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Week Ending FRIDAY March 27, 2009---------------------------News Archive

New Way to Make Stem Cells Avoids Risk of Cancer
A team of University of Wisconsin-Madison researchers reports that it has created induced human pluripotent stem (iPS) cells completely free of viral vectors and exotic genes.

By reprogramming skin cells to an embryonic state using a plasmid rather than a virus to ferry reprogramming genes into adult cells, the Wisconsin group's work removes a key safety concern about the potential use of iPS cells in therapeutic settings.

The new method, which is reported in today's (March 26) online issue of the journal Science, also removes the exotic reprogramming genes from the iPS equation, as the plasmid and the genes it carries do not integrate into an induced cell's genome and can be screened out of subsequent generations of cells. Thus, cells made using the new method are completely free of any genetic artifacts that could compromise therapeutic safety or skew research results, according to the Science report.

The new work was conducted in the laboratory of James Thomson, the UW-Madison scientist who was the first to successfully culture human embryonic stem cells in 1998 and, in 2007, co-discovered a way to make human-induced pluripotent stem cells. Thomson, a professor in the UW-Madison School of Medicine and Public Health, is also the director of regenerative biology for the Morgridge Institute for Research, the private, nonprofit side of the new Wisconsin Institutes for Discovery at UW-Madison.

"We believe this is the first time human-induced pluripotent stem cells have been created that are completely free of vector and transgene sequences," says Thomson.

The new study was led by geneticist Junying Yu, the Wisconsin researcher who, with Thomson, co-discovered a method for reprogramming adult skin cells to behave like embryonic stem cells, the master cells that arise at the earliest stages of development and that ultimately develop into all 220 cell types in the human body.

While the methods first devised for reprogramming adult cells yielded embryonic-like cells, the process resulted in the permanent integration of both viral genes and the genes used for reprogramming into the genomes of the newly induced cells. Such genetic baggage posed safety concerns for potential therapies like cell transplants, and confounded work in the lab, as the introduced genes sometimes spurred mutations that interfered with the normal function of induced cells.

The new work was accomplished using a plasmid, a circle of DNA, and cells from the foreskins of newborns. "The plasmids carry all the needed transgenes, but don't integrate into the host DNA, they just float around as episomes" in the cell, Thomson says.

The plasmids replicate, but they do so somewhat inefficiently, Thomson explains, so that after they perform the job of reprogramming, they can subsequently be weeded out, leaving the induced cells free of any exotic genetic material. "Once the transgenes have done their job and are no longer needed, one can merely recover induced pluripotent stem cells that have lost their episomes."

The resulting cells, says Thomson, are remarkably similar to embryonic stem cells and show the same capacity to proliferate indefinitely in culture and diversify into all the cell types of the human body.

"The recent discovery that adult cells could be reprogrammed to iPS cells that resemble embryonic stem cells opened up tremendous potential for regenerative medicine," says Marion Zatz of the National Institute of Health's National Institute of General Medical Sciences, which partially funded the new work. "However, the early methods posed significant risks in using iPS cells in a clinical setting. This latest discovery by Thomson's group of a new method for generating iPS cells without inserting viral vectors into the cells' genetic material is a major advance toward safely reprogramming cells for clinical use."

Thomson notes that researchers have developed other promising approaches using mouse cells, and previously had removed most of the vector and exogenous gene sequences from human-induced pluripotent stem cells. However, those efforts did not succeed in removing all of the genetic artifacts of reprogramming, which could still result in mutations in induced cells.

"Given the rapid pace of the field, it won't be surprising if there are several alternative methods for producing vector and transgene free cells very soon," says Thomson. "However, it will be essential to determine which of these methods most consistently produces induced pluripotent stem cells with the fewest genetic abnormalities. Any problems would impact research, drug development and possible transplantation therapies."

The Host Makes All the Difference
Braunschweig Helmholtz researchers demonstrate the role of the host in influenza illnesses

"Where there are many scientific works dealing solely with the flu virus, we have investigated how the host reacts to an infection," says Klaus Schughart, head of the Experimental Mouse Genetics research group. In infection experiments the researchers have now discovered that an excessive immune response is responsible for the fatal outcome of the disease in mice. This overreaction has genetic roots. The findings have now been published in the scientific magazine PLoS One.

For their investigations the researchers injected seven different inbred mouse strains with the same quantity of type Influenza A flu viruses. All of the animals within one mouse strain are genetically identical, like identical twins. However, one strain differs from another just like different individuals in the human population. To their surprise, the researchers were able to identify strong differences in the progression of the influenza between the seven strains. In five of the strains the illness was mild: the animals lost weight, recovering completely after seven to eight days. However, in two of the mouse strains the animals lost weight rapidly and died after just a few days.

The researchers looked for reasons for these differences: they investigated how the immune system of the animals responds to the virus. "The mice die from their own immune defences, which are actually supposed to protect them against the virus. The immune system produces too many messengers, which have a strong activating effect on the immune cells. These cells then kill tissue cells in the lungs that are infected with the virus," says Schughart. At the same time, these overactive cells also destroy healthy lung tissue. In mice that died the researchers also found one hundred times more viruses than in animals that survived. "It appears that the animals have specific receptors on their cells that make them more receptive to a severe viral infection." Flu infections in humans could take a similar course, here too, genetic factors could favour a severe progression of the illness. "It is only now that we are beginning to understand the role played by the genetic factors of the host and what increased receptiveness means in the case of influenza," says Schughart.

Every year between 10,000 and 30,000 people in Germany die from influenza, the majority via pathogens of the Influenza A type. There are various sub-types of the main type A, in which the composition of the virus envelope differs. H1N1 and H3N2 are the most widely-distributed flu strains amongst humans, H5N1 the familiar avian flu virus. The H stands for the protein haemagglutinin, with which the virus latches onto the cells of the airways, infecting them. In order for the newly-created flu viruses to leave the host cells, in turn, they require neuraminidase (N). To evade an immune response the virus changes the H and N characteristics constantly. Sometimes light, sometimes heavy: the result is a completely new virus type with a new number, with the consequences generally a severe global flu pandemic.

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Article: Srivastava B, Błażejewska P, Heßmann M, Bruder D, Geffers R, et al. 2009 Host Genetic Background Strongly Influences the Response to Influenza A Virus Infections. PLoS ONE 4(3): e4857. doi:10.1371/journal.pone.0004857


Discovery of Protein that Reactivates Herpes Simplex Virus
Landmark study provides molecular target for finding new and better cancer therapies

Research in PLoS Pathogens appears to solve a long standing medical mystery by identifying a viral protein, VP16, as the molecular key that prompts herpes simplex virus (HSV) to exit latency and cause recurrent disease.

Led by researchers at Cincinnati Children's Hospital Medical Center and the University of Cincinnati College of Medicine, the landmark study points to a molecular target for designing improved HSV vaccines and treatments. It also could direct refined engineering of HSV viruses used in cancer therapy, the investigators said.

The study was conducted in collaboration with the Medical Research Council Virology Unit of Glasgow, Scotland.

The two distinct lifestyles of HSV – active and latent – were first proposed 80 years ago. The virus replicates itself at the body surface, producing thousands of copies that can be transmitted to other people. In neurons, however, the virus can enter a silent state, where the viral genetic code can be maintained for the lifetime of the infected person.

"Our current findings show that, in elegant simplicity, the herpes simplex virus regulates this complex lifecycle through the expression of VP16," said Nancy Sawtell, Ph.D., author and researcher in the Division of Infectious Diseases at Cincinnati Children's Hospital Medical Center.

The study points to what causes the virus to periodically reactivate in latently infected neurons, prompting new rounds of virus replication at the body surface. By understanding how HSV achieves this complex interaction inside the human nervous system, researchers can gain crucial insight into how to control the spread of the virus. At present, there is no way to eliminate latent virus or prevent the virus from exiting latency. There also are no effective vaccines to protect people who are uninfected and transmission rates remain high, the researchers said.

In the study, the research team simulated high fever in a mouse model of HSV infection, demonstrating that VP16 must be produced before the virus can exit the latent state in neurons. Fever has long been known to induce HSV reactivation, and recurrent lesions are often called cold sores or fever blisters because of this association. In the vast majority of neurons, the virus remains latent. In a few neurons, however, the scientists observed that fever in the mice led to a stochastic, or random de-repression of VP16, causing the virus to exit latency and reactivate.

"This completely changes our thinking about how this virus reactivates from latency," said Richard Thompson, Ph.D., co-author and researcher in the Department of Molecular Genetics, Biochemistry and Microbiology at UC. "Instead of a simple positive switch that turns the virus on following stress, it appears instead to be a random de-repression of the VP16 gene that results in reactivation."

The leading infectious cause of blindness and acute sporadic encephalitis in the United States, HSV-1 is usually acquired during childhood. Both HSV-1 and HSV-2 can be sexually transmitted diseases that when passed to newborns during birth causes a severe and often fatal infection. As many as 80 percent or more of people are infected with HSV. Most of the time, people carrying the virus do not have symptoms, although they can still transmit the virus.

The researchers hypothesize that HSV usually remains latent because VP16, which normally enters the cell with the virus particle, does not make the long trip the virus takes through the nervous system and isn't transported efficiently to the nerve cell nucleus.

Future studies will use this new information to develop strategies to prevent or control herpetic disease, said Dr. Sawtell, who also is an associate professor of Pediatrics at UC.

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Funding support for the study came from National Institutes of Health.

Changes in Gene May Stunt Lung Development in Children
Mutations in a gene may cause poor lung development in children, making them more vulnerable to diseases such as chronic obstructive pulmonary disease (COPD) later in life, say researchers at the University of Pittsburgh Graduate School of Public Health and the German Research Center for Environmental Health

Their study, published online in Physiological Genomics, measured expression levels of the gene and its variants in both mouse lungs and children ages 9 to 11.

Study authors, led by George Leikauf, Ph.D., professor of occupational and environmental health at the University of Pittsburgh Graduate School of Public Health, and Holger Schulz, M.D., professor of medicine at the Institute of Lung Biology and Disease, German Research Center for Environmental Health, Munich, focused on a gene called superoxide dismutase 3 (SOD3), previously shown to protect the lungs from the effects of asbestos and oxidative stress.

“People lose lung function as they age, so it’s important to identify possible genetic targets that control healthy development of the lungs during childhood,” said Dr. Leikauf.

Drs. Leikauf, Schulz and colleagues compared SOD3 expression levels in strains of mice with poor lung function to one with more efficient airways and lungs two times the size. As with people, the lungs of mice fully form as they mature to adulthood. The better-functioning strain maintained higher levels of SOD3 – levels in these mice were four times higher at the final stage of lung development. They also found the presence of single nucleotide polymorphisms, or SNPs, variations in DNA sequences, in SOD3 that were linked to lung function in mice.

