Thursday, January 28, 2010

The LHC enters a new phase

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After achieving the world record energy of 1.18 TeV per proton beam last November, the LHC is now preparing for higher energy and luminosity.

Before the 2009 running period began, all the necessary preparations to run the LHC at the collision energy of 1.18 TeV per beam had been carried out. The goal of the technical stop, which will end mid-February, is to prepare the machine for running at 3.5 TeV per beam. In order to achieve that, a current as high as 6 kAmps will have to flow into the LHC magnets.

The main work is taking place on the new quench protection system (nQPS) where teams are improving the electrical reliability of the connection between the Instrumentation Feedthrough Systems (IFS) on the magnets and the nQPS equipment. There are around 500 of these connectors for each of the eight sectors in the LHC that need to be repaired. These operations are necessary to ensure that higher currents can be safely handled.

The interventions on the nQPS follow a very strict procedure that ensures an extremely accurate quality control of the repairs: after repairing the electrical connections, specialized teams make a first visual control to check that everything is properly assembled; after that, the Electrical Quality Assurance (ELQA) team performs the high voltage tests; finally, the Hardware Commissioning teams power the magnets, initially with low current and finally at 6 kAmps.

Several teams are taking advantage of the technical stop to carry on other technical verifications and efficiency tests, for instance on some vacuum pumping units, the kicker system, the Oxygen Deficiency Hazard detectors, some ventilation components and a few others.

At present, all teams are on schedule: the powering tests have just started in Sectors 8-1 and 1-2; Sectors 2-3, 3-4 and 4-5 will follow shortly. According to the planning Sector 1-2 will be the first sector ready for beam operation by the end of January.

At the same time as the LHC interventions, repairs are going on at CMS on the water cooling system. All work, both in the LHC and in the CMS experiment, is expected to be completed by mid-February.


Monday, January 25, 2010

In Journey from Maggot to Fruit Fly, a Clue About Cancer Metastasis

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ScienceDaily (Jan. 21, 2010) — Scientists trying to understand how cancer cells invade healthy tissue have used the fruit fly's metamorphosis from maggot to flying insect as a guide to identify a key molecular signal that may be involved in both processes.

The research by a team at the University of Rochester Medical Center, published as the cover article in the January issue of the journal Developmental Cell, identifies a molecule that is key for determining how cells invade and create new tissues. That process, which is what makes cancer so deadly, has proven very difficult for scientists to understand in enough depth to interrupt to stop the spread of cancer.

At a glance, the change from crawling maggot to flittering fruit fly seems a long way from the search for new treatments or cures for human health. But there are good reasons scientists study the fruit fly. Many processes in fruit flies are very similar to those in people, only simpler and thus much more approachable to the probing eyes of scientists. In the case of cancer, the action in fruit flies allows scientists to take a close look at molecular signals that may be involved in both development and disease.

"The principles that govern how organs are made in a fruit fly and in a person are more similar than most people ever believed," said Dirk Bohmann, Ph.D., professor of Biomedical Genetics and the leader of the team. "Many of the same signals that control the growth and organization of fruit fly organs control similar processes in people. If we can understand such signaling in fruit flies, it will help us understand what is happening in people, to try to prevent or stop diseases like cancer in which such signaling process go astray.

"It's surprising, but fruit flies -- which in nature actually never get cancer themselves -- have taught us more about that disease than many other animals that do," added Bohmann, who is also a scientist at the James P. Wilmot Cancer Center.

The first author of the current work is graduate student Qiong Wang. Also contributing to the research was former post-doctoral associate Mirka Uhlirova, Ph.D., who is now a faculty member at the University of Cologne in Germany.

In the current work, the team looked at the development of the air sac in a fruit fly. The air sac is a crucial organ that connects the flight muscles to an air supply, allowing the wings to operate and the insect to fly. The air sac arises late in development, at the same time when the wings and the flight muscle are added to an organism that did not need such structures during its life as a maggot. That's why the growing air sac has to burrow its way through already-existing tissues, a process that requires a good amount of cellular choreography and exactly the kind of molecular trickery that cancer cells need themselves.

During metamorphosis from maggot to fruit fly, complex molecular signals govern the change from rudimentary structures in the maggot to the air sac in the fruit fly.

For scientists like Bohmann who study cancer, understanding how a maggot develops the air sac is an exploration into how some cells develop, dominate, and push out of the way existing tissue while creating a whole new structure. That's what happens in cancer, where cells from a tumor of the prostate, breast, lung or other organ push their way through tissue and spread to other organs. The spread or metastasis of a tumor, not the first tumor itself, is usually what kills a patient.

"A maggot, of course, looks very different from a fruit fly. Generally when it becomes an adult, a maggot pretty much melts down all its material and starts over, building new cells and tissues. But not completely. A maggot has a rudimentary tubular structure that supplies oxygen, and that structure forms the basis for the air sac," said Bohmann.

