's group focused on a gene called transforming growth factor- beta. TGF-beta is known to play an important role in human and animal development.

To study the gene's effect on the skull, the researchers deleted TGF-beta in mice embryos, but only in the cranial neural crest cells that build facial bone and cartilage.

"If you knock out this gene in every single cell in the body, the embryos die very early. That doesn't help us figure out the role the gene plays in cranial development," Chai said.

The rest of the embryo's cells were allowed to retain the gene and grew normally.

Mice born from the treated embryos carried severe craniofacial defects, including cleft palate and skull malformations.

The results showed that TGF-beta is necessary for proper development of frontal bones.

In addition, the researchers found that they could rehabilitate embryos with missing TGF-beta by inoculating them with FGF, an intermediate protein in the "signaling cascade" that starts with TGF-beta and ends with healthy facial structures.

Chai's group concluded that TGF-beta acts through FGF to ensure proper development. This suggests a potential therapy for embryos that are missing TGF-beta in the neural crest cells.

"This might be useful to try out some possible rescue experiments," Chai said.

Although Chai's current results apply only to mice, a paper last spring in Nature Genetics (Loeys et al., 2005) identified a handful of human families with inherited mutations in TGF-beta receptors and with a high incidence of craniofacial defects, including cleft palate and skull malformations.

If the signaling mechanism in mice were to carry over to humans, pharmaceutical researchers could start to investigate FGF as a potential supplement for pregnant women, analogous to folic acid for prevention of spina bifida.

Chai noted that the FGF supplements in mice restored normal cell growth only in the skull region.

"Using FGF signaling we can actually rescue the cell proliferation defect, but so far we have not been able to do the same thing in the palate," Chai said.

TGF-beta also appears to work through different mechanisms in other parts of the body, Chai added. This suggests that no single treatment can correct all birth defects related to TGF-beta.

Scientists probe connection between regulatory DNA and disease

Sat, 17 Dec 2005 15:34:00 PST

( from http://www.rxpgnews.com ) Through the Human Genome Project, the HapMap Project and other efforts, we are beginning to identify genes that are modified in some diseases. More difficult to measure and identify are the regulatory regions in DNA the 'managers' of genes that control gene activity and might be important in causing disease.

Today, a team led by the Wellcome Trust Sanger Institute, together with colleagues in the USA and Switzerland, provide a measure of just how important regulatory region variation might be in a pilot study based on some 2% of the human genome. As many as 40 of 374 genes showed alteration in genetic activity that could be related to changes in DNA sequence called SNPs.

"We were amazed at the power of this study to detect associations between SNP variations and gene activity," commented Dr Manolis Dermitzakis, Investigator, Division of Informatics at the Wellcome Trust Sanger Institute. "We were even more amazed at the number of genes affected: more than 10% of our sample or perhaps 3000 genes across the genome could be subject to modification of activity in human populations due to common genetic variations."

The study combined the map of genetic variation developed through the HapMap with estimates of gene activity obtained from cell cultures from 60 individuals who provided samples for the HapMap. More than 630 genes were studied, of which 374 were active in the cell cultures. If gene activity in a cell culture was skewed from the average, it was investigated further.

These genes were correlated with more than 750,000 SNPs sequence differences between individuals in the sample collection. A series of statistical tests were carried out to provide increased confidence in the association between gene activity and sequence variation.

"Our sample size of 60 individuals is relatively small," continued Dr Dermitzakis, "and we might expect not to detect rare variations. However, our pilot project gives us greater confidence to take on a genome-wide survey of gene activity."

A global map of sequence variation and gene activity will be an important tool in the interpretation of variation and disease. Such genome-wide association studies will be able to identify some regions of the genome with strong disease effects.

"The HapMap is proving to be useful in a wide range of applications," commented Dr Panos Deloukas, Senior Investigator, Division of Medical Genetics, Wellcome Trust Sanger Institute. "The journey for our biomedical research is from DNA sequence to individual people and individual disease. The HapMap is a bridge from sequence data to the differences in individuals."

The project focused on three regions of the human genome. The first, called the ENCODE regions, and about 30 million base-pairs of DNA, are being intensively studied around the world as a group of 'typical' human genome regions. The second was 35million base-pairs of chromosome 21 sequence: three copies of chromosome 21 lead to Down Syndrome. The third was a region of chromosome 20 10 million base-pairs that is known to be associated with diabetes and obesity.

In comparison with gene sequences that contain the instructions to make proteins, regulatory regions that control genes are relatively poorly understood. Their structure is variable and their distance from the genes they control also varies among genes.

New tools are needed in the search of our genome for the sequences that contribute to disease, tools that will harness the massive amounts of DNA information and transform them into information of real biomedical utility. The methods described here, with the power of the HapMap data and the cell cultures available, will speed that transformation.

Detailed analysis of the dog genome published

Thu, 08 Dec 2005 20:05:00 PST

( from http://www.rxpgnews.com ) An international team, led by researchers at the Broad Institute of MIT and Harvard, today announced the publication of the genome sequence of the dog. In the Dec. 8 issue of the journal Nature, the researchers present a detailed analysis of the dog genome and describe how the data offer the potential for improving the health of man and man's best friend.

"When compared with the genomes of human and other important organisms, the dog genome provides a powerful tool for identifying genetic factors that contribute to human health and disease," said Francis S. Collins, M.D., Ph.D., director of the National Human Genome Research Institute (NHGRI), which supported the research. "This milestone is especially gratifying because it will also directly benefit veterinary researchers' efforts to better understand and treat diseases afflicting our loyal canine companions."

Efforts to create the genetic tools needed for mapping disease genes in dogs have gained momentum over the last 15 years, and already include a partial survey of the poodle genome. More than two years ago, Kerstin Lindblad-Toh, Ph.D., co-director of the genome sequencing and analysis program at the Broad Institute, and her colleagues embarked on a two-part project to assemble a complete map of the dog genome.

In the first phase, they acquired high-quality DNA sequence covering nearly 99 percent of the dog genome, from a female boxer named Tasha. The boxer was chosen as a representative of the average purebred dog to produce what has become a reference sequence for the dog genome community. Using the sequence information as a genetic "compass," they navigated the genomes of 10 different dog breeds and other related canine species, including the gray wolf and coyote. In this sampling, they pinpointed tiny spots of genetic variation, called single nucleotide polymorphisms (SNPs), which serve as recognizable signposts that can be used to locate the causes of genetic disease.

"Of the more than 5,500 mammals living today, dogs are arguably the most remarkable," said senior author Eric Lander, director of the Broad Institute, professor of biology at MIT and systems biology at Harvard Medical School, and a member of the Whitehead Institute for Biomedical Research. "The incredible physical and behavioral diversity of dogs from Chihuahuas to Great Danes is encoded in their genomes. It can uniquely help us understand embryonic development, neurobiology, human disease and the basis of evolution."

Humans domesticated the dog, Canis familiaris, from gray wolves as long as 100,000 years ago. As a result of selective breeding over the past few centuries, modern dog breeds present a model of diversity. From 6-pound Chihuahuas to 120-pound Great Danes, from high-energy Jack Russell Terriers to mild-mannered basset hounds, and from the herding instincts of Shetland sheepdogs to pointers pointing, humans have bred dogs for desirable physical and behavioral traits. While such breeding practices aimed to preserve the preferred traits of one generation in the next, they also predispose many dog breeds to genetic disorders, including heart disease, cancer, blindness, cataracts, epilepsy, hip dysplasia and deafness.

Elaine A. Ostrander, Ph.D., chief of NHGRI's Cancer Genetics Branch, co-authored the Nature paper, along with postdoctoral research fellows, Heidi G. Parker and Nate B. Sutter. Dr. Ostrander's laboratory maps genes responsible for cancer susceptibility in canines and humans, including breast and prostate cancers. In addition, Dr. Ostrander was the lead author of the white paper that provided the biomedical rationale for sequencing the dog genome.

"The leading causes of death in dogs are a variety of cancers, and many of them are very similar biologically to human cancers." said Dr. Ostrander. "Using the dog genome sequence in combination with the human genome sequence will help researchers to narrow their search for many more of the genetic contributors underlying cancer and other major diseases."

While dogs occupy a special place in human hearts, they also sit at a key branch point, relative to humans, in the evolutionary tree. It was already known that humans share more of their ancestral DNA with dogs than with mice; the availability of the dog genome sequence has allowed researchers to describe a common set of genetic elements -- representing about 5 percent of the human genome -- that are preferentially preserved among human, dog and mouse. Rather than being evenly distributed, some of these elements are crowded around just a small fraction of the genes in the genome. Future studies of these clusters may give scientists the critical insight needed to unravel how genomes work.

Other interesting observations emerged from this cross-genome analysis. For example, the research group found that while different breeds show amazing physical diversity, they often share large segments of their DNA, likely reflecting their recent shared origin. As a result, genetic tools being developed at the Broad Institute and NHGRI for any one breed of dog are likely to be useful in genetic experiments in nearly any breed.

The international team of researchers also identified roughly 2.5 million single nucleotide polymorphisms (SNPs) sprinkled throughout the dog genome. SNPs are variations in the DNA code, some of which contribute to diseases or the overall health of a dog. SNPs also can be used to create a set of coordinates with which to survey genetic changes, both within and across dog breeds. These efforts revealed that individual breeds have maintained a large amount of genetic variability, despite their long history of restrictive breeding. In practical terms, this means that future efforts to locate disease genes in dogs can be much narrower in scope than comparable human studies, requiring a smaller number of genetic markers and DNA samples collected from the blood or cheek from only a few hundred dogs.

Scientists in the canine genetics community worldwide are currently tackling this problem by applying the knowledge gained from SNP analysis to find disease genes. To this end, the dog-owner community is an essential collaborator. "We deeply appreciate the generous cooperation of individual dog owners and breeders, breed clubs and veterinary schools in providing blood samples for genetic analysis and disease gene mapping," said Dr. Lindblad-Toh, who is the paper's first author. "Without their interest and help we could not be doing this work."

Sequencing of the dog genome was conducted as part of NHGRI's Large-Scale Sequencing Research Network, at an approximate cost of $30 million. NHGRI is one of the 27 institutes and centers at the NIH, an agency of the Department of Health and Human Services (DHHS).

Bias in Reporting of Genetic Association Studies

Tue, 22 Nov 2005 16:01:00 PST

( from http://www.rxpgnews.com ) One of the tools in the scientist's armory for resolving a medical issue or consolidating a body of clinical trials is the systematic review of the published medical literature. This technique involves doing a literature search and critical appraisal of individual studies, and in addition, may also use statistical techniques to combine the results of these studies. One of the aims of such reviews is to assess and then, ideally, include all appropriate studies that address the question of the review. But finding all studies is not always possible, and researchers have no way of knowing what they have missed. But does it matter if some studies are left out?

It would definitely matter if the missing studies differed significantly from the included ones. And the worst-case scenario is that the accumulation of evidence might point to the wrong answer if the studies included are unrepresentative of all those that have been done.

Studies of publication bias have noted that papers with significant positive results are easier to find than those with nonsignificant or negative results. As a result, overrepresentation of positive studies in systematic reviews might mean that such reviews are biased toward a positive result. Publication bias is just one in a group of related biases, all of which potentially lead to overrepresentation of significant or positive studies in systematic reviews. Other types of bias include time lag bias (positive studies are more likely to be published rapidly); multiple publication bias (positive studies are more likely to be published more than once); citation bias (positive studies are more likely to be cited by others); and language bias (positive studies are more likely to be published in English).

In PLoS Medicine, John Ioannidis and colleagues have taken a closer look at bias in Chinese genetics studies. Research done in non-English-speaking countries has two outlets. A study might be published in English-language journals, which are usually indexed in major international bibliographic databases such as PubMed, or in domestic journals, many of which are not indexed in international databases. The Chinese literature is a prominent example of where domestic scientific journals are not catalogued in international databases. There is some evidence that the decision to publish in international versus domestic journals might be influenced by the results. For example, significant results are often published in international journals, whereas nonsignificant results appear in the local literature, resulting in a language biasalthough, the reverse situation has also been described.

Genetics studies pose particular problems for impartial reporting. There are millions of polymorphisms in the human genome, and an exponentially increasing number of studies are trying to associate genetic polymorphisms with risk of disease or treatment outcomes. Selective publication might invalidate the overall picture of genetic risk factors.

The authors examined 13 genedisease associations. Studies were more likely to be published when the disease was considered common in China. They found 161 Chinese studies on 12 of these genedisease associations, only 20 of which were indexed in PubMed. Chinese studies had significantly more prominent genetic effects than non-Chinese studies, and 48% were statistically significant per se, despite their smaller sample size. Moreover, the largest, most exaggerated genetic effects were often seen in PubMed-indexed Chinese studies. Chinese studies usually appeared several years after their equivalent was first postulated in the world literature.

The larger genetic effects in Chinese studies are unlikely to reflect genuine heterogeneity and are more likely to do with publication bias operating within the Chinese literature, say the authors. It is possible that there was reluctance to submit and publish negative or inconclusive results when a large body of English-language literature has shown the presence of genetic effects. However, such forced confirmation negates the importance of independent confirmation of research results. This problem is probably not limited to the Chinese literature. These phenomena haven't been noted in molecular medicine before, but could become a serious problem in such a fast-moving field. Moreover, the inclusion of poor-quality research and additional selectively reported data may contaminate the better literature rather than provide a more accurate, comprehensive picture

The findings have two broad implications. First, language bias might be important to consider in meta-analyses of observational studies, where its effect might be larger than its effect on randomized evidence. Second, because human genome epidemiology is a global enterprise, a comprehensive global view is important to help decipher artifacts from true genetic effects. The Chinese literature in particular will be essential for the evaluation of evidence on genetic risk factors. China is making rapid scientific progress in this field and joining in international collaborative projects, such as the Human Genome Project. To develop a global perspective, one way forward might be for all investigators working on the genetics of a specific disease to register with a common network, making it easier to trace additional unpublished or nonindexed data.

Japanese haplotypes would help in identifying genes associated with disease and drug response

Sat, 29 Oct 2005 16:29:00 PST

( from http://www.rxpgnews.com ) Researchers at Kyushu University of Japan, in collaboration with Perlegen Sciences, Inc., have identified common patterns of human DNA sequence variation, or haplotypes, in the Japanese population. By combining a unique set of DNA samples collected at the Medical Institute of Bioregulation at Kyushu University with Perlegens high throughput, whole-genome analysis approach, based on next generation Affymetrix GeneChip ® technology, the collaborators identified common haplotypes that can be used to conduct comprehensive genetic research on human disease and variable response to medicines.

Understanding the patterns of genetic variation in the Japanese population will greatly enhance our ability to identify genes associated with disease and drug response stated Dr. Kenshi Hayashi, Professor of the Research Center for Genetic Information, Medical Institute of Bioregulation at Kyushu University. The unique approach of this study complements the recently completed International HapMap project. Both studies make critical contributions to enabling personalized medicine in Japan.

The collaboration between Kyushu University and Perlegen Sciences was particularly effective in identifying long-range haplotypes (over one million bases long). With this study, Dr. Hayashi and his colleagues at Kyushu University have made an important contribution to medical research in Japan, stated David Cox, Chief Scientific Officer of Perlegen. It will enable researchers to identify the genetic factors that determine why drugs work well for some patients, but not for others.

"Collaborating with the top researchers in Japan such as Dr. Hayashi is a priority for Perlegen, stated Akira Usui, General Manager of Perlegen Japan, KK. We are committed to improving the therapeutic treatments available to Japanese patients.

Kyushu University and Perlegen Sciences will report their findings in the October 26 issue of Genome Research.

Combination Model Controls Alternative mRNA Splicing

Wed, 26 Oct 2005 16:02:00 PST

( from http://www.rxpgnews.com ) In 1977, a flurry of papers ushered in a radical new concept in molecular biologyâthe idea of RNA splicing. It had been known for some years that the information for building organisms is stored as DNA sequences, which are transcribed into messenger RNAs (mRNAs) before translation into proteins. Although it had been established that the DNA and mRNA sequences line up exactly in bacteria, molecular biologists began to suspect in the mid-1970s that the genomes of eukaryotes (organisms with nuclei) are organized somewhat differently. Eukaryotic genes, it turns out, are encoded in small sections scattered over enormous distances of DNA. To make proteins from these âsplit genes,â the whole length of DNA is transcribed into pre-mRNA and then converted into mRNA by spliceosomesâmolecular machines that remove the non-coding pieces of RNA (the introns) and splice together the protein-coding pieces (the exons).

One important consequence of RNA splicing is that one gene can produce several different mRNA variations, or isoforms, simply by stitching together different combinations of exons. For example, a single gene in vertebrates encodes calcitonin (a thyroid hormone that controls calcium levels) and calcitonin-gene-related peptide (a neuropeptide). Alternative splicing also contributes to human diseaseâfor instance, the selection of different splice sites generates aberrant ratios of mRNA isoforms in several neurological diseases.

But how are these alternative splice sites selected? One popular model proposes that alternative splicing in mammalian cells is largely controlled by binding of general splicing factors to pre-mRNA molecules during the formation of the spliceosome. The spliceosome contains many of these factors, including a class of proteins called SR proteins, which contain one or two RNA-binding domains and a proteinâprotein interaction domain that is rich in serine and arginine amino acids. An important prediction of the combinatorial model for control of alternative splicing is that alternatively spliced transcripts will recruit different combinations of pre-mRNA splicing factors in vivo. New data from Mabon and Misteli support this prediction.

