Two biologists at the University of California, San Diego have discovered the first of a new class of cellular motor proteins that “rewind” sections of the double-stranded DNA molecule that become unwound, like the tangled ribbons from a cassette tape, in “bubbles” that prevent critical genes from being expressed.

“When your DNA gets stuck in the unwound position, your cells are in big trouble, and in humans, that ultimately leads to death” said Jim Kadonaga, a professor of biology at UCSD who headed the study.  “What we discovered is the enzyme that fixes this problem.”

The discovery represents the first time scientists have identified a motor protein specifically designed to prevent the accumulation of bubbles of unwound DNA, which occurs when DNA strands become improperly unwound in certain locations along the molecule.

The UCSD researchers’ findings, detailed in the October 31 issue of Science, are also important because they provide biomedical scientists with a greater understanding of the molecular mechanisms that lead to a rare genetic disorder called Schimke immuno-osseous dysplasia.  The discovery will eventually allow medical researchers to design future treatments for this devastating genetic disorder, which causes strokes, congestive heart failure, kidney failure and death in young children.

“We knew this particular protein caused this disease before we started the study,” said Kadonaga.  “That’s why we investigated it.  We just didn’t know what it did.”

What this protein, called HARP for HepA-related protein, did astounded Kadonaga and Timur Yusufzai, a postdoctoral fellow working in his laboratory.  The two molecular biologists initially discovered that this motor protein burns energy in the same way as enzymes called helicases and, like helicases, attached to the dividing sections of DNA.  But while helicases use their energy to separate two annealed nucleic acid strands—such as two strands of DNA, two strands of RNA or the strands of a RNA-DNA hybrid— the scientists found to their surprise that this protein did the opposite; that is, it rewinds sections of defective DNA and thus seals the two strands together again.

As a consequence, the UCSD biologists termed their new enzyme activity an “annealing helicase.”

“We didn’t even consider the idea of annealing helicases before this study started,” said Kadonaga.  “It didn’t occur to us that such enzymes even existed.  In fact, we never knew until now what happened to DNA when it got stuck in the unwound position.”

Now scientists who study the action of helicases on DNA and RNA have an entirely new class of proteins to investigate.

“This will open up a whole new area of study,” said Kadonaga.  “There are very few enzymes known that alter DNA structure.  And we’ve discovered an entirely new one.  This was not expected to happen in the year 2008.  We should have found them all by now.”

“I believe it’s going to go beyond DNA.  Just as there are DNA-DNA helicases, there are RNA-DNA helicases and RNA-RNA helicases.  So it doesn’t take a lot of imagination to foresee that there are probably going to be RNA-DNA annealing helicases and RNA-RNA annealing helicases.  The field potentially can be fairly large.  And as more and more people discover additional annealing helicases, this field will expand.”

Kadonaga and Yusufzai are already searching for more annealing helicases, but they also plan to continue their studies of HARP.

“First, what we want to do is find more of these proteins, so we’re looking for more right now,” said Kadonaga.  “We also want to see what other specific processes are affected by this particular protein, HARP, in the cell.”

Scientists at the Gladstone Institute of Neurological Disease (GIND) have identified a new strategy to destroy amyloid-beta (AB) proteins, which are widely believed to cause Alzheimer’s disease (AD). Li Gan, PhD, and her coworkers discovered that the activity of a potent AB-degrading enzyme can be unleashed in mouse models of the disease by reducing its natural inhibitor cystatin C (CysC).

All of us produce AB proteins in the brain. However, in most people, the proteins never build up to dangerous levels because they are cleared away by enzymes that destroy them. Previously Dr. Gan’s laboratory had shown that cathepsin B (CatB) is such an AB-degrading enzyme. In the latest issue of the journal Neuron, the researchers report a highly effective approach to promote CatB-mediated clearance of AB .

“Many groups have developed drugs to block the production of AB, but the efficacy and safety of this approach remains to be demonstrated in clinical trials,” said GIND Director Lennart Mucke, MD “By identifying an effective strategy to enhance the removal of AB, this research provides a very promising alternative or complementary therapeutic avenue.”