"The formation of the air sac in a fruit fly is a great example of careful, planned invasive cell movement," added Bohmann. "It resembles, in many ways, the invasive growth carried out by a tumor in the body of a person with aggressive cancer. That's why we think that it's possible that cancer cells spread into healthy tissue by hijacking similar normal mechanisms of tissue growth."

Scientists have known that fibroblast growth factor or FGF plays an important role in directing tissues to migrate and grow through other tissues. FGF has a part in creating the air sac as well as other processes involving growth and spread of tissue.

Now Bohmann's team has identified a molecule that controls FGF. That important task falls to a protein known as a matrix metalloprotease or MMP. Scientists had known that this class of proteins has an important role directly clearing a path for one type of tissue to grow through another. MMPs play a role in a myriad of processes involved in tissue rearrangement, including the growth of lung, breast and kidney tissue.

Bohmann's team found that in the fruit fly, MMP2 controls FGF. While Bohmann says the involvement of an MMP in some aspects of invasive tissue growth has already been known, Bohmann's team found that the protein works by controlling FGF signaling, which is a surprise. If a similar mechanism is at work in humans, scientists might be able to exploit it as a new way to knock out FGF, which contributes to many types of cancer.

Journal Reference:
Qiong Wang, Mirka Uhlirova, Dirk Bohmann. Spatial Restriction of FGF Signaling by a Matrix Metalloprotease Controls Branching Morphogenesis. Developmental Cell, 2010; 18 (1): 157-164 DOI: 10.1016/j.devcel.2009.11.004


Studies Shed New Light on Early Transmembrane Signaling

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ScienceDaily (Jan. 25, 2010) — Two new studies by researchers at the University of Washington further our understanding of the molecular steps in the PLC cascade, a G protein-coupled receptor signaling mechanism that underlies a wide variety of cellular processes, including egg fertilization, hormone secretion, and the regulation of certain potassium channels.

The studies appear online January 25 in the Journal of General Physiology.

Falkenburger et al. take advantage of recent progress in fluorescence technology -- which allows for analysis of biochemical events in single living cells -- to perform a systematic analysis of the PLC signal transmission process.

According to Tamas Balla (National Institutes of Health) in a Commentary accompanying the articles, the new studies extend the kinetic model of the signaling cascade to cover the entire process, from the activation of the M1 muscarinic receptors to the regulation of the potassium channels. Specifically, Falkenburger et al. show the steps that link changes in PtdIns(4,5)P2 -- an important plasma membrane regulatory lipid -- to changes in KCNQ potassium channel activity.

Journal references:
Balla, T. 2010. J. Gen. Physiol. doi:10.1085/jgp.200910396.
Falkenburger, B.H., et al. 2010. J. Gen. Physiol. doi:10.1085/jgp.200910344.
Falkenburger, B.H., et al. 2010. J. Gen. Physiol. doi:10.1085/jgp.200910345.


Scientists Achieve First Rewire of Genetic Switches

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ScienceDaily (Jan. 25, 2010) — Researchers in Manchester have successfully carried out the first rewire of genetic switches, creating what could be a vital tool for the development of new drugs and even future gene therapies.

A team of scientists from the School of Chemistry and the Manchester Interdisciplinary Biocentre (MIB) at The University of Manchester have found a way of hijacking so-called 'riboswitches' and directing gene activity.

Working within cells of bacteria, chemical biologist Professor Jason Micklefield and his team have rewired these genetic switches so they are no longer activated by small naturally occurring molecules found in cells -- but through the addition of a synthetic molecule.

The work builds on the recent discovery that these naturally occurring molecules can turn genes on and off by triggering riboswitches found within a large molecule called 'messenger RNA'.

The research was funded by the Biotechnology and Biological Sciences Research Council (BBSRC) and Selective Chemical Intervention in Biological Systems Initiative.

In the latest research, when Manchester researchers added synthetic molecules, they bound to the riboswitches and caused the genes to spark into life.

The findings are reported in the latest edition of Proceedings of the National Academy of Sciences (PNAS).

The Manchester team monitored how successfully they had re-wired the cells by observing the creation of a gene product that makes the cells glow green.

Dr Neil Dixon, a senior researcher in the team, said: "Being able to selectively activate and regulate genes could have tremendous impact in drug discovery and the emerging field of synthetic biology.

"This technology could be used to turn on and off important biological pathways and processes, leading to a deeper understanding of how cells function.

"The next big challenge is to apply this technology to study biological processes within human cells. This could allow us to discover more about our hugely complex biological selves."
The Manchester team is now working on ways to simultaneously activate and control multiple genes using these re-wired riboswitches.