Pre-mRNA splicing factors accumulate at sites of active transcription, and splicing and can be detected and quantified in individual living cells by tagging the splicing factors with fluorescently labeled antibodies. So, to see whether different factors accumulate at alternatively spliced transcripts, the researchers developed stable cell lines carrying versions of the gene encoding a protein called tau designed to splice in different ways. In healthy people, exon 10 of the tau gene is included or excluded from tau mRNA with roughly equal probability during pre-mRNA splicing; in people with a rare Parkinsonism-like neurological disorder, mutations near one end of exon 10 result in its predominant inclusion. The researchers, therefore, examined cell lines carrying tau genes with and without a mutation of this type to change the ratios of mRNA transcripts including or excluding exon 10. Their results show that a subset of SR protein splicing factors is efficiently recruited to tau transcription sites that produce exon 10âcontaining mRNA, but less efficiently recruited to transcription sites where exon 10 is excluded.

These results provide the first in vivo evidence for the differential association of pre-mRNA splicing factors with alternatively spliced transcripts, and support a combinatorial mechanism for spliceosome formation. Exactly which splicing factors are recruited to each spliceosome would depend on both the concentration of each factor in individual cell types and the regulatory elements present in each pre-mRNA. The combinatorial mechanism for the control of alternative splicing, the authors suggest, could allow cells to adjust splicing outcome (and consequently which proteins they express) rapidly in response to intracellular or extracellular cues, as well as contributing to the generation of protein diversity. âJane Bradbury

Scientists find structure (Htz1 nucleosome) relevant to cell growth and cancer

Sat, 22 Oct 2005 02:40:00 PST

( from http://www.rxpgnews.com ) Researchers discovered a special type of molecular structure that helps keep genes properly turned off until the structure is ejected from those genes in a regulated manner to help turn the genes on.

The discovery is reported in the Oct. 21 issue of the journal Cell by scientists from the Huntsman Cancer Institute at the University of Utah.

In all organisms, the genome is split into chromosomes (compressed long strands of DNA) which are subdivided into functional DNA segments called genes. Genes function as the blueprints for building particular pieces of cellular machinery. However, different types of cells each require different types of cellular machinery, and must produce that machinery according to a biological timetable. A central issue in molecular biology is finding out how a cell regulates which genes are on, or active, and which genes are off, or repressed. This topic has direct relevance to human disease, as improper activation or repression of genes that regulate cellular growth is a common feature of cancer cells.

"We must understand how genes are activated or repressed in normal cells in order to understand how this process is misregulated in cancer cells," says Brad Cairns, Ph.D., lead scientist on the study and an investigator with Huntsman Cancer Institute. "We are beginning to understand how gene activation and repression is altered in cancer cells, and how that leads to tumor growth. However, the design of targeted treatments that can correct these alterations will require a deep knowledge of the basic cellular mechanisms that regulate gene expression."

The scientists studied a group of proteins known as histones, which form disk-like structures called nucleosomes when they are wrapped by genes. Under an electron microscope, the nucleosomes look like beads strung along the DNA strand. Normal nucleosomes block access to the cellular machinery that reads the blueprint stored in the gene, keeping the gene off or repressed.

Huntsman Cancer Institute investigators discovered that certain genes contain a special type of nucleosome bearing a protein called Htz1. This Htz1-containing nucleosome was shown to be "fragile," meaning it is ejected from the gene in a regulated manner, allowing reading of the gene's instructions by the cellular machinery. When the gene returns to its inactive or repressed state, the Htz1 nucleosome is reconstructed, again blocking the machinery from reading the gene.

Cairns, an associate professor in the Department of Oncological Sciences at the University of Utah School of Medicine and an investigator with the Howard Hughes Medical Institute, along with Huntsman Cancer Institute graduate students Haiying Zhang and Douglas N. Roberts, studied yeast cells to make the discovery.

"We and hundreds of other laboratories world-wide use yeast as a model system to study gene expression, as the analytical tools for studying yeast are actually more advanced than those available for human cells. However, all the factors that we study in yeast have virtually identical counterparts in human cells, so we fully expect the discovery to apply in humans as well," Cairns says.

Farnesyl Transferase Inhibitors in Hutchinson-Gilford progeria syndrome

Wed, 28 Sep 2005 13:20:00 PST

( from http://www.rxpgnews.com ) The new Hopkins research, and similar results from other labs, shows that a class of drugs known as farnesyl transferase inhibitors, or FTIs, can reverse an abnormality in laboratory-grown cells engineered to mimic cells from progeria patients. In the laboratory, however, treating these engineered cells with an FTI already in clinical trials in cancer patients restored the cells to a normal appearance, the researchers report Sept. 26 in the advance online section of the Proceedings of the National Academy of Sciences. The drug blocks the first step in processing the faulty protein that causes the syndrome.

"We've been hopeful that our two decades of research on how proteins are processed and modified in cells might ultimately help people with certain forms of cancer," says Susan Michaelis, Ph.D., professor of cell biology at Johns Hopkins' Institute for Basic Biomedical Sciences.

"But for progeria, we and others only recently learned that it involves the one of the modified proteins we've been studying, a nuclear protein called lamin A. As a basic scientist, it is really exciting to have leapfrogged from studying a fundamental process to finding evidence that an existing drug might be useful in treating a devastating disease in children," she says.

Michaelis emphasizes that no one knows whether making the cells' nuclei look normal will be enough to reverse the disease process or slow it down. The class of drugs they tested prevents the first step in cells' processing of certain critical proteins in yeast and mammals. For more than 20 years, Michaelis has been studying this complex process.

The process starts with a fully assembled protein, then adds a fatty appendage called farnesyl very close to the protein's end, and then a tiny modification called a methyl group to a nearby building block. In yeast, the protein that gets the full treatment helps the single-celled organisms reproduce -- and the useful protein is the smaller part with all the fancy modifications. In cells' processing of lamin A in mammals, however, the plain, big chunk is the active part, and it's critical for the proper function and organization of cells' nuclei.

In children with progeria, however, a genetic mutation causes a piece of the original lamin A protein to be deleted, a discovery made by National Institutes of Health researchers and reported in 2003. "The normal mammalian protein, lamin A, doesn't have all those modifications; the modified part is thrown away," says Michaelis. So Michaelis and postdoctoral fellow Monica Mallampalli, Ph.D., set out to test that idea. Mallampalli genetically engineered a human cell line (HeLa) to have either of two mutations in the gene for lamin A. One mutation halted the process at the very beginning, by preventing addition of the fatty farnesyl appendage. The other affected the end of the process by preventing cleavage of the otherwise normal, fully modified protein.

"Neither has the correct lamin A protein, but only one has a modified protein hanging around," says Michaelis. "We found that only the cells with the farnesyl-modified protein had the problems seen in cells with the HGPS mutation."
Mallampalli also altered the version of the gene that produces the abnormal, persistently modified, disease-causing protein, called progerin, to uncover the effect of preventing the addition of farnesyl. Sure enough, even though the cells still didn't have normal lamin A, their nuclei looked normal when the faulty protein couldn't get modified.

"We were thrilled that, as our genetic studies predicted, the experimental drug did the trick," says Michaelis. "Because FTIs are already in advanced clinical trials with cancer patients and seem to be quite well-tolerated, it's hopeful that they could be tested in patients with progeria fairly quickly."

FTIs prevent addition of farnesyl to all proteins that have a particular molecular tag. In cancer, the key target among these proteins is one called Ras, which is activated by the same farnesyl-triggered process as lamin A and which promotes cancerous growth when there's too much of it.

Gene Responsible for Chronic pain syndrome found

Tue, 27 Sep 2005 06:57:00 PST

( from http://www.rxpgnews.com ) In a significant advance toward understanding a perplexing and painful neurological disorder, an international team of researchers has discovered gene mutations associated with an inherited chronic pain and weakness syndrome known as hereditary neuralgic amyotrophy (also called HNA). No treatment is known for this disabling condition, which short-circuits a peripheral nerve center called the brachial plexus, a network of over 100,000 nerves, that branches from the spinal cord to supply muscular function and sensation to the shoulders, arms, and hands.

Episodes are often triggered by an infection, an immunization, childbirth, or overworking the arms and shoulders. Nerve inflammation and changes in the blood suggest that problems with the person's immune response are contributing to the episode. The on again/off again course of the condition, and the environmental triggers, are unusual among inherited nerve disorders.

An associated aspect of the disorder in some individuals is facial features -- a long, slender face and narrow, close-set eyes slanting upward -- reminiscent of portraits by the early 20th-century Italian painter Modigliani, according to Phillip F. Chance, MD, professor of pediatrics and neurology at the University of Washington in Seattle, whose laboratory first located the gene for this disorder to chromosome 17 in 1996.

Twenty-seven medical scientists at universities in Germany, Belgium, the United States, Finland, and Spain conducted the research to find the specific gene responsible for HNA. The lead authors of the study, which appears in the Sept. 25 edition of Nature Genetics, include Dr. Gregor Kuhlenbaumer of the University of Munster, Dr. Vincent Timmerman of the University of Antwerp, and Dr. Mark C. Hannibal and Dr. Phillip Chance, both from the Division of Genetics and Developmental Medicine at the University of Washington.
By studying several multigenerational families who had several relatives with HNA, the researchers identified mutations in a gene named septin-9 ( known as SEPT9). Cells from a variety of life forms, ranging from yeast to fruit flies to humans, contain septins. Septins form protein filaments that provide the internal scaffolding of cells, and play key roles in the process by which cells divide. Cells depleted of SEPT9 often fail to complete normal cell division. According to the authors of the SEPT9 gene mutations study, SEPT9 has particular structures that distinguish it from all other septins, but the significance and function of these structures is as yet unknown. Future research on the SEPT9 gene and its mutations may lead to a better understanding of the normal function of the gene and its protein products.

Clioquinol, an antibiotic shows new promise for Huntington's Disease

Mon, 12 Sep 2005 18:14:00 PST

( from http://www.rxpgnews.com ) Clioquinol, an antibiotic that was banned for internal use in the United States in 1971 but is still used in topical applications, appears to block the genetic action of Huntington's disease in mice and in cell culture, according to a study reported by San Francisco VA Medical Center (SFVAMC) researchers.

The study, led by principal investigator Stephen M. Massa, MD, PhD, a neurologist at SFVAMC, was reported in the August 16, 2005 issue of Proceedings of the National Academy of Sciences.

Huntington's disease is a hereditary, degenerative, and ultimately fatal disease of the brain that causes changes in personality, progressive loss of memory and cognitive ability, and a characteristic uncontrolled jerking motion known as Huntington's chorea. There is no known cure or effective treatment. A person who carries the mutant Huntington's gene may pass it on unknowingly because the disease often manifests in early to late middle age after the carrier's children have already been born.

During the course of the disease, the Huntington's gene causes the production of a toxic protein, mutant huntingtin, in neurons (brain cells). Eventually the protein kills the neurons, causing the disease's degenerative effects.

In Massa's study, Clioquinol appeared to interrupt the production of mutant huntingtin. In the first part of his study, Massa and his research team tested the effect of Clioquinol on neurons in cell culture that contained a form of the mutant Huntington's gene. "We found that not only did cells look better and survive a bit longer when exposed to the drug, but they also seemed to make less of the toxic protein," observed Massa, who is also a clinical assistant professor of neurology at the University of California, San Francisco (UCSF).

Based on the in vitro results, Massa decided to test the drug in vivo, on mice bred to express the toxic huntingtin protein. The mice were given approximately 1 milligram of Clioquinol per day in water. After eight weeks of treatment, they had accumulated four times less toxic protein in their brains than control mice given water alone. The experimental animals lived 20 percent longer than the control animals, did better on tests of motor coordination, and had less weight loss.

"It's a limited study, in that we used the same drug dose on all the animals as opposed to comparing different doses, but fairly convincing," Massa concluded. "Together, the in vitro and in vivo results suggest that Clioquinol has an effect of decreasing the symptoms of Huntington's, its pathology, and perhaps even the actual production of the toxic protein."

However, he noted, "the drug's mechanism of action remains unclear." The clearer the mechanism of the drug, he explained, the better the chance that researchers might eventually be able to create a medication that is both safe and effective.

Like some other antibiotics, Clioquinol is known to be a chelator -- that is, it binds metals in body tissues, particularly copper and zinc, and removes them when it is excreted. Massa and other researchers believe that this chelation effect may interfere with production of the mutant huntingtin protein in some way. "But there are still a couple of explanations we need to rule out," he said.

To that end, Massa's next studies will involve the creation of an in vitro system in which toxic and non-toxic forms of huntingtin are made in the same cell. He and his team will then evaluate the effects of Clioquinol on several phases of protein synthesis within the cell. Massa hopes these experiments will confirm initial indications that Clioquinol preferentially interferes with synthesis of the toxic form of the protein. "Then we can move on to trying to isolate the actual mechanism of the drug," he predicted.

"However," Massa cautioned, "the record of successfully translating drugs from animal to human use is not good."

Clioquinol has shown promise as a potential treatment for Alzheimer's disease in recent studies in mice and humans. Apparently through chelation, it interferes with the creation of beta-amyloid plaque in the brain, which has been implicated in the progression of Alzheimer's symptoms.

Currently, Clioquinol is banned for internal use in many countries because of its side effects. In Japan in the late 1950s and 60s, the drug was found to cause a neurologic condition called subacute myelo-optico-neuropathy (SMON), with symptoms including visual loss, muscle weakness, and numbness, in several thousand people. However, noted Massa, the doses given in current clinical trials are much smaller than were commonly prescribed in Japan. In addition, he explained, it has been found that vitamin B12, when taken along with the drug, protects against its potential toxic effects.

Chimp Genome Offer Clues To Human Diseases

Fri, 09 Sep 2005 17:52:00 PST

( from http://www.rxpgnews.com ) The recently published Chimp Genome Sequencing project highlights the similarities between humans and our closest genetic cousins, the great apes. But a researcher at UCSD suggests it is in the differences, rather than the similarities, that clues to understanding human disease might be found.

Two upcoming papers co-authored by Ajit Varki, M.D., Professor of Medicine at UCSD, describe genetic differences between the species: one, the discovery of the first human-specific protein that is also expressed in brain cells associated with human brain diseases; the second, a single oxygen atom difference that makes humans and chimpanzees resistant to each others malarial parasites.

This research provides examples of how studying the evolution of humans and apes from a common ancestor may yield clues to explaining human and chimpanzee diseases. Chimpanzees have long been thought of as a model for studying human diseases said Varki. In fact, what is most remarkable is that many of our diseases are rather different, either in incidence or in severity. Focusing on understanding these differences will eventually benefit both humans and chimpanzees.

Varki and colleagues at UCSDs Glycobiology Research and Training Center detail their finding of the first truly human-specific protein in the September 9 issue of Science.

Siglecs are molecules that serve as binding receptors for sialic acids, which are sugars found on the surface of all higher animal cells. Comparing human and chimpanzee genome sequences, the researchers noted that a human gene they had recently discovered called Siglec-11, was actually human-specific. The gene was generated by a mechanism called gene conversion that occurred in the human lineage sometime after our common ancestor with chimpanzees.

Nissi Varki, a professor of pathology at UCSD, found that while the protein encoded by the gene was expressed in human brain cells called microglia, it was not expressed in the brains of chimpanzees and other apes. Whats interesting is that microglia are also involved in Alzheimers disease, multiple sclerosis and HIV-induced dementia, conditions that have so far not been reported in chimpanzees said Varki, adding This study raises more interesting questions than answers.

By defining this human-specific genetic change, scientists may eventually better understand such neurological disorders, and possibly why humans brains are different from those of apes. The next step for the Varkis and their team is to explore the functional consequences and the mechanisms of brain expression. Causing Siglec-11 to be expressed in a mouse brains might help. Meanwhile, another scientist might find a person with a neurological disease, and discover that patient has a genetic mutation of Siglec-11, helping us understand the functions of this gene in the normal human brain, he said.

It has long been known that chimpanzees dont get sick from the human malaria parasite Plasmodium falciparum, nor are humans infected with the malarial species that affects chimpanzees, called Plasmodium reichenowi. But until now, the reason for this surprising difference has been a mystery.

Just as humans and chimps have been shown to be very close genetic cousins, the two malaria parasites are genetically very similar. It is these two seemingly coincidental, but surprising, similarities that piqued the interest of Varki, and UCSD colleague Pascal Gagneux, Ph.D., a scientist in the Department of Cellular and Molecular Medicine who is also affiliated with the Zoological Society of San Diego,

A paper to be published in the September 6 issue of Proceedings of the National Academy of Sciences and currently on line, shows that this mystery can be explained by a genetic change that occurred in our ancestors about three million years ago. In earlier work, Varki, Gagneux and colleagues had found that humans were genetically unable to make one kind of common sialic acid called Neu5Gc, a molecule commonly found in apes and many other mammals (see earlier press release) They have now found that this difference of a single oxygen atom on sialic acids can shed light on the puzzling discrepancy in malaria susceptibility between the two species.

To find the difference between the two malarial parasites, the researchers focused on how each invaded their target red blood cells. Both Plasmodium species uses molecular hooks on their surfaces to latch onto the sialic acids on the red blood cell.

The researchers detected differences in the red cell binding capabilities between the two malaria species that could be explained by the differences in sialic acids. Thus, they argue that in the course of evolution, humans first became resistant to the malaria parasite infecting great apes by loss of the target molecule Neu5Gc. However, in the bargain they gained an excess of another sialic acid called Neu5Ac, eventually facilitating the evolution of P. falciparum, a parasite that now causes more than 1.5 million deaths a year in humans.

Cloning after Dolly - Professor Ian Wilmut Talks

Tue, 06 Sep 2005 00:09:00 PST

( from http://www.rxpgnews.com ) The leader of the team that produced Dolly the sheep, the first animal to develop after nuclear transfer from an adult cell, will present a talk at the University of Glasgow on Wednesday 7 September about the future of this area of research.