High levels of AB in the brain may result from overproduction of AB or from an inability to eliminate it from the brain. While most work has focused on the first option, the latter has been problematic. For example, efforts to develop a vaccine that would trigger the immune system to eliminate AB have shown limited success and resulted in adverse side effects.

“Our strategy to harness the activity of a powerful AB-degrading enzyme takes advantage of the brain’s own defense system to remove the toxic AB build-up,” said Dr. Gan. “In principle, one could boost the activity of CatB by expressing more of it in the brain or by reducing the activity of CysC, its natural inhibitor. We focused on the latter strategy because it has greater long-term therapeutic potential.”

Many enzymes that degrade proteins are kept in check by regulators called protease inhibitors. The activity of CatB is regulated by the protease inhibitor CysC. By reducing CysC activity, the scientists were able to unleash the AB-degrading power of CatB, effectively preventing the build-up of AB in mouse models of AD.

To examine the impact of this manipulation on brain function, Dr. Gan’s team measured brain cell activities that relate closely to learning and memory. Increasing CatB activity by lowering CysC levels prevented AB-induced deficits in those cellular activities. The investigators also tested the modified AD mice for learning and memory in a water maze. Higher levels of CatB activity improved the ability of AD to learn the maze and to retain the new information. Increasing CatB activity also prevented the premature mortality that is typically seen in these Alzheimer models.

“Our results suggest that CysC reduction has major therapeutic potential,” Dr. Gan said. “The next step will be to develop pharmacological approaches to inhibit CysC in the human brain.”

Scientists at Dana-Farber Cancer Institute have identified a previously undetected trigger point on a naturally occurring “death protein” that helps the body get rid of unwanted or diseased cells. They say it may be possible to exploit the newly found trigger as a target for designer drugs that would treat cancer by forcing malignant cells to commit suicide.Loren Walensky, MD, PhD, pediatric oncologist and chemical biologist at Dana-Farber and Children’s Hospital Boston, and colleagues report in the Oct. 23 issue of the journal Nature that they directly activated this trigger on the “executioner” protein BAX, killing laboratory cells by setting in motion their self-destruct mechanism.

The researchers fashioned a peptide (a protein subunit) that precisely matched the shape of the newly found trigger site on the killer protein, which lies dormant in the cell’s interior until activated by cellular stress. When the peptide docked into the binding site, BAX was spurred into assassin mode. The activated BAX proteins flocked to the cell’s power plants, the mitochondria, where they poked holes in the mitochondria’s membranes, killing the cells. This process is called apoptosis, or programmed cell death.

“We identified a switch that turns BAX on, and we believe this discovery can be used to develop drugs that turn on or turn off cell death in human disease by targeting BAX,” said Walensky, who is also an assistant professor of pediatrics at Harvard Medical School.

BAX is one of about two dozen proteins known collectively as the BCL-2 family. The proteins interact in various combinations leading to either the survival of a cell or its programmed self-destruction. Cancer cells have an imbalance of BCL-2 family signals that drives them to survive instead of dying on command.

The late Stanley Korsmeyer, MD, an apoptosis research pioneer and Walensky’s Dana-Farber mentor, had suggested that killer proteins like BAX could be activated directly by “death domains,” termed BH3, contained within a subset of BCL-2 family proteins. He hypothesized that this activating interaction was a fleeting “hit-and-run” event, making it especially challenging for scientists to study the phenomenon.

As suspected, the proposed BAX-activating interactions could not be captured by traditional methods. “When you tried to measure binding of the BH3 subunits to BAX, you couldn’t detect the interaction,” explained Walensky. He recognized, however, that the BH3 peptides being used in the laboratory didn’t retain the coiled shape of the natural BH3 domains that participate in BCL-2 family protein interactions. Walensky and his colleagues pioneered the design of “stapled” BH3 peptides, which contain a chemical crosslink that locks the peptides into their natural coiled shape. With biologically active shape restored, the stapled BH3 peptides bound directly to BAX and triggered its killer activity.