Journal Reference:
Reengineering orthogonally selective riboswitches. Proceedings of the National Academy of Sciences, 2010; (in press)


Illuminating Protein Networks in One Step

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ScienceDaily (Jan. 24, 2010) — A new assay capable of examining hundreds of proteins at once and enabling new experiments that could dramatically change our understanding of cancer and other diseases has been invented by a team of University of Chicago scientists.

Described January 24 in the journal Nature Methods, the new micro-western arrays combine the specificity of the popular "Western blot" protein assay with the large scale of DNA microarrays. The technique will allow scientists to observe much of a cell's intricate protein network in one experiment rather than peeking at one small piece at a time.

"The proteins are the actual machines that are doing everything in the cell, but nobody's been able to examine them in depth because it's been too complicated. Now, we can begin to do that with this new method," said Richard B. Jones, senior author and assistant professor at and the University of Chicago's Ben May Department for Cancer Research and the Institute for Genomics and Systems Biology.

Since the 1970's, laboratories have used Western blots to measure proteins. Cellular material is loaded into a gel and proteins of different sizes are separated by an electric field. A protein is then targeted by an antibody, allowing scientists to measure the amount present in the cells.

The method has led to numerous findings across the field of cell biology, but is limited by a need for large amounts of cell material and expensive antibodies, and the inability to measure more than a handful of proteins at a time. With hundreds or even thousands of proteins involved in cellular networks, scientists were restricted to observing only a small fraction of protein activity with each experiment.

"When you walk into a dark room and don't have much information, it's difficult to predict where everything is going to be," Jones said. "If someone can simply turn on the light, you don't have to progress one step at a time by bumping into things. With this new technology, you can potentially see everything at the same time."

Micro-western arrays adapt the technology of the micro-array, typically used to assess the expression of thousands of genes in a single experiment, to proteins. With pre-printed micro-western array gels, essentially comprising 96 miniature Western blots, scientists can compare the levels of hundreds of proteins simultaneously, or compare dozens of proteins under dozens of treatment conditions in one shot. Mere nanoliters of cell material and antibodies are needed for the experiments, reducing cost and maximizing the information obtained from a single sample.

To demonstrate the potential of the micro-western array, Jones and colleagues from the University of Chicago and the Massachusetts Institute of Technology looked at the behavior of proteins in a cancer cell line with elevated amounts of epidermal growth factor receptor (EGFR).

"We started asking questions about what we could do that no one else could previously do," Jones said. "We could actually reproducibly see 120 things at a time rather than looking at 1 or 2 or 5."

The experiments found that activating EGFR simultaneously activated several other receptors in the cell -- a new discovery that may explain why some tumors become resistant to cancer therapies.

With more information, the method may potentially be used clinically for more precise diagnoses of cancer and other diseases that can direct individualized treatment.

"In the clinic, you're limited by the fact that typically most cancers are diagnosed by one or two markers; you're looking for one or two markers that are high or low then trying to diagnose and treat an illness," Jones said. "Here, we can potentially measure a collection of proteins at the same time and not just focus on one guess. We've never been able do that before."

Other scientists in the field of systems biology said that micro-western arrays would make possible experiments that were previously beyond the scope of laboratory methods.

"I think this is really a breakthrough technology that allows us to monitor in close to real time the activity profiles of modified signaling proteins, which is essentially impossible right now," said Andrea Califano, professor of biomedical informatics at Columbia University. "This opens up a completely new window in terms of the molecular profiling of the cell."

"One of the biggest hurdles for systems biology is the struggle for high density, dynamic and quantitative data, and the micro-western array method will go a long way to address this problem," said Walter Kolch, director of Systems Biology Ireland and Professor at University College Dublin. "It is a fine example of generating exciting new technology from applying a new idea to an old method."

The work was funded by The University of Chicago Comprehensive Cancer Center, the American Cancer Society, the Cancer Research Foundation, the Illinois Department of Public Health, the National Institutes of General Medical Sciences, the National Cancer Institute, and the National Science Foundation.

конецформыначалоформыJournal Reference:
Richard B. Jones, Mark F. Ciaccio, Chih-Pin Chuu, Joel P. Wagner and Douglas A. Lauffenburger. Systems analysis of EGF receptor signaling dynamics with micro-western arrays. Nature Methods, January 24, 2010


Biophysicists Manipulate 'Zipper,' Reveal Protein Folding Dynamics

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ScienceDaily (Jan. 20, 2010) — Biophysicists at TUM, the Technische Universitaet Muenchen, have published the results of single-molecule experiments that bring a higher-resolution tool to the study of protein folding. How proteins arrive at the three-dimensional shapes that determine their essential functions -- or cause grave diseases when folding goes wrong -- is considered one of the most important and least understood questions in the biological and medical sciences.

Folding itself follows a path determined by its energy landscape, a complex property described in unprecedented detail by the TUM researchers. In the Proceedings of the National Academy of Sciences, they report taking hold of a single, zipper-like protein molecule and mapping changes in its energy landscape during folding and unfolding.