Professor Ian Wilmut from the University of Edinburgh, set up a research group to determine the molecular mechanisms that are important for normal development of cloned embryos and to use that knowledge in biology, medicine and agriculture.

Over the past nine years, research has been focused on the factors regulating embryo development after nuclear transfer. The work led to the first birth of live lambs from embryo-derived cells and then to the birth of lambs derived from foetal and adult cells, including Dolly. Subsequently, genetic changes were introduced into sheep by nuclear transfer from cultured modified cells.

A considerable improvement in efficiency is required before wide scale use for livestock. The opportunity to introduce precise genetic changes to livestock is available for the first time through the use of gene targeting procedures in cultured cells that are used as nuclear donors. The research has the potential application in the production of organs for transplantation to humans, studies of human genetic disease and basic research in to the control of gene expression and function.

Scientists get look at genes defensive playbook

Tue, 06 Sep 2005 00:01:00 PST

( from http://www.rxpgnews.com ) Using a new method to identify networks of infection-fighting genes, scientists writing in todays (8-31) online edition of Nature say more than 15 percent of our genes are mobilized to defend against microbial attacks.

The bodys overwhelming genetic defense, which has implications for the survival of patients who are severely burned or injured, was revealed in a sweeping analysis of gene activity in volunteers who were injected with a bacterial product that temporarily created flu-like symptoms.

During a 24-hour period, the expression of more than 3,700 genes changed in blood leukocytes, said Lyle Moldawer, Ph.D., a surgery professor in the University of Florida College of Medicine, part of the national consortium that published the findings. It was a dramatic reprioritization of genes. But beyond individual genes, we were able to look at networks, or functional modules of different gene clusters, that change in concordance with one another. We have now identified previously unknown relationships among different genes that tell us in greater detail how blood cells respond to an infectious challenge.

Inflammation is part of normal healing when people are severely burned or injured, but in some patients, it can be fatal, causing bloodstream infections and multiple organ failure. Learning how and why inflammation becomes harmful will help doctors more accurately predict how each injured patient will fare.

This work represents a major step in understanding inflammation in severely injured or burned patients, said Jeremy M. Berg, Ph.D., director of the National Institute of General Medical Sciences, the component of the National Institutes of Health that funded the research. We hope this knowledge eventually will help physicians better predict patient outcomes and tailor treatments accordingly.

UF Genetics Institute researchers are part of a national group of scientists united by a five-year, $37 million glue grant from the NIGMS. Glue grants bring together scientists from diverse fields in this case surgery, critical care medicine, genomics, bioinformatics, immunology and computational biology to solve problems in biomedical science that no single laboratory could address.

Scientists injected healthy volunteers with a microbial product that temporarily causes nausea and fever, triggering natural immune responses.

The condition is similar to sepsis, which can happen when the bodys infection-fighting white blood cells spring into action, causing potentially harmful inflammation in the process.

Basically we made the volunteers appear septic for a couple of hours and examined changes in the gene expression from their white blood cells, Moldawer said. Such genomic analyses give us the ability to simultaneously survey the activity of every gene in the cell, giving us vast lists of genes that change in response to stimulation. It provides us with an unprecedented amount of data.

To make sense of the enormous amount of information, researchers plugged their list of nearly 4,000 gene changes into a database of interactions of known human and mouse genes developed by Ingenuity Systems Inc. of Mountain View, Calif. The results identified the networks of genes that helped the body respond to the challenge.

We were able to identify changes in functions that we never would have seen before, Moldawer said. For example, the ability of the infection-fighting cells to make energy appeared to be down-regulated, as if the cells were shutting down all other functions not required to rid the body of the bacteria. This may well be the signal that something is wrong with the cell and may be a reason why some patients who are injured or infected go on to develop organ failure.

With that knowledge, scientists may be able to look at new ways to re-establish stability within the cells and avert the negative consequences of infection fighting.

The apparent repression of genes that occurs has never been fully appreciated, said Henry Baker, Ph.D., associate director of the UF Genetics Institute and director of the UF lab that performs genomic analyses for the consortium.

Initially, more than half of the genes became less active, but over the long haul, they were more focused on the inflammatory response. By drawing samples for analysis over six time points in 24 hours, we were able to infer the sequence of events and how some changes in gene expression cause other changes.

Additional genomic analysis took place at the Stanford Genome Technology Center in Palo Alto, Calif., and the department of surgery at Washington University in St. Louis, Mo. The research is particularly valuable because it plots inflammatory response over time, according to Scott D. Somers, Ph.D., NIGMS program director of this glue grant.

In the case of injury, time is critical, Somers said. To provide the best treatment, doctors need to know how the human body responds in the moments and days after an injury. No other study of injury or inflammation has tracked changes to the entire human genome over time.

New method identified the specific gene involved with high cholesterol

Sun, 04 Sep 2005 10:08:00 PST

( from http://www.rxpgnews.com ) Until now, the genetic causes of diseases such as cancer, heart disease, and diabetes, have proven very elusive. A new approach, reported in Elsevier's journal Genomics, may soon put an end to this. Scientists have successfully used this method to identify the specific gene involved with high cholesterol.

The genetics of some human diseases can be quite easily defined: a simple mutation in a single gene causes it to malfunction or be inactivated. The gene's mutation can then be shown to be the direct cause of the disease. Unfortunately, however, the genetic basis of some of the deadliest human diseases is far more complex than this often involving subtle changes to several genes on different chromosomes.

A recent paper published by Dr. Alessandra Cervino and colleagues in the journal Genomics describes a new method for identifying the elusive genes involved in such complex diseases. Using a mouse model system for human diseases, scientists have successfully used this new method to identify a gene called Insig2, which is directly involved in the control of blood cholesterol levels. This gene has been demonstrably linked with the causes of obesity, diabetes and atherosclerosis in mice. As Dr. Eric Schadt, of Rosetta Inpharmatics, LLC, a subsidiary of Merck & Co. remarked, "Using this new method we have identified a gene involved with high blood cholesterol in mice, and we fully expect the human equivalent to be just as significant. This opens new doors in the potential treatment or prevention of cardiovascular disease in humans."

This new method is also directly applicable to many other diseases in human beings. According to Dr. Mark S. Boguski, Editor-in-Chief of Genomics, "The potential to use this approach to address the causes of serious human diseases is enormous. This might well be a landmark publication."

A Cobweb of Life?

Wed, 31 Aug 2005 02:16:00 PST

( from http://www.rxpgnews.com ) The tree of life has long served as a useful tool for describing the history and relationships of organisms over evolutionary time. One species is represented as a branching point, or node, on the tree, and the branches represent paths of descent from a parental node. The tree diagram carries an implicit assumption that genes are transferred vertically, from parent to child, and that all the genes in a new species come from the ancestral species. In theory, one should be able to trace the origin of each gene in a species back to its ancestor. In practice, however, the ancestral gene is rarely available, so researchers look for the gene in a closely related species. (These similar genes, which diverge slightly after a speciation event, are called orthologs.)

But as the tools of genome analysis became more refined, searches for similar genes sometimes turned up sequences that belonged to a species on a different branch of the evolutionary tree. Clearly, vertical gene transfer was not the only mechanism of genetic transmission. Organisms, it turns out, can acquire genes from non-ancestral species through a mechanism called horizontal gene transfer (HGT)think of it as acquiring genes from your neighbor instead of your parents. Such genetic exchanges, most common among bacteria and other microbes, are not represented in the tree of lifeno single branch connects the two unrelated species. Initial studies suggested that HGT events were extremely common, prompting some to say it was time to replace the tree with a netlike diagram. Other studies have since suggested that methods used to calculate HGT overestimated its frequency: researchers detect HGT events by finding inconsistencies between gene trees and organism, or whole-genome, trees, but statistical errors can artificially increase the number of HGT events.

In a new study, Fan Ge, Li-San Wang, and Junhyong Kim estimate the frequency of HGT events by using a novel statistical method to compare the gene trees and whole-genome trees of microbes. Their method solves the statistical problem by directly testing for discrepancies between trees that arise from statistical error versus true HGT events. Analyzing over 40 microbial genomes, Kim and colleagues estimate that HGT infiltrates just 2% of the average microbial genome. Even when relatively common, the authors conclude, HGT events do not disrupt the integrity of the tree of life, contributing just small bits of genetic material, much like cobwebs on tree branches.

To construct both gene trees and a whole-genome tree for the microbes, the authors selected core sets of orthologous gene groups from the NIH database of clusters of orthologous genes. (Clusters are derived by comparing protein sequences encoded in complete genomes, which represent major lineages on the evolutionary tree. Each cluster corresponds to an ancient, conserved protein domain.) Kim and colleagues created gene trees for each cluster of orthologous genes they selected, then created whole-genome trees from the gene trees and compared each gene tree to the whole-genome tree, using their new method. HGT events were detected when two species appeared close together on a gene tree but far apart on the whole-genome tree. Overall, just over 11% of the orthologous gene clusters showed statistically significant HGT events, with HGTs accounting for about 2% on average of each of the 40 microbial genomes.

Altogether, these results suggest that HGT is not as common as once thought. And even when large-scale HGT events do occurwhich Kim simulated in a previous studythey do not obscure the evolutionary path of most genes and lineages. If you imagine a tree with 10,000 taxa, the authors explain, and 1,000 HGTs per genome across all the taxa, the HGTs would form extremely thin connections like cobwebs, leaving the backbone of the tree intact. Infrequent though it may be, HGT likely has some impact on the evolutionary history of lifeimpacts that advances in genome analysis technology may help uncover. Liza Gross

Improved Statistical Tools Reveal Many Linked DNA Loci

Wed, 31 Aug 2005 02:06:00 PST

( from http://www.rxpgnews.com ) Using traditional statistical tools to analyze the modern wealth of biological data is a bit like trying to move a muscle car with a buggy whipyou're not likely to get anywhere very fast. The problem is perhaps most acute in the quest to understand how genes interact to regulate one another's expression. The amount of RNA made by any one gene is likely influenced by DNA at dozens of loci, or locations around the genome. Such loci are often situated within genes that participate in the same pathway as the gene being influenced, and a central goal is to understand this network of mutually influential genes and loci. Consider piecing together this puzzle for each of the many thousands of genes and many thousands of potentially influential loci, and the old analytical tools simply can't keep up. In this issue of PLoS Biology, John Storey and colleagues tackle the challenge with a new approach.

The authors began by mating two strains of yeast that have minor differences in their DNA at more than 3,000 locicreating over 3,000 markersand then tracking the inheritance of these markers in the yeast offspring. Because the two genomes randomly reshuffle upon mating, any single offspring will contain some random combination of marker outcomes from each parent. The authors also examined the amount of RNA produced by over 6,000 individual genes in each offspring. The next step was to determine how these two large sets of datavariations at specific loci and variations in expression of specific geneswere correlated.

Straightforward statistical tests performed on each gene's expression revealed the single most influential location in the DNA. But such tests don't reveal the more complicated reality that any single gene is likely to be influenced by more than one locus. Linking expression of a single gene (or expression trait, in genetic parlance) to more than one locus has been stymied by the inability of conventional statistical approaches to cope with the mountains of data involved. Not only is an exhaustive pair-by-pair testing of all possible interactions computationally demanding, but it can also be very difficult to distinguish whether elevated expression is due to one or both of the loci being tested. The problem becomes exponentially harder as more potentially linked loci are tested.

To overcome the limitations of standard approaches, Storey et al. used a novel statistical approach that exploited what they had the most ofdata. They began by determining the single most significant locus for each expression trait. They then moved on to the next most significant locus for that trait, but tested its linkage (that is, its influence on expression) with the assumption that the first locus was also linked. The ability to assign a probability of linkage to the first locus greatly simplified the calculations for the subsequent locus, reducing by almost a thousand-fold the number of possibilities that needed to be tested.

As opposed to standard methods, the authors show that their approach is able to assess true joint linkage of two loci to an expression trait, while requiring substantially less computation. In addition, they found that about one in seven expression traits is controlled by epistatic, or hierarchical, relationships among the two loci, while the standard method revealed none. This method can be adapted to search for even larger numbers of linked loci, to provide insights into the many interlocking pathways that make up the gene regulatory network, and ultimately result in the organism itself.

Anti-cancer drugs might work in aging disease

Tue, 30 Aug 2005 19:45:00 PST

( from http://www.rxpgnews.com ) Working together, scientists at the National Institutes of Health and the University of North Carolina at Chapel Hill have developed a promising new strategy for treating a form of progeria. That rare but deadly and heartbreaking genetic disease causes children to age remarkably fast and die almost always before they complete their teens.

The average lifespan of victims, who eventually resemble very old bald people or, some might say, Hollywood's conception of space aliens, is 12 years.

Along with their staffs and students, Dr. Francis S. Collins, director of the National Human Genome Research Institute, has collaborated with Drs. Channing J. Der and Adrienne D. Cox of the UNC School of Medicine in laboratory studies.

They have shown that certain anti-cancer drugs known as FTIs can block some of the complex biochemical processes that result in progeria's symptoms. The collaboration came about because the UNC scientists have been working on the drugs for more than a decade, and Collins' group has been actively investigating progeria of childhood, which also is known as Hutchinson-Gilford syndrome.

The potential treatment has not been used with patients yet but had a strong positive effect on progeria patients' cells, they said.

Respectively, Cox and Der are associate professor of radiation oncology and pharmacology and professor of pharmacology and members of UNC's Lineberger Comprehensive Cancer Center. Collins, a UNC medical graduate, attracted widespread attention in 1989 as the discoverer of the defective gene that causes cystic fibrosis, another fatal illness that afflicts children.

A report on the research appears in the Sept. 6 issue of the Proceedings of the National Academy of Sciences. Other authors of the report include M.D.-Ph.D. student Brian C. Capell of Collins' NIH laboratory; NIH staff members Drs. Michael R. Erdos, Renee Varga and Leslie B. Gordon (also medical director of the Progeria Research Foundation); doctoral student James P. Madigan of Cox's laboratory; Dr. James Fiordalisi, assistant professor of radiation oncology at UNC; and Dr. Karen N. Conneely of the University of Michigan School of Public Health.

"Fortunately, progeria is very rare, and only about one child in four million comes down with it," Der said. "It was first identified in the early 1900s. Since then only 100 or so cases have been found. Still, it is quite devastating for those children who have it and their families."

He and Cox concentrate on FTIs, or farnesyltransferase inhibitors, which block the action of the enzyme farnsyltransferase. Currently, several chemical variations are undergoing clinical trials with cancer patients.

"There's a lot of interest in FTIs now because they target an enzyme that's required for a protein called RAS to cause cancer," Der said. "The idea that these FTIs also might be useful in treating progeria came up because it turns out that the gene that is mutated in that rare illness also requires this enzyme for generating an active protein known as lamin A."

In progeria patients, he said, the process that results in the normal, mature form of lamin A doesn't work correctly because of the genetic mutation, so a damaged form of lamin A is made instead. Researchers reasoned that the anti-cancer drugs might block the enzyme and hence interfere with the mutated lamin A gene's haywire actions.

"In the paper, we describe experiments showing that the mutant form of lamin A was indeed sensitive to these drugs," Der said. "The second thing we found was that some of the aberrant biology that this mutant protein causes can be stopped when we treat the cells with the inhibitors."

Because the drugs already are undergoing clinical trials and much is known about their action and safety, scientists have a significant head start in getting the possible treatment to patients, Cox said.

"We are very excited about the possibility that a drug class whose actions we have been working so hard to understand in cancer might soon be useful for this devastating 'orphan disease,'" she said. "Progeria is clearly an illness that would otherwise get no attention from pharmaceutical companies due to the tiny numbers of children afflicted."

The next step will be to test the enzyme inhibitors in mouse models of the disease which already have been made, Cox said. If those experiments succeed, then scientists could start clinical trials with patients.

Lamin research project provides clues about premature aging

Tue, 30 Aug 2005 19:42:00 PST

( from http://www.rxpgnews.com ) A step towards understanding cell mutations that cause a variety of human diseases, particularly in children -- including that which brings about premature aging and early death -- has been taken by researchers at the Hebrew University of Jerusalem Silberman Institute of Life Sciences and the John Hopkins University School of Medicine.

The scientists have focused their research on a study of induced mutations in the nuclear envelope of cells from the tiny C. elegans worm. Their aim is to thus provide clues towards a better understanding of mutations in proteins of the envelope of the cell nucleus in humans.

Such mutations, particularly in lamin (nuclear envelope) proteins A and C, cause many different diseases, including Hutchison Gilford progeria syndrome. Children with this disease develop premature aging and die usually before the age of 13. Other diseases brought about by these mutations include a form of muscular dystrophy, cardiomyopathy (a weakening of the heart muscle), and various other forms of irregular or retarded growth in childhood.

A report on the lamin research project was published in a recent issue of the Proceedings of the National Academy of Sciences in the U.S. The project was carried out primarily by Ayelet Margalit, a doctoral student in genetics at the Hebrew University, working under the supervision of Prof. Yosef Gruenbaum, and in cooperation with Prof. Katherine L. Wilson and Dr. Miriam Segura-Totten of Johns Hopkins University.

Experimenting with removal of the worm's lamin protein or its interacting protein partners emerin, MAN1 or BAF, the researchers have described "down-the-line" consequences, including the disruption of various proteins necessary for normal cell reproduction. Even though the C. elegans worm has only one lamin protein and few proteins that interact with it, the processes that occur there are similar to what happens in humans and provide clues to the laminopathic diseases affecting people..