Defining how the activating peptides docked on BAX remained a formidable catch-22. In order to solve the structure of an interaction complex, it needed to be stable enough for analysis. In this case, the BH3 binding event itself triggers BAX to change its shape and self-associate to perform its killer function, rendering the activating interaction unstable by definition.

What if, Walensky proposed, you could set up the interaction of BH3 and BAX under laboratory conditions that caused it to be more stable or proceed in slow motion? The plan was to adjust the potency of the stapled BH3 peptide so that, according to Walensky, “it was good enough to bind BAX, yet activate it just a bit more slowly so that we could actually study the interaction.” The researchers would then look for any detectable shift in the three-dimensional structure of the BAX protein to help point them to the docking site.

The researchers used nuclear magnetic resonance (NMR) spectroscopy to monitor the arrangement of atoms in the protein. First authors of the Nature paper Evripidis Gavathiotis, PhD, of Walensky’s laboratory and Motoshi Suzuki, PhD, of Nico Tjandra, PhD,’s laboratory at the National Institutes of Health, succeeded in generating pure BAX protein that could be put into solution with the stapled BH3 peptide — the latter in increasing concentrations until it initiated a BH3-BAX interaction. Gavathiotis and Suzuki used the NMR technique to spot a group of BAX amino acids, the building blocks of proteins, which were affected by the addition of the stapled BH3 peptide.

“The discrete subset of amino acids that shifted upon exposure to the stapled BH3 peptide mapped to a completely unanticipated location on BAX,” said Walensky. The long-elusive binding site on BAX that initiates its killer activity was revealed. “Because BAX lies at the crossroads of the cell’s decision to live or die, drugs that directly activate BAX could kill diseased cells like in cancer and BAX-blocking drugs could potentially prevent unwanted cell death, such as in heart attack, stroke, and neurodegeneration,” said Walensky.

In a paper published today in Nature, the team led by Professor Nigel Robinson have revealed a mechanism that ensures the right metal goes to the right protein. Proteins are essential and involved in just about every process in living cells.

Life, microbe, plant or human, is a painstaking assembly of trillions of atoms. The atoms include metals such as copper and manganese which act as catalysts in proteins. The proteins wrap around the metal atoms.

The research team has shown that to ensure a copper and a manganese protein wrap around the correct metal atoms they do this in different parts of the cell, in zones which contain different metals. Therefore, which protein attaches to which metal is determined by where the folding action takes place in the cell.

Previously, a common view was that the right metals were simply those which were most attracted to the protein, but in this work that is not the case.

Professor Nigel Robinson at Newcastle University who led the research says: “This has taken us one step closer to understanding why metals and proteins assemble in the ways they do.”

“One motive behind the work is pure curiosity, but as so many proteins need metals this type of work has many potential uses - for example, in synthetic biology which is striving to produce green power from bacteria by using energy from sunlight to produce hydrogen gas, a process which needs nickel and iron.

“It may also help in diseases such as Alzheimers where there are unexplained links to proteins binding metals such as copper. There’s also application in controlling infections by Staphylococcus aureus; a bacterium which our bodies defences succeed - or sometimes fail - in killing by removing manganese and zinc from abscesses.”

The researchers have shown that the way the metals attach is identical for a protein that binds manganese to one that binds copper. In both cases the metals bind inside protein barrels with the same type of metal-attractions.

Carrying out the work in a blue-green algae, a cyanobacterium, the team has been able to show that a protein requiring copper transports to the periplasm, the outer area of the cell, where it then folds around the available metal, which is copper.

Conversely, manganese but not copper atoms are found in the cytosol, in the middle of the cell. The team has demonstrated that a protein requiring manganese folds in the cytosol. The manganese protein is then transported to the periplasm having first trapped its manganese.

The cyanobacterium organism was chosen because it has a high demand for these two metals which are required for proteins involved in photosynthesis. These metals were chosen because they lie towards opposite ends of a chemical series called the Irving-Williams series, such that selecting these metals for proteins should be especially demanding.