The researchers went on to assess SOD3 mutations in children ages 9 to 11 by testing for SNPs linked to lung function. After analyzing DNA from 1,555 children in Munich and Dresden who were part of the International Study of Asthma and Allergy in Children, they discovered two common SNPs associated with poorer lung function. One of these SNPs likely alters the expression levels of SOD3. Lung function was tested with spirometry, which measures the amount and speed of exhaled air.

Previously, genetic variants in SOD3 have been associated with loss of lung function in COPD, which is mainly caused by cigarette smoking. “We know SOD3 protects the lung against injury caused by chemicals in cigarette smoke, and it could be a link between childhood exposure to environmental tobacco smoke and poor lung development,” said Dr. Leikaf. In the future it might be possible to identify at-risk children and to develop a medication that would foster optimal lung development, he added. The researchers also are exploring sex differences in SOD3 gene expression and lung development, and girls appear to be at greater risk than boys.

COPD is the fourth leading cause of death in the United States, accounting for more than 120,000 deaths annually and costing more than $30 billion per year. It is estimated that more than 16 million Americans have COPD.

The study was funded by the National Institutes of Health and the German Research Center for Environmental Health.

Scientists Reveal Mechanism Regulating Cancer-Causing Gene
Two University of Rhode Island scientists have revealed how a cancer causing protein is regulated by reactive oxygen species (ROS) - a type of stress signal

Their findings provide new insight into how this protein normally behaves in human cells and may help in the design of drugs targeting specific cancers.

Doctoral student David J. Kemble and Professor Gongqin Sun in the URI Department of Cell and Molecular Biology are the first to provide a biochemical mechanism describing how certain protein tyrosine kinases sense and respond to oxidation. This sensing system was found to uniquely apply to two families of proteins implicated in numerous cancers: the Src and Fibroblast Growth Factor Receptor families of tyrosine kinases.

Their results were published online March 9 in the Proceedings of the National Academy of Sciences.

Src was the first enzyme identified as a cancer-causing gene in the early 1900's. For years scientists have been studying how the enzymes are expressed in cancer cells – what do they do and what controls them.

According to Kemble and Sun, Src is a master regulator of cell function, controlling cell metabolism, division, and death. In normal cells, the function of Src is turned off, and it is turned on only when certain stimulatory signals activate it. When the regulatory mechanisms that control Src activity are disrupted, Src may be turned on all the time, which turns the host cell into a cancer cell. Thus, it is crucial to understand how Src function is controlled.

Reactive oxygen species have long been viewed as damaging byproducts of oxygen-based metabolism. However, it is now recognized that ROS are produced when the cells are under growth stimulation, and they in turn regulate other cellular events. Accumulating evidence indicates that ROS can directly regulate the function of Src function, and thus indirectly control many cellular processes. Yet how Src responds to this regulation has remained elusive.

The URI scientists took a systematic approach, examined all the potential mechanisms, and identified the sensor that enables Src to respond to ROS regulation. They further found that the sensor is also present in several other similar enzymes, mostly in the FGFR family.

"Our results were surprising at first, given that the results contradict some reports in the literature," Kemble said. "But there was always a very clear answer to each question we asked. It was both unusual and exciting to see things progress as smoothly as it did."

According to Sun, this mechanism of regulation represents just a small piece of the large puzzle of how Src is controlled in the cells. "Src function is under the control of several different mechanisms; each one needs to fit in with the others to form a seamless regulatory system." Sun said.

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The URI scientists next plan to study how these pieces fit together to control Src function in the cells.

DNA Repair Mechanisms Relocate in Response to Stress
Like doctors making house calls, some DNA repair enzymes can relocate to the part of the cell that needs their help, a collaborative team of scientists at Emory University School of Medicine has found

The signal that prompts relocation is oxidative stress, an imbalance of cellular metabolism connected with several human diseases.

The study integrated the expertise of three Emory groups and resulted in a new level of understanding of the cell's response to genetic damage. The finding could lead to new targets for anti-cancer drugs that interfere with DNA repair, says Paul Doetsch, PhD, professor of biochemistry, radiation oncology, and hematology and oncology at Emory University School of Medicine.

The results were published in the February 1 issue of Molecular and Cellular Biology. The journal's editors chose an image of yeast cells with fluorescent DNA repair enzymes for the cover.

"DNA damage and oxidative stress are very closely related," Doetsch says. "For example, the way radiation inflicts most of its damage on DNA is through oxidative stress. The more we know about how cells respond to oxidative stress, the more chances there could be to influence those responses for diagnostic or therapeutic purposes."

The DNA inside cells is continually under assault by heat, radiation and oxygen. Cells have an extensive set of repair enzymes that comb through DNA, continually excising and re-copying damaged segments. To complicate matters, mitochondria (cells' miniature power plants) have their own DNA.

Working with Doetsch, Emory graduate students Lyra Griffiths and Dan Swartzlander, and biochemists Anita Corbett and Keith Wilkinson, genetically modified strains of yeast so that two different DNA repair enzymes would be fluorescent. They were able to follow the enzymes around the cell when yeast was exposed to hydrogen peroxide, causing oxidative stress, or to other chemicals causing DNA damage.

One DNA repair enzyme they studied, Ntg1, moves to the nucleus or the mitochondria depending on where DNA damage is concentrated, the authors found. In contrast, a related enzyme, Ntg2, stays in the nucleus under all conditions.

Cells appear to direct Ntg1's relocation by briefly attaching a small protein called SUMO to what needs to be moved around, the authors found. SUMO is found in fungi, plants and animals and is already being investigated by several research groups as a possible target for anti-cancer drugs.

Hormone-Mimics in Plastic Bottles - Just Tip of the Iceberg?
InStudy shows drinking water contaminated with potent estrogen

Plastic packaging is not without its downsides, and if you thought mineral water was ‘clean’, it may be time to think again. According to Martin Wagner and Jörg Oehlmann from the Department of Aquatic Ecotoxicology at the Goethe University in Frankfurt am Main, Germany, plastic mineral water bottles contaminate drinking water with estrogenic chemicals. In an analysis1 of commercially available mineral waters, the researchers found evidence of estrogenic compounds leaching out of the plastic packaging into the water. What’s more, these chemicals are potent in vivo and result in an increased development of embryos in the New Zealand mud snail. These findings, which show for the first time that substances leaching out of plastic food packaging materials act as functional estrogens, are published in Springer’s journal Environmental Science and Pollution Research.

Wagner and Oehlmann looked at whether the migration of substances from packaging material into foodstuffs contributes to human exposure to man-made hormones. They analyzed 20 brands of mineral water available in Germany – nine bottled in glass, nine bottled in plastic and two bottled in composite packaging (paperboard boxes coated with an inner plastic film). The researchers took water samples from the bottles and tested them for the presence of estrogenic chemicals in vitro. They then carried out a reproduction test with the New Zealand mud snail to determine the source and potency of the xenoestrogens.

They detected estrogen contamination in 60% of the samples (12 of the 20 brands) analyzed. Mineral waters in glass bottles were less estrogenic than waters in plastic bottles. Specifically, 33% of all mineral waters bottled in glass compared with 78% of waters in plastic bottles and both waters bottled in composite packaging showed significant hormonal activity.

By breeding the New Zealand mud snail in both plastic and glass water bottles, the researchers found more than double the number of embryos in plastic bottles compared with glass bottles. Taken together, these results demonstrate widespread contamination of mineral water with potent man-made estrogens that partly originate from compounds leaching out of the plastic packaging material.

The authors conclude: “We must have identified just the tip of the iceberg in that plastic packaging may be a major source of xenohormone* contamination of many other edibles. Our findings provide an insight into the potential exposure to endocrine-disrupting chemicals due to unexpected sources of contamination.”
*man-made substance that has a hormone-like effect


THURSDAY March 26, 2009---------------------------News Archive

The Egg Makes Sure That Sperm Don't Get Too Old
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.


Researchers Uncover Stem Cell Pathway
The discovery of a mechanism that regulates movement of blood-forming stem cells may help scientists increase the effectiveness of bone marrow transplants

Researchers at the Keck School of Medicine of USC have identified a signaling pathway that helps regulate the movement of blood-forming stem cells in the body - a finding that provides new insight into how stem cells move around the body and which may lead to improvements in the efficiency of bone marrow transplants.

The study will appear in the journal Nature.

“By identifying the key mechanism by which these stem cells engraft to the bone marrow, it may be possible to pharmacologically treat the cells to activate this pathway and thus increase the effectiveness of bone marrow transplants,” said lead author Gregor Adams, assistant professor of cell and neurobiology at the Keck School and a researcher at the Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC.

Hematopoietic stem cells are blood-forming cells that circulate through the body, shifting back and forth between the bloodstream and bone marrow, Adams explained.

When patients receive a bone marrow transplant, healthy blood stem cells are injected into their veins. Unless those stem cells can find their way into a specific site known as the stem cell niche, they cannot develop properly to replenish the white cells, red cells and platelets in the patient’s blood.

The mechanisms that guide the cells during this migration have not been well understood. However, in this study the researchers found that blood-forming stem cells that lacked a specific signaling molecule, called GalphaS, did not home to or engraft in the bone marrow of adult mice, Adams said.

“Here we show that the GalphaS pathway is a critical intracellular pathway involved in this process,” he said. “Currently, large numbers of blood-forming stem cells are required in bone marrow transplantation due to the limited efficiency of the homing process. This study opens up the possibility of treating bone marrow cells with GalphaS pathway activators as a means to increase the effectiveness of bone marrow transplants.”

Improving the efficiency with which stem cells colonize the bone marrow following transplantation could have far-reaching implications for disease treatment, according to Martin Pera, director of the Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC.

“For example, such a discovery might enhance the utility of umbilical cord blood, which contains only limited numbers of stem cells, for the treatment of cancer and blood disorders in children and adults,” Pera said.

The study was funded by the National Heart, Lung & Blood Institute.

Stopping Autoimmunity Before it Strikes
Current research describes a new method to track the development of autoimmune diseases before the onset of symptoms

The related report by Zangani et al, "Tracking early autoimmune disease by bioluminescent imaging of NF-κB activation reveals pathology in multiple organ systems," appears in the April 2009 issue of The American Journal of Pathology.

Autoimmune diseases such as lupus, multiple sclerosis, rheumatoid arthritis and diabetes are caused when the immune system attacks the body's own cells. Normally, immune cells are prevented from attacking normal cells; however, in patients with autoimmune disease, this "tolerance" is lost. The immediate causes of autoimmune diseases remain unknown, partially due to the inability to detect disease before the onset of symptoms. Early detection of autoimmune disease is critical for assessing new treatments.