Previous studies, including atomic force microscopy experiments by the same Munich laboratory, have gone a long way toward characterizing energy thresholds or barriers that stand between a protein's unfolded and folded states. Detailed observations of the quick transition from one state to the other have remained elusive. The new results open the door to higher-resolution, direct measurements. Better characterization of the folding process is seen as a vital link in understanding the chain of events leading from DNA coding for a protein to that protein's biological function. Another motivation for research in this field is the search for new drugs and therapies, because malfunctions in protein folding are implicated in a number of serious diseases -- including diabetes, cancer, cystic fibrosis, prion diseases, and Alzheimer's.

This is the latest in a long series of single-molecule biophysical experiments carried out by Professor Matthias Rief and colleagues in the TUM Department of Physics. Co-authors Christof Gebhardt and Thomas Bornschloegl are members of Rief's lab; Gebhardt also is a member of the Munich Center for Integrated Protein Science.

As a model system for studying real-time protein folding dynamics, the TUM scientists chose a so-called leucine zipper found in yeast. It offers, as proteins go, a relatively simple "coiled coil" structure and zipper-like folding action: Picture two amino acid strings side by side, joined at the bottom, open at the top, and made essentially to zip together.

The researchers extended this structure so that they could make independent measurements at the top, bottom, and middle parts of the zipper. They took hold of the free ends at the top of the zipper with handles made of double-stranded DNA. These DNA handles in turn were attached to tiny beads that could be directly manipulated by "optical tweezers" -- a tool based on the ability of laser beams with a certain kind of profile to pin down nanoscale objects. One end of the protein molecule was held fixed, and the other was held under tension but with some freedom to move, so that folding dynamics could be measured directly, in real time, as the protein zipped and unzipped. This arrangement enabled measurements with high resolution in both space and time.

"What I consider the major improvement is that the new experiments allow the observation of thousands of transitions between the folded and the unfolded state," Rief said. "This enables us to detect not only the folded and unfolded states but also, directly, the excursions of the large energy barriers separating those states. This has previously been impossible, and it now allows direct insight into the precise energy profile of this barrier."

Journal Reference:
J. Christof M. Gebhart, Thomas Bornschloegl, and Matthias Rief. Full distance resolved folding energy landscape of one single protein molecule. Proceedings of the National Academy of Sciences, 2010; (in press)


Sunday, January 24, 2010

Inflammation 'on Switch' Also Serves as 'Off Switch'

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ScienceDaily (Jan. 21, 2010) — In a surprising finding, researchers at North Carolina State University have discovered the critical importance of a protein previously believed to be a redundant "on switch" for certain immune-system responses.

Scientists previously understood that the protein called TAB2 activates inflammation, an important biological process that stimulates wound-healing and prevents invasion of harmful organisms. But scientists considered TAB2 nonessential to the process due to the redundant function of a cousin protein, called TAB3, which has no trouble serving as an "on switch" to activate the inflammation process in TAB2's absence.

In a study published in the Jan. 22 edition of the Journal of Biological Chemistry, the NC State researchers show that underestimating TAB2 can be dangerous. Rather than merely serving as an "on switch," TAB2 also serves as an "off switch" that turns off the inflammation process. When TAB2 is absent or knocked out in cell cultures, the inflammation process continues unabated.

Too much inflammation can be a really bad thing. It is associated with human diseases including certain cancers, inflammatory bowel syndrome and psoriasis.

Knowing more about the regulatory mechanisms in cells may one day lead to drugs that can target excessive inflammation, say NC State's Dr. Jun Ninomiya-Tsuji, associate professor of environmental and molecular toxicology, and her graduate student, Peter Broglie, the lead authors of the paper describing the study.

In the study, Ninomiya-Tsuji and Broglie show that cells lacking TAB2 had a prolonged inflammation response. Normally, TAB2 can be counted on to bring a protein called TAK1 close to tumor necrosis factor, or TNF, a circulating molecule that is a normal component of the immune system. Bringing TAK1 close to TNF activates TAK1, thereby starting the inflammatory response.

In normal systems, this inflammatory response would be quickly regulated to prevent too much inflammation. This is done by a regulating molecule called PP6, which deactivates TAK1, and, therefore, the inflammation process. When TAB2 was absent or knocked out, however, PP6 did not shut down TAK1. The NC State scientists infer, then, that TAB2 has a heretofore unknown function -- it brings TAK1 close enough to PP6 to halt the inflammation process.

The NC State scientists were so surprised by the finding that, Broglie says, "Dr. Ninomiya-Tsuji made me replicate the study three times."

The study was funded by a grant to Ninomiya-Tsuji from the National Institutes of Health. Co-authors of the paper included scientists from the University of Virginia and two Japanese universities -- Nagoya University and Osaka University.