In the work funded by the BBSRC, the Newcastle University team first developed a new approach to discover metal-binding proteins. This is now being swiftly applied to lots of other types of living cells and other essential metals (zinc, nickel, cobalt, iron). Unexpectedly, x-ray crystal structures showed that the identified proteins, MncA for manganese and CucA for copper, were both cupins (Latin for barrels) with identical sets of atoms for binding to the metals. Consistent with the chemical series, a ten-thousand times excess of manganese over copper was needed to fill the MncA barrel with manganese when folding is done in the laboratory.

Once folded, the manganese site is buried, the metal trapped inside the protein, and so the manganese protein can subsequently co-exist with the copper protein because its’ metal becomes impervious to replacement by metals further up the Irving-Williams series.

The work exemplifies a cell overcoming the metal binding preferences of proteins.

The new discipline of synthetic biology aims to engineer cells to carry out useful tasks, for example to generate valuable compounds. Because metals are the catalysts for so much of biology, knowing how to engineer a supply of the right metals to the right proteins will be important to the success of these ventures.

The cause of diseases such as BSE in cattle and Creutzfeld–Jakob disease in humans is a prion protein.  This protein attaches to cell membranes by way of an anchor made of sugar and lipid components (a glycosylphosphatidylinositol, GPI) anchor.  The anchoring of the prions seems to have a strong influence on the transformation of the normal form of the protein into its pathogenic form, which causes scrapie and mad cow disease.  A team headed by Christian F. W. Becker at the TU Munich and Peter H. Seeberger at the ETH Zurich has now “recreated” the first GPI-anchored prion in the laboratory.  As they report in the journal Angewandte Chemie, they have been able to develop a new general method for the synthesis of anchored proteins.

The isolation of a complete prion protein that includes the anchor has not yet been achieved, nor has it been possible to produce a synthetic GPI-anchored protein.  The function of the GPI anchor has thus remained in the dark.  A new synthetic technique has now provided an important breakthrough for the German and Swiss team of researchers.

The sugar component of natural prion GPI anchors consists of five sugar building blocks, to which further sugars are attached through branches.  Details of the lipid component have not been determined before.  As a synthetic target, the researchers thus chose a construct made of the five sugars and one C18-lipid chain and worked out the corresponding synthetic route.  First, the anchor was furnished with the sulfur-containing amino acid cysteine.  The prion protein was produced with the use of bacteria and was given an additional thioester (a sulfur-containing group).  The centerpiece of the new concept is the linkage of the protein and anchor by means of a native chemical ligation, in which the cysteine group reacts with the thioester.  This allowed the prion protein to firmly attach to the vesicle membranes by way of the artificial anchor.

This new concept will allow production of sufficient quantities of proteins modified with GPI anchors for in-depth studies.  Experiments with the artificial GPI prion protein should help to clarify the influence of membrane association on conversion of the protein into the pathogenic scrapie form.  This should finally make it possible to track down the infectious form of the prion.

A normal developmental protein that sometimes goes awry has been implicated in breast cancer.  This discovery indicates the mechanism by which inappropriate expression of the Notch pathway may contribute to breast cancer.

The breast cancer team at WEHI, led by Drs Jane Visvader and Geoff Lindeman from the Victorian Breast Cancer Research Consortium, have identified important roles for Notch genes in regulating breast development and function.

This discovery has important implications for breast cancer, since elevated levels of Notch have been linked to breast cancer.  The advance builds on the group’s 2006 discovery of the breast stem cell in mice.

Research carried out by Drs Toula Bouras and Bhupinder Pal has uncovered dual functions for Notch in breast tissue.

First, Notch helps restrict breast stem cell number, so that when Notch is ’switched off’, there is a resultant expansion in breast stem cells.

Second, Notch is important for ensuring that stem cells produce the sleeve of cells that normally line breast ducts.  These ‘luminal’ cells may be the cells that give rise to common types of breast cancer.

Thus, Notch helps to orchestrate the formation of breast tissue: it plays an important role in controlling stem cell number and instructs stem cells to produce luminal cells.

Significantly, Dr Bouras and colleagues found that errant activation of Notch resulted in uncontrolled growth of luminal precursors, leading to the formation of breast tumours.

The work has spotlighted the potential importance of deregulated Notch in ductal precursor cells as a forerunner to breast cancer.