Stopping Autoimmunity

The molecule NF-κB is activated by inflammation, which plays a key role in autoimmune disease development, making NF-κB a prime candidate to track autoimmune activity. Researchers at the University of Oslo led by Drs. Ludvig Munthe and Bjarne Bogen in collaboration with Rune Blomhoff engineered NF-κB such that it would emit light when activated. Using a mouse model of systemic autoimmunity with features of lupus, they found that NF-κB activation signals were present in affected organs several weeks before the clinical manifestations of disease. The light signal intensity correlated with disease progression. NF-κB tracking may therefore provide a new tool in the evaluation of early autoimmune therapies.

The article from Zangani et al "indicate[s] that NF-κB mediated bioluminescence is a very sensitive and early indicator of inflammation and disease", allowing precise identification of incipient disease sites for biomedical and pathogenetic studies. In future studies, Drs. Munthe, Bogen, and colleagues will utilize this new model "for studies on early intervention, e.g. drug treatment, to prevent or treat autoimmune disease", and for studies of the development of B cell lymphoma.

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The Norwegian Research Council, The University of Oslo, Medinnova and Rikshospitalet Medical Center funded the work.

Zangani M, Carlsen H, Kielland A, Os A, Hauglin Harald, Blomhoff Rune, Munthe L A, Bogen B: Tracking early autoimmune disease by bioluminescent imaging of NF-κB activation reveal pathology in multiple organ systems. Am J Pathol 2009 174: 1358-1367


Therapeutic Cloning Gets Boost With New Research Findings
San Antonio and Honolulu researchers make important discoveries about point mutation rates in cloned mouse fetuses

Germ cells, the cells which give rise to a mammal's sperm or eggs, exhibit a five to ten-fold lower rate of spontaneous point mutations than adult somatic cells, which give rise to the body's remaining cell types, tissues and organs. Despite their comparatively higher mutation rates, however, adult somatic cells are used as the donor cells in a cloning process called somatic cell nuclear transfer (SCNT). This made researchers wonder if cloning by SCNT leads to progeny with more mutations than their naturally conceived counterparts. Also, would cloned fetuses receive DNA programming predisposing them to develop mutations faster than natural fetuses of the same age?

Those scenarios are simply not likely, say researchers at The University of Texas at San Antonio, The University of Texas Health Science Center at San Antonio and The University of Hawaii at Honolulu's John A. Burns School of Medicine. The team, which spent more than five years analyzing mutation rates and types in cloned Big Blue® mouse fetuses recently published its findings in the online Early Edition of the Proceedings of the National Academy of Sciences in a paper titled "Epigenetic regulation of genetic integrity is reprogrammed during cloning."

The paper offers the first direct demonstration that cloning does not lead to an increase in the frequency of point mutations.

John McCarrey, professor of cellular and molecular biology at UTSA and the study's principal investigator, suggests a "bottleneck effect" is partially responsible for the observations his team recorded. "To create a cloned fetus by somatic cell nuclear transfer, only one adult somatic cell -- one donor cell -- is needed," he explains. "Because a random cell population exhibits a low mutation rate overall and only one cell from that population is used for cloning, the likelihood is remote that the cell chosen to be cloned will transfer a genetic mutation to its cloned offspring. Therefore, the bottleneck effect limits the transfer of mutations from donor cells to cloned offspring."

Not only did the researchers find that SCNT does not lead to an increase in the frequency of point mutations in cloned mice, the team also found that naturally conceived fetuses and cloned fetuses that are the same age have similar rates of spontaneous mutation development. They attribute this finding to epigenetic reprogramming.

It is known in the scientific community that germ cells contain an epigenome, a programmed state of the genome, that keeps mutation rates low. They suggest this type of epigenome is found in germ cells because those cells are responsible for contributing genetic information to subsequent generations. Adult somatic cells (the donor cells in SCNT) have higher mutation rates and less stringent epigenetic programming to avoid mutations than germ cells, but offspring produced from somatic cells by cloning have mutation rates similar to those in offspring produced by natural reproduction, suggesting that the epigenome of an adult somatic cell is reprogrammed during cloning to maintain the genetic integrity of that cell's progeny.

The Matchmaker that Maintains Neuronal Balance
A protein identified by researchers at Baylor College of Medicine helps maintain a critical balance between two types of neurons, preventing motor dysfunction in mammals

In a report in the current edition of the journal Neuron, Dr. Soo-Kyung Lee, assistant professor of molecular and human genetics, molecular and cellular biology and neuroscience at BCM, and her colleagues describe the protein LMO4 as critical in allowing progenitor cells to choose their fates – between the V2a neurons that are excitatory and the V2b neurons that are inhibitory. Excitatory neurons encourage the activity of neurons on which they act. Inhibitory neurons act in an opposite manner.

In previous work, Lee and members of her laboratory identified the double-barreled or dimerized complex containing the protein Lhx3 that pushes the progenitor cells to become V2a excitatory neurons. In this paper, she notes the LMO4 not only forms a complex that binds to DNA and promotes the choice of cell fate to the V2b inhibitory neurons, it also blocks the path to becoming a V2a excitatory neuron.

Because LMO4 cannot bind directly to DNA, it plays matchmaker instead, building a complex of DNA-binding components that allow the cells the choice to become inhibitory neurons.

"These individual DNA-binding components are present in the neurons," she said. "But they do not have the ability to find their DNA partners. LMO4 'glues' these proteins together and makes them functional."

She and her colleagues have demonstrated these both in the laboratory and in mice bred to lack LMO4. Without the protein, the balance becomes tipped in favor of excitatory neurons, which would result in motor dysfunction.

Others who took part in this research include Kaumudi Joshi, Seunghee Lee, Bora Lee and Jae W. Lee, all of BCM.

Lee credits graduate student Kaumdi Joshi with much of the laboratory work in accomplishing this understanding.

Funding for this research came from the National Institute of Neurological Disorders and Stroke, the Pew Scholars in Biomedical Science Program, the March of Dimes Foundation and the BCM Intellectual and Developmental Disabilities Research Center.

The paper is available at http://www.cell.com/neuron/home.


WEDNESDAY March 25, 2009---------------------------News Archive

UK to Begin Trial of Blood From Embryonic Stem Cells
UK scientists plan a major research project to see if synthetic human blood can be made from embryonic stem cells

Led by the Scottish National Blood Transfusion Service, the three year trial could provide an unlimited supply of blood for emergency transfusions. The blood should be free of infections like the human form of mad cow disease.

Teams will test human embryos left over from IVF treatment to find those destined to develop into the universal "O-negative" blood donor group.

O-negative blood can be transfused into anyone without fear of tissue rejection and is the only safe option when a patient's blood group is unknown or not immediately available. This precious blood is in limited supply because only 7% of the population belongs to this blood group.

The Wellcome Trust is understood to have promised £3m towards the cost of the multimillion-pound project, with further funding coming from the blood transfusion services of Scotland, and England and Wales. The Irish government is also believed to be involved.

The project will be led by Professor Marc Turner of Edinburgh University who is the director of the Scottish National Blood Transfusion Service. He said the work would begin in the next few weeks after final approval had been gained from the relevant research bodies.

Making blood from embryos
1. Embryo created from IVF is tested for O-negative blood group, then allowed to develop for several days until stem cells can be extracted

2. Stem cells are cultured in laboratory with nutrients to stimulate red blood cell creation

3. Nuclei are removed in final stage to produce oxygen-carrying mature blood cells. Trillions of these will be needed to build up a blood bank

Stem cells are the body's master cells, with the ability to transform into any type of tissue.

Scientists have already shown it is possible to take a single stem cell from an early human embryo and encourage it to develop into mature blood cells in the laboratory. And a US firm called Advanced Cell Technology has managed to produce billions of red blood cells from embryonic blood cells in this way. The challenge now is to scale up the production and move the science from the lab to the bedside, which will take years.

Professor Turner said: "We should have proof of principle in the next few years, but a realistic treatment is probably five to 10 years away. "In principle, we could provide an unlimited supply of blood in this way." However, many groups object to the use of embryonic stem cells on the grounds that it is unethical to destroy embryos in the name of science.

Josephine Quintavalle of the public interest group Comment on Reproductive Ethics said: "Like so many of the claims associated with embryonic stem cells, this is first steps research rather than a cure around the corner, and just as hypothetical as the rest of the claims which try to justify destroying the human embryo for the benefit of mankind.

"Associating this controversial research with a National Blood Transfusion service may even end up contaminating the feel-good image of blood banks.

"Those who donate blood but who defend the right to life of the human embryo may be reluctant to continue giving their blood."

FROM THE BBC WORLD SERVICE
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Diabetics on High-Fiber Diets Might Need Extra Calcium
The amount of calcium your body absorbs might depend, in part, on the amount of dietary fiber you consume

Researchers at UT Southwestern Medical Center report that patients with noninsulin-dependent diabetes (type 2) excreted less calcium through their urine when they consumed 50 grams of fiber a day than when they ate 24 grams a day. Excreting less calcium indicates that they absorbed less of the mineral.

“We already know that fiber helps improve your cholesterol and glucose control and improves your bowel regularity. Our new findings suggest that dietary fiber reduces the body’s capacity to absorb calcium,” said Dr. Abhimanyu Garg, professor of internal medicine and an investigator in the Center for Human Nutrition at UT Southwestern. He is senior author of a study appearing online in Diabetes Care. “Because more calcium equals better bone health, we recommend that people on high-fiber diets talk to their physician about increasing their dietary calcium as well, in order to get the most benefit from both.”

Dr. Garg said it’s important to speak with a physician or a registered dietitian before increasing your calcium intake because excessive levels may cause kidney stones.

The American Diabetes Association (ADA) recommends a daily intake of 24 grams of dietary fiber, but the average American consumes about 14 to 15 grams of fiber a day.

Sometimes called “roughage,” dietary fiber is the indigestible portion of plant foods that pushes food through the digestive system, absorbing water and easing defecation. Calcium is a nutrient found in food that is absorbed by the body and then excreted in urine, feces or sweat. It is the most abundant mineral in the human body.

Prior research at UT Southwestern has shown that a high intake of dietary fiber, mostly from fruits and vegetables, lowers blood glucose levels and leads to decreased insulin levels in the blood, as well as lowering blood lipid concentrations in patients with type 2 diabetes, the most prevalent type of diabetes.

For the current study, 13 patients with type 2 diabetes ate either a high-fiber diet (50 grams per day) or the moderate-fiber diet (24 grams per day) recommended by the ADA for six weeks, then switched to the other diet for six weeks. All participants stayed at UT Southwestern’s Clinical and Translational Research Center (CTRC) for the final week of each six-week period.

