Thursday, February 25, 2010

Scientists Use Dental Stem Cells to Create New Bone Tissue in Humans

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GEN News Highlights

Italian scientists claim to be the first to have succeeded in using implants of dental pulp stem/progenitor cells (DPCs) for autologous oromaxillofacial bone regeneration in humans. Their technique was used to repair bone defects due to wisdom tooth problems in 17 patients.

The procedure and results are reported in European Cells and Materials in a paper titled, “Human Mandible Bone Defect Repair by Grafting of Dental Pulp Stem/Progenitor Cells and Collagen Sponge Biocomplexes.” The researchers suggest that the approach could also be applied to any other area of reconstructive and orthopedic surgery.

The human trial, conducted by Professor Gianpaolo Papaccio, Ph.D., and colleagues at the Second University of Naples, involved the extraction and expansion of DPCs from the maxillary third molars (wisdom teeth) of 17 patients requiring wisdom tooth extraction. The cells were then seeded onto a collagen sponge scaffold. The resulting biocomplex was used to fill in the injury site left by the removed tooth.

X-ray evaluation three months after autologous DPC grafting confirmed that the alveolar bone of treated patients had optimal vertical repair and complete restoration of periodontal tissue back to the second molars. Histological observations also demonstrated the complete regeneration of bone at the injury site. Optimal bone regeneration was evident one year after grafting.

The Naples team conclude that the autologous DPC technique represents a new tool for bone tissue engineering. “Stem cells represent an easy and natural alternative to repair/regenerate damaged tissues,” they note. “This is essential, especially when bone loss subsequent to degenerative or traumatic diseases cannot be amended through conventional therapies. The procedure is efficient, exhibits low morbidity of the collection site, and is free from diseases incurred by transmission of pathogens. The regeneration process is fast and efficient.”

Dental biobanking company, BioEden, welcomed the achievement, claiming the success “yields a vast number of medical possibilities for dental stem cells and for those people who store them for future use.” BioEden collects, assesses, and cryogenically stores living tooth cells from deciduous baby teeth.

The company holds a global patent for the extraction, cryopreservation, and storage of dental stem cells for medical use. Founded in Austin, TX, BioEden has laboratories in the U.K. (serving Europe) and Thailand (serving South East Asia). Further bases are planned for Russia, Australia, India, and the Middle East.


Bioengineering - Japanese researchers grow teeth in mice

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Japanese scientists have grown teeth in mice. This research is expected to advance the development of "tooth regenerative therapy" which have the potential for replacing diseased or damaged teeth with bioengineered teeth. The research is expected to evolve into a wide variety of organ regenerative technologies for liver, kidneys and other organs.

A research group led by Takashi Tsuji (Professor in the Research Institute for Science and Technology, Tokyo University of Science, and Director of Organ Technologies Inc.) has demonstrated in growing new organs in adult mice. Tsuji is a research team member in “Health Labor Sciences Research Grant: Research on Regenerative Medicine for Clinical Application (Domain Leader: Professor Akira Yamaguchi of Tokyo Medical and Dental University)”, and “Priority Domain Research: Bio-engineering (Domain Leader: Professor Toshio Fukuda of Nagoya University)”. In transplantation experiments using the tooth as a model, a bioengineered tooth germ develops into a fully functioning bioengineered tooth with sufficient hardness for mastication and a functional responsiveness to mechanical stress in the maxillofacial region. The research also provided the results that the nerve fibers that have re-entered the pulp and periodontal ligament (PDL) tissues of the bioengineered tooth have proper perceptive potential in response to noxious stimulations such as orthodontic treatment and pulp stimulation.

This research is expected to substantially advance in the development of “tooth regenerative therapy”, which have potential as next-generation regenerative therapies for replacing diseased or damaged teeth with bioengineered teeth. Specifically it will not only promote “tooth regenerative therapy”, whereby organ germs of bioengineered teeth are transplanted into the jaw bone to grow “3rd generation tooth”, but is expected to evolve into a wide variety of organ regenerative technologies for liver, kidney and other organs.

This research outcome was the fruit of joint research with Professor Teruko Takano-Yamamoto (Division of Orthodontics and Dentofacial Orthopedics, Graduate School of Dentistry, Tohoku University, Japan) and Professor Shohei Kasugai (Oral and Maxillofacial Surgery, Department of Oral Restitution, Division of Oral Health Sciences, Graduate School, Tokyo Medical and Dental University, Japan). It was announced in an Advance Online Publication of the US scientific journal “Proc. Natl. Acad. Sci. USA” at 17:00hrs (Eastern US Time) on Aug. 03, 2009.

Background to the research

Regenerative therapy is expected to be one of the novel clinical systems in the 21st century. One of the more attractive concepts in regenerative therapy is stem cell transplantation of enriched or purified tissue-derived stem cells, or in vitro manipulated embryonic stem cells. This therapy has the potential to restore the partial loss of organ function. The ultimate goal of regenerative therapy is to develop fully functioning bioengineered organs that can replace lost or damaged organs following disease, injury or aging. We have recently developed a method for creating three-dimensional bioengineered organ germ, which can be used as an ectodermal organ such as the tooth or whisker follicle (Nature Methods 4, 227-30, 2007). Our analyses have provided a novel method for reconstituting this organ germ and raised the possibility of tooth replacement with integrated blood vessels and nerve fibers in an adult oral environment. However, it remains to be determined whether a bioengineered tooth can achieve full functionality.

Outline of the research outcome

We analyzed the development of a bioengineered tooth germ transplanted into adult oral environment. To develop a tooth regenerative method for a clinical application, a bioengineered tooth should achieve full functionality, including sufficient masticatory performance, biomechanical cooperation with tissues in the oral and maxillofacial regions, and proper responsiveness via sensory receptors to noxious stimulations in the maxillofacial region. Recently, we developed an adult murine lost tooth transplantation model. After the extraction of first molar, its cavity was repaired by osteogenesis for a month. Then, a hole was prepared in alveolar bone using drill. Then, a bioengineered molar tooth germ was transplanted into the hole with correct orientation and gingival transplantation area was sutured (Fig. 1a, b).

1. The eruption and masticatory potential of the bioengineered tooth

At 16 days after transplantation, eruption of the bioengineered tooth could not be observed. At 37 days after transplantation, exposure of the cusp tip occurred in the gingival area of transplantation, indicating an eruption of the bioengineered tooth. At 49 days and thereafter, this bioengineered tooth was observed to have erupted to reach the occlusal plane and was seen to achieve opposing tooth occlusion.

Following the achievement of occlusion, there was no excessive increase in the tooth length at up to 120 days after transplantation (Figure. 2). However, the crown width of the bioengineered tooth was smaller than that of other teeth, since at present we are not able to regulate the crown width and cusp position. We are now trying to develop a novel cell manipulation technology to regulate the tooth size and morphology.

We next performed a Knoop hardness test, which is a test for mechanical hardness. The hardness of these mineralized tissues of the bioengineered tooth, not only enamel but also dentin, were equivalent with those of normal tooth. The microhardness test suggested that the bioengineered tooth have sufficient masticatory performance as well as the normal, mature tooth

2. Bioengineered tooth response to mechanical stress

It is well known that an alveolar bone remodeling was induced via the response of periodontal ligament according to a mechanical stress such as the treatment of orthodontic movement. Previous studies demonstrated that the localizations of osteoclast for bone absorption in compression side and osteoblast for bone formation in tension side were observed. Thus, we analyzed the tooth movement and localizations of osteoclast and osteoblast for bone remodeling in alveolar bone by an experimental orthodontic movement. A
normal tooth and the bioengineered tooth was moved buccally for 17 days with a mechanical force. At 6 days after the treatment, histological analysis revealed that morphological changes of periodontal ligament were observed in both the lingual tensioned side and buccal compressed side. The bioengineered tooth could successfully moved in response to the mechanical stress as well as normal tooth

3. Perceptive potential of neurons entering the tissue of the bioengineered tooth

Perception of noxious stimulations such as mechanical stress and pain, are important for the protection and proper functions of teeth. Neurons in the trigeminal ganglion, which innervate the pulp and PDL, can detect these stress events and transduce the corresponding perceptions to the central nervous system. In our current experiments, we evaluated the responsiveness of nerve fibers in the pulp and PDL of the bioengineered tooth to induced noxious stimulations. Nerve fibers were detected in the pulp, dentinal tubules, and PDL of the bioengineered tooth as in a normal tooth. We also found in our current analyses that the nerve fibers innervating both the pulp and PDL of the bioengineered tooth have perceptive potential for nociceptive stimulations and can transduce these events to the central nervous system (the medullary dorsal horn).