The researchers say that it is too early to speculate on whether the design of anti-Notch therapies could help patients facing breast cancer.

Individuals with a number of clinical conditions, including pneumonia, and those treated by mechanical ventilation for a prolonged period of time are at risk of acute lung injury, a life-threatening disorder for which there is no treatment. It is hoped that understanding the natural processes by which acute lung injury spontaneously resolves in some individuals might provide new therapeutic targets. Thus, Holger Eltzschig and colleagues, at the University of Colorado Health Sciences Center, Denver, suggest that their observation in mice with ventilator-induced lung injury (VILI) implicate the protein A2BAR as a potential therapeutic target for acute lung injury.

In the study, mice lacking A2BAR were found to have reduced survival time and more severe VILI, when compared with normal mice. Consistent with this, normal mice treated with an A2BAR antagonist exhibited more severe lung damage than untreated mice, whereas an A2BAR agonist attenuated the severity of VILI. Further analysis revealed that one way in which the A2BAR agonists helped was by enhancing the clearance of fluid in the lungs (i.e., they helped dry out the lungs). These data indicate that agonists of A2BAR are likely to be part of the natural mechanism by which acute lung injury spontaneously resolves and might make good therapeutics.

Imagine having to copy an entire book by hand without missing a comma. Our cells face a similar task every time they divide. They must duplicate both their DNA and a subtle pattern of punctuation-like modifications on the DNA known as methylation.

Scientists at Emory University School of Medicine have caught in action one of the tools mammalian cells use to maintain their pattern of methylation. Visualized by X-ray crystallography, the SRA domain of the protein UHRF1 appears to act like a bookmark while enzymes are copying a molecule of DNA.

The team’s description of the protein’s structure while bound to DNA is published this week in Nature.

Scientists refer to methylation, the addition of a methyl group to DNA, as an “epigenetic” modification because it adds a layer of information on top of the genetic sequence of the DNA itself. It marks genes for silencing, which means they do not manufacture proteins.

“The processes that copy the methylation pattern have to be faithful,” says senior author Xiaodong Cheng, PhD, professor of biochemistry and a Georgia Research Alliance eminent scholar. “Otherwise, losing DNA methylation marks can have serious consequences, causing genes to become active at the wrong places and times.”

“Gene silencing via DNA methylation is critical for normal development and for curbing the runaway cell division that characterizes cancer,” said Peter Preusch, PhD, who oversees biophysics grants at the National Institute of General Medical Sciences of the National Institutes of Health. “Alterations in methylation patterns are also important for generating embryonic stem-like cells from differentiated cells.”

In mammalian cells, methylation usually appears on double stranded DNA where the nucleotide Cytosine (C) is followed by Guanine (G). The complementary sequence on the opposite strand is also C then G, and the methylation appears on both Cs.

When a cell is copying its DNA, a set of enzymes duplicates the DNA sequence from the parental strand to the new “daughter” strand but not the methylation. Each new daughter strand of the DNA molecule is left with the previously methylated Cs unmethylated. UHRF1 recognizes this “hemi-methylated” DNA and calls in a methyltransferase enzyme to add a second methyl group onto the daughter strand.

“UHRF1 has the important task of making sure the methyltransferase enzyme does its job in the right place and right time,” Cheng says.

Mouse cells that have deleted the UHRF1 gene are more sensitive to DNA-damaging agents such as radiation, and mouse embryos without the gene cannot complete development. Other studies have found that cancer cells produce more UHRF1 than non-cancerous cells.

What was an unexpected finding was how the SRA domain of UHRF1 recognizes the hemi-methylated DNA, Cheng says. It flips the methylated nucleotide out of the DNA helix, which only had been seen previously in enzymes that physically modify the DNA.

Cheng says the flipping mechanism could prevent the protein from sliding away once it has found a hemi-methylated site.

“It suggests that it serves as a placeholder, where it recruits other enzymes for faithful DNA methylation or repair enzymes if the DNA has been damaged,” he says.
Hashimoto H., et al Nature, advance online publication, Sept. 3, 2008
The SRA domain of UHRF1 flips 5-methylcytosine out of the DNA helix.