CTRC staff prepared both diets so that they contained the same number and proportion of calories from carbohydrates, fats and proteins, as well as an equal amount of minerals such as calcium, phosphorous, magnesium, sodium and potassium. The high-fiber diet included numerous fiber-rich foods including cantaloupe, grapefruit, papaya, okra, winter and zucchini squash, granola and oatmeal. No supplements were used.

“The reduction of urinary calcium excretion on high-fiber diets tells us that the amount of dietary fiber has a direct impact on calcium absorption,” Dr. Garg said. “In other words, the participants excreted less calcium on the high-fiber diet because the additional fiber caused their bodies to absorb less calcium.”

Though most of the additional fiber in the high-fiber diet was soluble fiber, Dr. Garg said he cannot say for sure whether soluble or insoluble fiber affects calcium absorption.

“Generally, more fiber of either type is beneficial,” he said. “We should encourage people to try food sources rich in fiber and calcium such as spinach, broccoli, figs, papaya, artichoke, okra, beans, mustard and turnip greens, and cactus pads.”

Other UT Southwestern researchers involved in the study were Dr. Meena Shah, lead author and clinical associate professor of clinical nutrition; Dr. Manisha Chandalia, clinical associate professor of internal medicine with the Center for Human Nutrition; Beverley Adams-Huet, assistant professor of clinical sciences; Linda Brinkley, former research dietitian; Dr. Khashayar Sakhaee, chief of mineral metabolism; and Dr. Scott Grundy, director of the Center for Human Nutrition.

The work was funded by the National Institutes of Health and Southwestern Medical Foundation.

Visit http://www.utsouthwestern.org/endocrinology to learn more about UT Southwestern’s clinical services in endocrinology, including diabetes treatment and nutrition.

Life and Death Protein Regulates Immune System Response
White blood cells called neutrophils and macrophages are the first responders of the immune system. They serve as the first line of defense against invading microbes—identifying them, engulfing them and eliminating them. However, once these protective cells have obliterated their quarry, they must quickly commit suicide, so the immune system can return to normal and the body can dispose of the toxic microbial waste and damaged cells

St. Jude researchers have now established a central regulator of this knife-edge balance between life and death of myeloid lineages. They have found that a gene called MCL-1 produces a protein that protects neutrophils against cell suicide, or apoptosis, as they mature in the bone marrow. However in macrophages, the MCL-1 protein appears to be dispensable for maturation, but instead governs the macrophages’ life-and-death balance, promoting survival while the macrophage works to eliminate extracellular microbes.

“The fine-tuned balance mediated by MCL-1 is critical because you want to protect the macrophages when they encounter a pathogen and allow them to do their job,” said Joseph Opferman, PhD, Biochemistry. “But after the clearance of the microbe, you need to rapidly downregulate the immune response by eliminating the cells that have been recruited to the infection site and return the immune system to normal homeostasis. Otherwise, an accumulation of active inflammatory cells can lead to tissue destruction.”

Opferman is the senior author of a report on this work published in the December 8, 2008, advanced online issue of the journal Blood.

The researchers’ basic findings of MCL-1’s function could yield insights into its role in such disorders as sepsis, an often lethal inflammation in which the immune system goes out of control. Also, the finding could yield insight into leukemias in which MCL-1 levels are known to increase, contributing to the abnormally prolonged life of the malignant cells, Opferman said.

Previous studies in the Opferman lab indicated that MCL-1 was protective during the birth stages of white blood cells in the bone marrow. In the scientists’ new experiments, they sought to understand whether MCL-1 was also important later, as neutrophils and macrophages differentiate into mature cells and carry out their function.

To study MCL-1’s role in myeloid function, the researchers used a conditional knockout technique that allowed them to switch off the MCL-1 gene specifically in the neutrophil and macrophage lineages. They needed to use a conditional knockout technique because MCL-1 is so critical throughout development that simply deleting it in fertilized mouse eggs results in death at an extremely early embryonic stage.

These knockout experiments establish that in neutrophils MCL-1 is critical to their differentiation into mature cells. The researchers’ experiments yield clues about how MCL-1 functions with other known components of the cell death machinery in neutrophils.

In studies with macrophages, the researchers found that while MCL-1 was not necessary for development and basic function, its loss rendered the macrophages sensitive to death when they ingested microbes.

“This is one of the most striking findings of our study,” Opferman said. “In all other blood cell lineages, if you delete MCL-1, those cells are basically gone. But macrophages survive, and we want to find out why they survive, and why losing MCL-1 only makes them more susceptible to cell death.”

Besides yielding insights into inflammation, deeper understanding of MCL-1’s normal role may also help in developing treatment strategies for myeloid leukemia, Opferman said.

“MCL-1 is highly expressed in a lot of different leukemias and lymphomas, and many groups are developing treatments to antagonize MCL-1 function in order to kill cancer cells,” he said. “As we come to understand the primary, normal functions of MCL-1, we appreciate that simply eliminating its function might have significant detrimental side effects. So, our data suggest that treatment strategies should aim at modulating MCL-1 protein levels, without completely getting rid of its function.”

In further studies, Opferman and his colleagues will trace how MCL-1 is regulated in the immune cells to aid development of strategies to precisely adjust levels of the protein.

Other authors of this paper include Kelli Boyd, DVM, PhD, Veterinary Pathology Core; Gerard Zambetti, PhD, Biochemistry; and Desiree Steimer, formerly of St. Jude.

This research was supported in part by the Pew Scholars Program in the Biomedical Sciences, the National Cancer Institute and ALSAC.


Human Cytomegalovirus Can Stimulate Telomerase Activity
Infection with human cytomegalovirus (HCMV) triggers expression of the telomerase reverse transcriptase (hTERT) gene in both normal human cells and tumor cells grown in culture

HCMV has been suspected of causing or promoting cancer, but the mechanism by which the virus acts has been unclear. Several other cancer-related viruses stimulate telomerase activity, which promotes cell proliferation and immortalization.

To learn whether HCMV induces telomerase activity, Dawei Xu, M.D., Ph.D., of the Karolinska Institute in Stockholm, Sweden, and colleagues infected normal diploid fibroblasts and malignant glioma cells with HCMV or introduced a copy of the viral immediate early antigen 72 (IE-72) gene into the cells.

Infection with HCMV induced telomerase expression in both normal and malignant human cells. Introduction of the viral IE-72 gene also induced telomerase activity in both cell types.

"In this study, we reveal a novel mechanism through which HCMV may be linked to or modulate oncogenesis by demonstrating that HCMV stimulates hTERT transcription, thereby activating telomerase, which is essential for the immortalization and transformation of human cells," the authors write.

In an accompanying editorial, Jindrich Cinatl Jr., Ph.D., of the Universität Frankfurt in Germany and colleagues note that they hypothesized previously that HCMV may increase tumor malignancy rather than instigate malignancy. The results by Xu and colleagues may provide important mechanistic insight as to how such a modulatory role could occur.

"…HCMV-induced telomerase activation represents a mechanism that is of possible relevance for both (initiation of) malignant trans¬formation and oncomodulation by HCMV infection," the editorialists write.

Contacts
Article: Dawei Xu, dawei.xu@ki.se
Editorial: Jindrich Cinatl, cinatl@em.uni-frankfurt.de, +49 (0)69 6301 6409

Genetic Changes Outside Nuclear DNA Suspected to Trigger More Than Half of All Cancers
A buildup of chemical bonds on certain cancer-promoting genes, a process known as hypermethylation, is widely known to render cells cancerous by disrupting biological brakes on runaway growth. Now, Johns Hopkins scientists say the reverse process — demethylation — which wipes off those chemical bonds may also trigger more than half of all cancers

One potential consequence of the new research is that demethylating drugs now used to treat some cancers may actually cause new cancers as a side effect.

“It’s much too early to say for certain, but some patients could be at risk for additional primary tumors, and we may find that they need a molecular profile of their cancer before starting demethylating therapy,” says Joseph Califano, M.D., professor of otolaryngology–head and neck surgery and oncology at Johns Hopkins.

The findings, based on studies of normal and cancer cells from human mouth, nose and throat tissue, provide more evidence that important regulators of gene activity occur outside as well as inside DNA in a cell’s nucleus.

“While cancer-causing and other mutations alter vital protein-making pathways by rewriting the gene’s DNA code, epigenetic changes affect genes without changing the code itself. The new studies tell us that such changes occur not only when methyl groups bond to a gene’s on-off switch, but also when they come unglued,” says Califano.

Califano says sporadic reports of demethylation as a tool in activating cancer-promoting genes led his team to develop a systematic way to discover these epigenetic changes and show how the process is linked to cancer.

To gather their evidence, Califano and his group treated two cell lines from normal oral tissue with the demethylating drug 5-azacytidine and collected a list of genes that were activated as a result. They used special silicon chips carrying pieces of genetic material that allow thousands of genes to be analyzed at one time to locate genes activated by demethylation.

The list was cross-referenced with genes “turned on” in 49 head and neck cancer samples and 19 normal tissue samples. In all, Califano and his team found 106 genes specific to head and neck cancer that were activated by the demethylation process. “Some of the genes regulate growth, others metabolize sugars and some have already been linked to cancer development,” says Califano. A report on this work appears on March 23 in PLoS One.

Further analysis by the Johns Hopkins team revealed a single connection among 106 genes: methylation within them is regulated by another gene called BORIS. BORIS acts as a “master regulator,” recruiting other proteins to demethylate a coordinated set of genes and signaling the development of cancer. According to the scientists, nearly 60 percent of a wide range of cancers, including head and neck and lung cancer, have high levels of BORIS expression.

He envisions that agents like 5-azacytidine may need to be combined with a “BORIS blocker,” a drug that has yet to be developed to protect patients who need demethylating therapies.

Dr. Califano maintains a clinical practice at Johns Hopkins Head and Neck Surgery at Greater Baltimore Medical Center and the Milton J. Dance Jr. Head and Neck Center.

The research is funded by the Flight Attendant Medical Research Institute, the National Institute of Dental and Craniofacial Research, and the National Cancer Institute.

Research participants included Ian M. Smith, Chad A. Glazer, Suhail K. Mithani, Michael F. Ochs, Wenyue Sun, Sheetal Bhan, Andrew Gray, Chunyan Liu, Steven S. Chang, Kimberly L. Ostrow, William H. Westra, Shahnaz Begum and Mousumi Dhara from Johns Hopkins; and Alexander Vostrov, Ziedulla Abdullaev and Victor Lobanenkov from the National Institutes of Health.

Johns Hopkins Kimmel Cancer Center.

Therapeutic Cloning Gets Boost With New Research Findings
San Antonio and Honolulu researchers make important discoveries about point mutation rates in cloned mouse fetuses

Germ cells, the cells which give rise to a mammal's sperm or eggs, exhibit a five to ten-fold lower rate of spontaneous point mutations than adult somatic cells, which give rise to the body's remaining cell types, tissues and organs. Despite their comparatively higher mutation rates, however, adult somatic cells are used as the donor cells in a cloning process called somatic cell nuclear transfer (SCNT). This made researchers wonder if cloning by SCNT leads to progeny with more mutations than their naturally conceived counterparts. Also, would cloned fetuses receive DNA programming predisposing them to develop mutations faster than natural fetuses of the same age?