In this study, we successfully demonstrated that our bioengineered tooth germ develops into a fully functioning tooth with sufficient hardness for mastication and a functional responsiveness to mechanical stress in the maxillofacial region. We also show that the neural fibers that have re-entered the pulp and PDL tissues of the bioengineered tooth have proper perceptive potential in response to noxious stimulations such as orthodontic treatment and pulp stimulation. These findings indicate that bioengineered tooth generation techniques can contribute to the rebuilding of a fully functional tooth. Our study provides the first evidence of a successful replacement of an entire and fully functioning organ in an adult body through the transplantation of bioengineered organ germ, reconstituted by single cell manipulation in vitro. Our study therefore makes a substantial contribution to the development of bioengineering technology for future organ replacement therapy

Research Institute for Science and Technology, Tokyo University of Science

●Adress: 2641 Yamazaki, Noda, Chiba, 278-8510, JAPAN

●Project Leader: Takashi Tsuji (Professor of the Research Institute for Science and Technology, Tokyo University of Science and Director of Organ Technologies Inc.)


■ Organ Technologies Inc.

●Adress: 2-2 Kandatsukasamachi, Chiyoda, Tokyo, 101-0048, JAPAN

●President: Hiroaki Asai


■ Health Labor Sciences Research Grant from the Ministry of Health, Labor and Welfare
“Research on Regenerative Medicine for Clinical Application”
・Research Domain Leader: Akira Yamaguchi (Professor, Oral Pathology, Department of Oral
Restitution, Division of Oral Health Sciences, Graduate School, Tokyo Medical and Dental University)

■ Grants-in-Aid for Scientific Research Project of the Ministry of Education, Culture, Sports,
Science and Technology “Scientific Research for Priority Area: System cell engineering (bio-engineering) by multiscale manipulation”

・Research Domain Leader: Toshio Fukuda (Professor, Institute for Advanced Research / Graduate School of Engineering, Nagoya University)

■ Grants-in-Aid for Scientific Research Project of the Ministry of Education, Culture, Sports,
Science and Technology “Scientific Research (A)”
・Principal investigator: Takashi Tsuji (Professor of the Research Institute for Science and Technology, Tokyo University of Science and Director of Organ Technologies Inc.)

Associated links


Stem Cells From Monkey Teeth Can Stimulate Growth And Generation Of Brain Cells

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ScienceDaily (Nov. 12, 2008) — Researchers at the Yerkes National Primate Research Center, Emory University, have discovered dental pulp stem cells can stimulate growth and generation of several types of neural cells. Findings from this study, available in the October issue of the journal Stem Cells, suggest dental pulp stem cells show promise for use in cell therapy and regenerative medicine, particularly therapies associated with the central nervous system.

Dental stem cells are adult stem cells, one of the two major divisions of stem cell research. Adult stem cells have the ability to regenerate many different types of cells, promising great therapeutic potential, especially for diseases such as Huntington’s and Parkinson’s. Already, dental pulp stem cells have been used for regeneration of dental and craniofacial cells.

Yerkes researcher Anthony Chan, DVM, PhD, and his team of researchers placed dental pulp stem cells from the tooth of a rhesus macaque into the hippocampal areas of mice. The dental pulp stem cells stimulated growth of new neural cells, and many of these formed neurons. “By showing dental pulp stem cells are capable of stimulating growth of neurons, our study demonstrates the specific therapeutic potential of dental pulp stem cells and the broader potential for adult stem cells,” says Chan, who also is assistant professor of human genetics in Emory School of Medicine.

Because dental pulp stem cells can be isolated from anyone at any age during a visit to the dentist, Chan is interested in the possibility of dental pulp stem cell banking. “Being able to use your own stem cells for therapy would greatly decrease the risk of cell rejection that we now experience in transplant medicine,” says Chan.

Chan and his research team next plan to determine if dental pulp stem cells from monkeys with Huntington’s disease can enhance brain cell development in the same way dental pulp stem cells from healthy monkeys do.


Genetic Link Between Misery and Death Discovered; Novel Strategy Probes 'Genetic Haystack'

Interaction between nerves (red) and tumor cells (blue) in an ovary provides one way by which stress biochemistry signals can be distributed to sites of disease in the body. (Credit: Image courtesy of University of California - Los Angeles)

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ScienceDaily (Feb. 25, 2010) — In ongoing work to identify how genes interact with social environments to impact human health, UCLA researchers have discovered what they describe as a biochemical link between misery and death. In addition, they found a specific genetic variation in some individuals that seems to disconnect that link, rendering them more biologically resilient in the face of adversity.

Perhaps most important to science in the long term, Steven Cole, a member of the UCLA Cousins Center for Psychoneuroimmunology and an associate professor of medicine in the division of hematology-oncology, and his colleagues have developed a unique strategy for finding and confirming gene-environment interactions to more efficiently probe what he calls the "genetic haystack."

The research appears in the current online edition of Proceedings of the National Academy of Sciences.

Using an approach that blends computational, in vivo and epidemiological studies to focus their genetic search, Cole and his colleagues looked at specific groups of proteins known as transcription factors, which regulate gene activity and mediate environmental influences on gene expression by binding to specific DNA sequences. These sequences differ within the population and may affect a gene's sensitivity to environmental activation.

Specifically, Cole analyzed transcription factor binding sequences in a gene called IL6, a molecule that is known to cause inflammation in the body and that contributes to cardiovascular disease, neurodegeneration and some types of cancer.

"The IL6 gene controls immune responses but can also serve as 'fertilizer' for cardiovascular disease and certain kinds of cancer," said Cole, who is also a member of UCLA's Jonsson Comprehensive Cancer Center and UCLA's Molecular Biology Institute. "Our studies were able to trace a biochemical pathway through which adverse life circumstances -- fight-or-flight stress responses -- can activate the IL6 gene.

"We also identified the specific genetic sequence in this gene that serves as a target of that signaling pathway, and we discovered that a well-known variation in that sequence can block that path and disconnect IL6 responses from the effects of stress."

To confirm the biochemical link between misery and death, and the genetic variation that breaks it, the researchers turned to epidemiological studies to prove that carriers of that specific genetic variation were less susceptible to death due to inflammation-related mortality causes under adverse social-environmental conditions.

They found that people with the most common type of the IL6 gene showed an increased risk of death for approximately 11 years after they had been exposed to adverse life events that were strong enough to trigger depression. However, people with the rarer variant of the IL6 gene appeared to be immune to those effects and showed no increase in mortality risk in the aftermath of significant life adversity.

This novel method of discovery -- using computer modeling and then confirming genetic relationships using test-tube biochemistry, experimental stress studies and human genetic epidemiology -- could speed the discovery of such gene and environmental relationships, the researchers say.

"Right now, we have to hunt down genetic influences on health through blind searches of huge databases, and the results from that approach have not yielded as much as expected," Cole said. "This study suggests that we can use computer modeling to discover gene-environment interactions, then confirm them, in order to focus our search more efficiently and hopefully speed the discovery process.

"This opens a new era in which we can begin to understand the influence of adversity on physical health by modeling the basic biology that allows the world outside us to influence the molecular processes going on inside our cells."

Other authors on the study were Jesusa M. G. Arevalo, Rie Takahashi, Erica K. Sloan and Teresa E. Seeman, of UCLA; Susan K. Lutgendorf, of the University of Iowa; Anil K. Sood, of the University of Texas; and John F. Sheridan, of Ohio State University. Funding was provided by the National Institutes of Health, the UCLA Norman Cousins Center and the James L. Pendleton Charitable Trust. The authors report no conflict of interest.

Journal Reference:
Steven W. Cole, Jesusa M. G. Arevalo, Rie Takahashi, Erica K. Sloan, Susan K. Lutgendorf, Anil K. Sood, John F. Sheridan, and Teresa E. Seeman. Computational identification of gene-social environment interaction at the human IL6 locus. Proceedings of the National Academy of Sciences, 2010; DOI: 10.1073/pnas.0911515107


Biologists Use Mathematics to Advance Our Understanding of Health and Disease

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ScienceDaily (Feb. 21, 2010) — Math-based computer models are a powerful tool for discovering the details of complex living systems. John Tyson, professor of biology at Virginia Tech, is creating such models to discover how cells process information and make decisions.

"Cells receive information in the form of chemical signals, physical attachments to other cells, or radiation damage, for instance," Tyson said. "On the basis of this information, the cells must make the correct response, such as to grow and divide, or to stop growing and repair damage, or to commit suicide."

The question for a molecular biologist is: What are the underlying molecular mechanisms that implement these information processing systems? "Just as computer is an information processing system, with silicon chips, wires, mother board, clock, and power source, a cell is a an information processing system made of genes, messenger RNAs, proteins, and enzymes," Tyson said. "Somehow these molecules interact with each other to detect signals, make decisions, and implement the proper response."