Scientists from the Structural Genomics Consortium (SGC) have determined the 3D structure of a key protein component involved in enabling “epigenetic code” to be copied accurately from cell to cell.Epigenetic code is a series of chemical switches that is added onto our DNA in order to ensure that the cells in our body can form different types of tissue, for example liver and skin, despite having identical DNA genetic code.

When DNA is copied from cell to cell, it is essential that the epigenetic code is also copied accurately. If not, a liver cell may divide into another type of cell, such as a nerve or eye cell. A breakdown in this system might also mean that a gene for cell growth is accidentally switched on, for example, leading to unregulated cell growth and the development of tumours.

Research published in 2007 showed the importance of the nuclear protein UHRF1 in ensuring that the epigenetic code is accurately copied. Epigenetic switches are created by the addition of a chemical group (methyl) to DNA in a process known as methylation, facilitated by the enzyme DNMT1. The researchers believe that when this code is copied, UHRF1 ensures the accuracy of the process, like a proof-reader checks a typeset article before printing.

The key element of UHRF1 involved in this “proofreading” process is known as the Set and Ring Associated (SRA) domain, but the exact mechanisms by which the SRA domain accomplishes this task were unclear. Today, in three different articles, the journal Nature publishes the structure of the key element of UHRF1 that facilitates this process.

“Given the increasing focus on epigenetics as a mechanism behind cancer, elucidating the structure of UHRF1 may provide crucial insights into what goes wrong,” says Professor Sirano Dhe-Paganon from the Structural Genomics Consortium laboratories at the University of Toronto, Canada.

The structural papers not only represent an advance for the epigenetics field, but also an advance for how the science was done. The concurrent publication of the three papers highlights the competitive nature of this field, but in fact these papers were made possible because the SGC, in keeping with its policy of making its data freely and immediately available, made the underlying information available in the Protein Data Bank late in 2007. The availability of this information allowed the other groups to make more rapid progress in their own work.

“By releasing the structural information into the public databases as soon as it was available, we have ensured that other research groups could make immediate and maximum benefit from the shared knowledge,” says Professor Dhe-Paganon.

Professor Masahiro Shirakawa from Kyoto University, Japan, openly acknowledges that the SGC data was crucial to his team’s paper, which also appears in today’s edition of Nature.

“We would like to express our gratitude to the researchers at the SGC for making their available on net,” says Professor Shirakawa. “Structural biology is a complex, but very important field, with the potential to drive forward important research in many areas. The information provided by the SGC significantly speeded up our own work.”

The SGC’s “open source” policy contrasts with the accepted practice in the structural biology field, which is to make the underlying data available only after the work appears in print. However, Professor Al Edwards, Director of the SGC, believes strongly that data such as the 3D structure of proteins should be made freely available as soon as they are discovered.

“From the outset, it’s been important to us to release our structural data immediately,” says Professor Edwards. “This is contrary to the way many scientists work, but we believe it is crucial for facilitating scientific and medical progress, and our policy has not inhibited our ability to publish our work in the top journals. All the protein structures studied by the SGC have medical relevance and making them freely available ensures that scientists are able to use them to make progr

Although every cell of our bodies contains the same genetic instructions, specific genes typically act only in specific cells at particular times. Other genes are “silenced” in a variety of ways. One mode of gene silencing depends upon the way DNA, the genetic material, is packed in the nucleus of cells.

When packed very tightly around complexes of proteins called histones, the DNA double helix is rendered physically inaccessible to molecules that mediate gene expression. Now, a research team that includes Michael Q. Zhang, Ph.D., a professor at Cold Spring Harbor Laboratory (CSHL), has published a comprehensive analysis of modification patterns in histones.

Using a new technology called ChIP-Seq, the team identified 39 histone modifications, including a “core set” of 17 modifications that tended to occur together and were associated with genes observed to be active.

Modification Patterns With Different “Personalities”

Scientists have long known that chemical changes at particular locations in histone complexes influence how tightly the DNA is wrapped around the histones. “But it is important to know whether particular modifications occur together in characteristic patterns, or if these patterns can predict gene activities,” Dr. Zhang explained.