Those scenarios are simply not likely, say researchers at The University of Texas at San Antonio, The University of Texas Health Science Center at San Antonio and The University of Hawaii at Honolulu's John A. Burns School of Medicine. The team, which spent more than five years analyzing mutation rates and types in cloned Big Blue® mouse fetuses recently published its findings in the online Early Edition of the Proceedings of the National Academy of Sciences in a paper titled "Epigenetic regulation of genetic integrity is reprogrammed during cloning."

The paper offers the first direct demonstration that cloning does not lead to an increase in the frequency of point mutations.

John McCarrey, professor of cellular and molecular biology at UTSA and the study's principal investigator, suggests a "bottleneck effect" is partially responsible for the observations his team recorded. "To create a cloned fetus by somatic cell nuclear transfer, only one adult somatic cell -- one donor cell -- is needed," he explains. "Because a random cell population exhibits a low mutation rate overall and only one cell from that population is used for cloning, the likelihood is remote that the cell chosen to be cloned will transfer a genetic mutation to its cloned offspring. Therefore, the bottleneck effect limits the transfer of mutations from donor cells to cloned offspring."

Not only did the researchers find that SCNT does not lead to an increase in the frequency of point mutations in cloned mice, the team also found that naturally conceived fetuses and cloned fetuses that are the same age have similar rates of spontaneous mutation development. They attribute this finding to epigenetic reprogramming.

It is known in the scientific community that germ cells contain an epigenome, a programmed state of the genome, that keeps mutation rates low. They suggest this type of epigenome is found in germ cells because those cells are responsible for contributing genetic information to subsequent generations. Adult somatic cells (the donor cells in SCNT) have higher mutation rates and less stringent epigenetic programming to avoid mutations than germ cells, but offspring produced from somatic cells by cloning have mutation rates similar to those in offspring produced by natural reproduction, suggesting that the epigenome of an adult somatic cell is reprogrammed during cloning to maintain the genetic integrity of that cell's progeny.


TUESDAY March 24, 2009---------------------------News Archive

Redefining DNA: Darwin From the Atom Up
In a dramatic rewrite of the recipe for life, scientists from Florida today described the design of a new type of DNA with 12 chemical letters instead of the usual four

Presented here at the 237th National Meeting of the American Chemical Society (ACS), this artificial genetic system already is helping to usher in the era of personalized medicine for millions of patients with HIV, hepatitis and other diseases.
The research may also shed light on how life arose on Earth, by producing a self-sustaining molecule capable of Darwinian evolution and reproduction, much like one that many scientists suggest arose at the dawn of life on Earth nearly four billion years ago.

Led by Steven Benner, Ph.D., this team is rewriting the rulebook that Nobel laureates James Watson and Francis Crick started when they described DNA's structure in 1953. One of the crowning discoveries of 20th century science, Watson and Crick's discovery established how the four chemical "letters" of DNA — A, T, C and G — pair up.

"This is a man on the moon goal," says Steven Benner, Ph.D. "It has dragged us kicking and screaming into uncharted territory. But we've learned all sorts of reasons about how the Watson and Crick rules don't enable technology to do useful things like highly parallel amplification of DNA or highly parallel diagnosis of human diseases. These things are worth a lot of money."

These pairing rules, for instance, make it very difficult for researchers to develop multiplexed diagnostic tests for viral diseases — tests that require identification and tagging of viral DNA. Old methods used regular DNA to bind and tag foreign genetic material. But natural DNA would often bind with non-disease DNA and generate confusing false positive and false negative results.

Benner's artificial genetic system does not operate under Watson-Crick rules, so the tagging gives accurate results. Benner's artificial alphabet already has been applied commercially. It is the basis of a viral load detector, which helps personalize the health care of those 400,000 patients annually infected with hepatitis B, hepatitis C, and HIV, the cause of AIDS.
"This is a hundred million dollar product right now," Benner noted. "It's used to manage cystic fibrosis, as well. We can also use this technology to go into biological samples and extract known genes with cancer-causing mutations. We can do all of this because we have an artificial DNA system.

For patients with HIV and hepatitis, the viral load detector can mean the difference between life and death.

Modern drug cocktails for these diseases are highly effective, reducing the viral load in the bloodstream to nearly zero. But at some point, the virus mutates, enabling it to evade the drugs and repopulate. As the viral tide rises, there are no outward symptoms in the patient, so the mutated strain is often discovered long after the virus has spread again.

The viral load detector, which relies on Benner's 12 letter system to tag DNA, may change that.

"What we want to do with personalized care is to give you a cocktail, and then monitor you and discover when the virus becomes resistant to it," explains Benner. "Now we don't want to do that too soon – that would waste a lifetime of good viral inhibitors — but not too late, of course. The patient would go in once a month to get their viral load measured. At some point the virus mutates and its viral load goes up. Then you know you better change the cocktail."

Benner says that the artificial DNA system is poised to become an essential tool in genomics research. The 12 letter alphabet already underlies new work at the National Human Genome Research Institute to connect large quantities of genomic data with human medicine.

The 12 letter system might also shed light on one of most mysterious times in Earth's history — the dawn of life nearly four billion years ago. Many scientists believe that this might have occurred when DNA's ancient cousin, RNA, began to act like a living organism.

"The idea has been that life originated on earth as RNA molecules assembled randomly and spontaneously in the prebiotic soup," says Benner. "Then, one of them found the ability to make copies of itself. In doing so, it made those copies with imperfections, so that some of its 'kids' were a bit better. Most were worse, so the better ones took over more resources. That started Darwinian processes. The rest is history."

Benner's ultimate goal is to synthesize a similar life form in his lab at the Foundation for Applied Molecular Evolution. His 12 letter genetic system is capable of nearly all of the actions that define a living thing — reproduction, growth and response to its environment — all without the benefit of genes refined over billions of years of evolution.

"But it still isn't self-sustaining," Benner explains. "You need a graduate or post-doc to come in the morning and feed it. It doesn't look for its own food. No one has gotten that first step to work. If you start making estimates of how many molecules you have to look for in order to find one that does this, you're talking about 10,000,000,000,000,000,000,000,000,000,000,000 molecules."

While Benner continues to pursue a chemical system fully capable of Darwinian evolution, he emphasized the lessons already learned from the development of the 12 letter system.
"We haven't just taken things from nature, but we've actually understood something about how chemical structure is related to genetic behavior. With that, we've been able to make new versions of it," says Benner.

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The American Chemical Society is a nonprofit organization chartered by the U.S. Congress. With more than 154,000 members, ACS is the world's largest scientific society and a global leader in providing access to chemistry-related research through its multiple databases, peer-reviewed journals and scientific conferences. Its main offices are in Washington, D.C., and Columbus, Ohio.


DNA Duplication: A Mechanism for 'Survival of the Fittest'
The end of an era
Some 65 million years ago, the earth's most recent 'mass extinction' took place. One or more catastrophic events - such as a comet strike or increased volcanic activity - produced widespread fires and clouds of dust and smoke that obstructed sunlight for a long period of time. These adverse conditions killed off about 60% of the plant species and numerous animals, including the dinosaurs. Only the most well-adapted plants and animals were able to survive this mass extinction - but what is 'most well-adapted'?

A role for DNA duplication?
effrey Fawcett, Steven Maere and Yves Van de Peer (VIB-UGent) have been working as bioinformatics specialists to decode various plant genomes - the complete content of a plant's DNA - ranging from small weeds to tomatoes and rice to trees. Time and again, they have been confronted with the fact that, over the course of the history of these plants, their entire DNA was duplicated one or more times. By means of sophisticated research techniques, they have dated these duplications as closely as possible.

Yves Van de Peer's group then noticed that the most recent duplications occurred at approximately the same time in all of the plants. But, in terms of evolution, 'the same time' is relative: the DNA duplications occurred between 40 and 80 million years ago. So, the bioinformaticians worked to refine the dating. Thanks to their expertise in comparative genome studies and their extensive database, they were able to make a very precise dating of the duplications on the basis of standard evolution trees. This indicated that, in all of the plants under study, the most recent genome duplication occurred some 65 million years ago - thus, at the time of the last mass extinction.

A universal mechanism
From these results, the VIB researchers concluded that plants with a duplicated genome were apparently the 'most well-adapted' for survival in the dramatically changed environment. Normally, in unaltered circumstances, duplications of DNA are disadvantageous. In fact, they cause very pronounced properties that are not desired in an unaltered environment. However, in radically changed circumstances, these very properties can make the organism better adapted to the new climate.

In previous research, Yves Van de Peer had discovered very old genome duplications in early ancestors of vertebrates and fish. At that time, he showed that these duplications were probably crucial for the development of vertebrates and thus of human beings as well. So, genome duplication is probably a universal mechanism that has ensured that the role of our planet's vertebrates and flowering plants has become much greater over time.s.

Tsai and her colleagues then dissected the precise molecular role that DISC1 plays in brain stem cells. She found that the protein acts like lithium, a drug commonly prescribed as a mood stabilizer for patients with bipolar disorder. In particular, DISC1 inhibits the enzyme GSK3beta, the same enzyme inhibited by lithium. “Lithium is known to inhibit GSK3beta directly and indirectly, so it looks like DISC1 behaves like endogenous lithium,” says Tsai. Bipolar disorder and schizophrenia are often diagnosed together, and many psychiatrists consider the two disorders intimately linked, perhaps even lying along a spectrum.

In a final set of experiments, Tsai’s team depleted DISC1 activity in the brain stem cells of adult mice, and then treated the mice with a molecule that inhibits GSK3beta much like lithium does. The brain stem cells returned to normal, revving up production of new neurons. Further, the previously agitated and depressed animals recovered and began behaving normally.

“This drug-like small molecule completely reverses the deficiencies seen in the neural progenitor cells while alleviating the abnormal behavior in the mice,” says Tsai.

Tsai’s team also identified the key segment of the DISC1 protein that inhibits GSK3beta. Knowing the structure of this protein segment may help drug developers as they pursue better treatments for schizophrenia and related disorders, Tsai says. “We hope this information will lead to better treatments for schizophrenia, which are sorely needed.”

Tsai is now working with geneticists at the Stanley Center for Psychiatric Research in Cambridge, Massachusetts, where she holds a joint appointment, to identify additional variations in the DISC1 gene. “We need to get a handle on the genetics of schizophrenia,” says Tsai. “But now we know how DISC1 probably contributes to the disorder, which is a big step.”