Journal Reference:
Peter Broglie et al. A TAK1 kinase adaptor, TAB2, plays dual roles in TAK1 signaling by recruiting both an activator and an inhibitor of TAK1 kinase in TNF signaling pathway. Journal of Biological Chemistry, Jan. 22, 2010 DOI: 10.1074/jbc.M109.090522


Zebrafish Swim Into Drug Development

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ScienceDaily (Jan. 21, 2010) — By combining the tools of medicinal chemistry and zebrafish biology, a team of Vanderbilt investigators has identified compounds that may offer therapeutic leads for bone-related diseases and cancer.

The findings, reported in ACS Chemical Biology, support using zebrafish as a novel platform for drug development.

In 2007, Charles Hong, M.D., Ph.D., and colleagues described using fish embryos to screen for compounds that interfere with signaling pathways involved in early development -- pathways known to play roles in a variety of disease processes. They discovered the compound "dorsomorphin" and demonstrated that it blocked BMP (bone morphogenetic protein) signaling, which has been implicated in anemia, inflammatory responses and bone-related disorders.

But in examining dorsomorphin further, the investigators found that it had other "off-target" effects -- it also blocked the VEGF (vascular endothelial growth factor) receptor and disrupted zebrafish blood vessel development, a process called angiogenesis.

"Off-target effects contribute to side effects and limit the therapeutic potential of small molecule signaling inhibitors," said Hong, assistant professor of Medicine and Pharmacology.

To find compounds that were more selective BMP inhibitors (didn't have the off-target effects), Hong and colleagues opted to use their zebrafish drug discovery screen as a drug development/optimization tool.

Craig Lindsley, Ph.D., director of Medicinal Chemistry for the Vanderbilt Program in Drug Discovery, Corey Hopkins, Ph.D., associate director, and their colleagues used the dorsomorphin "backbone" as a starting point to synthesize many different analogs -- subtly different dorsomorphin-like compounds.

Then Hong and his team tested these compounds for their effects on zebrafish embryonic development.

"We quickly discovered that the two effects of dorsomorphin could be separated -- some analogs only affected patterning and some only affected angiogenesis," Hong said. The investigators biochemically characterized compounds of both types and found very selective and potent BMP inhibitors and selective VEGF inhibitors.

The zebrafish embryo, Hong said, is very good at assessing a compound's selectivity for a certain signaling pathway. Mixed signals from compounds that are not selective (they hit multiple targets) are toxic to the embryo -- it "shuts down development."

The team identified a VEGF inhibitor, for example, that outperformed an existing VEGF inhibitor that was being developed for cancer therapy (blocking angiogenesis cuts off the "supply lines" for a growing tumor) but was pulled from development during a Phase III trial.

"If they (the pharmaceutical company) had tested that compound in zebrafish, they would have quickly learned that it wasn't potent or selective," Hong said.

"Using zebrafish is a novel way to do a structure-activity relationship study" -- a study that examines a series of analog compounds to determine which is the most selective and most potent, he added.

Traditionally, pharmaceutical companies perform these types of studies in vitro, with isolated proteins or cells. But Hong points out that in vitro studies assess only "one dimension" of the biology. Compounds that have great activity in vitro often fail later because they have poor selectivity or because they do not have chemical properties that make them good drugs (they are not "bioavailable").

"The zebrafish assesses selectivity and bioavailability all at the same time," Hong says. "What the traditional approach takes months to do, the zebrafish does in a day."

Because BMP and VEGF inhibitors have therapeutic potential for a variety of diseases, the investigators will begin to test the drug candidates in mouse models.

Hong praised Vanderbilt leaders for putting into place the drug discovery infrastructure that made the work possible.

"Having medicinal chemists and zebrafish biologists together in the same building really fostered our collaboration," he said. "This kind of collaboration would not be likely at the majority of medical institutions."

The research was supported by the Veterans Administration, the Center for Research in Fibrodysplasia Ossificans Progressiva and Related Disorders, the National Institutes of Health and the GSK Cardiovascular Research and Education Foundation.


How Plants 'Feel' the Temperature Rise

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ScienceDaily (Jan. 10, 2010) — Plants are incredibly temperature sensitive and can perceive changes of as little as one degree Celsius. Now, a report in the January 8th issue of the journal Cell, a Cell Press publication, shows how they not only 'feel' the temperature rise, but also coordinate an appropriate response -- activating hundreds of genes and deactivating others; it turns out it's all about the way that their DNA is packaged.

The findings may help to explain how plants will respond in the face of climate change and offer scientists new leads in the quest to create crop plants better able to withstand high temperature stress, the researchers say.

"We've uncovered a master regulator of the entire temperature transcriptome," said Philip Wigge of John Innes Centre in the United Kingdom in reference to the thousands of genes that are differentially activated under warmer versus cooler conditions.