The results seen from these lamin complex disruptions are a halted process of cell division, resulting in a static "bridge" structure between cells that should have separated, plus damage to the gonad cell structure. In both cases, the ability of the organism to grow and to reproduce is severely impaired.

The researchers hope that through further laboratory experimentation with the worm they will be able to better understand the functions of lamin-based complexes, and why mutations in these proteins cause a variety of different laminopathic diseases, such as progeria and muscular dystrophy in humans.

Drug prevents cell abnormality leading to progeria

Tue, 30 Aug 2005 19:39:00 PST

( from http://www.rxpgnews.com ) BACKGROUND: One in 4 million children are born with progeria, a genetic disease marked by accelerated aging and early cardiovascular disease. The children suffer from dwarfism, baldness, wrinkles, hardened arteries and osteoporosis. Most die from heart disease before age 15.

The rare disorder stems from a mutation in a gene that produces an abnormal cellular protein, which attaches itself to structures in the cell's nucleus. The accumulated protein deforms the nucleus, sparking miscommunications with other cells and leading to the genetic disease.

FINDINGS: UCLA scientists studied cells isolated from people with progeria and cultured the cells with a drug that blocked the mutant protein from attaching to the cells' nuclei. The drug significantly reduced the number of human cells with misshapen nuclei. The UCLA team earlier used the same approach to improve the shape of abnormal nuclei from mice genetically engineered to develop progeria.

IMPACT: The findings offer new clues into how progeria develops and could lead to new drugs to treat the disease and its related disorders, including osteoporosis and hardening of the arteries. UCLA's next step will be to test whether returning the shape of the nuclei to normal stops development of progeria in mice.

Farnesyltransferase inhibitors (FTIs) might be useful in Hutchinson-Gilford Progeria Syndrome

Tue, 30 Aug 2005 19:33:00 PST

( from http://www.rxpgnews.com ) In a surprising development, a research team led by the National Human Genome Research Institute (NHGRI), part of the National Institutes of Health (NIH), has found that a class of experimental anti-cancer drugs also shows promise in laboratory studies for treating a fatal genetic disorder that causes premature aging.

In a study published Monday in the online edition of the Proceedings of the National Academy of Sciences (PNAS), Brian Capell and his colleagues at NHGRI reported that drugs known as farnesyltransferase inhibitors (FTIs), which are currently being tested in people with myeloid leukemia, neurofibromatosis and other conditions, might also provide a potential therapy for children suffering from Hutchinson-Gilford Progeria Syndrome, commonly referred to as progeria. A related study from Stephen Young, M.D., and colleagues at the University of California at Los Angeles is being published in the same issue of PNAS.

There are currently no treatments for progeria, which is a genetic disorder estimated to affect one child in 4 million. When they are born, children with progeria appear normal. But, as they grow older, they experience growth retardation and show dramatically accelerated symptoms of aging -- namely hair loss, skin wrinkling and fat loss. Accelerated cardiovascular disease also ensues, typically causing death from heart attack or stroke at about the age of 12.

"Our findings show that FTIs, originally developed for cancer, are capable of reversing the dramatic nuclear structure abnormalities that are the hallmark of cells from children with progeria. This is a stunning surprise, rather like finding out that the key to your house also works in the ignition of your car," said NHGRI Director Francis S. Collins, M.D., Ph.D., who is the study's senior author.

The new work involved using FTIs to treat skin cells taken from progeria patients and grown in laboratory conditions. If upcoming studies in a mouse model validate the results of the cell experiments and translate into improvements in the animals' conditions, a clinical trial of FTIs in children with progeria may begin as early as next spring, researchers said.

Dr. Collins and his colleagues discovered in April 2003 that mutations in the lamin A (LMNA) gene cause progeria, spurring renewed interest among researchers to study this rare syndrome. Among those were Capell, a New York University medical student participating in the Howard Hughes Medical Institute/NIH (HHMI/NIH) Research Scholars Program. In July 2004, he joined Dr. Collins' lab and immediately set his sights on understanding the molecular basis of progeria.

"What really interested me in this research in the first place were the potential links to aging and atherosclerotic disease," said Capell. Indeed, understanding progeria at the molecular level may illuminate the general processes involved in normal human aging.

The LMNA gene codes for a protein called lamin A, which constitutes a major component of the scaffold-like network of proteins just inside the cell's nuclear membrane, called the lamina. The gene mutation implicated in progeria causes a section of 50 amino acids within the lamin A protein to be deleted, resulting in a mutated protein that is called progerin. This protein fails to integrate properly into the lamina, thereby disrupting the nuclear scaffolding and causing gross disfigurement of the nucleus. Cells with progerin have a nucleus with a characteristic "blebbed," or lobular, shape.

To find its way to the lamina, lamin A carries two tags, rather like ZIP codes, that help to direct the protein's travels. One tag at the end of lamin A instructs another protein to modify it through a process called farnesylation. Farnesylation tethers lamin A to the inner nuclear membrane. Once there, a second tag within the protein signals an enzyme to cleave off the terminal portion of the protein, including the farnesyl group, freeing lamin A to integrate properly into the nuclear lamina.

Because progerin carries the farnesylation tag but lacks the second cleavage tag, Capell speculated that progerin was becoming permanently stuck to the inner nuclear membrane. There, he suspected, it enmeshed other scaffolding proteins, preventing their proper integration into the lamina. If progerin's tendency to stick to the inner nuclear membrane is indeed the culprit in nuclear blebbing and the root of the progeria defect, Capell and his colleagues reasoned that they could prevent these defects by blocking farnesylation of progerin.

The researchers' hunch proved correct. When they changed one amino acid within progerin's farnesylation tag to prevent the addition of a farnesyl group and tested the effect in cells grown in the laboratory, progerin did not anchor itself to the inner nuclear membrane and instead clumped within the nucleus. Moreover, they observed no nuclear blebbing.

The researchers then tried treating the cells carrying progerin with FTIs, which are drugs originally developed to inhibit certain cancer-causing proteins that require farnesylation for function. FTIs are now being tested in phase III clinical trials of patients with myeloid leukemia. So far, clinical trials using FTIs have found little toxicity, even when the drug treatment significantly raises levels of unfarnesylated proteins.

After FTI treatment, the progerin-carrying cells showed no blebbing. More importantly, researchers saw the same effect when they used FTIs to treat cells grown from skin biopsies of progeria patients: Cell blebbing decreased to near normal levels.

Cell-Autonomous Death of Cerebellar Purkinje Neurons with Autophagy in Niemann-Pick Type C Disease

Mon, 25 Jul 2005 17:13:00 PST

( from http://www.rxpgnews.com ) Niemann-Pick disease type C is a deadly neurodegenerative disease that is most often due to mutations in a gene called npc1. As a consequence of intracellular lipid trafficking defects, patients with Niemann-Pick type C, and mice with the same disease, lose an important class of cerebellar neurons called Purkinje cells (PCs).

Npc1 (the protein coded by npc1) might be needed in other cell types to produce substances that nourish PCs or within the PCs themselves. To see which is true, the researchers constructed genetically mosaic mice in which some cells have mutant Npc1 and some have normal Npc1 function.

In the cerebella of these mosaic mice, PCs lacking Npc1 continued to die even while surrounded by normal cells, while normal PCs appeared unaffected by their partially mutant surroundings. From these findings, the researchers concluded that the neurodegeneration is due to a problem within PCs and not due to a lack of supporting factors provided by other cells or an extrinsic toxic or inflammatory insult. Npc1 probably functions within PCs to allow critical transport processes necessary for cell survival.

The researchers also found that the degenerating PCs undergo a complex process called autophagy in which the cells sense a lack of key nutrients and start to break down their own structures to feed themselves.

By identifying exactly which cells require Npc1 function, the researchers set the stage for investigating the exact molecular roles of Npc1 protein in the cells where it is most needed.

A Human-Curated Annotation of the Candida albicans Genome

Mon, 25 Jul 2005 17:13:00 PST

( from http://www.rxpgnews.com ) Candida albicans is a commonly encountered fungal pathogen usually responsible for superficial infections (thrush and vaginitis). However, an estimated 30% of severe fungal infections, most due to Candida, result in death. Those who are most at risk include individuals taking immune-suppressive drugs following organ transplantation, people with HIV infection, premature infants, and cancer patients undergoing chemotherapy.

Current therapies for this pathogen are made more difficult by the significant secondary effects of anti-fungal drugs that target proteins that are also found in the human host.

Recent sequencing and assembly of the genome for the fungal pathogen C. albicans used simple automated procedures for the identification of putative genes. Here, we report a detailed annotation of the 6,354 genes that are present in the genome sequence of this organism, essentially writing the dictionary of the C. albicans genome.

Comparison with other fungal genomes permitted the identification of numerous fungus-specific genes that are absent from the human genome and whose products might be targeted for antifungal therapy. The results of these efforts will thus ensure that the Candida research community has uniform and comprehensive genomic information for medical research, for the development of functional genomic tools as well as for future diagnostic and therapeutic applications.

Lorenzo's oil (LO) reduced the risk of developing severe X-linked adrenoleukodystrophy (ALD)

Tue, 12 Jul 2005 12:26:00 PST

( from http://www.rxpgnews.com ) Treatment of boys with X-linked adrenoleukodystrophy (ALD) with Lorenzo's oil (LO) reduced their risk of developing the severe debilitating form of the disease, according to a study in the July issue of the Archives of Neurology, one of the JAMA/Archives journals.

Individuals with ALD accumulate high levels of saturated very long-chain fatty acids (VLCFA) in their brains. The course of the disease results in a number of different manifestations [phenotypes], according to background information in the article. The rapidly progressive cerebral ALD (CERALD) type typically begins between ages four and eight and progresses rapidly to total disability within a few years. An adult form is non-inflammatory, progresses slowly and is far less disabling. Children who do not develop abnormalities as measured by magnetic resonance imaging (MRIs) by age seven or clinical symptoms by age 10, have greatly diminished risk of developing cerebral ALD.

In 1989, one of the authors of this study, Augusto Odone, pioneered a treatment (Lorenzo's oil), which was shown to normalize the levels of saturated very long-chain fatty acids within four weeks in most patients with ALD. "The striking effect of LO on plasma C26:0 [a saturated very long-chain fatty acid] levels engendered the hope that it would be of clinical benefit for patients with ALD," the authors write. However, previous clinical trials led to the conclusion that Lorenzo's oil did not alter the rate of progression of the disease in patients who already had neurological symptoms.

Hugo W. Moser, M.D., of the Kennedy Kreiger Institute, Baltimore, and colleagues treated 89 boys with ALD who had no neurological symptoms and normal brain MRIs with moderate dietary fat restriction and Lorenzo's oil between 1989 and 2002. Sixty-four of the patients were younger than seven years old when they began treatment and all were followed up for an average of approximately seven years. Because of the devastating nature of cerebral ALD, and the hope that the striking reduction of very long chain fatty acid levels would lead to clinical benefit, none of the boys were given placebo. Fatty acids blood levels were assessed every month for the first six months after enrollment in the study and every three to six months thereafter. Neurological examinations and MRIs were scheduled every six to 12 months.

Sixty-six patients (74 percent) were well at last follow-up. Twenty-one patients (24 percent) developed MRI abnormalities and 10 patients (11 percent) developed neurological abnormalities. The researchers found a significant association between the development of MRI abnormalities and an increase in the levels of the saturated very long chain fatty acid C26:0. "Patients who had a neurological abnormality had significantly higher weighted average C26:0 levels than those who did not have an abnormality, suggesting that an LO-induced decrease in the C26:0 level could protect against the inflammatory cerebral disease," the authors report.

"We recommend that LO therapy be offered to male patients with ALD who are neurologically asymptomatic, have normal brain MRI results, and are at risk of developing CERALD," the authors conclude. "This recommendation is based on strongly suggestive, albeit not fully definitive, evidence of a preventive effect combined with our awareness of the severe prognosis of the untreated patients with CERALD. ...The patients who are younger than seven years represent prime candidates for this therapy. We hypothesize that intensive LO therapy during the ages at which the risk for CERALD is greatest may protect against this phenotype until they reach the ages at which the risk for CERALD diminishes."

In an editorial accompanying the article, Raymond Ferri, M.D., Ph.D. and Phillip F. Chance, M.D., of the University of Washington, Seattle, write "In recent years, extraordinary progress has also been made in developing effective treatments, and ALD serves as an excellent model for the treatment of neurometabolic diseases. ...Current treatment includes hematopoietic stem cell transplantation (HSCT) to stabilize neurologic progression, steroid therapy for adrenal insufficiency, and symptomatic treatments. The article by Moser et al in this issue may establish new standards for the treatment of this degenerative disorder."

"Therefore, Moser and colleagues propose LO therapy for all asymptomatic patients with biochemical evidence of ALD to slow the progression of disease and to prevent symptoms until the child is past the age for the development of the childhood cerebral form of the disorder," the authors write. "Successful implementation of this practice requires early identification of at-risk patients. However, because almost 20 percent of the patients are either asymptomatic or have Addison disease only, at-risk children may not be identified. As also mentioned in the article, neonatal screening would identify more at-risk patients at a very early age. This would allow for further studies to examine very early treatment with LO for affected children, and dietary therapy can be studied in other ALD phenotypes [manifestations]. Also, this study can be extended to follow patients for an even greater duration to establish the full treatment effects of LO."

"X-linked ALD is a rare, progressive neurometabolic disorder, but coordinated, worldwide research efforts have made it a treatable disease," the authors conclude. "Dietary therapy started early in life and HSCT have markedly improved the longevity and quality of life for affected people, and new standards for treatment have been established."

Genes play an overriding role in cholesterol response

Sun, 10 Jul 2005 15:11:00 PST

( from http://www.rxpgnews.com ) Why does it seem like some people can eat all the ice cream they want without increasing their cholesterol or gaining much weight, while others with high cholesterol have to watch their diets like a hawk? Because no matter what their lifestyle, people's genes play an overriding role in their cholesterol response.

So says a new study by researchers at the Department of Energy's Lawrence Berkeley National Laboratory and the Children's Hospital Oakland Research Institute (CHORI), conducted by Paul Williams of Berkeley Lab's Life Sciences Division in collaboration with Robin Rawlings and Patricia Blanche of CHORI and Ronald M. Krauss of CHORI and Berkeley Lab's Genomics Division. They report their findings in the July 8, 2005, issue of the American Journal of Clinical Nutrition.

The investigators analyzed how "bad" cholesterol (low-density lipoprotein, or LDL, cholesterol) responded to diets that were either high or low in fat in 28 pairs of identical male twins one twin a vigorous exerciser, the other a comparative couch potato.

"Although identical twins share exactly the same genes, we chose these twins because they had very different lifestyles," says Williams. "One member of each pair was a regular long-distance runner, someone we contacted through Runner's World magazine or at races around the country. His brother clocked 40 kilometers a week less, at least, if he exercised at all."

For six weeks the twins ate either a high-fat diet (40 percent of its calories from fat) or a low-fat diet (only 20 percent of its calories from fat); then the pairs switched diets for another six weeks. After each six-week period the twins' blood cholesterol levels were tested.

The researchers were interested in learning if blood cholesterol changes due to the different diets would be the same or different in each pair of genetically identical twins, even though their lifestyles were very different. A correlation of zero between the two would mean that their responses to the diets had no relation to each other, while a correlation of 1.0 would mean that their responses were identical.

The researchers found an astounding 0.7 correlation in responses to the change in diet, an incredibly strong similarity in the way each pair of twins responded even though the responses themselves among different pairs of twins differed considerably.

"If one of the twins could eat a high-fat diet without increasing his bad cholesterol, then so could his brother," says Williams. "But if one of the twins' LDL cholesterol shot up when they went on the high-fat diet, his brother's did too."

The correlations showed that the twins had very similar changes in LDL cholesterol because they had the same genes. Some twins had one or more genes that made them very sensitive to the amount of fat in their diets. Other twins had genes that made them insensitive to dietary fat, no matter how much they exercised.

"Our experiment shows how important our genes are," says Williams. "Some people have to be careful about their diets, while others have much more freedom in their dietary choices."

He adds, "This type of experiment allows us to test whether genes are important without having to identify the specific genes involved." Although several specific genes have been associated with cholesterol changes in response to changes in diet, these cannot account for the large correlations seen in this study. Williams hopes his findings will inspire additional research to identify the specific genes involved.

"Concordant lipoprotein and weight responses to dietary fat change in identical twins with divergent exercise levels," by Paul T. Williams, Patricia J. Blanche, Robin Rawlings, and Ronald M. Krauss, appears in the July 8, 2005, issue of the American Journal of Clinical Nutrition. The work was supported by Dairy Management Incorporated, with additional support from the National Institutes of Health.

Master switch in cell death discovered

Fri, 01 Jul 2005 12:52:00 PST

( from http://www.rxpgnews.com ) Researchers at UT Southwestern Medical Center have found an enzyme vital for controlling the early stages of cell death - a beneficial and normal process when it works right, but malignant in a variety of cancers when it malfunctions.

The researchers are now examining tissue from cancer patients to try to determine how mutations in the enzyme's gene may relate to cancer.

"We think this gene will really be a hot spot in research," said Dr. Qing Zhong, postdoctoral researcher in biochemistry at UT Southwestern and lead author of a paper to be published in the July 1 issue of the journal Cell.

The life and death of cells is a complex avalanche of reactions, controlled by a few molecules that sit atop a biochemical "pyramid."

The newly discovered enzyme, which the researchers have named Mule, destroys a key molecule at the top of the pyramid, thus leading to the cascading disintegration of the cell. Their findings also suggest a new drug target for controlling tumor formation.