Tyson and other biologists want to know how jumbles of molecules can figure out how a cell should respond to its environment in order to survive, grow, and reproduce. "So we do what any good engineer would do. We create a mathematical model of the components and their interactions, and let the computer work out the details."

Tyson presented his findings at the American Association for the Advancement of Science meeting February 18-22 in San Diego, as part of a session on "Moving Across Scales: Mathematics for Investigating Biological Hierarchies," which includes talks ranging from "HIV interventions in Africa" to the "Neural Dynamics of Decision Making." Tyson will talk about "Molecular Network Dynamics and Cell Physiology," or the cell as an information-processing system.

The speakers in this session will illustrate how math models help scientists reason across scales in biology, such as from interactions between sick and healthy people to the spread of global pandemics. Whereas models of this sort can inform public health decisions on a global scale, Tyson's research addresses basic science at the smallest scale -- bridging the gap from molecules to cells. "We have to first understand the molecular basis of normal cell behavior; then we have a chance of figuring out how the system is broken in diseased cells," said Tyson.

"What decision-making processes tell a cell when to grow and divide and when to just hang-out? It is mistakes in this decision process that cause cancer. Tumors are cells growing when and where they shouldn't. Cancer is a collection of diseases caused by faulty decision-making at the cellular level. The cells are no longer obeying the rules. We know the cause is in the molecules that are supposed to be enforcing these rules."

During the course of his research, Tyson and colleagues have used computer simulations to test their math models. "If the math model behaves in the computer the way cells behave in the lab, we gain confidence that we understand the molecular interactions correctly. If not, we can be sure that our models are missing something important."

Tyson will talk about the control of cell division in yeast and in mammalian cells. "Yeast cells are easy to work with in the lab, and their molecular control systems are very similar to the control systems in mammalian cells," he said As a result of the success that Tyson and his colleagues have had in modeling yeast cell growth and division, they are now making the transition to mammalian cells and cancer.

"We do not yet have an engineer's understanding of normal mammalian cell proliferation and of what goes wrong in cancer cells," Tyson said. "Cancer treatment is still a matter of cutting out, blasting, or poisoning cancer cells -- and any normal cells that get in the way. We could be more subtle and perhaps more effective in treating cancers if we had a systematic insider's understanding of the molecular networks that control cell growth, division and death, and an ability to manipulate this control system with a new array of drugs and procedures."


Friday, February 19, 2010

Genome data

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Where to download genome data:

Nice flash movie (sequencing):

List of medical databanks:

Saturday, February 13, 2010

Towards the longest run in CERN’s history

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Following the decisions taken at the Chamonix meeting, the teams are preparing the LHC to run at collision energies of 7 TeV during the coming 18-24 months. The consolidation of the nQPS connectors has been successfully completed and the magnet powering at high current has begun.

Since the last issue of the Bulletin, all of the approximately 4000 high-voltage electrical connectors of the new Quench Protection System (nQPS) have been replaced in record time. Teams are now upgrading the software on some of the electronic circuit boards. This work is already complete in half the machine.

At the same time, the Hardware Commissioning group has started to power up the magnets in all the sectors. This is a lengthy process that involves gradually increasing the current to reach the 6 kA needed to steer beams at an energy of 3.5 TeV/beam. Currently, all the sectors have passed the tests at low current, and Sectors 1-2 has been successfully tested at 5 kA.

Over the coming days, the teams plan to power all the sectors to high current in order to make them ready for “machine check-out” – the process that prepares the machine for beam operation. According to the present schedule, beams should be circulating again in the LHC towards the end of February.


Wednesday, February 10, 2010

Virology: Some Viruses Use Fats to Penetrate a Cell

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ScienceDaily (Feb. 8, 2010) — SV40 viruses use an amazing means of communication, in order to be able to penetrate into a cell: fats, whose structure must fit like a key in a lock.

Just like a ball, driven into the goal, causing the net to bulge out and wrap itself closely around the leather: This is how it appears when the Simian Virus 40 (SV40) penetrates into a cell. The virus docks onto the cell membrane, which in turn invaginates deeply, wraps itself tightly around the intruder and buds into a vesicle that is finally pinched off inside the cell.

Suitable Trojans required

Until recently, scientists were unable to explain how this dramatic rearrangement of the cell membrane took place in order to make it possible for the virus to penetrate, since only few proteins seem to be involved in this process. A new work, which has just appeared in Nature Cell Biology, now throws new light on the mechanism by which the SV40 outwits its host: it exploits the components of the cell membrane itself, fats.

If a virus wants to reproduce itself, the same question generally arises: How does it get into a cell in order to use the latter's reproductive mechanism for its own purposes? After all, although viruses carry with them a short piece of genetic information depending upon their type, they need to penetrate into the cell and its nucleus in order to propagate their genome. There, the cells own replication machinery is reprogrammed to produce new viruses, which finally abandon the cell and infect further cells.

Fats as a Velcro fastener

SV40 has now developed a unique strategy. Instead of binding to a protein receptor in the plasma membrane and entering vesicles created by an apparatus around the protein molecule clathrin, this polyomavirus attaches itself to lipids. It does however not bind to one receptor, but many lipid molecules, in a similar way to a Velcro fastener. The individual connections are weak, but many connections taken together are strong. As soon as the virus has connected itself to many fats, the plasma membrane of the cell changes dramatically: It undergoes deep invagination and, in the course of time, completely surrounds the virus and finally forms a vesicle, which is pinched off inside the cell.

Interestingly the many connections are not only important for virus binding, but also for this membrane deformation process. The researchers could show that different molecules binding to the same fat cannot deform the membranes. At least five fat connections were required for membrane deformation. The membrane is then organized so closely around the virus that hardly any space remains between its surface and the virus. The virus in this way optimizes the number of connections with the membrane and can exert a strong force on the membrane sufficient to deform it without the help of cellular proteins.

Short chains do not bind

In addition, the correct fats must be present on the surface of the virus. The carbon chains forming part of the fat must be of the correct length. If they are too short, then the membrane does not invaginate. This has been demonstrated by experiments with structurally altered fats.

"It surprised us that there is a relationship between structure and function even with fats," observed Helge Ewers, ETH Group-Leader, who signs as primary author of the paper. With proteins, such key/lock principles are common. "With this work we have proven that it can also be the same for fats," says the former graduate student of Ari Helenius, Professor of Biochemistry.

Widely distributed mechanism

In co-operation with the Curie Institute in France the ETH researchers were able to show that SV40 is not the only pathogen, which gains entry to cells via multi-lipid connections. This route is also taken by bacterial toxins, such as, for instance, the cholera toxin or mouse polyomavirus. It thus seems to be a widespread mechanism.

It is not yet possible, however, to use this knowledge therapeutically. Antiviral medicines continue to eliminate the infected cells. Finding active substances, which block the viral fat connection, is considered by Ewers and Helenius to be a difficult task.


SV40 naturally infects Asian apes, such as macaques and rhesus monkeys. It can also be passed on to humans. In the '60s it was discovered in cultures of kidney cells from rhesus monkeys. The cells were used for the production of vaccines against polio, the childhood paralysis. During the inoculation from 1955 to 1963 several million humans were probably infected with SV40. Like other polyomaviruses, SV40 can cause tumours under certain conditions. However, in most cases the infection remains symptom-free. In humans no direct connections between an SV40-infection and the emergence of cancer could be proven. Oncogenes of this virus, however, play a role in the emergence of cancer cells from human cells in cell culture.

Journal Reference:
Ewers H et al. GM1 structure determines SV40-induced diaphragm invagination and infection. Nature Cell Biology, 20 December 2009 DOI: 10.1038/ncb1999


Scientists find survival factor for keeping nerve cells healthy

Site of the day: (Jan.26, 2010) — Scientists at the Babraham Institute have discovered a novel survival factor whose rapid transport along nerve cells is crucial for keeping them alive. The same factor seems likely to be needed to keep our nerves healthy as we age. These findings, published today in the online, open-access journal PLoS Biology, show that a molecule known as Nmnat2 provides a protective function; in its absence healthy, uninjured nerve cells start to degenerate and boosting levels of Nmnat2 can delay degeneration when the cells are injured. This suggests an exciting new therapeutic avenue for protecting nerves from disease and injury-induced degeneration.

This breakthrough by Drs Jon Gilley and Michael Coleman at Babraham, an institute of the Biotechnology and Biological Sciences Research Council (BBSRC), furthers our understanding of the basic biology of our nerves and provides new insight into the factors causing neurodegenerative diseases like Motor Neurone Disease and Multiple Sclerosis.

Neurodegenerative diseases are characterised by a loss of viable nerve cells, which in many cases has been shown to be preceded by degeneration of the axon. Axons are the long, slender projections from nerve cells, sometimes over a metre long, that carry messages to target cells such as other nerve or muscle cells, rather like a fibre-optic cable carrying outgoing messages. Although the disintegration and collapse of axons is seen in many neurodegenerative diseases, the factors driving this have remained elusive.