Common Gene Variants Influence Risk Factor for Sudden Cardiac Death
Study identifies gene variants that influence QT interval

A new study has identified several common genetic variants related to a risk factor for sudden cardiac death. The report receiving early online release in the journal Nature Genetics identifies variants in genes, some known and some newly discovered, that influence the QT interval measured on the electrocardiogram (EKG) performed routinely in doctors' offices. These findings could eventually help to prevent sudden cardiac death and arrhythmia by limiting the use of medications that affect QT interval in people with these variants.

The QT interval is the time from the beginning of electrical activation of the heart to the end of electrical relaxation. "It is well established that prolongation of the QT interval in the general population is a potent and heritable risk factor for sudden death," said Christopher Newton-Cheh, MD, MPH, of the Massachusetts General Hospital (MGH) Center for Human Genetic Research and Cardiovascular Research Center (CVRC) and lead author of the Nature Genetics article. "In addition, QT prolongation results from medications leading to drug-induced cardiac arrhythmias and sudden death. This is a cardiotoxic side effect of scores of medications in widespread use and has been a major barrier to the development of novel drugs. From studies of families with congenital long-QT syndrome, we know that rare mutations with strong effects on ion channel function lead to QT prolongation and sudden death. But the common genetic basis for QT prolongation has been very difficult to establish."

To search for QT-associated variants, the investigators formed the QTGEN consortium, assembling more than 13,000 individuals from three studies – including the National Heart, Lung and Blood Institute's and Boston University's Framingham Heart Study, the Rotterdam Study and the Cardiovascular Health Study. All individuals had undergone testing of hundreds of thousands of common gene variations called single-nucleotide polymorphisms (SNPs).

By pooling results from the three studies, investigators identified 14 common variants in 10 different gene regions (some regions had more than one variant) that were related to QT interval duration. A separate companion paper from the QTSCD consortium in the same issue of Nature Genetics – led by Arne Pfeufer, MD, of the Technical University of Munich and Aravinda Chakravarti, PhD, Johns Hopkins University School of Medicine – included more than 15,000 individuals from Europe and the US. This independent study strongly confirmed 12 out of 14 of the QTGEN variants and identified two additional gene regions. "We were very reassured to see such strong replication in two independent studies" said Newton-Cheh.

The QTGEN investigators examined the effect of a genotype score comprised of all 14 variants tested together. The top 20 percent of the population with the highest genotype scores had 160 to 210 percent higher risk of prolonged QT interval compared to the 20 percent of the population with the lowest scores. They also had around 10 millisecond longer QT intervals, which is approximately the degree of QT interval prolongation observed for some drugs pulled from the market for arrhythmias.

"While it is commonly a combination of risk factors that contributes to drug-induced arrhythmias – such as older age, female sex, use of other medications, or heart disease – it is certainly possible that common genetic variants will add incrementally to risk prediction," said Newton-Cheh. "It's currently premature to advocate screening gene variants for risk assessment, but someday it may be possible to identify individuals who are at particularly high risk and should avoid such medications."

Newton-Cheh adds, "It's likely that many more genes will be found to contribute to changes in QT interval, and the real challenge will be understanding the mechanism behind their effects. Five of the gene regions we identified had never before been implicated in QT interval physiology and these genetic observations may thus provide key insights into normal and abnormal human biology." He is an assistant professor of Medicine at Harvard Medical School.


Time (and PPAR-beta/delta) Heals All Wounds
Organotypic cultures help to unravel how a transcription factor modulates crosstalk between different layers of the skin

Mammalian skin requires constant maintenance, but how do skin cells know when to proliferate and at what rate? In the March 23, 2009 issue of the Journal of Cell Biology, Nguan Soon Tan and colleagues reveal that skin fibroblasts use a protein called PPARβ/δ to make sure overlying epithelial cells don't proliferate too quickly. Their results highlight how communications between different cell types are critical to maintain the skin as a barrier against the outside world.

Skin has two main layers: the underlying dermis, made up of fibroblasts and other cells, and the outer epidermis, containing epithelial keratinocytes. Signals are exchanged between these layers to coordinate their function, but dissecting these signals is tricky. For example, PPARβ/δ is an important protein for maintaining healthy skin, but its precise function remains controversial.

PPARβ/δ is a nuclear hormone receptor that regulates gene expression. In mice lacking PPARβ/δ, epidermal cells proliferate excessively after wounding (1). But cultured keratinocytes from these mice don't proliferate any faster than normal cells and, in fact, are more susceptible to apoptosis (2). According to Nguan Soon Tan, this discrepancy was the first indication that PPARβ/δ might regulate crosstalk between layers of the skin—the epidermal hyperproliferation seen in the knockout mice could be due to faulty signals from the dermal cells.

But this couldn't be studied further in mice, as it is not yet possible to delete a gene exclusively from the dermis. "We had to look at a situation where the different types of cells were not in isolation but could communicate with each other," says Tan. "Organotypic skin cultures are a really good technique for this."

First developed in the 1980s (3), organotypic skin cultures (OTCs) are made by embedding dermal fibroblasts in a gel of extracellular matrix proteins. Keratinocytes are seeded on top of this gel and the two cell types develop into an in vitro version of skin that looks remarkably like the real thing. The fibroblasts and keratinocytes can therefore be manipulated separately—knocking down or overexpressing proteins— before the skin is reconstructed.

Chong et al. found that PPARβ/δ-deficient fibroblasts made wild type keratinocytes hyperproliferative in OTCs by secreting extra doses of several growth factors. The fibroblasts were stimulated to produce these growth factors by keratinocyte-released cytokine IL-1 - underscoring the reciprocity between the two cell types. Blocking either the IL-1 signal or any of the growth factors released by the fibroblasts returned the OTCs to normal.

So why do fibroblasts lacking PPARβ/δ send out more growth factors in response to IL-1? The authors discovered that PPARβ/δ stimulates the production of sIL-1ra, a protein that inhibits IL-1 signaling by competing for the IL-1 receptor. Normally, this would decrease the IL-1 signal received by fibroblasts and therefore reduce the growth factor signals sent back to the keratinocytes. But in PPARβ/δ's absence, fibroblasts keep stimulating keratinocyte division. Similarly, PPARβ/δ knockout mice expressed less sIL-1ra after wounding and produced more growth factors that stimulate the epidermis. "Proliferation is important in early stages of wound healing," explains Tan. "But excessive proliferation isn't good: you can end up with hypertrophic scarring."

This may also be critical to prevent tumor development. Contradictory reports exist on whether PPARβ/δ promotes or inhibits epithelial cancers (4-6). Tan's group has already found that fibroblasts lacking the protein can increase the proliferation of squamous carcinoma cells; they now plan to investigate PPARβ/δ's expression in tumor-associated fibroblasts. "Fibroblasts often play important roles by communicating with epithelial cells," says Tan. "But dissecting these networks has been very difficult. We've managed to show how one particular nuclear factor in fibroblasts can have a wide ranging effect on a tissue."

Research Links Evolution of Fins and Limbs With That of Gills
The genetic toolkit that animals use to build fins and limbs is the same genetic toolkit that controls the development of part of the gill skeleton in sharks, according to research to be published in Proceedings of the National Academy of Sciences on March 23, 2009, by Andrew Gillis and Neil Shubin of the University of Chicago, and Randall Dahn of Mount Desert Island Biological Laboratory


The shark arch gill skeleton ((above) shows primitive gill rays that are found only in sharks and other cartilaginous fishes. The gills of other fishes (right) are also arched but lack gill rays. This primitive feature of sharks allowed the researchers to link the developmental genetic program for fins and limbs to the more primitive one for gill rays.

"In fact, the skeleton of any appendage off the body of an animal is probably patterned by the developmental genetic program that we have traced back to formation of gills in sharks," said Andrew Gillis, lead author of the paper and a graduate student in the Department of Organismal Biology and Anatomy at the University of Chicago. "We have pushed back the evolutionary origin of the developmental genetic program that patterns fins and limbs."

This new finding is consistent with an old theory, often discounted in science textbooks, that fins and (later) limbs evolved from the gills of an extinct vertebrate, Gillis added. "A dearth of fossils prevents us from definitely concluding that fins evolved from gills. Nevertheless, this research shows that the genetic architecture of gills, fins and limbs is the same."

The research builds on the breakthrough discovery of the fossil Tiktaalik, a "fish with legs," by Neil Shubin and his colleagues in 2006. "This is another example of how evolution uses common developmental programs to pattern different anatomical structures," said Shubin, who is the senior author on the PNAS paper and Professor and Associate Dean of Organismal and Evolutionary Biology at the University of Chicago. "In this case, shared developmental mechanisms pattern the skeletons of vertebrate gill arches and paired fins."

The research also showed for the first time that the gill arch skeleton of embryonic skates (a living relative of sharks that has gill rays) responds to treatment with the vitamin A derivative retinoic acid in the same way a limb or fin skeleton does: by making a mirror image duplicate of the structure as the embryo develops. According to the researchers, the genetic circuitry that patterns paired appendages (arms, legs and fins) has a deep evolutionary origin that actually predates the origin of paired appendages themselves.

"These findings suggest that when paired appendages appeared, the mechanism used to pattern the skeleton was co-opted from the gills," Gillis said. "Perhaps we should think of shark gills as another type of vertebrate appendage—one that's patterned in essentially the same way as fins and limbs."

The deep structural, functional, and regulatory similarities between paired appendages and developing gill rays, as well as the antiquity of gills relative to paired appendages, suggest that the signaling network that is induced by retinoic acid had a patterning function in gills before the origin of vertebrate appendages, the research concludes. And this function has been retained in the gill rays of living cartilaginous fishes.

Plastic Protein Protects Bacteria from Stomach Acid
A tiny protein helps protect disease-causing bacteria from the ravaging effects of stomach acid, researchers at the University of Michigan and Howard Hughes Medical Institute have discovered

Their findings were scheduled to be published online in the Proceedings of the National Academy of Sciences the week of March 23.

Stomach acid aids in food digestion and helps kill disease-causing bacteria. One way that acid kills bacteria is by causing the proteins in them to unfold and stick together in much the same way that heating an egg causes its proteins to form a solid mass. Just as it is virtually impossible for a cook to unboil an egg, it is also very difficult for bacteria to dissolve these protein clumps, so bacteria and most living things can die when exposed to acid or heat.

However, disease-causing bacteria such as the notorious E. coli are protected from stomach acid by a tiny protein called HdeA. In the PNAS paper, James Bardwell and coworkers describe how this protein works to protect bacteria. Like other proteins, HdeA unfolds and becomes more flexible when exposed to acid. But in a clever twist, the unfolding process that inactivates most other proteins activates HdeA. Once unfolded, this plastic protein molds itself to fit other bacterial proteins that have been made sticky by acid- induced unfolding.