Dr. Xiaodong Wang, professor of biochemistry at UT Southwestern and a researcher with the Howard Hughes Medical Institute, said the discovery of Mule will open up a whole field of research to study the enzyme's role in normal cell death and cancer.

"We think these findings are very significant," said Dr. Wang, senior author of the Cell study. "This is the first enzymatic step that regulates the degradation of proteins that control cell death."

The beneficial side of cell death - known as apoptosis - occurs when it kills cells at appropriate times, as is the case, for example, when it removes the webbing from the fingers of an embryo or shapes a developing brain. But the darker side of this complex process manifests itself in cancers when cells don't die when they're supposed to.

The key to the researchers' finding was the interaction between the Mule enzyme and a major player in cell death, the protein Mcl-1. Dr. Wang said that while there are many possible routes a cell may take toward apoptosis, this interaction serves as one of the "master switches" controlling whether or not those other pathways are triggered.

Normally, Mcl-1 keeps cells alive by protecting them against apoptosis. For a cell to die, Mcl-1 has to be disabled. "It's just like a guardian," Dr. Zhong said.

A healthy organism needs just the right amount of Mcl-1. Too little Mcl-1 can lead to a damaged immune system or even death. Too much, and cells stay alive when they shouldn't, leading to cancers such as lymphomas.

Using human cell extracts, the researchers found that Mule caused a protein called ubiquitin to bind to several sites on Mcl-1. When ubiquitin binds to a molecule, it serves as a flag for that molecule to be destroyed.

"If you have too much Mule in a cell, Mcl-1 will degrade tremendously," Dr. Zhong said.

The search for Mule took more than two years, as the UT Southwestern researchers specifically searched for an enzyme that controls Mcl-1.

The interaction between Mule and Mcl-1 might someday be manipulated to help cancer patients, Dr. Wang said. For instance, a tumor may contain cells with a deficit of Mule, making the tumor more likely to grow and perhaps be resistant to chemotherapy. Treatment might then focus on the biochemistry of Mule and Mcl-1, he said.

"We might be able to see if there's a problem with Mule, or perhaps we could screen beforehand," Dr. Wang said.

New Gene Regulation System Revealed

Wed, 22 Jun 2005 12:50:00 PST

( from http://www.rxpgnews.com ) By comparing 140 sequenced bacterial genomes, researchers have uncovered a system for regulating genes essential to bacterial replication - and they did it solely by computer keystrokes and mouse clicks.

Mikhail Gelfand, a Howard Hughes Medical Institute international research scholar at the Institute for Information Transmission Problems (IITP) in Moscow, and his postdoctoral fellow, Dmitry Rodionov, used comparative genomics to identify a new transcription factor system in bacteria that represses expression of genes involved in DNA replication. They scanned gene sequences and proteomes of several taxonomic groups of bacteria, identifying not only a highly conserved signal sequence, but also the regulatory transcription factor that bound it, the repressor nature of the signal, and other genes also regulated by this system.

We provided a very detailed description of a system just by doing bioinformatics alone, says Gelfand, director of the IITP's research and training center of bioinformatics. It's a proof of principle that you can go a very long way by comparative genomics now. Their findings will be published in the July issue of Trends in Genetics, with early publication now online. Gelfand is presenting the work on June 24, 2005, at the annual meeting of HHMI international research scholars in Mérida, Mexico.

Gelfand and Rodionov started their search using a technique called phylogenetic footprinting to review the upstream DNA sequences of a group of genes that code for ribonucleotide reductase enzymes. These enzymes convert the ribonucleotide building blocks of RNA into the deoxyribonucleotides used to build DNA. This conversion is critical for duplicating a bacterium's entire genome before it divides to reproduce.

The search revealed a conserved palindromic sequence occurring upstream of many ribonucleotide reductase (Nrd) genes. A genetic palindrome is a sequence of nucleotides on one strand of DNA that reads the same as the sequence on the opposite strand, only backwards - a common feature of DNA sequences that are recognized by regulatory molecules. They designated the sequence the NrdR-box.

Because the signal was found in so many diverse groups of bacteria, the researchers thought it might represent a universal regulatory mechanism. The next question was whether the signal was promoting or repressing expression of Nrd genes.

The team observed that their signal always overlapped with the promoter signal, the region of DNA required for the initiation of the conversion of gene to protein. Molecules that promote transcription recognize and bind to this sequence, which lies just outside of the gene. Repressor signals commonly work by allowing other proteins to bind on top of the promoter sequence and physically block promoters. Therefore, the duo predicted that the NrdR-box functioned as a repressor sequence.

Next, the researchers identified the transcription factor protein that binds to the NrdR-box. To do this, they used a bioinformatics approach they call phylogenetic profiling, compiling a list of genomes that clearly contained the NrdR-box and those that clearly did not have it. Then they searched the proteomes of 63 bacteria species, looking for proteins that strictly followed the same present-or-absent pattern as the NrdR-box. Only one protein cluster matched the pattern, and it represented a family of proteins that shared traits of transcription factors.

To strengthen the prediction that these proteins were the transcription factors that bind the NrdR-box, the team used another comparative genomic tool called positional clustering. Positional clustering takes advantage of the fact that functionally related gene sequences (such as the genes for Nrd and its transcription factor) frequently inhabit the same `neighborhood' of the chromosome.

If you are looking in one genome, many genes will be neighbors by coincidence, Gelfand noted. But if two genes are neighbors in many diverse genomes, then they are likely to be related. Indeed, the Nrd genes and the transcription factor genes clustered together, providing additional evidence that the regulatory picture drawn by the team was correct.

Israeli researchers simultaneously showed through `wet' biology experiments in Streptomyces bacteria that a transcription factor from this family represses Nrd gene expression in the living bacterial cell, confirming the Russian researchers' predictions. Confident that they had identified a new repressor of bacterial genes, Gelfand and Rodionov searched genomes for other upstream sites where the NrdR-box occurred. They found that it regulates other genes related to DNA replication, such as the enzymes that cut, paste, and untangle new DNA as it is synthesized, and enzymes that are involved in recycling nucleotide building blocks.

Although the work does not have direct application to human medicine, Gelfand pointed out that many antibiotics work by attacking the process of bacterial DNA replication. So, he said, this work has identified potential targets for designing new antibiotic drugs. But more importantly, the work shows how molecular discoveries of whole regulatory systems can be made through careful analysis of genomeswithout ever lifting a pipette, he said.

There are 100 enzymes functioning at the core of bacterial metabolism for which the genes are still unknown, said Gelfand. Using multiple bioinformatics tools can uncover cell systems that might have escaped experimental detection, he suggested. By comparing hundreds of genomes, you can see patterns that are not seen when looking at just a couple of them.

Lack of coherent cloning policies reflects polarized debate

Fri, 20 May 2005 02:31:00 PST

( from http://www.rxpgnews.com ) The confusing welter of state laws regarding human cloning for reproductive purposes and for research uses reflects a national political impasse on regulating cloning, according to a new report by The Genetics & Public Policy Center, a project of The Pew Charitable Trusts and Johns Hopkins University. This lack of a national consensus comes at a time when rapid advances in cloning technology make crafting broader public policy increasingly urgent, the report notes. "While human cloning technology is still in its infancy, the science is outpacing the public's understanding and the formulation of coherent public policy," it warns.

"Scientists have cloned cows, cats, and human embryos," says Kathy Hudson, director of the Center. "Meanwhile, the public and policymakers have reached a political impasse we're embroiled in a complex and divisive ethical and policy debate that too often is rushed and emotionally charged. We hope this report will contribute to public understanding and to the development of sound science policy."

The Center's report, "Cloning: A Policy Analysis" is the most recent comprehensive analysis of the science of cloning (including stem cell research), the moral and ethical arguments that frame the cloning debate, and rapidly changing policies at the state, federal, and international levels. The report also details new public opinion data that reflect high awareness about cloning in general but limited understanding of its scientific feasibility.

RAPIDLY EVOLVING SCIENCE LANDSCAPE

Somatic cell nuclear transfer (SCNT) is the technique currently in use to create embryos that are genetically identical to another animal or person, the report explains. Research cloning has been used in the laboratory to derive human stem cells from cloned embryos; their nuclear genome is identical to that of the source of the donated nucleus. Therapeutic cloning refers to the potential use of stem cells from cloned embryos to treat degenerative diseases through the transplantation of genetically matched cells or tissues; although this has not to anyone's knowledge yet been successfully attempted in humans. Reproductive cloning has been demonstrated in animals, Dolly the sheep in 1996 being the most publicized example. Claims of human reproductive cloning are made periodically but none have been substantiated, the report notes.

Studies have shown that many cloned animals die in utero or soon after birth, or survive with severe birth defects. In fact, few cloning attempts are successful. Abnormalities are wide-ranging and can include heart, liver, kidney, immune system, and brain defects. Using a nucleus from an adult cell requires it to be "reprogrammed" into behaving like the nucleus of a very early embryo, meaning genes that were turned on in the adult need to be turned off and genes that are required for embryonic development need to be turned on. Currently, it is not clear to what extent reprogramming errors lead to SCNT failure; more animal research will be needed to better understand these and similar cloning problems, the report says.

Cloned non-human primate embryos have been obtained using SCNT, the report notes, but so far no pregnancy has been reported. Cloned primate embryos have exhibited misaligned chromosomes and the absence of structures necessary for cell division, but some scientists argue that these are technical barriers that can be overcome, the report observes.

The report notes that many scientists and patients, among others, support the use of cloning because of its potential to better understand human development and treat disease, and a small minority of the public also would support its use to help couples who otherwise could not to bear genetically related children. At the same time, some fear "a Brave New World-like civilization in which people intentionally are designed for the use and control of those more powerful," it explains. Some oppose cloning because it requires destruction of human embryos, while others view the cloning of a human being as an ill-conceived attempt to usurp Divine authority or second-guess Nature, the report explains.

These divergent views in turn are based on different underlying values, including the view of the moral worth of a human embryo, conceptions of human personhood and human dignity, the importance of human individuality, the imperative to heal the sick, the right to reproductive autonomy, and the proper role of government in socially charged and ethically complex issues, the report points out.

NATIONAL GRIDLOCK; STATES CHARTING NEW TERRITORY

"These widely disparate concerns, fears, and hopes have created a political impasse at the federal level," it explains; no federal laws have been passed regulating cloning, although Congress has considered numerous bills in every session since 1997. "Congress currently is gridlocked on cloning issues," says the report's principal analyst, Gail Javitt. "The controversy about human embryo research, on the one hand, and the specter of human reproductive cloning, on the other, have kept the cloning debate an emotionally charged and highly controversial fixture of the political landscape for the past several years."

However, at the state level -- as well as in other countries -- numerous laws have been passed either banning or promoting certain uses of cloning. These existing regulations, as well as policy statements by many scientific and advocacy organizations, are catalogued in the report.

The report notes that five U.S. states ban all forms of cloning outright, four expressly permit SCNT for research or therapeutic purposes while banning reproductive cloning, and three restrict the use of state funds for research or therapeutic cloning. Several other states have existing laws pertaining to the use of embryos in research that may impact regulation of cloning. By contrast, a few states -- notably California -- have passed laws that actively promote stem cell research. "It's a real-life example of our federalist system in action -- the states are, literally, serving as the 'laboratories' of democracy," explains Javitt. "At the same time, lack of federal involvement could leave gaps in oversight."

DIVERSITY OF PUBLIC OPINION ON CLONING

The report notes that public opinion has been tapped many times by groups either for or against cloning, often with very different outcomes. "Wording really does matter," Hudson says. "You can get a very different answer depending on how you ask the question, what kind of promises you imply about health benefits when you ask the question, or how you phrase the source of the embryonic material to be used." Poll results also can be skewed by recent media coverage or political activities, the report notes.

That's why, Hudson explains, a recent Gallup poll could show only 38 percent of respondents supportive of cloning embryos specifically for research, while a different recent survey -- when the word "cloning" is not used in survey questions, but the alternative term "somatic-cell nuclear transfer" is used and medical value of the procedure is stressed -- found public support for research cloning at 72 percent.

In the Center's own 2004 survey looking at a variety of reproductive genetic technologies, Americans opposed human cloning for both reproductive and research uses by margins of more than 3-1, the report explains. The survey, conducted by Knowledge Networks, Inc., also found that most Americans appear to have a very poor understanding of the science surrounding cloning technologies, noting that:

* Overall, only 56 percent of those surveyed correctly responded yes when asked whether it was "scientifically possible today to produce a cloned embryo." Roughly a third reported they did not know if it was possible, while slightly fewer than 10 percent indicated they believed it was not possible.
* When asked whether it was "scientifically possible today to produce a cloned human baby," only 18 percent correctly reported that it was not possible, while 38 percent indicated they didn't know. Slightly more than 45 percent said it was possible to produce a cloned human baby.

Regarding research cloning, the Center's survey also found that:

* 76 percent of Americans surveyed did not approve of "scientists working on ways to create a cloned human embryo for research."
* More adults over age 50 disapproved of scientists working to create a cloned human embryo for research (81 percent) compared to adults aged 18-29 (71 percent)
* Forty-two percent of religiously unaffiliated respondents approved of scientists working to create a cloned human embryo for research versus only 7 percent of Fundamentalist or Evangelical Christians
* Approval of research cloning was greater among participants with higher levels of education

On issues related to reproductive cloning, the Center found that:

* Eighty-eight percent disapproved of "scientists working on ways to create a cloned human baby"
* Twice as many men as women approved of cloning to create a human baby (16 percent versus fewer than 8 percent). In addition, men also were twice as likely as women to answer yes when asked "if you could clone yourself, would you?"
* Fewer Evangelical or Fundamentalist Christians (4 percent) said they approved of scientists working on ways to clone a human baby compared to other religions and those with no affiliation

Of the minority of respondents who answered yes when asked whether human cloning --whether for research or reproduction -- "should be allowed at all," 85 percent stated that the government should regulate cloning "based on quality and safety," and 54 percent responded that the government should regulate cloning "based on ethics and morality."

The report also concludes that Americans don't form their opinions about cloning in a vacuum. "American's opinions about cloning are not firmly held and likely are being influenced by their positions on more familiar issues such as abortion and the value of biomedical research to develop new therapies and treatments for the sick," the report says. "Given this situation, it is not surprising that lawmakers in Washington and in various state legislatures have not been able to reach consensus on laws to regulate cloning, or how cloning ultimately might be used in medicine."

Exercise Training in Ordinary People Activates 500 Genes

Tue, 03 May 2005 13:21:00 PST

( from http://www.rxpgnews.com ) A new study from Karolinska Institutet in Stockholm shows that hundreds of genes in the thigh muscle are activated in regular cycle training. The study also reveals that great differences in training response may be due to the ability in some people to activate their genes much more forcefully.

It is common knowledge that it is very dangerous to be inactive and that regular physical activity brings health, improves quality of life and extends life span. How these positive effects are created in the body is not known. Influences on gene activity in the heart, vessels and muscles are probably immensely important.

In this study, the first of its kind, Drs James Timmons, Carl J Sundberg and co-workers show that hundreds of genes are activated by regular cycle training for six weeks in young healthy men. Some of these genes are most likely linked to diabetes and cardiovascular disease. These training study findings can therefore be important for the development of new treatment strategies for such diseases.

Some people respond more easily to training than others. It is not known what explains this. The results from the training study show that those individuals that improved their performance most also activated several genes in the muscles markedly more. This has not been shown before.

Finally, the researchers made a comparison between the effects of endurance training and the situation in patients with Duchennes muscle dystrophy, a muscle wasting disease. Most of the muscle genes previously claimed to be specific for Duchenne were also activated with endurance training. Maybe the musculature in Duchenne patients strive to adapt in part similar to what happens in training. The results from this study will help to clarify which genes are uniquely affected in Duchenne.

A 3D Map of Human Chromosomes

Wed, 27 Apr 2005 02:27:00 PST

( from http://www.rxpgnews.com ) On this, theologians, philosophers, and biologists can agree: we are more than the sum of our genes. Biological complexity arises not from gene number but from patterns of gene expression, which change under the direction of both genetic and so-called epigenetic mechanisms. Epigenetics, broadly defined, concerns heritable changes in gene function that dont involve changes in DNA sequence. Until recently, studies of heritable traits have focused largely on mutations in DNA. But its become increasingly clear that how DNA is packaged in the nucleus also impacts heritability.

Epigenetic changes are mediated largely by proteins that shape and remodel chromatinthe association of DNA and histone proteins that condenses the genome into compact bundles inside the nucleus. Different cell types have different chromatin arrangements during development and cell differentiation that appear to regulate gene expression, which possibly accounts for the unique gene expression patterns associated with specific cell types. Such phenomena have been well-studied for specific genes or chromosomal regions, but to understand the full impact of epigenetic mechanisms on gene regulation, we need a more panoramic view of gene organization within the nucleus.

In a new study, Thomas Cremer together with Andreas Bolzer and an interdisciplinary team of German physicists, bioinformaticians, and geneticists created 3D positional maps of each human chromosome simultaneously in a single nucleus to investigate the link between chromatin structure and cell-specific gene expression. Working with human fibroblasts, cultured from a skin biopsy from a two-year-old boy, the authors were able to visualize and study the order of the full genetic complement within a human nucleus.