Unravelling the processes initiating axon degeneration is helping to understand mechanisms of disease progression. It also increases our potential to protect synapses and axons in disease using Nmnat2 as a therapeutic target.

"What is really exciting here is how a single, intrinsic protein affects nerve cell survival," explained Dr Coleman, a Group Leader at Babraham. "It offers a new approach to treating axonal disorders by specifically targeting this protein, or by targeting other steps in the same pathway that we hope to work on next."

Axonal transport is a remarkable process that traffics thousands of biochemical compounds needed for axon survival and function along every one of our hundred billion nerve cells, day and night, across distances that dwarf any other mammalian cell. We are not aware of it until it goes wrong but then the results can be devastating. Alzheimer's disease, glaucoma, motor neuron disease and multiple sclerosis are some of the neurodegenerative disorders that involve a block of axonal transport. Even the healthy ageing process shows a dramatic decline in axonal transport that may predispose us to these and other age-related disorders.

Coleman continued, "Think about the fate of a flower after its stem is cut. Without water it quickly wilts and dies. In water it lasts much longer but still dies earlier than on the plant, so water is the limiting factor for survival even if the flower needs other essential substances in the longer term.

There are some similarities when a nerve is injured. If a nerve is cut, axons beyond the injury site die within a couple of days because they lack essential proteins that are normally transported along the nerve.

"Like the flower's critical need for water, we found that one protein seems to be a limiting factor for axon survival by a large margin", explained Coleman. "Other missing proteins have little effect on this timescale. Nerve cells do differ in that they die through an active process rather than withering away, but the process may still be triggered by one factor, or at most just a few."

Cultured nerve cells were used to find which of the many biochemical factors limit axon survival. This builds on earlier work in the Coleman lab, which revealed that a single, harmless genetic variation, the slow Wallerian degeneration (WldS) gene, can extend the survival of a cut axon tenfold. However, this cannot provide what axons normally need to survive because most animals and probably all people lack the WldS gene. Nevertheless, its identity provided vital clues.

The new research, supported by the MRC and BBSRC, has identified a key axon survival factor present in all of us, Nmnat2, without which axons quickly degenerate. Nmnat2 is metabolic enzyme situated in part of the cell known as the Golgi, and now the Babraham group also finds it in axons. This raises the possibility of manipulating its activity with drugs in order to protect or delay axons from degeneration.

"As Nmnat2 is present in all our nerves it could be modulated directly, whereas WldS would first have to be introduced to our nerves." Coleman said. "By understanding how Nmnat2 is trafficked along nerves, what regulates its stability, and what it does when it gets there, novel treatments could now be developed for thus far incurable neurodegenerative diseases."

More information: "Endogenous Nmnat2 is an Essential Survival Factor for Maintenance of Healthy Axons". DOI: 10.1371/journal.pbio.1000300



Monday, February 8, 2010

Scientists ID a Protein That Splices and Dices Genes

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ScienceDaily (Feb. 5, 2010) — A novel finding offers a clue as to how genes can have what you might call multiple personalities.

The research was recently described online in the journal Science by teams from the National Cancer Institute, The University of Texas Health Science Center at San Antonio and the University of Toronto.

Genes are long strings of DNA letters, but they can be cut and spliced to make different proteins, something like the word "Saskatchewan" can have its middle cut out to leave the word "Swan," its front, middle and end deleted to leave the word "skate," or its front and back chopped off to make the word "chew."

This latest discovery reveals that the protein MRG15, which previously had been known to affect cell growth and aging, also directs the gene-splicing machinery. Olivia Pereira-Smith, Ph.D., a professor in the Department of Cellular and Structural Biology and the Sam and Ann Barshop Institute for Longevity and Aging Studies at the UT Health Science Center San Antonio, has studied the function of MRG15 for more than 10 years.

As people or animals age, this gene-splicing machinery can go awry, producing nonsense proteins ("Sskt" instead of "Swan," for instance) rather than the proper ones. These aberrant proteins can damage cells, possibly leading to cancer or other diseases of aging. The finding thus has potential implications for therapies to treat both cancer and aging, a Texas researcher said.

The Science paper's lead author is Reini F. Luco, Ph.D., a fellow in the laboratory of senior author Dr. Tom Misteli, Ph.D., at the National Cancer Institute (NCI). Other co-authors include Kaoru Tominaga, Ph.D., from the UT Health Science Center, and Benjamin J. Benclowe, Ph.D., and Qun S. Pan, Ph.D., from the University of Toronto.

"We've known for three or four years, from other analyses, that this protein was also involved in splicing, but we needed the expertise of Dr. Misteli's lab," Dr. Smith said. "Dr. Luco led the splicing studies on this project."

Dr. Tominaga, a faculty member of the Department of Cellular and Structural Biology and the Barshop Institute in San Antonio, said it may be possible to design cancer drugs to regulate MRG15's activity.


Link Between Human Birth Defect Syndrome, Cancer Metastasis Explored

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ScienceDaily (Feb. 4, 2010) — Some cells are natural rule-breakers. Neural crest cells for example, not only migrate throughout the body during development (most cells are more selective in their wandering), they are also more developmentally flexible than their predecessors (a no-no for nearly all cell types). Now researchers at the Stanford University School of Medicine have shown that a protein that controls DNA accessibility is responsible for the cells' unruly ways.

The finding not only offers a better understanding of the molecular basis of a spontaneous genetic disease in humans called CHARGE syndrome, it may also be important in understanding how cancer cells gain the ability to migrate, or metastasize.

"Most cells lose developmental potential as they differentiate," said Joanna Wysocka, PhD, assistant professor of developmental biology and of chemical and systems biology. "But neural crest cells are a spectacular example of migratory cells that are capable of becoming over 100 different cell types, including neurons, the bone and cartilage of the face, jaw and teeth, pigment cells and certain heart structures." Wysocka is the senior author of the research, which will be published online Feb. 3 in Nature.

Wysocka, who studies how chromatin modification affects development, became interested in the cells when it became apparent that mutations in a protein called CHD7 were responsible for CHARGE syndrome. The condition's name is an acronym for a constellation of associated birth defects that affect about one in 10,000 children. Children with the disorder have a combination of craniofacial malformations; eye, ear and heart defects; and other abnormalities. The unusual combination of this wide array of symptoms led physicians and researchers to speculate that the problem arose early in development in the neural crest cells.

Most DNA in a cell is tightly wrapped around proteins and compacted into what is called chromatin. CHD7 belongs to a class of proteins called ATP-dependent chromatin remodelers, which orchestrate the movement of the DNA packing proteins to provide or restrict access to particular genes. Choosing which portions of DNA to expose and which to keep tightly bundled can control cell fate.

"This was fascinating to me because next to nothing is known about chromatin regulation in neural crest cells, which are multipotent by nature," said Wysocka, who is also a member of Stanford's Cancer Center. "And yet, CHD7's involvement in CHARGE indicated that this chromatin remodeler is a critical component of the proper migration and specialization of the neural crest."

The neural crest forms early in development (in humans, at three to five weeks of gestation) when a portion of the cells that will become the embryo folds inward into a tube that will become the brain and the spinal cord. Neural crest cells form at the seam of this tube and rapidly migrate throughout the body to form the bones and cartilage of the face; the neurons and glia of the peripheral nervous system; heart structures; a portion of the gut; and many other important components of the developing organism.

Ruchi Bajpai, PhD, a postdoctoral scholar in Wysocka's lab and first author of the study, coaxed human embryonic stem cells to become what resembles functional neural crest cells in a laboratory dish. These cells could become neurons and many other cell types derived from the neural crest. When the researchers suppressed CHD7 expression, they saw that fewer neural crest cells formed and migrated across the surface of the dish.

For obvious ethical reasons, the researchers couldn't study the effect of tweaking CHD7 levels in human embryos. Because the problems occur so early in development, Wysocka and her colleagues turned to frog embryos to test how CHD7's activity affected neural crest cells in a living animal. Unlike mice, frog embryos develop outside of the body and can be easily monitored. Researchers found that blocking CHD7 expression or its activity in frog embryos interfered with the ability of the neural crest cells to migrate during development. What's more, the resultant tadpoles also exhibited many of the major clinical features of human CHARGE syndrome.

"This gave us confidence that we were on the right track," said Wysocka. "It's apparent that CHD7 is required for the reprogramming and migration of the neural crest cells, which is when one would predict major changes in chromatin organization would be taking place."

Further research showed that CHD7 works with another protein called PBAF to bind areas of DNA associated with, but far from, genes involved in neural crest cell specialization and migration. These so-called distal DNA elements control the expression of faraway genes. "It's a long-distance relationship," said Wysocka.