Using the model plant Arabidopsis thaliana the researchers show that a key ingredient for plants' temperature sensing ability is a specialized histone protein, dubbed H2A.Z, that wraps DNA into a more tightly packed structure known as a nucleosome. Wigge likens nucleosomes to compact balls of string. As temperatures rise, H2A.Z histones allow DNA to progressively unwrap, leading nucleosomes to loosen up, they show.

"When it gets warmer, the DNA unwraps," he said, which allows some genes to switch on and others to switch off. They aren't yet sure exactly how all that happens, but Wigge suspects the altered nucleosome structure gives access to sites on the DNA where activators of some genes can bind along with repressors of other genes.

"In addition to H2A.Z containing nucleosomes having more tightly wrapped DNA, our results suggest that the degree of unwrapping may also be responsive to temperature," the researchers wrote. "This result suggests a direct mechanism by which temperature may influence gene expression, since it has been shown that RNA Pol II [the enzyme responsible for transcribing DNA into messenger RNA] does not actively invade nucleosomes, but waits for local unwrapping of DNA from nucleosomes before extending transcription. In this way, genes with a paused RNA Pol II will show increased transcription with greater temperature as local unwrapping is increased." The basic discovery could ultimately prove to have important implications for world food security, the researchers said.

As the number of people and affluence around the world continues to grow, "it is projected that world agriculture will have to increase yields by 70 to 100 percent in the next 100 years," Wigge said. "Under climate change it will be challenging simply to maintain present yields, let alone increase them." Crops such as wheat are particularly vulnerable to very hot and dry summers, he added, as evidenced by the fact that wheat reserves recently fell to their lowest level in 30 years.

He says the new understanding of plants' temperature sensitivity may prove to be critical for breeding more temperature-resistant crops. His team plans to explore this possibility by studying the role of these H2A.Z histones in a model plant that is more closely related to crops.

"We'd like to engineer a plant where we can control the histones in particular tissues such that it is selectively 'blind' to different temperatures," Wigge said. "Obviously you can't make a completely temperature-proof plant, but there is a lot of scope to develop crops that are more resilient to the high temperatures we are increasingly going to experience."

The researchers include S. Vinod Kumar and Philip A. Wigge, of John Innes Centre, Norwich, UK.


Protein Dynamics: Hidden, Transient Life of a Protein Between Active States Illuminated

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ScienceDaily (Jan. 8, 2010) — Understanding the incredibly speedy atomic mechanisms at work when a protein transitions from one shape to another has been an elusive scientific goal for years, but an essential one for elucidating the full panoply of protein function. How do proteins transition, or interconvert, between distinct shapes without unfolding in the process? Until now, this question has been a hypothetical one, approached by computation only rather than experimentation.

In a groundbreaking study this week in Cell, Brandeis researchers reveal for the first time computationally and experimentally the molecular pathway that a protein takes to cross the energy barrier, the "climb over the mountain." The study reports how folded proteins can efficiently change shape while avoiding unfolding, a critical requirement for any protein in the cell.

Using computation and nuclear magnetic resonance (NMR) spectroscopy, the researchers were able to experimentally measure how fast the signaling nitrogen regulatory protein jumps from one shape to another, and to shed light into the atomistic pathway.

"If you think of crossing the energy barrier as reaching the summit of a mountain, what we revealed is the molecular "hiking" path the protein follows from a deep valley, to the area around the summit, and then back into another not quite-as-deep valley," said Brandeis biophysicist and Howard Hughes Medical Institute (HHMI) Investigator Dorothee Kern.

Historically, scientists had proposed that proteins must break apart, or partially unfold, between distinct active shapes. "That never made sense to me," said Kern, "because if you break the shape of the protein, you have to build it new again and that is too complicated and energy-inefficient; it would take too long."

Kern said they discovered that the signaling protein unfolds on the minute timescale, once every five minutes, while interconversion between the functionally active states happens on the microsecond timescale (10,000 times per second).

First computationally and then experimentally, Kern and her colleagues showed that the protein actually never unfolds on the way, but rather goes through transient, or bridging, states that last less than a nanosecond. The proof? In the transient states, hydrogen bonds, which do not exist in the protein's ground states (the valleys), enable function while circumventing the risk of unfolding.

"This is a proof of principle paper; it changes the paradigm of protein dynamics because these transient nonnative atomic interactions were really hidden before," said Kern. "This paper underscores the need to develop an iterative approach between computation and experimentation."

Understanding protein dynamics is essential to improving protein design for all kinds of applications, including engineering, materials science, and pharmaceuticals. "We don't understand how to make proteins change their shape; the missing link is understanding how nature very efficiently and specifically changes the shape of proteins," said Kern. "If we knew the physics of proteins better, it would help us design functional proteins."

The study was funded by grants from HHMI, National Institutes of Health, the National Science Foundation, the Department of Energy, and the Keck Foundation.