Cremer and colleagues first produced a 3D topological map of all 46 chromosomes in different cell types at key points in the cell cyclea landmark achievementusing a fluorescent staining technique that preserves chromosome shape during visual inspection under the microscope. Next, they established that small chromosomes in quiescent (nondividing) fibroblasts hewed close to the center of the nucleus while the large chromosomes were preferentially found at the nuclear rim, regardless of their gene density. Nuclei from cells entering the prometaphase stage of the cell cyclejust before chromosomes are aligned along the center of the nucleus prior to segregationrevealed a size-correlated chromosomal distribution akin to that seen in the quiescent nuclei. Statistical modeling analyses indicated that these size correlations do not simply reflect the geometric constraints of fitting into the nucleus, but likely hint at some degree of functional order within the nucleus.

Because previous studies of cells with sphere-like nuclei correlated chromosomal arrangements with gene density, the authors investigated how shape affects chromosome position along the nuclear radius. Fibroblast nuclei are somewhat flat and ellipsoidal. Chromosomes in similarly shaped amniotic fluid cells assumed the same size-related positions taken by chromosomes in fibroblast nuclei. But when the authors examined the higher-order chromatin arrangements in fibroblasts and lymphocytes, they found that, even though the cell types differ in nuclear shape and radial chromosomal arrangements, they both show a nonrandom higher-order chromatin architecture correlated with gene density. Many questions remain concerning the functional and physiological significance of these observations: Do shape changes produce changes in chromosomal arrangements and vice versa? Do shape changes produce changes in gene expression patterns?

Cremer and colleagues conclude that, although nonrandom chromosome positions occur, these appear to be governed by a degree of uncertainty and more likely reflect probabilistic preferences inside the nucleus. Still, deterministic mechanisms in higher-order chromatin structure may existsequestering gene-rich chromatin areas in the nuclear interior, for example, protected from malevolent agents entering the nucleus. And given the coexistence of size-correlated features with gene-density-correlated features seen in this study, it may well be that both random and deterministic factors combine to create the nuclear landscape.

An Enzyme That Oversees RNA Quality Control

Tue, 19 Apr 2005 17:12:00 PST

( from http://www.rxpgnews.com ) The path from DNA to RNA to protein sounds straightforward enough, but few processes in the life of a cell could be called simple. Gene expression is a complex, highly regulated affair that involves the activity of discrete teams of molecular manipulators at key steps in the protein production pathway. As soon as transcription begins, RNA processing machinery sets to work on messenger RNA (mRNA) precursors in the nucleus, proofreading the RNA copy, stabilizing the elongating transcripts, and making sure mRNAs reach the translation machinery in the cytoplasm.

A key step in RNA processing involves the addition of a string of adenine nucleotides to the 3' end of the growing transcript, a modification called polyadenylation. (Each RNA chain has whats called a 5' end and a 3' end, which relates to the chemical polarity of the nucleotides.) In eukaryotes like yeast and humans, polyadenylation helps to stabilize the mRNA transcript and to ensure its export from the nucleus. Once in the cytoplasm, the mRNAs poly(A) tails interact with components of the translation apparatus and facilitate protein synthesis. Polyadenylation is mediated by a class of enzymes called poly(A) polymerases (PAPs), which typically act in concert with other proteins in the cells nucleus.

Just three years ago, a new class of PAPs was discovered that belong to the Trf4/5 family of yeast proteins and are found in the cytoplasm, rather than the nucleus, of the cell. Unlike the nuclear PAPs, in which one protein contains both catalytic activity and an RNA-binding domain, the new class relies on two or more protein subunits to carry out these tasks. Its been suggested that these enzymes stabilize specific mRNAs in the cytoplasm by extending their poly(A) tails. Its long been known that adding poly(A) tails to RNAs in prokaryotic bacteria promotes the degradation of defective RNAs, but the existence of a similar mechanism in yeast has just recently come to light. Previous genetic experiments on live yeast cells by James Anderson and collaborators suggested that when the protein Trf4p polyadenylated a particular type of abnormal transfer RNA (tRNA)the intermediary that translates the nucleotide code into the amino acid codethe tRNA was destroyed. In a new study, Walter Keller and colleagues investigate the biochemistry, composition, and function of Trf4p in the brewers yeast Saccharomyces cerevisiae, and find evidence that Trf4p-mediated polyadenylation plays a role in RNA quality control in eukaryotes.

After showing that mutating two amino acid residues in the predicted catalytic center of Trf4p eliminates its activity, Keller and coworkers determined that the enzyme forms a PAP complex with three other proteins (including two putative RNA-binding proteins, Air1p and Air2p, and a putative RNA-unwinding enzyme called Mtr4p that most likely functions to unwind structured regions in the RNAs). They also showed that the Trf4p complex selectively targets and successfully polyadenylated only tRNA molecules that were either lacking the chemical modifications required for normal folding or that were misfolded by mutation. Polyadenylation appears to tag the aberrant RNA as damaged goods, signaling the cells nuclear molecule-degradation complex, the exosome, to initiate destruction.

Keller and colleagues propose that the Trf4p complex recognizes structural defects in RNA, prompting the Trf4p subunit to add poly(A) tails to the RNA, which initiates RNA degradation. In this model, Trf4p, along with either Air1p or Air2p, interacts with the RNA enzyme Mtr4p, which physically connects the tRNA-Trf4-PAP complex to the exosome. These results suggest that Trf4-PAP monitors the quality of tRNAs by detecting misfolded RNAs and engineering their destruction before they can gum up the works of protein assembly.

That the polyadenylation pathway for discarding defective tRNA appears in both bacteria and yeast suggests that this quality-control mechanism represents the ancient role for polyadenylation, the authors propose; the stabilization function of adding poly(A) tails may have arisen as eukaryotes evolved a nucleus and other organelles. Whether this notion or the model described here proves correct remains to be seen, and the authors outline a number of avenues for further study. Determining the structure of RNAprotein complexes, for example, and their binding properties and interactions, the authors argue, should elucidate the mechanism by which this RNA surveillance complex operates and what features of its RNA substrates it recognizes.

Exon Silencing Regulated by a Trio of Short RNA Motifs

Tue, 19 Apr 2005 17:12:00 PST

( from http://www.rxpgnews.com ) Our cells make many more kinds of proteins than can be accounted for by the relatively modest number of genes in our genome. The key to this protein-coding bounty is alternative splicing, in which one or more transcribed exonsnucleotide sequences that code for a specific segment of the proteinare excluded from the final messenger RNA before it is translated into protein. While the majority of human genes are alternatively spliced, little is known about specific RNA sequences that dictate exclusion of these exons. In a new study, Paula Grabowski and colleagues show that three short sequences, two within the excluded exon and one in an adjacent intron, or non-coding nucleotide sequence, trigger exclusion in at least one gene, and probably a large handful of others as well.

Grabowski and colleagues studied this process in a class of proteins essential for brain function called glutamate receptors. As the name implies the glutamate receptors bind to glutamate, the principal excitatory neurotransmitter in the brain. NMDA glutamate receptors, which play a role in memory formation and neuronal development, are composed of multiple subunits. Within the NR1 subunit, exclusion or inclusion of the CI cassette exon has dramatic functional consequences. The CI exon appears in the forebrain but is virtually absent in the hindbrain. How this differential splicing is regulated is poorly understood.

The authors noted an atypical but highly conserved GGGG motif in the intron just downstream from the splice site that ends the CI exon. When they introduced point mutations in this motif, the exon was included up to four times as often. The rate of exclusion, or silencing, could be dramatically increased by the addition of another GGGG tetrad farther inside the intron.

Systematic mutation within the exon identified a pair of UAGG motifs that also promoted exon silencing, an effect that could be enhanced even further by introducing a third, artificial, UAGG. The pair of UAGG tetrads appears to work in combination with the GGGG tetrad, since without the former sequences, the latter had little power to silence CI expression. Silencing is mediated by binding of UAGG to the ribonucleoprotein hnRNP A1, which also apparently interacts with the GGGG within the intron.

The authors next did a series of genomic database searches, to identify these motifs in other genes. They reasoned that if the triad was a common means of exon silencing, it should be overrepresented among genes known to undergo alternative splicing. In more than 90,000 exons in human and mouse genomes, they found 16 with the motif pattern, of which three (19%) were known skipped exons. In contrast, among those without the pattern, the proportion of skipped exons was only 5%. They also found that the GGGG motif by itself was overrepresented among skipped exons, indicating it probably plays a significant role in exon exclusion even without its UAGG partners.

These results alone cannot explain why one cell type includes an exon while another excludes it, since the primary transcript in different cell types is the same. Instead, these differences are likely explained by tissue-specific differences in levels of splicing factors or binding proteins. With such small absolute gene numbers, it is clear that the specific trio identified by Grabowski and colleagues is only one of many likely to regulate exon inclusion. In the search for others, this study indicates the value of bioinformatics strategies that employ not only specific sequences, but also spatial configurations.

Understanding the interaction of Fragile X mental retardation protein and kissing complex RNAs

Mon, 18 Apr 2005 04:57:00 PST

( from http://www.rxpgnews.com ) Fragile X syndrome is the most common inherited form of mental retardation, affecting approximately 1 in 3600 males and 1 in 4000-6000 females. Fragile X syndrome results from loss of expression of the Fragile X mental retardation protein (FMRP), the product of the FMR1 gene. Now, Drs. Robert and Jennifer Darnell and colleagues, from The Rockefeller University, report the uncovering of a new interaction between FMRP and messenger RNAs (mRNAs) containing a tertiary RNA structure termed a "kissing complex".

Their studies, published in the April 15th issue of Genes & Development, provide a new direction for efforts to understand how the loss of FMRP function leads to the complex behavioral and cognitive defects characteristic of Fragile X syndrome.

While the importance of identifying a function for FMRP has been clear for some time, what this function actually is has continued to evade researchers. FMRP is a protein characterized by the presence of three RNA binding domains: two tandem KH-type RNA binding domains and an RGG box. Scientists have focused on the identification of FMRP RNA ligands in an effort to understand FMRP function. This effort is particularly meaningful since FMRP is believed to regulate mRNA translation in the brain, and identifying the mRNA targets of this regulation would be a huge step in understanding how loss of this protein results in the varied and complex phenotypes of Fragile X syndrome.

In most Fragile X patients, loss of FMRP is due to silencing of FMR1 resulting from the unusual amplification of a CGG repeat (over 200 copies in affected patients versus less than 60 copies in unaffected individuals) that leads to hypermethylation of FMR1 and shut down of transcription of the gene. However, Fragile X patients expressing mutations or deletions within the FMR1 gene have also been described, including a severely affected patient harboring a missense mutation that resulted in a one amino acid change, isoleucine at position 304 for asparagine, in one of the KH domains of FMRP, KH2.

Dr. Darnell and colleagues focused on understanding how this specific mutation leads to loss of FMRP function. They first screened an RNA library to identify what RNA motif is recognized by the KH2 domain. They found that the KH2 domain of FMRP recognizes a loop-loop pseudoknot, or "kissing complex" structure in the RNA, and that this recognition is abrogated by the isoleucine to asparagine mutation. Notably, they show that the association of FMRP with the translation machinery (in brain polyribosomes) can be competed out with kissing complex RNA, an important finding since previous biochemical studies have reported altered polyribosome distribution of mRNAs in Fragile X patients.

These findings will redirect the search for the RNA targets of FMRP whose misregulation is responsible for the disease, to those containing kissing complex motifs.

Though much remains to be understood in the biology leading to Fragile X syndrome and the function of FMRP, Dr. Darnell is confident that "these findings may provide a crucial link between the association of FMRP in brain polyribosomes, its proposed role in regulation mRNA translation, and neurologic dysfunction in the Fragile X syndrome".

New Gene for Charcot-Marie-Tooth Disease Involved in Cell Transport

Wed, 13 Apr 2005 00:21:00 PST

( from http://www.rxpgnews.com ) Charcot-Marie-Tooth disease (CMT), which affects the peripheral nerves, comes in several forms and is due to mutation in a variety of genes.

Autosomal dominant CMT of the so-called intermediate form has previously been linked to chromosome 19.

In this study, an international team of researchers identified the responsible gene as Dynamin 2 (DNM2), whose protein helps cell membranes fuse together and separate. The mutation, found in several families from North America, Australia, and Belgium, impairs a variety of critical cell transport processes.

In addition, neutropenia, a white blood cell disorder, was found to be inherited along with the disease, which has not been seen in CMT families before. The same protein function that is impaired by the mutation--the ability to bind a high-energy molecule called CTP--is also impaired by other CMT gene mutations, suggesting this may be a central pathway for a large class of peripheral neuropathies.

Roberts Gene ESCO2 Discovered to be behind "PSEUDOTHALIDOMIDE" Syndrome

Tue, 12 Apr 2005 13:02:00 PST

( from http://www.rxpgnews.com ) A team of scientists from Colombia, the United States and elsewhere has successfully completed a 15-year-plus search for the genetic problems behind the very rare Roberts syndrome, whose physical manifestations often include cleft lip and palate and shortened limbs that resemble those of babies whose mothers took thalidomide during pregnancy.

The discovery, which is reported in the April 10 advance online section of Nature Genetics, proves that genes behind very rare inherited diseases can now be found, offering excellent opportunities to strengthen understanding of craniofacial and limb development, health and disease beyond the rare disease itself, say the researchers.

Because of advances in technology and computer analysis, the researchers were able to find the Roberts gene, called ESCO2, by studying samples from just 15 Roberts syndrome families from Colombia, Turkey, Canada and Italy and to provide insight into its biological effect.

"For decades now, we've known that the appearance and number of chromosomes were abnormal in people with Roberts syndrome, but we hadn't been able to figure out why or how," says Ethylin Jabs, M.D., a professor in the McKusick-Nathans Institute of Genetic Medicine at Johns Hopkins. "Just within the last few years have the genetic techniques, the genomic information, and the computer analysis become powerful enough to find the genetic mutations behind a disease as rare as Roberts."

Some of the techniques they used -- such as that to make many copies of DNA from a small sample -- have been around in some form for more than a decade. But others are much more recent developments. For example, the researchers found important genetic changes in part by comparing different species' genetic sequences, most of which were published only within the last four years.

"In 1989, we were collecting samples and characterizing the chromosome problem in cells from people with Roberts syndrome," Jabs remembers. "We knew it would be really important to find the gene, but it just wasn't practical at that time."

A few years later, in 1995, two Colombian geneticists started their quest to fully understand Roberts syndrome. Without their push, the gene for Roberts might still be unknown.

Colombian Hugo Vega had noticed an unusual number of patients with Roberts syndrome in the clinic at the University of Bogotá. Fairly quickly, he tracked down seven families with Roberts syndrome in two villages outside Bogotá. Four of the families share an 18th-century ancestor, he and Miriam Gordillo, then an undergraduate, discovered.

"The families have really collaborated with us, they've worked with us quite closely to help us uncover the gene behind the syndrome," says Gordillo. "Now we have about 10 affected families from outside Bogotá, and we can offer a genetic test to families at risk of Roberts syndrome."

Vega and Gordillo, a husband-and-wife team, criss-crossed the globe to continue their work and find better funding opportunities. In Japan, Vega tied the Colombian families' syndrome to a large region of chromosome 8. In The Netherlands, a post-9/11 detour, he added to his analysis samples from Turkish and Italian families with Roberts syndrome.

In 2004, Gordillo got a student visa to work with Jabs and to study for her doctorate in human genetics at Johns Hopkins. Over the past year, Gordillo analyzed the chromosome 8 region in samples from 15 families (consisting of 18 affected members and 33 unaffected members) and tied the condition to one of 6 genes.

Then, the international team compared the human sequence of the genes to those from chimpanzee, mouse, rat, chicken and zebrafish, and to the gene sequences of the affected family members. One segment of a gene called ESCO2 that was identical in all the animals contained changes that disrupted the gene's protein-making instructions in people with the syndrome. Knocking out the equivalent gene in yeast and fruit flies led to the same chromosome problems, says Gordillo.

"Comparative genomics didn't really exist even five years ago," says Jabs. "Techniques to genetically engineer yeast, fruit flies and even mice have dramatically improved in the last 15 years. And we were also able to look at when and where the gene is expressed during human development. Without these techniques, and without the powerful computer programs, we wouldn't have been able to identify this gene and confirm its role in Roberts syndrome."

The physical similarities of people with Roberts syndrome and those whose mothers took thalidomide suggest similar underlying biology, Jabs notes. Although there's some evidence that thalidomide prevents blood vessel growth, it's not clear why. If the underlying biology is related somehow, then thalidomide might affect chromosomes and cell division like ESCO2 in Roberts syndrome, Jabs speculates.

During normal cell division, every chromosome is copied, and each of the "original" chromosomes is attached to its "new" copy. While there are attachment points along the entire chromosome, the bulk of the connection is at the centromere, a chromosome's functional hub.

The chromosomes' connection allows the cell to move them together, ensuring that the two copies are lined up together at the center of the dividing cell. Once lined up, tiny molecular "motors" attach to the centromere of each copy and pull the original and the new copy away from each another as division proceeds.

However, in cells from people with Roberts syndrome, the chromosome copies are frequently not attached to each other at their centromeres and the chromosomes don't get lined up properly. As a result, the cell doesn't divide or divides very slowly, and the new cells can end up with too many or too few chromosomes (a problem also seen in cancer cells). In Roberts syndrome, the cells tend to stop growing or die, precluding proper development of the limbs, palate and other structures.

Common Factor Discovered behind Myocardial Infarction, Rheumatism and MS

Mon, 11 Apr 2005 20:58:00 PST

( from http://www.rxpgnews.com ) A common gene variant has been identified as the risk factor behind a number of common diseases by research scientists at Karolinska Institutet and the Centre for Molecular Medicine (CMM), Stockholm, Sweden. Up to a quarter of the population could be affected.

Researchers in the fields of cardiovascular disease, rheumatism and MS have together shown that there is a common risk factor for these conditions. It is the first identified gene to link autoimmune diseases with cardiovascular diseases.