The finding may not only lead to a new understanding of CHD7's role in CHARGE syndrome, it also suggests that CHD7 and PBAF may be involved in the reprogramming and migration of other types of cells, such as cancer cells. Two genes controlled by CHD7 and PBAF -- called Twist and Slug -- have been implicated in metastasis in many human cancers.

"If we can cause a CHARGE syndrome in tadpoles simply by reducing CHD7 levels by twofold," said Wysocka, "it's possible that increases in CHD7 levels in cancer may significantly enhance the metastasis program. Interestingly, CHD7 duplications have been recently associated with small-cell lung cancer, one of the most highly metastatic and aggressive types of cancer. "

In addition to Wysocka and Bajpai, other Stanford scientists involved in the research include graduate student Denise Chen; postdoctoral researchers Alvaro Rada-Iglesias, PhD, and Yiqin Xiong, PhD; associate professor of surgery Jill Helms, DDS, PhD; assistant professor of medicine Ching-Pin Chang, MD, PhD; and senior research scientist Tomek Swigut, PhD. The research was supported by a SEED grant from the California Institute for Regenerative Medicine, the W.M. Keck Foundation, the Searle Scholars Program and the National Institutes of Health.

More information about Stanford's departments of developmental biology and chemical and systems biology, which supported the work, is available at and

Journal Reference:
Ruchi Bajpai, Denise A. Chen, Alvaro Rada-Iglesias, Junmei Zhang, Yiqin Xiong, Jill Helms, Ching-Pin Chang, Yingming Zhao, Tomek Swigut & Joanna Wysocka. CHD7 cooperates with PBAF to control multipotent neural crest formation. Nature, 2010; DOI: 10.1038/nature08733


Nuclear Pore Complexes Harbor New Class of Gene Regulators

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ScienceDaily (Feb. 4, 2010) — Nuclear pore complexes are best known as the communication channels that regulate the passage of all molecules to and from a cell's nucleus. Researchers at the Salk Institute for Biological Studies, however, have shown that some of the pores' constituent proteins, called nucleoporins, pull double duty as transcription factors regulating the activity of genes active during early development.

This is the first time nucleoporins' gene regulatory function has been demonstrated in multicellular organisms, and these findings, reported in the Feb. 5, 2010 issue of Cell, not only reveal a new class of transcription factors but may offer new insights into the mechanisms behind cancer.

"Nuclear pores are not only transport channels but play a role in the organization of the genome and a very direct role in gene expression," says senior author Martin Hetzer, Ph.D., Hearst Endowment associate professor in Salk's Molecular and Cell Biology Laboratory. "NPC components, called nucleoporins, are also present in the nuclear interior and bind to certain genes, which puts them in a new class of gene regulators."

Despite evidence in yeast that nuclear pores are required for expression of certain genes, it was unknown how the connection worked. Since nuclear pore complexes are part of the nuclear membrane, which delineates the cell nucleus, it had been assumed that when a gene was regulated by a nucleoporin, it somehow had to make its way to the periphery of the nucleus, adjacent to the nuclear pore.

It was also unknown whether similar regulatory properties were present in multicellular organisms. For more than a decade, however, scientists had known that when the protein called Nup98, a nucleoporin, abnormally fuses with certain proteins that regulate gene expression, the marriage causes leukemia. What is more, nucleoporins Nup214 and Nup88 are highly overexpressed in other cancers, including colon cancer and very aggressive forms of lung cancer. Scientists had long questioned why these components of the cell's transport channel are implicated in cancer and had theorized that the connection might stem from a problem related to the conveyance of molecules in and out of the nucleus.

To probe the role of these proteins, Hetzer and his group studied the development of salivary glands in fruit flies. The chromosomes in Drosophila salivary glands are polytenized -- meaning that they undergo multiple rounds of replication without cell division, resulting in giant chromosomes, which can be easily visualized by a light microscope. Using antibodies, Capelson and her collaborators were able to detect the binding of Nup98, SEC13, and FG-containing nucleoporins to specific genes on the polytene chromosomes. When they did a three-dimensional reconstruction of the nuclei, they found the nucleoporins inside the nucleus.

"Very few studies have looked at nuclear pore components for their potential role in gene regulation in animal cells," says Hetzer. "The fact that NPC components can interact with genes inside the nucleus makes a lot more sense in how they can regulate gene activity. The gene doesn't go to the pore; the pore protein goes to the gene."

"There had been a longstanding mystery about nuclear pores and active genes," adds first author Maya Capelson, Ph.D., a postdoctoral fellow in Hetzer's lab. "The ability of the nuclear pore proteins to come off the pore and go inside the nucleus provides a global mechanism for the involvement of pores in gene regulation."

The nucleoporins don't regulate all genes, but are required for a subset of genes, including developmentally regulated genes, which are turned on and off in a controlled manner during cell differentiation and tissue development. "What is exciting to us is that they are key regulators for developmental genes and also potential markers for causes of cancer," Hetzer explains.

This finding is significant in the context of Nup98's relationship to leukemia when it fuses with other DNA binding proteins. Why Nup98's translocation causes leukemia is unknown, but the discovery of Nup98 as bona fide transcription regulator has major implications for cancer and the cause of leukemia.

Previously, Hetzer's lab had shown that nuclear pore components deteriorate during aging in post-mitotic cells. His team is now investigating whether age-dependent deterioration in long-lived cells, such as neurons and muscles, could also explain why certain genes are misregulated. Damage to those components, he suggests, may help explain why gene expression changes as we age -- a phenomenon that is very well known but not understood. "Our hypothesis is that target genes may be misexpressed in aging cells," he says.

This study was partly supported by grants from the National Institute of General Medical Sciences and the Damon Runyon Cancer Research Foundation. Researchers who also contributed to this study included Yun Liang, Roberta Schulte, William Mair of the Salk Institute and Ulrich Wagner from Bing Ren's laboratory at UCSD.


Cell Growth Regulates Genetic Circuits

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ScienceDaily (Feb. 3, 2010) — Genetic circuits control the activity of genes and thereby the function of cells and organisms. Scientists from the Max Planck Institute of Colloids and Interfaces in Potsdam and the University of California at San Diego have shown how various genetic circuits in bacterial cells are influenced by growth conditions. According to their findings, even genes that are not regulated can display different activities -- depending on whether they are translated into proteins in slow- or fast-growing cells.

The results provide researchers with new insights into gene regulation and will help them in the design of synthetic genetic circuits in the future (Cell 139, 1366-1375, 2009).

Control circuits do not only exist in CD players, coffee makers, or cars, but also in living cells -- in this case as "genetic circuits." They consist of a network of different genes which can mutually stimulate or inhibit each other. With the help of these circuits, a cell can switch genes on or off and thus control what proteins it produces. However, genetic circuits also depend on the functioning of the cell as a whole to provide sufficient resources needed for building proteins. For example, the standard laboratory bacterium, Escherichia coli, can adjust its generation time to anywhere between 20 minutes (under optimal conditions) and several hours (e.g. when food is scarce). The change in generation time or growth rate is accompanied by changes of almost all properties of the cells such as their size and their chemical composition.

Protein concentration of unregulated genes decreases at faster growth

By combining theoretical circuit models and experiments with simple synthetic circuits in bacteria the scientists demonstrated that growth rate decisively influences the activities of genes and thus genetic circuits. "We wanted to know how the activity of a hypothetical gene that is not regulated at all would depend on the growth rate of the bacteria. This dependence needs to be taken into account when interpreting changes in gene expression in experiments," says Stefan Klumpp, a Research Group Leader at the Max Planck Institute of Colloids and Interfaces in Potsdam.

Changes within the cell affect protein concentration in various ways. For example, in faster-growing cells more RNA polymerases are available for the transcription of the gene, so the gene is read out more frequently. But there is also less time to accumulate the protein before the next cell division. In addition, faster-growing cells are bigger, so making the same number of protein molecules amounts to a smaller concentration. Assembling all that information in their model, the researchers were able to predict how the protein concentration would depend on the bacterial growth rate. What they discovered was that protein concentration decreases at faster growth rates -- a result that matched well with experimental data for unregulated genes.

That the activity of genes and genetic circuits depends on how fast the cells grow makes life more complicated for scientists interested in quantitative characteristics of genetic circuits. Different quantities used to characterize the activity of a gene such as its mRNA and protein concentrations depend on the growth rate in different ways. "For example if the concentration of a particular mRNA is increased by a factor 3 in one condition compared to another, this would usually be taken as a solid indication that the gene is up-regulated," Stefan Klumpp explains.

"But if the cells with the higher mRNA level also grow faster, they could even have a smaller concentration of the corresponding protein." Changes in protein concentration are not necessarily a consequence of regulated gene expression. Such changes can also be the result of a slowdown or acceleration in cell growth.