At the heart of the team’s efforts to determine this, Keji Zhao, Ph.D., of the National Heart, Blood, and Lung Institute of the National Institutes of Health, and his colleagues developed a method to map modifications in human white blood cells known as CD4+ T cells. First they used an enzyme to cut the DNA into short segments, which remained attached to histone “spools.” For each of 39 distinct histone modifications, the scientists used an antibody to extract matching histone-DNA combinations. Finally, they used the ChIP-Seq DNA-sequencing technology to determine which parts of the genome were bound to each type of modified histone.

The team’s most recent research, published in the July 2008 issue of Nature Genetics, maps the DNA locations that bind to histones containing molecular configurations called acetyl groups at 18 different positions in the “tails” of the histone proteins. The scientists combined this information with earlier maps for 19 different changes called methylation modifications, and for replacement of one of the histone proteins with a well-known variant.

The various modifications showed distinctive “personalities,” each preferentially associating with particular regulatory regions of genes.

Looking for Patterns

Mapping many modifications enabled the researchers to explore whether different types tend to appear together in the same type of DNA regulatory regions. They found that some recurring combinations did occur frequently at “promoter” and “enhancer” regions in DNA, which are known to increase the activity of nearby genes. In particular, the team identified one combination of 17 modifications that was present in more than a quarter of the more than 12,000 promoter regions they examined.

On average, the genes corresponding to this “backbone” set were expressed more actively. That is to say, they were activated, setting the cellular machinery in motion to produce specific proteins, the workhorses of most life processes.

The rich relationships detected by the researchers among the various histone modifications suggests that specific combinations might carry specific meanings. Previous researchers have proposed a “histone code” hypothesis, which posits that a particular combination of modifications may be recognized by a particular protein module. Some scientists believe such histone code may determine the activity of a given gene.

But, cautions Dr. Zhang, while there are patterns, like the backbone, that are highly correlated, “none of them has exact predictive value.” He maintains “there must be something else” that also affects gene activity.

Since genes with higher or lower expression levels may have the same patterns of modification, and not all active genes share a common pattern, the reality is likely more complex than a universal histone code that predicts exact gene expression level. Nonetheless, the new research provides a rich data source for understanding how specific combinations of histone modifications modulate the effects of many genes, in turn helping to modify activity within and among cells. “Critical future research should focus on finding proteins that target histone modifications to genetic regions with particular sequences,” Dr. Zhang emphasized.

T-cell acute lymphoblastic leukemia (T-ALL)

The white blood cells in our body combat foreign intruders, such as viruses and bacteria.  However, in leukemia, the formation of white blood cells is disturbed: the cells that should develop into white blood cells multiply out of control without fully maturing.  This process disrupts the production of normal blood cells, making patients more susceptible to infections.  T-ALL, a particular form of leukemia, is the most prevalent cancer in children under 14 years of age and occurs predominantly between the ages of two and three.  At the moment, with an optimal treatment using chemotherapy, over half of the children are cured.  But scientists hope to be able to develop targeted therapies that are less toxic than chemotherapy, based on knowledge of the biological processes behind T-ALL.

Importance of the location

Oncogenes are often at the root of cancer.  So, scientists around the world are concentrating on identifying oncogenes and their related proteins.  Recent research by Kim De Keersmaecker and colleagues in Jan Cools’ research group (VIB-K.U.Leuven) indicates that the location in the cell where these proteins are found plays an important role in the entire carcinogenic mechanism.  In collaboration with Maarten Fornerod (Nederlands Kanker Instituut, Amsterdam) and Gary Gilliland (Harvard Medical School, Boston), the VIB researchers have demonstrated that NUP214-ABL1, a fusion of two proteins, is carcinogenic only when it is in a protein complex near the nucleus of the cell.  Located at another place in the cell, NUP214-ABL1 does not lead to cancer.  This finding sheds new light on the study of carcinogenic processes.

A new therapeutic approach?