"This gene variant can therefore be one of the single largest genetic causes of complex diseases with inflammatory components," says Fredrik Piehl, associate professor at Karolinska Institutet and researcher at the CMM. "There is also a chance that other diseases are also affected by this gene variant. The discovery can now lead to more reliable diagnostics and better treatments for a great number of patients."

The gene variant was first identified in an animal model and then studied in a number of patient groups to ascertain if there was a link to human diseases. The researchers discovered that people with the variant ran a 2040 per cent greater risk of developing rheumatism, MS or a myocardial infarction. The gene variant is also common: an estimated 2025 per cent of the population carry it.

The discovery reveals a new area of application for statins, drugs usually taken to lower cholesterol levels. Statins have been shown to reduce activity in this gene and thus produce anti-inflammatory effects. Statins have now been tested on MS patients and have been demonstrated to be beneficial in this very way.

The disease-associated gene variant leads to a reduction in the production of a number of immune defence proteins. Some viruses and bacteria have also been observed to influence the gene in an attempt to evade the immune defence system, a strategy employed, for example, by the viruses that cause AIDS, herpes and hepatitis.

Researchers Discover Largest "Gene Deserts"

Fri, 08 Apr 2005 03:59:00 PST

( from http://www.rxpgnews.com ) A detailed analysis of chromosomes 2 and 4 has detected the largest "gene deserts" known in the human genome and uncovered more evidence that human chromosome 2 arose from the fusion of two ancestral ape chromosomes, researchers supported by the National Human Genome Research Institute (NHGRI), part of the National Institutes of Health (NIH), reported today.

In a study published in the April 7 issue of the journal Nature, a multi-institution team, led by Washington University School of Medicine in St Louis, described its analysis of the high quality, reference sequence of chromosomes 2 and 4. The sequencing work on the chromosomes was carried out as part of the Human Genome Project at Washington University; Broad Institute of MIT, Cambridge, Mass.; Stanford DNA Sequencing and Technology Development Center, Stanford, Calif.; Wellcome Trust Sanger Institute, Hinxton, England; National Yang-Ming University, Taipei, Taiwan; Genoscope, Evry, France; Baylor College of Medicine, Houston; University of Washington Multimegabase Sequencing Center, Seattle; U.S. Department of Energy (DOE) Joint Genome Institute, Walnut Creek, Calif.; and Roswell Park Cancer Institute, Buffalo, N.Y.

"This analysis is an impressive achievement that will deepen our understanding of the human genome and speed the discovery of genes related to human health and disease. In addition, these findings provide exciting new insights into the structure and evolution of mammalian genomes," said Francis S. Collins, M.D., Ph.D., director of NHGRI, which led the U.S. component of the Human Genome Project along with the DOE.

Chromosome 4 has long been of interest to the medical community because it holds the gene for Huntington's disease, polycystic kidney disease, a form of muscular dystrophy and a variety of other inherited disorders. Chromosome 2 is noteworthy for being the second largest human chromosome, trailing only chromosome 1 in size. It is also home to the gene with the longest known, protein-coding sequence - a 280,000 base pair gene that codes for a muscle protein, called titin, which is 33,000 amino acids long.

One of the central goals of the effort to analyze the human genome is the identification of all genes, which are generally defined as stretches of DNA that code for particular proteins. The new analysis confirmed the existence of 1,346 protein-coding genes on chromosome 2 and 796 protein-coding genes on chromosome 4.

As part of their examination of chromosome 4, the researchers found what are believed to be the largest "gene deserts" yet discovered in the human genome sequence. These regions of the genome are called gene deserts because they are devoid of any protein-coding genes. However, researchers suspect such regions are important to human biology because they have been conserved throughout the evolution of mammals and birds, and work is now underway to figure out their exact functions.

Humans have 23 pairs of chromosomes - one less pair than chimpanzees, gorillas, orangutans and other great apes. For more than two decades, researchers have thought human chromosome 2 was produced as the result of the fusion of two mid-sized ape chromosomes and a Seattle group located the fusion site in 2002.

In the latest analysis, researchers searched the chromosome's DNA sequence for the relics of the center (centromere) of the ape chromosome that was inactivated upon fusion with the other ape chromosome. They subsequently identified a 36,000 base pair stretch of DNA sequence that likely marks the precise location of the inactived centromere. That tract is characterized by a type of DNA duplication, known as alpha satellite repeats, that is a hallmark of centromeres. In addition, the tract is flanked by an unusual abundance of another type of DNA duplication, called a segmental duplication.

"These data raise the possibility of a new tool for studying genome evolution. We may be able to find other chromosomes that have disappeared over the course of time by searching other mammals' DNA for similar patterns of duplication," said Richard K. Wilson, Ph.D., director of the Washington University School of Medicine's Genome Sequencing Center and senior author of the study.

In another intriguing finding, the researchers identified a messenger RNA (mRNA) transcript from a gene on chromosome 2 that possibly may produce a protein unique to humans and chimps. Scientists have tentative evidence that the gene may be used to make a protein in the brain and the testes. The team also identified "hypervariable" regions in which genes contain variations that may lead to the production of altered proteins unique to humans. The functions of the altered proteins are not known, and researchers emphasized that their findings still require "cautious evaluation."

Potential therapeutic target for Huntington's disease

Thu, 07 Apr 2005 18:14:00 PST

( from http://www.rxpgnews.com ) Researchers studying yeast cells have identified a metabolic enzyme as a potential therapeutic target for treating Huntington's disease, a fatal inherited neurodegenerative disorder for which there is currently no effective treatment.

The group performed a genetic experiment known as a loss-of-function suppressor screen, which searches for genes that, when switched off, reduce the toxic effects of the mutant protein associated with Huntington's. One of the genes they identified encodes an enzyme, called KMO, that has been previously implicated in the disease. The enzyme functions in a metabolic pathway that is activated at early stages of the disease in people with Huntington's, as well as in animal models of the disease.

"The nice thing about this finding is that there is a chemical compound available that inhibits KMO activity," said Dr. Paul Muchowski, assistant professor of pharmacology at the UW, who led the study. "We're in the midst of testing that compound in a mouse model of Huntington's disease."

Further support for KMO as a therapeutic target for Huntington's disease comes from a recent study led by Dr. Aleksey G. Kazantsev of Harvard Medical School. In this study, researchers used cell-based experiments to screen about 20,000 chemical compounds, and identified one that suppresses neurodegeneration in a fly model of the disease. That compound has a very similar chemical structure as the drug that inhibits the target identified by Muchowski's group. The results appeared in the Jan. 18, 2005, issue of the Proceedings of the National Academy of Sciences.

In addition to finding a potential drug target for future Huntington's treatment, the study by Muchowski and his colleagues could take research on the disease in a new direction: towards microglial cells, which are immune cells in the brain. Previous research has focused exclusively on neuronal cells, but the enzyme KMO is found predominantly in microglial cells. Since inhibiting KMO activity has a direct effect on toxicity of the mutant protein associated with Huntington's, that could mean microgial cells are home to an important step in progression of the disease.

Huntington's affects an estimated 30,000 people in the United States. It is characterized by loss of motor control and cognitive functions, as well as by depression or other psychiatric problems.

Where Do All Those Genes Come From?

Wed, 06 Apr 2005 16:28:00 PST

( from http://www.rxpgnews.com ) An important source of genetic novelty is the introduction of new genes. Since most genes in an organisms genome are under selective constraint, opportunities for the evolution of new gene functionswhich in turn might confer selective advantagemost often arise when new genes enter the genome. In eukaryotesa category that includes humans and ricenovel genes typically arise when existing genes undergo duplication. Extra copies of genes can be created when normal DNA replication hiccups and erroneously duplicates entire regions of DNA. These extra gene copies reside in species genomes for generations and might eventually mutate to code for novel proteins, adding new genes to the species repertoire. The new genes, along with the rest of the genome, are passed down from one generation to the next in a process known as vertical transmission.

In prokaryoteswhich include unicellular organisms in the bacteria and archaea domainsnovel genes can appear through multiple routes. In addition to gene duplication, prokaryote genomes can change when DNA fragments are taken up directly by cells, passed from cell to cell, or transferred to new cells with the help of viruses. All three scenarios provide a means for whole genes to move directly from one bacterial genome to another, a process called lateral gene transfer (LGT) or horizontal transmission.

Until now, the importance of vertical versus horizontal transmission in the evolution of any large prokaryote group was unknown. In a new study, Emmanuelle Lerat et al. capitalized on the availability of complete genome sequences within the diverse γ-Proteobacteria, a group of prokaryotes that includes Escherichia coli, Salmonella spp., and some nitrogen-fixing bacteria, to pursue that question.

Sorting out the issue is no simple task. If the same gene is present in more than one species, it could have been inherited from a common ancestor or it could have jumped from one lineage to another by LGT. Even if the same gene appears twice in one species genome, the copies could have different historiesone copy could have been acquired vertically from its ancestors, while the other could have come from a different species.

Though previous studies have looked at the distributions of genes across species phylogenies, information about gene origin appeared contradictory. To create a clearer picture, Lerat et al. accounted for the possibility of widespread LGT by statistically comparing the phylogenies of many different gene families with a benchmark phylogenetic tree that reflected the accepted evolutionary history of γ-Proteobacteria.

The authors found that LGT plays a substantial role in generating the diversity of genes found in γ-Proteobacteria genomes. Members of the group are constantly acquiring and losing genes, although the extent of LGT can vary greatly among species. In contrast, gene duplications play a much smaller role in explaining γ-Proteobacteria genome diversity, although duplications have been shown to be important for short-term adaptation.

Genes that have arrived by LGT within a single genome do not necessarily share a common history with each other. Many of the genes that are found only in a single genome and are not widely distributed across the γ-Proteobacteria were recently acquired from distant sources. Most of these acquired genes will likely be lost soon after joining a genome; those that persist are then inherited vertically. This helps to reconcile why gene trees tend to provide valid phylogenetic inferences about the relationships among different bacterial lineages, despite the potential mixing that could result from LGT. Phylogeneticists aiming to reconstruct a phylogeny for a group look at variations in genes distributed in the species, and these are largely vertically transmitted.

Lerat et al. propose that LGT is a common source of genes in γ-Proteobacteria because it has the potential to introduce functionally different genes into the genome with immediate contributions to fitness. Gradual evolution of gene duplicates doesnt provide the same type of immediate reward.

Single Quantitative Trait Loci (QTLs) with a Large Effect on Body Size

Wed, 06 Apr 2005 16:28:00 PST

( from http://www.rxpgnews.com ) The presence of a small number of discrete formsas you find in a classic Mendelian trait like eye colorsuggests that the phenotype is controlled by a very small number of genes. In contrast, a complex trait such as body size is influenced by multiple genes as well as environmental factors, giving rise to a continuous spectrum of phenotypes. This causal complexity makes discovery of the genetic determinants of the traitso-called quantitative trait loci (QTLs)very difficult. In this issue, Fiona Oliver, Julian Christians, and colleagues extend their work on a single QTL with a large effect on body size variation in mice, and show that the responsible gene is one already linked to a Mendelian growth disorder in humans.

In previous work, mice were divergently selected for large or small body size, revealing a QTL on the X chromosome that causes a 20% difference in growth rate. In this study, the authors further refined the map of the area to 660 kilobases, and found it contains only four genes: glypican-3 and glypican-4 (Gpc3 and Gpc4), and two other genes of unknown function. The glypicans are membrane-bound growth regulators; loss of function of Gpc3 in humans causes a rare syndrome of overgrowth, skeletal and other abnormalities, and neonatal death. Since none of the four genes showed coding-region variations that might explain the differences between the large and small mice, the authors examined expression levels. In Gpc3, but not the others, size correlated with a significant difference in gene expression: larger mice had low levels of the messenger RNA, and smaller mice had high levelsthe same pattern seen in the human loss-of-function disorder. The authors identified several non-coding polymorphisms in Gpc3 that differed between the two forms, although it remains to be seen whether the differential effect on growth is due to these or other DNA differences nearby.

The results from this study point out an important feature of inheritance, namely, that a gene implicated in a Mendelian trait can also contribute to quantitative variation. While catastrophic expression failure, such as a loss of function, can cause disease, smaller changes in expression of the same gene may simply help fill out the bell curve of normal variation.

Novel Data-Mining Approach Systematically Links Genes to Traits

Wed, 06 Apr 2005 16:28:00 PST

( from http://www.rxpgnews.com ) With exponential advances in computing power over the past ten years, data-generating capacity has far outpaced anyones ability to mine the rich seams of information. This is especially true in the field of genomics.

So far, over 222 prokaryote (bacteria) genomes have been sequenced, 21 archaea (primitive bacteria-like extremophiles), and 17 eukaryotes (from yeast to fly and rat to human), according to the Center for Biological Sequence Analysis in Denmark (http://www.cbs.dtu.dk/services/GenomeAtlas/). All these genomes promise to provide powerful insights into the biological processes of life, but such insights come with painstaking analysis by trained experts. Matching genotype to phenotypethe visible or measurable characteristics of speciesis a major challenge in what Francis Collins, Director of the United States National Human Genome Research Institute, has called the post-genomic era.

In a new study, Peer Bork and a team of bioinformatics-savvy molecular biologists tested a new approach to extracting biologically meaningful information from the massive MEDLINE database. The US National Library of Medicines MEDLINE contains over 12 million abstracts from thousands of publications dating back to 1965. Combining automated literature mining with comparative genomicswhich compares genome sequences of different organisms to discern differences and similarities in gene contentthe authors conducted a systematic search for associations between genes and phenotypic traits. Their approach automates tasks that typically require human curation.

Recognizing that the best source of information on species phenotypic traits is the scientific literature where biologists describe them, the authors first ran a search to identify associations between species and traits in MEDLINE abstracts. Words that tended to occur with subsets of species, the authors reasoned, were more likely to reflect particular traits. From a total of 255,249 MEDLINE abstracts showing any connection to 92 prokaryotic species with sequenced genomes, 172,967 nouns showed meaningful associations related to the species traits. Flagellum and motility showed up more often in self-propelling species, for example, and endosymbiont aptly appeared with the intracellular bacteria (Buchnera aphidicola) that inhabits aphids.

Next, Bork and colleagues detected the presence or absence of over 200,000 evolutionarily conserved genes across the 92 species and sorted the results into speciesword and speciesgene groups. The analysis revealed a number of words and genes with similar distribution in related species, leading to over 2,700 significant associations between trait-descriptive words and orthologous (evolved from a common ancestor) groups of genes. These genes encode over 28,000 proteins. Many were already knownincluding genes involved in pathogenicity, biodegradation and biosynthesis, and photosynthesisbut many, the authors note, are novel or of unexpected character and complexity.

And it is the ability to uncover unexpected relationships across numerous genes and genomespatterns likely to escape human analysisthat makes this approach so powerful. Among these unexpected match-ups, Bork and colleagues linked a number of food and food-poisoning-related terms with metabolic-enzyme-coding genes. All 37 genes predicated to play a role in food spoilage and toxicity are present in food-borne pathogens but not in most other prokaryotes. By assigning functions to these previously uncharacterized genes, the authors could also assign new roles for pathways that use the genes. For example, by linking two genes with pathways that metabolize propanediol and ethanolaminecompounds found almost exclusively in highly hazardous food-borne pathogensthe authors predict that propanediol and ethanolamine pathways are crucial genomic determinants of pathogenicity associated with food poisoning.

That their analysis linked so many predicted genes with bacterial pathogenicity might be expected, the authors note, since both genome sequencing and biological research are heavily focused on human health. Given the weekly increase in the number of genomes sequenced and in MEDLINE entries, the method outlined here should provide a valuable tool to help researchers narrow the gap between the promise and payoff of the genomic revolution.

New Gene Therapy using Homologous Recombination

Wed, 06 Apr 2005 16:23:00 PST

( from http://www.rxpgnews.com ) Harnessing the strength of a natural process that repairs damage to the human genome, a researcher from UT Southwestern Medical Center has helped establish a method of gene therapy that can accurately and permanently correct mutations in disease-causing genes.

By artificially initiating a DNA repair process known as homologous recombination, Dr. Matthew Porteus of UT Southwestern, working with scientists from Richmond, Calif.-based Sangamo Biosciences, was able to replace a mutated version of the gene that encodes a portion of the interleukin-2 receptor (IL-2R) in human cells, restoring both gene function and the production of the IL-2R protein.

Mutations in the IL-2R gene are associated with a rare immune disease called severe combined immunodeficiency disease, or SCID. Children with SCID are unable to successfully fight off infections, and must constantly live in a germ-free environment. Their lifespans are usually shortened by systemic infection, and while bone marrow transplants can be used to treat the disease, they are not always successful.

SCID is ideal for this sort of therapy because you only need to correct the defect in a small number of immune cells to fix the problem, said Dr. Porteus, assistant professor of pediatrics and biochemistry at UT Southwestern. This is called selective advantage; the healthy cells grow and divide preferentially over the mutant ones.

Previous gene therapy attempts for SCID have been only moderately successful because of technical difficulties in the delivery method. In one instance, the correct IL-2R gene was delivered to mutant cells of SCID children by a disabled virus, but some subsequently developed leukemia because the virus inadvertently turned on a cancer gene.

Dr. Porteus strategy differs fundamentally from previous gene therapies because it essentially replicates the natural process, which is more accurate. This accuracy means that, in practice, the gene therapy only affects the mutant gene.

Homologous recombination is a fairly rare event that occurs when DNA strands of one chromosome break, creating a damaged section. Cells have two copies of nearly every chromosome (one each from the mother and father) and they must duplicate these during cell division so that the subsequent cells will also have two each. The wounded chromosome takes advantage of the healthy copies created in preparation for division, and uses them as a template to repair the break in the DNA strands.