Feedback between regulated genes and cell growth

The study also shows how such growth effects interact with gene regulation. For example, protein concentration can be made independent of the growth rate if the gene is controlled by negative feedback: Negative feedback stops synthesis of the protein when a certain desired concentration is reached. If the cell continues to grow, protein concentration is initially decreased, so that further protein is built up until the desired concentration is achieved.

Growth effects can also provide a feedback mechanism if cell growth depends on the concentration of a protein, which in turn depends on the cellular growth rate. If, for example, a high concentration of a certain protein inhibits cell growth while at the same time slow-growing cells are producing more of this protein (positive feedback), one part of a population of genetically identical cells may grow faster than the rest. This is only due to the fact that a protein inhibiting cell growth is more frequently produced in some cells than in others. This leads to a slowdown of growth of these cells, which in turn increases the protein concentration, resulting in a further slowdown of growth.

Researchers believe that these growth effects may be actively used in nature and could possibly even be beneficial for bacteria. When bacteria acquire new functions such as new metabolic abilities or tolerance to antibiotics, growth effects may regulate such functions without direct gene regulation. This could serve as a base from which a regulatory circuit may evolve later.

Journal Reference:Stefan Klumpp, Zhongge Zhang, Terence Hwa. Growth Rate-Dependent Global Effects on Gene Expression in Bacteria. Cell, 2009; 139 (7): 1366 DOI: 10.1016/j.cell.2009.12.001


3-D Scaffold Provides Clean, Biodegradable Structure for Stem Cell Growth

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ScienceDaily (Feb. 3, 2010) — Medical researchers were shocked to discover that virtually all human embryonic stem cell lines being used in 2005 were contaminated. Animal byproducts used to line Petri dishes had left traces on the human cells. If those cells had been implanted in a human body they likely would have been rejected by the patient's immune system.

Even today, with new stem cell lines approved for use in medical research, there remains a risk that these cells will be contaminated in the same way. Most research labs still use animal-based "feeder layers" because it remains the cheapest and most reliable way to get stem cells to multiply.

Materials scientists at the University of Washington have now created an alternative. They built a three-dimensional scaffold out of a natural material that mimics the binding sites for stem cells, allowing the cells to reproduce on a clean, biodegradable structure. Results published in the journal Biomaterials show that human embryonic stem cells grow and multiply readily on the structure.

"The major challenge for stem cell therapy today is it's very difficult to make a lot of them with high purity," said lead author Miqin Zhang, a UW professor of materials science and engineering. "So far it seems like this material is very good for stem cell renewal."

Medical researchers hope to someday use stem cells to grow new tissues and organs. Key to the research is the fact that new cells maintain the property that holds medical promise -- the ability to differentiate into any of the more than 220 cell types in the adult human body.

Growing the cells in three dimensions better resembles conditions in the human body. It also allows mass production, which will be needed for any clinical applications.

"Three-dimensional scaffolds are an active area of research," said Carol Ware, a UW professor of comparative medicine and expert on stem cells. "They are not commonly used yet, but will be important to transition embryonic stem cells to the clinic. To date, nobody has found a perfect matrix."

Zhang's cylindrical scaffold is made of chitosan, found in the shells of crustaceans, and alginate, a gelatinous substance found in algae. Chitosan and alginate have a structure similar to the matrix that surrounds cells in the body, to which cells can attach. Different processing techniques can make the scaffold out of interconnected pores of almost any size, Zhang said.

Researchers first seeded the scaffold with 500,000 embryonic stem cells, and after 21 days the scaffold was completely saturated. The cells infiltrated the structure, Zhang added, unlike other materials where cells often grow only on the surface.

"This scaffold mimics the extracellular matrix at the atomic level, and so the cells are able to grow in this environment," Zhang said.

To retrieve the cells, researchers immersed the scaffold in a mild solution. The structure is biodegradable and so dissolved to release the stem cells. One also could implant the stem cell-covered scaffold directly into the body.

Analysis of gene activity and testing in the lab and in mice showed that the new stem cells retained the same properties as their predecessors.

Other researcher groups are also looking for alternatives to feeder layers. The leading contenders are scaffolds coated with custom proteins designed to mimic the key properties of the animal cells in the feeder layer. Such products are expensive and difficult to produce in a consistent manner, Zhang said. The proteins also get used up in a few days and have to be replaced, making them costly and time-consuming for everyday use.

"Our scaffold is made of natural materials that are already FDA-approved for food and biomedical applications. Also, these materials are unlimited, and the cost is cheap," she said.

Zhang's group is now working to build a scaffold larger than the current dime-sized prototype, and is collaborating with the UW's Institute for Stem Cells and Regenerative Medicine and UW School of Medicine to try growing different types of stem cells, including those from umbilical cord blood and bone marrow, in the material. They will try to get the resulting cells to differentiate into bone, neuron, muscle and liver cells.

Co-authors are Zhensheng Li and Matthew Leung, UW doctoral students in materials science and engineering; Dr. Richard Hopper, an associate professor at the UW School of Medicine; and Dr. Richard Ellenbogen, professor and chair of neurological surgery at the UW School of Medicine.


Microbes Produce Fuels Directly from Biomass

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ScienceDaily (Jan. 31, 2010) — A collaboration led by researchers with the U.S. Department of Energy's Joint BioEnergy Institute (JBEI) has developed a microbe that can produce an advanced biofuel directly from biomass. Deploying the tools of synthetic biology, the JBEI researchers engineered a strain of Escherichia coli (E. coli) bacteria to produce biodiesel fuel and other important chemicals derived from fatty acids.

"The fact that our microbes can produce a diesel fuel directly from biomass with no additional chemical modifications is exciting and important," says Jay Keasling, the Chief Executive Officer for JBEI, and a leading scientific authority on synthetic biology. "Given that the costs of recovering biodiesel are nowhere near the costs required to distill ethanol, we believe our results can significantly contribute to the ultimate goal of producing scalable and cost effective advanced biofuels and renewable chemicals."

Keasling led the collaboration, which was was made up of a team from JBEI's Fuels Synthesis Division that included Eric Steen, Yisheng Kang and Gregory Bokinsky, and a team from LS9, a privately-held industrial biotechnology firm based in South San Francisco. The LS9 team was headed by Stephen del Cardayre and included Zhihao Hu, Andreas Schirmer and Amy McClure. The collaboration has published the results of their research in the January 28, 2010 edition of the journal Nature. The paper is titled, "Microbial Production of Fatty Acid-Derived Fuels and Chemicals from Plant Biomass."

A combination of ever-increasing energy costs and global warming concerns has created an international imperative for new transportation fuels that are renewable and can be produced in a sustainable fashion. Scientific studies have consistently shown that liquid fuels derived from plant biomass are one of the best alternatives if a cost-effective means of commercial production can be found. Major research efforts to this end are focused on fatty acids -- the energy-rich molecules in living cells that have been dubbed nature's petroleum.

Fuels and chemicals have been produced from the fatty acids in plant and animal oils for more than a century. These oils now serve as the raw materials not only for biodiesel fuel, but also for a wide range of important chemical products including surfactants, solvents and lubricants.

"The increased demand and limited supply of these oils has resulted in competition with food, higher prices, questionable land-use practices and environmental concerns associated with their production," Keasling says. "A more scalable, controllable, and economic alternative route to these fuels and chemicals would be through the microbial conversion of renewable feedstocks, such as biomass-derived carbohydrates."

E. coli isa well-studied microorganism whose natural ability to synthesize fatty acids and exceptional amenability to genetic manipulation make it an ideal target for biofuels research. The combination of E. coli with new biochemical reactions realized through synthetic biology, enabled Keasling, Steen and their colleagues to produce structurally tailored fatty esters (biodiesel), alcohols and waxes directly from simple sugars.

"Biosynthesis of microbial fatty acids produces fatty acids bound to a carrier protein, the accumulation of which inhibits the making of additional fatty acids," Steen says. "Normally E. coli doesn't waste energy making excess fat, but by cleaving fatty acids from their carrier proteins, we're able to unlock the natural regulation and make an abundance of fatty acids that can be converted into a number of valuable products. Further, we engineered our E. coli to no longer eat fatty acids or use them for energy."

After successfully diverting fatty acid metabolism toward the production of fuels and other chemicals from glucose, the JBEI researchers engineered their new strain of E. coli to produce hemicellulases -- enzymes that are able to ferment hemicellulose, the complex sugars that are a major constituent of cellulosic biomass and a prime repository for the energy locked within plant cell walls.

"Engineering E. coli to produce hemicellulases enables the microbes to produce fuels directly from the biomass of plants that are not used as food for humans or feed for animals," Steen says. "Currently, biochemical processing of cellulosic biomass requires costly enzymes for sugar liberation. By giving the E. coli the capacity to ferment both cellulose and hemicellulose without the addition of expensive enzymes, we can improve the economics of cellulosic biofuels."