"Just as plastic wrappers prevent candies from sticking together, HdeA prevents the unfolded proteins from sticking together and forming clumps," said Bardwell, a professor of molecular, cellular and developmental biology and of biological chemistry, as well as a Howard Hughes Medical Institute Investigator.

Postdoctoral fellow Tim Tapley, who spearheaded the research, said: "HdeA directly senses acid and changes from its inactive to active form within a fraction of a second." Instead of becoming completely unfolded in response to acid and sticking to itself, HdeA is only partially unfolded. It then uses the flexibility it gains through partial unfolding to rapidly become plastic enough to adapt to and bind various damaged proteins. This helps E. coli evade the otherwise deadly effects of stomach acid.

###
In addition to Bardwell and Tapley, the paper's authors are undergraduate students Jan Körner and Julia Hupfeld, graduate student Madhuri Barge, research investigator Joseph Schauerte, professor of biological chemistry Ari Gafni and associate professor of molecular, cellular and developmental biology Ursula Jakob. The research was funded in part by the National Institutes of Health.


MONDAY March 23, 2009---------------------------News Archive

1st Automated Carbohydrate 'Assembly line
Scientists from Germany today reported a major advance toward opening the doors of a carbohydrate-based medicine chest for the 21st Century. Much more than just potatoes and pasta, these carbohydrates may form the basis of revolutionary new vaccines and drugs to battle malaria, HIV, and a bevy of other diseases

Speaking at the 237th National Meeting of the American Chemical Society, Peter H. Seeberger, Ph.D., described development of an automated carbohydrate synthesizer, a device that builds these intricate molecules in a few hours — rather than the months or years required with existing technology.

"Our automated synthesizer is now the fastest method to make complex carbohydrates," says Seeberger, principal investigator for the research. "There are currently no competitive methods available. Today, if people working in biology run into a problem related to carbohydrates, they usually drop it because there are no tools available. They can't buy anything from a catalogue. It becomes a royal pain in the neck."

Scientists trying to synthesize DNA and protein-based molecules experienced a similar pain-in-the neck decades ago, until the invention of automated DNA and protein synthesizers. These devices helped kick start a revolution in genetics and proteomics. The carbohydrate synthesizer may do the same thing for the emerging fields of glycochemistry and glycobiology — named for carbohydrate sugar chains known as "glycans."

In 2001, Seeberger and colleagues reported the design of a prototype synthesizer. Revealed for the first time at the ACS National Meeting, the latest version is now fully automated, much faster, and can be operated by a non-expert, says Seeberger. Developed at the Swiss Federal Institute of Technology in Zurich, the instrument produces significant quantities of carbohydrate molecules that were nearly inaccessible until now.

Carbohydrates are tough molecules to build because of their complicated, branched structure. So instead of trying to build carbohydrates from scratch, scientists today use molecules isolated from nature, a painstaking process that could take months.
"We make things chemically that people used to isolate," explains Seeberger. "The automated synthesizer puts single sugars, the building blocks of carbohydrates, together like beads on a string."

Carbohydrates play crucial roles in the immune system, especially in the body's defenses against disease-causing viruses and bacteria. Most of these microbes have unique carbohydrate markers on their surfaces. The immune system recognizes these carbohydrates as foreign material, and creates antibodies that launch an immune response to battle the infection.
"Vaccines 'educate' the immune system to recognize a specific molecule on the surface of infectious organisms," explains Seeberger. "The synthesizer allows us to make not one but many carbohydrate structures from a particular organism and test those to see if they protect against the microbe. Synthetic carbohydrates that show promising protective qualities then may become the basis for new vaccines.

In a recent finding, the team discovered a carbohydrate on the surface of the malaria parasite P. falciparum that enables the parasite to infect human red blood cells, thus solving a long-standing mystery about how infection happens.

Seeberger's group used the carbohydrate synthesizer to develop a malaria vaccine. Clinical trials for the vaccine are scheduled for 2010 in Mozambique and Tanzania. Its unique "anti-disease" mechanism makes it the only vaccine of its kind, he says.
"To my knowledge, ours is the first attempt at an anti-disease vaccine. It doesn't actually kill the malarial parasite; it blocks its toxic action. You create antibodies against the sugar structure, and these antibodies block the carbohydrate toxin from leading to inflammation and anemia, the hallmarks of malarial infection," says Seeberger. He explained that they will pair the carbohydrate vaccine with a traditional, protein-based one to create a "conjugate vaccine," which is best suited to immunize the most vulnerable group of potential malaria victims — children under the age of two.

The malaria vaccine is only one of almost a dozen vaccines from Seeberger's lab headed for clinical trials. Carbohydrate-based vaccines could target some of today's most serious infectious diseases, including antibiotic-resistant infections and HIV.
Seeberger is commercializing the carbohydrate synthesizer through his start-up company, Ancora Pharmaceuticals, based in Medford, Mass. Looking ahead, Seeberger discussed the other major obstacle facing carbohydrate research.

"In the area of glycobiology, there are two technological hurdles right now. One is to get access to molecules, which we have now addressed. The second one is sequencing. If you look at the human genome project, or genomics and proteomics, sequencing and synthesis were always the key issues," says Seeberger.

Seeberger saw firsthand the profound effect that automated DNA synthesizers had on genetics and biotechnology. His doctoral advisor, Marvin H. Caruthers, Ph.D., of the University of Colorado, helped develop the first model in 1980.

"We hope that we have the same effect on carbohydrate research," says Seeberger.

###
The American Chemical Society is a nonprofit organization chartered by the U.S. Congress. With more than 154,000 members, ACS is the world's largest scientific society and a global leader in providing access to chemistry-related research through its multiple databases, peer-reviewed journals and scientific conferences. Its main offices are in Washington, D.C., and Columbus, Ohio.


Schizophrenia-linked Gene Controls Birth of New Neurons
“Lithium is known to inhibit GSK3beta directly and indirectly, so it looks like DISC1 behaves like endogenous lithium.” Li-Huei Tsai

A gene strongly implicated in schizophrenia is essential for normal brain development and the growth of new neurons in the adult brain, according to new research by Howard Hughes Medical Institute (HHMI) scientists.

A research team led by HHMI investigator Li-Huei Tsai at the Massachusetts Institute of Technology found that a mutated form of the gene disrupts the growth and development of brain cells. Their findings may provide new targets for the development of novel drugs to treat schizophrenia.

The researchers also showed that the gene DISC1 is part of the signaling pathway targeted by the mood stabilizer lithium. “For the first time, we have linked an evolutionarily conserved signaling pathway with schizophrenia,” says Tsai. “The beauty of knowing that this is the signaling pathway is that researchers now have many new targets to aim for as they develop drugs to treat schizophrenia.”

Tsai and her colleagues published their studies on March 20, 2009, in the journal Cell.

Schizophrenia is a common mental illness, affecting up to one percent of adults worldwide. Symptoms begin in late adolescence or early adulthood and can include delusions, hallucinations, paranoia, depression, and cognitive impairment.

Scientists have only just begun to unravel the complicated genetics of schizophrenia, says Tsai. Mutations in a variety of genes appear to increase risk for the disorder. In the early 1990s, researchers linked DISC1 to mental illnesses prevalent in a large Scottish family. Over five generations, many members of the family had developed schizophrenia, bipolar disorder, and other mood disorders. Each family member diagnosed with mental illness also carried a broken copy of the DISC1 gene. “DISC” stands for “disrupted in schizophrenia.”

“Most people in the field consider DISC1 to be very important in schizophrenia and related disorders,” says Tsai.

When Tsai set out to understand DISC1’s role in brain development and how defects in the gene might lead to schizophrenia, she first studied the cells of embryonic mice to see where the protein was produced in abundance. She found large amounts of DISC1 in brain stem cells, which are crucial for proper brain growth.

Next, Tsai’s team examined DISC1 in the brain stem cells of adult mice. Also called neural progenitor cells, brain stem cells are active in only a few regions of the adult brain, including the hippocampus, a seahorse-shaped structure previously implicated in mood disorders. Throughout life, these cells continually grow, divide, and spin off new neurons, a process dubbed neurogenesis.

Tsai and he colleagues also found large amounts of DISC1 in the brain stem cells of adult mice. But when the scientists reduced the amount of DISC1 in those adult cells – simulating what occurs in people carrying a broken version of the gene – she found that the cells failed to grow and divide. “We show that this one gene product is clearly a key regulator of the proliferation of neural progenitors during embryonic brain development and adult neurogenesis,” says Tsai.

Furthermore, the researchers found that animals that had no DISC1 in their brain stem cells displayed behaviors that mimic schizophrenia in people. The mice skittered around their cages as if agitated, behavior considered a parallel to mania in people. When the mice were given a forced-swim test, which is commonly used by researchers to measure how antidepressant drugs affect the behavior of mice, the DISC1-deficient mice seemed depressed and did not swim for long. “When we downregulated DISC1 in the dentate gyrus, which is part of the hippocampus, the mice displayed abnormal behavior consistent with schizophrenia,” Tsai says.

Tsai and her colleagues then dissected the precise molecular role that DISC1 plays in brain stem cells. She found that the protein acts like lithium, a drug commonly prescribed as a mood stabilizer for patients with bipolar disorder. In particular, DISC1 inhibits the enzyme GSK3beta, the same enzyme inhibited by lithium. “Lithium is known to inhibit GSK3beta directly and indirectly, so it looks like DISC1 behaves like endogenous lithium,” says Tsai. Bipolar disorder and schizophrenia are often diagnosed together, and many psychiatrists consider the two disorders intimately linked, perhaps even lying along a spectrum.

In a final set of experiments, Tsai’s team depleted DISC1 activity in the brain stem cells of adult mice, and then treated the mice with a molecule that inhibits GSK3beta much like lithium does. The brain stem cells returned to normal, revving up production of new neurons. Further, the previously agitated and depressed animals recovered and began behaving normally.

“This drug-like small molecule completely reverses the deficiencies seen in the neural progenitor cells while alleviating the abnormal behavior in the mice,” says Tsai.

Tsai’s team also identified the key segment of the DISC1 protein that inhibits GSK3beta. Knowing the structure of this protein segment may help drug developers as they pursue better treatments for schizophrenia and related disorders, Tsai says. “We hope this information will lead to better treatments for schizophrenia, which are sorely needed.”

Tsai is now working with geneticists at the Stanley Center for Psychiatric Research in Cambridge, Massachusetts, where she holds a joint appointment, to identify additional variations in the DISC1 gene. “We need to get a handle on the genetics of schizophrenia,” says Tsai. “But now we know how DISC1 probably contributes to the disorder, which is a big step.”