Researchers Identify Scaffold Regulating Protein Disposal

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ScienceDaily (Jan. 5, 2010) — How does a cell manage to identify and degrade the diverse types of defective proteins and thus protect the body against serious diseases? The researchers Sabine C. Horn, Professor Thomas Sommer, Professor Udo Heinemann and Dr. Ernst Jarosch of the Max Delbrück Center for Molecular Medicine (MDC) Berlin-Buch, Germany, have now found a crucial piece in this puzzle.

In an enzyme complex that plays a critical role in the quality control of proteins, they discovered a scaffold regulating the identification and disposal of various defectively produced proteins.

Proteins are the building materials and the machinery of life. They are found by the thousands in a cell and carry out vital tasks in the organism.

The production site of many of the proteins is located in a cell organelle called the endoplasmic reticulum (ER). Here the proteins are produced, folded and routed to their destination.

However, during protein production errors can occur: during the process proteins can be folded in the wrong way. Older proteins may also accumulate defects due to environmental stress.

They can lose their original structure and thus fail to carry out their function and may possibly even cause damage. Diseases can develop such as Alzheimer's, Parkinson's or cystic fibrosis. Defective proteins must therefore be detected in the cell and disposed of.

Protein quality control: rejects receive a molecular tag

Proteins run through a quality control process in the cell. For the identification of defective proteins, an enzyme complex -- the HRD-ubiquitin ligase -- plays a key role.

It functions like a kind of tagging machine: If it recognizes the protein as defective, it tags it with a molecule, the protein ubiquitin, thus marking it for disposal.

Great demands are placed on the HRD-ubiquitin ligase, because proteins adapt to their cellular locations and functions and thus have quite different structures.

For instance, there are water-soluble proteins inside the cell as well as water-insoluble proteins that are situated on or in the cell membrane.

Until now it remained unclear how the enzyme complex manages to recognize and mark such different types of proteins.

Flexible scaffold makes tagging machine universally usable

The study of the MDC researchers has now shed light on this puzzle. The researchers have discovered the central and flexible scaffold of the enzyme complex, the subunit Usa1. Depending on what is required, it tethers specific modules of the complex, connecting them with each other.

When identifying and tagging soluble proteins, Usa1 establishes the contact between the subunits Der1 and Hrd1.

Furthermore, the researchers discovered that the HRD-ubiquitin ligase binds with other HRD-ubiquitin ligases to form a larger enzyme complex in order to degrade insoluble membrane proteins. This process is also regulated by the subunit Usa1.

Journal Reference:Horn et al. Usa1 Functions as a Scaffold of the HRD-Ubiquitin Ligase. Molecular Cell, 2009; 36 (5): 782 DOI: 10.1016/j.molcel.2009.10.015


How Precursors of Gene-Regulating Small RNAs Are Sorted by Cellular Machinery

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ScienceDaily (Jan. 5, 2010) — A team of scientists at Cold Spring Harbor Laboratory (CSHL) has determined a hierarchical set of criteria that explain how the molecular precursors of gene-regulating small RNAs are sorted by the cellular machinery.

Led by Benjamin Czech, a group working in the laboratory of CSHL Professor Gregory Hannon posed the question: can distinct patterns be observed in the process that unfolds when double-stranded RNAs enter the RNAi pathway? Shorthand for RNA interference, RNAi is a biological response to double-stranded RNA that can culminate in the regulation of gene expression. It has been observed in a vast range of organisms ranging from plants to worms to flies to man.

An enzyme called Dicer cuts double-stranded RNAs into smaller double-stranded pieces called duplexes. Czech, Hannon and colleagues propose rules governing the next step in the RNAi pathway, in which duplexes are sorted to proteins called Argonautes which are at the core of a molecular complex called RISC (the RNA-Induced Silencing Complex).

"Only one strand of each duplex is chosen," explains Czech, "and which one makes all the difference. In the fruit flies that we used as models for this series of experiments, the selection of one or another strand effectively determines whether the short RNA will seek out and regulate a gene, or whether it will perform another function such as protecting a cell against a viral invader."

The rules determining how a duplex is processed and sorted are discussed in a paper the team published recently in Molecular Cell. These include the overall arrangement of the nucleotides in the duplex; how many bases are paired; where they're paired and unpaired; and how tightly the ends of the duplex are stuck together.

"These rules for sorting are important for two reasons," according to Hannon, who is also an Investigator of the Howard Hughes Medical Institute. "One is that since small RNAs play critical biological roles in nearly every process, understanding which strands of the small RNAs entering RISC act as regulators of gene expression is critical for our fundamental understanding.

"The rules are also important because scientists are hoping to use small RNAs one day as therapeutics. By understanding the rules by which small RNAs are processed and sorted, we move closer to the goal of being able to manipulate the RNAi pathway, bend it to the purpose of addressing disease."