The JBEI team is now working on maximizing the efficiency and the speed by which their engineered strain of E. coli can directly convert biomass into biodiesel. They are also looking into ways of maximizing the total amount of biodiesel that can be produced from a single fermentation.

"Productivity, titer and efficient conversion of feedstock into fuelare the three most important factors for engineering microbes that can produce biofuels on an industrial scale," Steen says. "There is still much more research to do before this process becomes commercially feasible."

This research was supported by funds from LS9, Inc., and the UC Discovery Grant program. LS9 is using synthetic biology techniques to develop patent-pending UltraClean™ fuels and sustainable chemicals. The UC Discovery Grant program is a three-way partnership between the University of California, private industry and the state of California that is aimed at strengthening and expanding California's economy through targeted fields of research.


Breakthrough Could Lead to New Treatment for Malaria

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ScienceDaily (Jan. 29, 2010) — Malaria causes more than two million deaths each year, but an expert multinational team battling the global spread of drug-resistant parasites has made a breakthrough in the search for better treatment. Better understanding of the make-up of these parasites and the way they reproduce has enabled an international team, led by John Dalton, a biochemist in McGill's Institute of Parasitology, to identify a plan of attack for the development of urgently needed new treatments.

Malaria parasites live inside our red blood cells and feed on proteins, breaking them down so that they can use the proceeds (amino acids) as building blocks for their own proteins. When they have reached a sufficient size they divide and burst out of the red cell and enter another, repeating the process until severe disease or death occurs. Dalton and his colleagues found that certain "digestive enzymes" in the parasites enable them to undertake this process. Importantly, the researchers have also now determined the three-dimensional structures of two enzymes and demonstrated how drugs can be designed to disable the enzymes.

"By blocking the action of these critical parasite enzymes, we have shown that the parasites can no longer survive within the human red blood cell," Dalton explains. The discovery will be published in the Proceedings of the National Academy of Sciences, and is the result of collaboration including Australia's Queensland Institute of Medical Research, Monash University and the University of Western Sydney, Wroclaw University of Technology in Poland and the University of Virginia in the U.S. The team is putting their findings into action immediately and is already pursuing anti-malarial drug development.


Secrets of Immunologic Memory: New Understanding of CD44 Receptor's Role in Immune Cell Survival

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ScienceDaily (Jan. 28, 2010) — Investigators at Sanford-Burnham Medical Research Institute (Sanford-Burnham) have discovered a new way the cell surface protein, CD44, helps specific T helper (Th1) cells develop immunologic memory. Linda Bradley, Ph.D., Bas Baaten, Ph.D., and colleagues determined that without CD44, Th1 cells died off during their initial immune response and were unable to generate immunologic memory. This is the first time scientists have identified this unique CD44 function on Th1 cells, making the protein a potential target to treat a variety of diseases.

The study was published online on January 14 in the journal Immunity.

CD44, a protein found on many cell types throughout the body, binds to the glycan hyaluronic acid (HA) in the extracellular matrix. When T helper cells are activated by infection, they upregulate (increase the activity of) CD44. Though CD44 is a marker for these "experienced" cells, its function has remained elusive. T cells are important components in the body's defense against diseases and, as memory cells, provide immunity to subsequent infections.

"In various infections and autoimmune conditions, Th1 cells are often the bad guys," said Dr. Bradley. "They can contribute to disease by overproducing cytokines and are often responsible for the disease pathology. Our findings reveal an opportunity to harness CD44 to control this pathogenesis."

The Bradley laboratory used T cells lacking CD44 that recognized a protein fragment in the influenza virus. Noting that the T cells did not survive and were unable to generate immunologic memory, the laboratory determined that CD44 protected the cells from programmed cell death initiated by the Fas receptor. This protective effect was specific to a subset of T cells, the Th1 cells, and mediated by the PI 3 kinase pathway. The researchers also demonstrated that survival of the cells could be controlled by antibodies capable of modulating CD44 signaling.


Lack of Cellular Enzyme Triggers Switch in Glucose Processing

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ScienceDaily (Jan. 27, 2010) — A study investigating how a cellular enzyme affects blood glucose levels in mice provides clues to pathways that may be involved in processes including the regulation of longevity and the proliferation of tumor cells. In their report in the January 22 issue of Cell, a Massachusetts General Hospital (MGH)-based team of researchers describes the mechanism by which absence of the enzyme SIRT6 induces a fatal drop in blood sugar in mice by triggering a switch between two critical cellular processes.

"We found that SIRT6 functions as a master regulator of glucose levels by maintaining the normal processes by which cells convert glucose into energy," says Raul Mostoslavsky, MD, PhD, of the MGH Cancer Center, who led the study. "Learning more about how this protein controls the way cells handle glucose could lead to new approaches to treating type 2 diabetes and even cancer."

SIRT6 belongs to a family of proteins called sirtuins, which regulate important biological pathways in organisms from bacteria to humans. Originally discovered in yeast, sirtuins in mammals have been shown to have important roles in metabolic regulation, programmed cell death and adaptation to stress. SIRT6 is one of seven mammalian sirtuins, and Mostoslavsky's team previously showed that mice lacking the protein die in the first month of life from acute hypoglycemia. The current study was designed to investigate exactly how lack of SIRT6 causes this radical drop in blood sugar.

Normally cells convert glucose into energy through a two-step process. The first stage called glycolysis takes place in the cytoplasm, where glucose is broken down into an acid called pyruvate and a few molecules of ATP, the enzyme that provides the energy to power most biological processes. Pyruvate is taken into cellular structures called mitochondria, where it is further processed to release much greater amounts of ATP through a process called cellular respiration.

In a series of experiments in mouse cells, the researchers showed that SIRT6-deficiency hypoglycemia is caused by increased cellular uptake of glucose and not by elevated insulin levels or defects in the absorption of glucose from food. They then found increased levels of glycolysis and reduced mitochondrial respiration in SIRT6-knockout cells, something usually seen when cells are starved for oxygen or glucose, and showed that activation of the switch from cellular respiration to glycolysis is controlled through SIRT6's regulation of a protein called HIF1alpha. Normally, SIRT6 represses glycolytic genes through its role as a compactor of chromatin -- the tightly wound combination of DNA and a protein backbone that makes up chromosomes. In the absence of SIRT6, this structure is opened, causing activation of these glycolytic genes. The investigators' finding increased expression of glycolytic genes in living SIRT6-knockout mice -- which also had elevated levels of lactic acid, characteristic of a switch to glycolytic glucose processing -- supported their cellular findings.

Studies in yeast, worms and flies have suggested a role for sirtuins in aging and longevity, and while much of the enzymes' activity in mammals is unclear, SIRT6's control of critical glucose-metabolic pathways could signify a contribution to lifespan regulation. Elevated glycolysis also is commonly found in tumor cells, suggesting that a lack of SIRT6 could contribute to tumor growth. Conversely, since knocking out SIRT6 causes blood sugar to drop, limited SIRT6 inhibition could be a novel strategy for treating type 2 diabetes.

"There's a lot we still don't know about SIRT6," adds Mostoslavsky, who is an assistant professor of Medicine at Harvard Medical School. "We need to identify the factors that interact with SIRT6 and determine how it is regulated; investigate whether it acts as a tumor suppressor and how it might help lower glucose levels in diabetes; and determine its target organs in living animals, all of which we are investigating."

Lei Zhong of the MGH Cancer Center is lead author of the Cell report. Co-authors are Agustina D'Urso, Debra Toiber, Carlos Sebastian, Douangsone Vadysirisack, Othon Iliopoulos, and Leif Ellisen, MGH Cancer Center; Alexander Guimaraes, Brett Marinelli, and Ralph Weissleder, MGH Center for Systems Biology; Ryan Henry and Joaquin Espinosa, Howard Hughes Medical Institute; Jakob Wikstrom and Orian Shirihai, Boston University School of Medicine; Tomer Nir and Yuval Dor, Hebrew University-Hadassah Medical School; Clary Clish, Broad Institute; and Bhavapriya Vaitheesvaran, Albert Einstein College of Medicine. The study was supported by grants from the V Foundation, the Sidney Kimmel Cancer Research Foundation, the American Federation for Aging Research, Massachusetts Life Sciences Center, Joslin Diabetes Center and the Boston Area Diabetes Endocrinology Research Center.


Unwanted Guests: How Herpes Simplex Virus Gets Rid of the Cell's Security Guards

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ScienceDaily (Jan. 26, 2010) — A viral infection is like an uninvited, tenacious houseguest in the cell, using a range of tricks to prevent its eviction. Researchers at the Salk Institute for Biological Studies have identified one of the key proteins allowing herpes simplex virus (HSV) DNA to fly under the radar of their hosts' involuntary hospitality.