Genetic Sleuth Solves Glaucoma Mystery
Michael Walter is one good gumshoe. The University of Alberta medical geneticist has cracked the case of WDR36, a gene linked to glaucoma.

Glaucoma is a leading cause of blindness in which cells in the optic nerve die, preventing the brain from understanding what patients see. Scientists have long suspected a link between WDR36 and glaucoma, but have been unable to determine what the gene does and why some people with variations of the gene get glaucoma while others don't.

Walter unravels this mystery in an article, published in the April print edition of the journal Human Molecular Genetics, based in Oxford, England.

Walter and his team investigated a yeast gene that is extremely similar to WDR36 but much easier to experiment with. They introduced the suspected WDR36 variations into the yeast gene and tested its ability to function, and discovered that WDR36 wasn't working alone. The gene variations only affected the yeast when there were simultaneous changes to another gene called STI1. Walter thinks that STI1 is only one of many other genes in which mutations must take place in order for WDR36 to cause glaucoma.

"Our results suggest that glaucoma is polygenetic, which means there have to be changes in several different genes in order for WDR36 to cause the disease," said Walter, a professor and chair of the Department of Medical Genetics in the Faculty of Medicine & Dentistry.

This discovery explains why only some people who have WDR36 gene variations get glaucoma and may also lead to further research to uncover the other genetic accomplices.

"Only 10 per cent of glaucoma cases are caused by known genes, so the genes involved in this polygenetic interaction may help to explain the other 90 per cent," said Walter, who is also a professor in the Department of Ophthalmology.

In addition, Walter uncovered what WDR36 does in normal function. The gene helps make ribosomes, specialized molecules that make the proteins necessary to keep the cell functioning. Walter suspects that changes to WDR36 will affect ribosome production, and in turn affect the cell's ability to function.

But this mutation alone isn't enough to cause glaucoma. Changes also have to happen to the gene's partner in crime, the STI1 gene, which normally packages the proteins produced by WDR36's ribosomes. Walter says these findings explain the mechanics of glaucoma and how changes in these two genes could lead to the illness.

"Glaucoma happens when WDR36 isn't producing ribosomes properly and STI1 isn't packaging those proteins properly. You need at least these two mutations to cause the disease."

Walter says this DNA detective work may have a tangible impact on preventing and treating glaucoma. "Glaucoma is one of the few blinding eye diseases that we can actually treat. But right now we're only treating the symptoms, not the disease."

"If we can understand who gets glaucoma, then we're in a much better place to prevent it, and if we can understand why they get glaucoma, then we have some important clues to use in developing second-generation medications that treat the disease itself."


Human Adult Testes Cells Can Become Embryonic Stem-like, Capable of Treating Disease
Using what they say is a relatively simple method, scientists at Georgetown University Medical Center have extracted stem/progenitor cells from adult testes and have converted them back into pluripotent embryonic stem-like cells

Researchers say that the naïve cells are now potentially capable of morphing into any cell type that a body needs, from brain neurons to pancreatic tissue.

And because they produced these stem-like cells without the use of additional genes, the technology should be safe for human use, the researchers say in a paper published online in the journal Stem Cells and Development.

“Given these advances, and with further validation, it is possible that in the not–too-distant-future, men could be cured of disease with a biopsy of their own testes,” says the study’s senior investigator, Martin Dym, PhD, a professor in the Department of Biochemistry and Molecular & Cellular Biology.

The Georgetown researchers are among the first scientists to show that human testes stem cells can become embryonic stem-like cells, and they have done this work using testis tissue from organ donors, which they say has provided enough valuable tissue to allow them to make their discoveries. While they have published their preliminary results before, they are now disclosing a new and simpler method to isolate the testes stem/progenitor cells than has not been seen in other published procedures in humans and rodents.

Being able to use adult stem cells for this type of cell-based therapy offers a number of advantages over other strategies currently being explored, says Dym. The use of embryonic stem cells is controversial because it necessitates destruction of an embryo, and pushing fully mature cells, such as skin cells, back into a stem-like state requires use of cancer genes, and has therefore been viewed as potentially risky for human treatment, he says.

The idea with this approach is that men with an incurable disorder or disease could have a biopsy of their testes, which Dym says is a common procedure in patients suspected of having testicular cancer. Testes stem/progenitor cells – those cells that can go on to produce sperm – would be removed from the biopsy tissue, and grown in the laboratory with the addition of certain chemicals and growth factors. This causes the cells to revert back into a pluripotent state, which could then be driven into chosen cell types.

“We are taking adult spermatogonial stem/progenitor cells, which in the body are unipotent, capable of only making sperm, and coaxing them back to embryonic stem-like cells, which are pluripotent,” Dym says.

Once these new cell types are produced – several weeks after initial collection – they can be frozen and used at any point in the future, the researchers say.
He and the research team conducted the study using testes donated to GUMC from four organ donors, aged 16-52 years old.

“This is novel data which strengthens the argument for carrying out further research on pluripotent cells derived from human testes,” Dym says.

The next step, he says, is to get differentiated cells to cure disease in animal models and the researchers are now working on a project that uses testes spermatogonial stem/progenitor cells that morphed into pancreatic cells to treat diabetes in mouse models of human diabetes.

The study was funded by a grant from the National Institutes of Health. The authors report no potential financial conflicts.

One Gene Decides Whether Coral Relative Will Fuse or Fight
When coral colonies meet one another on the reef, they have two options: merge into a single colony or reject each other and aggressively compete for space. Now, a report in the March 19th Current Biology, a Cell Press publication, has found a gene that may help to decide that fate

"We have identified a gene that controls how a colonial animal recognizes a member of its own species based on cell-cell contact," said Leo Buss of Yale University. "The ability to recognize individuals implies a capacity to categorize such encounters and, in this case, it allows colonies to discriminate between those individuals with which they will fuse or fight."

The researchers made their discovery by studying a colonial cnidarian called Hydractinia symbiolongicarpus in the laboratory. (Cnidarians are most familiarly represented by corals, sea anemones and jellyfish.) Perhaps best known to Atlantic basin and western Pacific beachcombers as the distinctive white fuzz growing on the top of hermit crab shells, Hydractinia have become a model for scientific exploration of such so-called allorecognition phenomena, which define self versus non-self.

Despite the ubiquity of allorecognition in colonial organisms and its ecological and evolutionary importance, its molecular basis had not been thoroughly defined. Such interactions in nature are also of interest because of their resemblance to those that occur in pregnancy and following organ transplantation.

Now, Buss, Stephen Dellaporta and their interdisciplinary colleagues have identified a key invertebrate allorecognition gene. The gene they identified appears to encode a transmembrane receptor expressed in all tissues capable of allorecognition. It also includes a hypervariable domain and exists in many different varieties that predict how Hydractinia colonies will interact with one another, they show. The gene sequence is most closely related to the immunoglobulin (Ig) superfamily of proteins, which include antibodies of the mammalian immune system.

"Relationships have often been suggested between cnidarian and protochordate allorecognition systems or between invertebrate allorecognition systems and elements of the vertebrate immune system..." the researchers wrote. However, while Ig-like domains are found in vertebrate immune molecules and potentially also in the Hydractinia allorecognition gene, there appears to be no additional similarity between the known surface molecules in these systems.

"Indeed," they concluded, "growing evidence suggests that animals have evolved a variety of unique molecular mechanisms to distinguish self from non-self, including the MHC in vertebrates, VCBPs in protochordates, VLR immune molecules in jawless fish, FREP proteins in molluscs, and the FuHC in tunicates. We can now add the Hydractinia allorecognition system to this diversity."


Monoclonal Antibodies Primed to Become Potent Immune Weapons against Cancer
New research suggests that monoclonal antibody therapy of cancer can be improved to be much more powerful than it is today, says the director of Georgetown University Medical Center’s Lombardi Comprehensive Cancer Center in the March 21 issue of The Lancet

“We believe that antibody therapy has the capacity to immunize people against cancer,” says Louis Weiner, MD, director of the cancer center at GUMC and an internationally recognized expert in development and use of monoclonal antibodies. “Treatment modifications might be able to prolong, amplify, and shape a continuous immune response to cancer cells.”

Weiner was asked by Lancet editors to write a review article discussing the newest research in this field. His co-authors are Madhav Dhodapkar, MD, of Yale University and Soldano Ferrone, MD, of the University of Pittsburgh.

Their analysis, based on reviewing the last eight years of research on monoclonal antibody treatment, suggests that a new era in use of these therapies is just around the corner. “Scientists have been able to use new tools to measure effectiveness of these therapies, and have found that antibodies are capable of stimulating the immune system in ways that had not been appreciated to date, and which we can now take advantage of,” Weiner says.

Antibodies are immune system proteins that seek out and neutralize molecules they recognize as foreign to a body, such as viruses and bacteria. Monoclonal antibodies are proteins crafted in a laboratory to recognize specific receptors, or antigens, on cancer cells; some antigens promote uncontrolled growth. These antibodies are designed to both attach to cancer receptors to inhibit their function and to alert and activate the immune system to the presence of these receptor proteins.

Monoclonal antibodies already offer effective treatment for a wide range of cancers, including breast cancer (Herceptin®, Avastin®), colorectal cancer (Erbitux®, Avastin), lung cancer (Avastin), and blood cancers (Rituxan®, Campath®), but they have appeared to primarily work by forcing tumor related receptors to shut down pro-growth signals, Weiner says.

“For years it has been presumed that the ability of antibodies to interfere with malignant cell-related signaling is the dominant mechanism of anticancer activity, but we have also known that the normal job of an antibody is to deliver an antigen to the body’s immune system which then destroys the target,” Weiner says.

Recent research by Weiner and others, however, now shows that antibodies can inhibit function not only as signaling manipulators but also as initiators of immune responses that leads to control of cancer, the authors say.

“We believe that Herceptin and Rituxan, as examples, work in part by immunizing people against cancer, but at this point, the magnitude of that response is variable and is frequently very small,” Weiner says.

Scientists now believe that it will be possible to alter the antibodies so that they induce both kinds of human immunity – the innate immune response that is short-lasting and which directly kills tumor cells, and a long-lasting “memory” response that comes from the adaptive immune response. “We have long thought that monoclonal antibodies are capable of stimulating the innate immune system, but we now have evidence that the therapy can prime an adaptive response as well. Such responses would make the treatment much more powerful, capable of keeping cancer under control,” he says.

“For the first time we are using technology that can measure the immune response that is occurring in monoclonal antibody treatment, and which will help us build better antibodies that amplify and shape that immune response to become more powerful,” Weiner says.

And in the future, it may be possible to build antibodies that are targeted to existing targets on a patient’s tumor, as well as to targets that may appear as the cancer mutates. “This one-two punch would anticipate how the tumor changes over time and cut off the cancer’s escape route,” Weiner says. “These new directions are very exciting.”

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