In the new gene therapy technique, researchers took advantage of homologous recombination by introducing a man-made enzyme that recognizes and binds DNA at specific points into human immune cells harboring the mutant IL-2R gene. Once bound to the mutant gene, the enzyme creates a break in the DNA sequence, initiating the recombination process.

In SCID, as well as many other diseases, both copies of the disease gene are mutated so there is no correct version naturally available. To overcome this, researchers also supplied the cells with a correct version of the IL-2R gene along with the enzyme.

Given the correct copy as a template, the recombination event occurred at the break site and 11 percent of the cells tested had traded one copy of the mutant gene for the correct version. In addition, 6 percent to 7 percent of cells had traded both copies of the bad gene for the correct version.

With the correct IL-2R gene in place, levels of the IL-2R protein also were restored.

The change to the cells seems permanent, Dr. Porteus said, and the correct gene is easily maintained after many cell divisions.

The rates of correction that we see are extremely exciting, said Dr. Porteus, who was recently awarded a 2004 Distinguished Young Researcher Award from the Presidents Research Council at UT Southwestern for his early contributions to the gene therapy technique. That we can fix mutations in human immune cells makes us very optimistic that this therapy will work in several other diseases, such as sickle cell disease.

Sickle cell disease is a disorder in which blood cells carry a mutation in hemoglobin. The mutation causes the blood cells to change shape, preventing them from flowing through the blood vessels efficiently. One out of every 12 African-American individuals carries this mutation, and one out of every 500 African-American births is afflicted with disease.

In theory, blood stem cells with the sickle cell mutation could be removed, treated with the enzyme and the correct version of the mutant gene, and eventually given back to the patient, spawning the growth of healthy blood cells, Dr. Porteus said.

RNA interference (RNAi) improved Huntington's disease Symptoms

Tue, 05 Apr 2005 17:14:00 PST

( from http://www.rxpgnews.com ) Researchers at the University of Iowa Roy J. and Lucille A. Carver College of Medicine have taken another step toward a potential treatment for Huntington's disease (HD). Using an approach called RNA interference (RNAi), the scientists reduced levels of the disease-causing HD protein in mice and significantly improved the movement and neurological abnormalities normally associated with the disease.

HD is a devastating, inherited, neurodegenerative disease that is progressive and always fatal. The disease-causing gene produces a protein that is toxic to certain brain cells, and the subsequent neuronal damage leads to the movement disorders, psychiatric disturbances and cognitive decline that characterize this disease.

"Many of the current approaches aimed at treating HD are indirect and target the symptoms of the disease. RNA interference gives us the first opportunity to attack the fundamental problem and reduce protein expression from the disease gene," said Beverly L. Davidson, Ph.D., the Roy J. Carver Chair in Internal Medicine and UI professor of internal medicine, physiology and biophysics, and neurology. "Our study is the first demonstration that a therapy designed to inhibit protein production has a beneficial effect."

The study will appear this week in the Online Early Edition of the Proceedings of the National Academy of Sciences (www.pnas.org). Davidson is the senior author and Scott Harper, Ph.D., a postdoctoral researcher in Davidson's lab, is lead author.

Harper, Davidson and their colleagues used RNAi to treat a mouse model of HD. Viral vectors (stripped-down viruses) carrying the genetic instructions to make a RNA interference molecule were injected into the brains of genetically engineered mice before the disease symptoms appeared. The treated mice showed nearly normal movement, and the characteristic neurological damage also was significantly improved in comparison to untreated mice.

Detailed examination of the protein levels in the treated mice showed that levels of the toxic HD protein were reduced to about 40 percent of the level seen in untreated mice.

"It is very exciting that a partial reduction is sufficient to produce a very beneficial effect in the animal. It means that we don't have to turn the gene off completely," Davidson said. "For a disease that takes decades to develop, a partial reduction may slow down the disease-causing copy of the gene to such an extent that either disease progression is delayed or possibly even disease onset is prevented."

It may even be the case that a partial reduction of toxic protein levels allows the brain cells' machinery to "catch up" with the disease-causing protein and clear out the damage caused by the mutant protein.

The genetically engineered or transgenic mouse model used by the UI team carries a section of the human HD gene. These mice quickly develop movement and coordination abnormalities and they die young. Aggregates, or clumps of protein, also develop in certain brain cells.

Davidson explained that this mouse is very good for proof-of-principle experiments, allowing the researchers to ask a very pointed question can RNAi improve HD-like symptoms in a mouse model in short order?

"Since our results are positive, we can now repeat the experiment in mouse models that develop disease more slowly and more closely resemble HD in humans," Davidson said.

Most genes are inherited as a pair, one from either parent. In HD, one mutated copy of the gene is sufficient to cause the disease. However, the normal Huntington gene produces a protein that is known to be critical in embryonic development. It is not known if the protein is critical in adult brain cells.

The RNAi molecule used in Davidson's current study would silence both the mutant and the normal gene. So, an important question that still needs to be addressed is whether adult neurons can tolerate and benefit from a partial reduction of both the toxic and the normal protein. If the normal protein is critical, then RNAi will need to be specifically targeted against the disease-causing gene.

Fortunately, RNAi is exactly the right tool to provide an answer regarding whether the normal gene is critical by silencing the normal gene in adult brain cells of HD models.

Despite the remaining hurdles, Davidson is optimistic about the potential of RNAi to treat HD and similar neurodegenerative diseases.

"If the benefit is confirmed in other mouse models of Huntington's disease, and it appears that we don't need to target the RNAi specifically to the disease-causing mutant gene, then I would think it might move to human testing within several years," she said.

Putting a finger on shortened digits

Sun, 03 Apr 2005 13:44:00 PST

( from http://www.rxpgnews.com ) Brachydactyly is a group of inherited disorders of the hands that are characterized by shortened fingers and abnormal joint formation.

In a paper appearing in the April 1 issue of The Journal of Clinical Investigation, Stefan Mundlos and colleagues from the Max Planck Institute for Molecular Genetics describe the analysis of a mouse model with limb mutations called short digits (Dsh).

The mice have disrupted Shh expression a factor that helps skeletal formation. The result is that the mice have symptoms similar to human brachydactyly type A1. This is because the misexpression of Shh disrupts other factors with normally regulate joint development as well as the growth and patterning of the digits

Luis de la Fuente and Jill Helms write, in an accompanying commentary, that this study shows "that removal or expansion of one of the factors that contributes to the establishment of a boundary can cause a multitude of processes, including those that shape and control development of the skeleton, then go awry." The developmental pathology associated with Shh misexpression extends our understanding of the developmental pathology of digit development and thus of human brachydactyly.

Analysis of X chromosome Completed

Sun, 03 Apr 2005 11:34:00 PST

( from http://www.rxpgnews.com ) By intensely and systematically comparing the human X chromosome to genetic information from chimpanzees, rats and mice, a team of scientists from the United States and India has uncovered dozens of new genes, many of which are located in regions of the chromosome already tied to disease.

Regions of the X chromosome, one of the two sex chromosomes (Y is the other), have been linked to mental retardation and numerous other disorders, but finding the particular genetic abnormalities involved has been difficult.

"To our knowledge, this is the first time critical analysis of an entire chromosome has been done by a group that wasn't involved in determining the chromosome's genetic sequence," says study leader Akhilesh Pandey, M.D., Ph.D., an assistant professor in the McKusick-Nathans Institute of Genetic Medicine at Johns Hopkins and chief scientific adviser to the Institute of Bioinformatics (IOB) in Bangalore, India, where the analyses took place. "We didn't start small. We wanted to prove that complete annotation can be done, and done in a way that lets you find new and unexpected things."

For 18 months, 26 Indian scientists pored through the publicly available sequence of the X chromosome (information generated by the Wellcome Trust Sanger Institute in England and others) to identify genes and other important parts of its DNA.

But unlike other efforts, the team didn't just "mine the data" by using computers to search for known patterns in the genetic sequence. Instead, Pandey decided they would look for similarities between the human X chromosome's protein-encoding instructions and corresponding regions in the mouse. Regions that were identical or nearly so were then examined carefully by IOB biologists.

"We didn't want to start out by saying that genes had to look a certain way," says Pandey. "So our only initial assumption was that if a genetic region is important and codes for a protein, the sequence will be conserved at the protein level. Thus, even if the genetic sequence is different here and there, the protein sequence could still be the same."

Essentially, the researchers took advantage of the redundancy inherent in the genetic code. DNA's four building blocks -- A, T, C and G -- act as instructions for proteins in select three-block sets. These three-block sets each "code" for just one of the 20 possible protein building blocks, or amino acids, but some of the sets code for the same amino acid. For example, the DNA sequences TTGAGGAGC and CTACGATCA are quite different, but both specify the same three amino acids -- leucine, arginine and serine, in that order.

"Instead of telling the computer what to look for, we let nature tell the computer what was important," says Pandey. "When you align the protein-encoding instructions of the human and mouse, the genes jump out at you."

In the regions that were the same between species, the scientists found 43 new "gene structures" that encode proteins. Some of the newly identified genes sit in regions long tied to X-linked mental retardation syndromes, which appear only in boys, or other disorders. Quite remarkably, Pandey says, almost half of the new genes don't look like any previously known genes, nor do they look like each other.

"These would not be found any other way, because no one knew to look for them," he says. "No one had ever identified any aspect of their sequences as being important."

The IOB scientists and the U.S. members of the team experimentally investigated a few of the new genes to confirm the comparative approach's validity. Their results, as well as data created by other scientists since the U.S-India team started working, confirm the existence of some of the newly identified genes. The team's work also showed that some so-called pseudogenes on the X chromosome are actually expressed, or transcribed, which contradicts the widespread idea that they are functionless.

"We're really trying to show that complete annotation of chromosomes can be done, and that doing it this way means you can find things you don't expect to find," says Pandey. "It's long, painstaking work, but it's worth it."

Pandey hopes that researchers will take the initiative to annotate sequenced genetic information and validate regions used in their work.

The Fate of Duplicated Genes

Thu, 31 Mar 2005 22:07:00 PST

( from http://www.rxpgnews.com ) Providing insight into the evolution of biological complexity, Steven Maere et al. have developed a computer model that simulates the fate of genes that arise from small-scale local duplications and duplications involving the entire genome.

Applying this model to the Arabidopsis thaliana genome revealed that 60% of the duplicate genes were survivors of three ancient genome duplications, whereas the remaining 40% arose through small-scale events.

The simulations also showed that the decay rate for different types of genes differed dramatically depending on the size of the duplication event. Genes involved in kinase activity, transcription, protein binding/modification, and signal transduction demonstrated low decay rates if they were produced by large-scale duplication events. Similar genes created through small duplications tended to decay more rapidly.

Based on these results, Maere et al. estimate that 90% of transcription factors in Arabidopsis and higher plants arose during the three genome duplications that occurred in the last 350 million years. Genes involved in secondary metabolism or that respond to biotic stimuli-like pathogen attack, drought, or salinity tend to be preserved regardless of the mode of duplication, according to the authors.

Rare Mutation that Speeds up the Biological Clock

Thu, 31 Mar 2005 15:35:00 PST

( from http://www.rxpgnews.com ) Scientists have identified a gene and mutation within it that causes a rare sleep behavior, in which individuals have a "fast" biological clock. The gene's enzyme could lead to a therapeutic target for the disrupted sleep patterns seen in such groups as those facing jet lag or nighttime work shifts.

More broadly, the gene provides a probe for exploring the regulatory mechanisms of the body's internal biological clock, or circadian rhythms -- a waxing and waning of genetic, biochemical and physiological processes that occurs in a 24 hour period -- about which little is known in humans at the molecular level.

As the findings hint that the genetic mutation might play a role in depression, the scientists are now exploring this possibility, as well.

"Evidence suggests that circadian rhythms may have a fundamental role in numerous behaviors," says Ying-hui Fu, PhD, associate professor of neurology at University of California, San Francisco and the senior author of the paper. "As the enzyme produced by the gene modulates many proteins, we may test for its impact on novelty seeking and learning and memory, too."

"The discovery of the gene opens the window just a crack, but it could let in a lot of light for probing the neurobiology of the brain," says co-senior author Louis Ptacek, MD, a Howard Hughes Medical Institute investigator and UCSF professor of neurology.

The finding, published in the March 31 issue of Nature, builds on previous research led by Ptacek and co-author Christopher Jones, PhD, associate professor of neurology at University of Utah. In 1999, in a study of three families with the unusual sleep pattern, known as familial advanced sleep phase syndrome (FASPS), the scientists created a family tree to map the incidence of inheritance, and made the seminal finding that the behavior was a single-gene trait. (Nature Medicine, Sept. 1999). In 2000, the team discovered the gene (hPer2) responsible for the condition in one particular family, the first report of a human circadian rhythm gene. (Science, Jan. 12, 2001).

The breadth of the knowledge coming out of the current study is somewhat unique, says Ptaceck. "I'm not aware of another study that has gone from the human to the fly to the mouse in a single study. It demonstrates of the combined power of human molecular genetics research and studies in animals."

The study began with the scientists isolating, or cloning, the mutant gene responsible for causing FASPS in five members of an extended family. The gene is known as casein kinase1 delta (ck1 delta); the mutated form is designated CK1 delta-T44A. They made the discovery by taking blood samples of the individuals and then using linkage analysis to hone in on the region of DNA within the chromosomes most likely to reveal the relevant gene, based on what was known about other genetic markers related to circadian rhythms. The gene was found on chromosome 17. (The mutated form was compared to the gene in family members who did not have FASPS.)

Next, they determined that, in a test tube, the activity of the enzyme (CK1 delta) encoded by the mutated gene was decreased.

Finally, to explore the effect of the genetic mutation on circadian activity, the scientists inserted the mutated human gene into the nerve cells of the circadian clock in the fruit fly Drosophila melanogaster and the mouse. Scientists have learned much about human gene function by studying animals, given the many genes that have been conserved through evolution, and both animals have been a key source of information regarding the genetic and molecular biology of circadian rhythms.

As expected, the transgenic mice displayed the same abnormal sleeping pattern seen in the human cases of FASPS, as measured in the decrease in their wheel-running activity.

Unexpectedly, however, the result was the opposite in the transgenic flies, as displayed by its extended locomotor activity. The scientists do not know what accounts for the difference.

"The findings suggest that, although many individual parts of the circadian rhythm clock are conserved across species, fundamental differences exist," says the first author of the study, Ying Xu, PhD, a visiting postdoctoral fellow in the Fu lab. "These models provide the opportunity to begin study of the similarities and differences between circadian clocks in model organisms and humans."

While the internal clock of most people operates in just over a 24-hour time period, the clock of those with FASPS advances an average of 45 minutes a day. If left unchecked, it would continue to speed up. Due to the social demands and needs of those with the disorder, it is kept in check to a relative degree.

The creation of the transgenic mouse model provides a tool for exploring how the CK1 delta-T44A enzyme causes FASPS. It also offers a model for testing promising compounds directed at the enzyme to treat a variety of sleep disorders, whether resulting from biological or environmental factors.

While FASPS is caused by an individual gene, other forms of sleep-pattern disturbances, such as the tendency for some elderly to wake up earlier than normal ("advanced sleep phase syndrome"), could be caused by "epigenetic" factors, which involve changes in gene expression rather than a mutation in a gene.

Finally, the potential insights extend beyond those involving sleep. Four of the five individuals in the study have clinical features or a history of depression. While the scientists say that this could be a coincidence, or that the depression is situational, i.e., due to being "out of phase with the rest of the world," the more provocative possibility, they say, is that circadian rhythm variants contribute to psychiatric disorders.

Now, in collaboration with UCSF's Larry Tecott, MD, PhD, UCSF associate professor of psychiatry and a member of the UCSF Center for Neurobiology and Psychiatry, the team is planning to use a mouse model of depression to study the gene.

"Whether or not it is in fact caused by this particular mutation remains to be tested, but the creation of animal models of the human circadian gene variant will allow direct testing of this fascinating and important hypothesis," says Fu.

Other hypotheses are also on the horizon: The team noted that five of the family members who have FASPS, as well as a sixth relative with possible FASPS, have asthma and migraine with aura (a neurological sensation that foretells the onset of a migraine). While they have not investigated a possible link, the scientists have taken note.

"We're going to follow the same approach we always have," says Ptacek. "If we find something interesting in any of these domains, we'll pursue."

Transcription Factors Control Snapdragon Asymmetry

Wed, 30 Mar 2005 06:45:00 PST

( from http://www.rxpgnews.com ) RAD, a transcription factor expressed in the dorsal region of a developing snapdragon (Antirrhinum majus), helps coordinate the genes that control the flower's unique asymmetrical shape, according to Susie Corley et al.

Four transcription factors, CYC, DICH, DIV, and RAD, determine the formation of dorsoventral asymmetry in the snapdragon, but how they interact is unclear.

Corley et al. cloned RAD and looked for its expression in the developing flowers of both wild-type and mutant plants. The authors found that RAD encodes a small protein with a MYB-like domain, and is thus a member of one of the largest transcription factor families in plants.

RAD was activated in the dorsal region of developing flowers by CYC and DICH, and once activated, RAD antagonized DIV, preventing its activity in the dorsal regions. DIV produces a transcription factor that leads to petals with ventral characteristics. RAD transcription factor is similar in sequence to the N-terminal domain of DIV, also a MYB transcription factor.

This similarity suggests that the two transcription factors compete for binding sites on DNA or interacting proteins. The authors suggest that RAD evolved from DIV or a common precursor via C-terminal deletion.