Their findings, to be published in a forthcoming issue of The EMBO Journal, reveal one of the tactics that HSV employs in order to overcome its hosts' defenses and may suggest a common mechanism by which viruses can successfully infect host cells.

HSV, like all viruses, requires a living host in order to multiply. But before it can hijack the cellular machinery to produce scores of copies of itself, it needs to evade the cell's security system. "We found that detection of the viral DNA by the host cell is an important barrier that the virus must overcome in order to achieve its goal," says Matthew Weitzman, Ph.D., associate professor in the Laboratory of Genetics, who led the study. "For this purpose, it brings along a protein that shuts down the normal cellular responses that would otherwise recognize and silence it."

To the host cell, invading viral DNA looks just like the product of DNA damage, which must be repaired or removed in order for the cell to stay healthy. As a result, DNA "security guards" continuously patrol our cells looking for unusual bits of DNA. "We reasoned that viral DNA would be recognized by the cell's DNA repair machinery and that the virus must somehow manipulate the cell's response to this foreign DNA," explains Weitzman.

To test this hypothesis, Weitzman and his team looked at what happens in a virus- infected cell when its DNA is damaged. In a normal cell, DNA damage sensor proteins rush to the site of damage. In cells infected with HSV, however, the cells' emergency repair teams don't respond correctly. "The virus effectively overrides the cell's DNA damage response in order to prevent its own DNA from being recognized," says Weitzman.

The team went on to identify a single viral protein that is to blame for knocking out the cell's security system, a protein called ICP0. They discovered that it flags for destruction two important DNA "security guards," the proteins called RNF8 and RNF168, thereby taking out the DNA damage response in human cells in one big swipe.

ICP0 attaches so-called ubiquitin marks, which instruct the cell to get rid of the very proteins that protect it. With RNF8 and RNF168 safely out of the way, the virus can begin to take over.

Delving deeper, the team looked at the role of these DNA "security guards" that are singled out by ICP0. Surprisingly, RNF8 and RNF168 also leave ubiquitin tags, but in this case, they mark regions of damage. They tag a protein called histone H2A, which directs DNA damage response proteins to accumulate at the sites of damage. The Salk team discovered that by removing RNF8 and RNF168, the viral ICP0 protein results in a decrease to the tag on the cellular H2A protein.

"We found that HSV targets the mark that is required to keep DNA damage sensors at damage sites," says postdoctoral researcher and first author Caroline Lilley, Ph.D. "We now think that HSV deliberately removes this mark so that the virus can infect cells without any trouble from its new host."

The findings highlight the importance of these histone marks in DNA damage. "By identifying how HSV dismantles the host's defense systems, we are shown the key steps, not only in viral infections, but also in the human DNA damage response," Weitzman explains.

HSV may have evolved this weapon because our cells use histone ubiquitination to try to silence gene expression from the viral DNA. "Ubiquitination would be a great way for the cell to silence the viral genome, and ICP0 provides the counterattack by the virus, so the virus and the cell are battling it out at that point," says Mira Chaurushiya, a graduate student in the lab and contributor to the study.

This work may point to a general mechanism viruses use to overcome the cell's defense. "DNA damage signaling and ubiquitination may be part of an anti-viral defense mechanism," explains Lilley. "Part of the cell's defense is to try to silence viral genomes, and viral proteins have to prevent this in order to achieve infection."

Along with Weitzman, Lilley and Chaurushiya, other contributors to this work were Sebastien Landry and Junghae Suh from the Salk Institute's Laboratory of Genetics (J.S. is now at Rice University, Houston); Stephanie Panier and Daniel Durocher from the Samuel Lunenfeld Research Institute, Mount Sinai Hospital in Toronto, Canada; Chris Boutell and Roger D. Everett at the University of Glasgow, UK; and Grant S. Stewart at Birmingham University, UK.


Saturday, February 6, 2010

New Insight Into Reprogramming of Cell Fate

Site of the day:

ScienceDaily (Feb. 1, 2010) — A discovery by Babraham scientists brings new insight into how cells are reprogrammed and a greater understanding of how the environment, or factors like nutritional signals, can interact with our genes to affect health. As an embryo develops, cells acquire a particular fate, for example becoming a nerve or skin cell. The findings, reported online in the journal Nature, pinpoint a protein called AID as being important for complete cellular reprogramming in mammals. In addition, these findings may advance the field of regenerative medicine, by potentially enhancing our ability to guide the reversal of cell fate, and pave the way for novel therapeutics.

Cell fate is governed not only by the genome, but also by chemical changes to DNA and its associated proteins, a research field called epigenetics. Modifying DNA by methylation for example, alters the DNA structure but not its sequence. These 'epigenetic' tags are one of the ways that genes get switched on or off in different places at different times, enabling different tissues and organs to arise from a single fertilised egg. When epigenetic processes go awry, diseases may occur. Epigenetics is therefore emerging as an important research area with relevance to understanding many adult conditions like heart disease, diabetes, obesity, cancer and autoimmune disorders.

Professor Wolf Reik, Associate Director at the Babraham Institute and Professor of Epigenetics at the University of Cambridge who led the research said, "With numerous human, animal and plant genomes now sequenced a key question is how genomes are regulated in normal development, health and disease. Altered regulation of the epigenome is likely to underlie many human diseases so unlocking the principles of reprogramming can be harnessed to benefit regenerative medicine and stem cell therapy."

This research at Babraham, an institute of the Biotechnology and Biological Sciences Research Council (BBSRC), reveals that AID plays an intriguing role in erasing the chemical marks that appear on the genome as an embryo develops and determine what a cell's identity will be. AID appears to be involved in removing the epigenetic tags from DNA by a process called demethylation, which has long been known to be a critical component of cellular reprogramming. A study published recently in Nature from Helen Blau's lab in Stanford backs up the findings that AID is important for reprogramming.

While it has been known that epigenetic modifications to the genome get erased and re-established in the early embryo, precisely how and the extent to which this occurs had remained elusive. This collaboration between scientists at Babraham, the Howard Hughes Medical Institute and University of California at Los Angeles (UCLA) reveals for the first time the massive extent to which erasure of epigenetic tags occurs in mammals, erasing the epigenome between generations.

They discovered that methylation levels drop from 80% to a staggering 7% before being re-established again. This defines the level of epigenetic inheritance of DNA methylation patterns between generations and is identifying parts of the genome apparently more resistant to reprogramming than others. Reik explained, "Whole epigenomes can now be unravelled and understood thanks to Next Generation Sequencing technology which we used in collaboration with the UCLA team, and which we also have at the Babraham, a partner in the East Anglia Sequencing and Informatics Hub."

The Aid gene is normally switched on early as the embryo develops, however, the Babraham team found that if the AID protein is missing in cells, the methylation patterns are not thoroughly wiped clean and an epigenetic 'memory' is inherited. Commenting on the discovery Reik said, "Clear mechanisms for DNA demethylation have been elusive for some time. The body of evidence is now pointing to indirect demethylation through the action of key enzymes such as AID."

Environmental factors can also affect the genome, producing epigenetic changes that influence cell behaviour. Reik added, "It is now well established that epigenetics is the 'integrator' between the environment and the genome and that external factors like nutritional signals may have consequences later in life or on future generations. There is also the possibility that epigenetic information could be inherited across generations, providing a shorter term and flexible type of inheritance in response to environmental signals. The ability to unravel whole epigenomes during normal development and healthy ageing, and to understand how epigenomes are modified by the environment is extremely exciting."

It is known that removing epigenetic information from the genome can induce adult cells to regain stem-cell like properties (induced pluripotent stem cells, iPS cells). Inducing 'pluripotency' is of direct relevance to regenerative medicine as it enables specific cell populations and tissues to be generated from and for patients. Currently reprogramming is inefficient because of the memory imparted by DNA methylation tags. These new findings pinpointing how DNA demethylation can be driven, may overcome a significant barrier in producing iPS cells.

The identification of proteins like AID, that drive epigenetic signalling, is an important advance in basic biomedical research, which may help define new targets and therapeutics for diseases including cancer. The Babraham team are pursuing commercial applications in collaboration with the company CellCentric.

"Epigenetics is a growing area of academic research and commercial development. By understanding what proteins cause cell fate change, new tools and methods can be designed for both regenerative medicine and the treatment of intractable diseases. Specifically, the identification of AID and its activity may offer the ability to test the importance of gene-specifc demethylation, as well as the potential to overcome a pivotal epigenetic barrier in reprogramming cells for induced pluripotent cell production," explained Dr Will West, CEO of CellCentric.

Journal Reference:
Popp et al. Genome-wide erasure of DNA methylation in mouse primordial germ cells is affected by AID deficiency. Nature, 2010; DOI: 10.1038/nature08829