Journal Reference:Benjamin Czech, Rui Zhou, Yaniv Erlich, Julius Brennecke, Richard Binari, Christians Villalta, Assaf gordon, Norbert Perrimon and Gregory J. Hannon. Hierarchical Rules for Argonaute Loading in Drosophila. Molecular Cell, 2009; 36 (3): 445 DOI: 10.1016/j.molcel.2009.09.028


New Molecule Identified in DNA Damage Response

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ScienceDaily (Jan. 1, 2010) — In the harsh judgment of natural selection, the ultimate measure of success is reproduction. So it's no surprise that life spends lavish resources on this feat, whether in the courtship behavior of birds and bees or replicating the cells that keep them alive. Now research has identified a new piece in an elaborate system to help guarantee fidelity in the reproduction of cells, preventing potentially lethal mutations in the process.

In experiments to be published in the December 18 issue of the Journal of Biological Chemistry, researchers at The Rockefeller University identified the molecule SMARCAL1 as part of cells' damage control response to malfunctioning DNA replication. In typical cell division, many different molecules have roles in guaranteeing the daughter strands of DNA are as identical as possible to their parent. Some molecules check for errors or 'proofread' the offspring for typos, for instance; others, when alerted to a problem, arrest the replication process and conduct repairs.

Lisa Postow, a postdoctoral fellow in Hironori Funabiki's Laboratory of Chromosome and Cell Biology, used mass spectroscopy to identify SMARCAL1 as involved in this intricate quality control process. Working with Brian T. Chait's Laboratory of Mass Spectrometry and Gaseous Ion Chemistry, Postow found the protein in a proteomics screen for molecules that were drawn to a dangerous DNA repair problem called a double-strand break.

In both human cells and in cells from African clawed frog egg extract, Postow found that at double-strand breaks, SMARCAL1 gathered with another molecule called RPA, which is known to coat broken strands of DNA and protect them while damage is repaired. SMARCAL1 had an added interest, too: A mutation in the gene that produces it is involved in a rare but lethal disease called Schimke immuno-osseous dysplasia, a disorder that causes wide-ranging problems including kidney malfunction, immunodeficiency and growth inhibition.

To Postow's surprise, she found that removing SMARCAL1 had little effect on double-strand break repair. However, it did facilitate a different aspect of the DNA damage response called replication fork stabilization, a process that holds steady the junction between parental and daughter strands -- the replication fork -- when replication is stalled because a problem has been detected. "For a mutation that causes such wide-ranging and severe physiological effects, it is surprising that the protein has such a relatively small effect at the cellular level," Postow says.
Postow's findings were largely corroborated by independent new research into SMARCAL1, which was published this fall in four back-to-back papers in Genes & Development. The work reveals another piece of the complex safeguards the body has in place to protect against dangerous mutations.

"This study also proves that the proteomic approach that Lisa has developed with Dr. Chait can efficiently identify proteins involving the DNA-damage recognition and repair process," says Funabiki. "Many more excitements are ahead of us."

Journal Reference:
Postow et al. Identification of SMARCAL1 as a Component of the DNA Damage Response. Journal of Biological Chemistry, 2009; 284 (51): 35951 DOI: 10.1074/jbc.M109.048330


Molecular Security System That Protects Cells from Potentially Harmful DNA Discovered

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ScienceDaily (Jan. 14, 2010) — Researchers at the University of Minnesota have discovered a molecular security system in human cells that deactivates and degrades foreign DNA. This discovery could open the door to major improvements in genetic engineering and gene therapy technologies.

Led by Reuben Harris, associate professor of biochemistry, molecular biology and biophysics in the College of Biological Sciences, the report's findings will be published online by Nature Structural and Molecular Biology on Jan. 10.

In the study, Harris and colleagues show how APOBEC3A, an enzyme found in human immune cells, disables double-stranded foreign DNA by changing cytosines (one of the four main bases in DNA) to uracils (an atypical DNA base). Persisting DNA uracils result in mutations that disable the DNA. In addition, the authors show that other enzymes step in to degrade the uracil-containing foreign DNA and sweep its remains out of the cell.

"Scientists have known for a long time that some human cells take up DNA better than others, but we haven't had good molecular explanations," Harris says. "This is definitely one of the reasons. Foreign DNA restriction is a fundamental process that could have broad implications for a variety of genetic diseases."

By understanding how the mechanism works, scientists can develop ways to manipulate it to enable more effective methods to swap bad genes for good ones. Harris is also intrigued to learn why the mechanism doesn't affect a cell's own DNA.

The discovery of an analogous foreign DNA restriction mechanism in bacteria launched the field of genetic engineering during the 1970s. Once bacterial DNA restriction enzymes were understood, their power was harnessed to cut and paste segments of DNA for a wide variety of therapeutic and industrial purposes.


Friday, January 1, 2010

Happy New Year !

Happy New Year !