Many forms of cancer are caused by genetic defects in which a certain kinase becomes too active and this is the case with NUP214-ABL1.  The most obvious solution is to make the carcinogenic kinase inactive, and so kinase inhibitors are usually used to combat these kinds of cancers.  However, the carcinogenic kinase often becomes resistant to these inhibitors which is certainly true for T-ALL.  So, scientists are actively seeking alternative approaches.

De Keersmaecker’s recent research results now offer a possibility.  Indeed, the scientists have shown in cells that NUP214-ABL1 is no longer carcinogenic when it cannot bind with the protein complex in the vicinity of the cell nucleus.  On the basis of these results, the researchers want to further investigate the therapeutic possibilities of compounds that render binding between the complex and NUP214-ABL1 impossible.  This study also indicates that the location of proteins can play an important role in other forms of cancer/leukemia as well.

Against currently held dogma, scientists at the Universities of Cambridge and Bristol have revealed that the interactions within membrane complexes can be maintained intact in the vacuum of a mass spectrometer. Their research is published in this week’s edition of Science Express.

The researchers were surprised to discover that membrane complexes could remain associated as it has always been assumed that they would not survive once transferred to the alien conditions inside the mass spectrometer.

“Even if interactions between proteins within the membrane could be maintained we would not have expected them to remain associated with proteins in the cell’s interior,” says Carol Robinson, Principal Investigator and Royal Society Research Professor at the University of Cambridge’s Department of Chemistry.

Cellular membranes surround cells and provide the ultimate in cellular security; nothing can get into a cell without the say so of membrane proteins – the worker molecules that reside in the membrane wall and provide tightly regulated entry points. This natural home of membrane proteins excludes water, yet methods available to study proteins at high resolution revolve round aqueous environments. The ability to “fly” intact membrane proteins in a mass spectrometer paves the way for weighing the proteins and identifying the molecular partners they work with in nature.

The new research, funded by the Biotechnology and Biological Sciences Research Council, will enable scientists to investigate membrane complexes with from a variety of sources and with a range of small molecules. Since about 60% of all drug targets are membrane proteins this is a significant discovery.

Ever since Professor Robinson first flew soluble protein complexes in a mass spectrometer in 1996, she has wanted to do the same with membrane complexes. Collaborating with a membrane biochemistry group in Bristol, led by Professor Paula Booth, she began to think of ways of studying these most challenging assemblies.

Dr Nelson Barrera a post-doctoral researcher in Chile, though experienced in membrane biochemistry, was a new recruit to mass spectrometry. He was largely unaware of the difficulties that had previously been encountered and approached the problem in a new way. Rather than trying to remove the detergent (used to keep the protein intact in solution once outside the natural membrane) he maintained the detergent in unusually high amounts. He then deliberately destroyed this protective detergent layer once in the gas phase. This allowed him to liberate the intact assembly. He was also able to remove units from the modular assembly in the gas phase, just as in solution.

Professor Robinson adds: “I am very excited by this finding given the importance of membrane complexes in guarding the entrance and exit to cells. The type of proteins we have been studying, for example, are involved in drug resistance in cancer cells and antibiotic resistance of bacteria.

“I look forward to exploiting this discovery to the full; not only in characterising the many membrane complexes for which controversy exists but also in discovering new assemblies and in investigating the potential of this approach in drug discovery.”

Professor Paula Booth, at the University of Bristol added: “This is a major advance that helps us understand how nature constructs cellular life. The membrane wall of cells is a precision-made, complex and highly regulated structure. We are now much better equipped to understand this incredible, natural self-assembly feat.”

Uncontrolled blood vessel growth is a key feature of many pathological conditions, including the degenerative diabetic eye disease known as diabetic retinopathy.  Understanding the factors involved in the process is vital to developing treatments for the disease.  In a new study, a team of researchers at Keio University, Japan, has revealed a role for the protein LIF in blood vessel growth in mice.

Specifically, mice lacking LIF were observed to have increased blood vessel growth in many regions of the body, but as this study was focused on the eye, the authors homed in on the increased blood vessel growth in the retina of the eye.  Further analysis showed that mice lacking LIF developed more aberrant blood vessels in a model of retinopathy.  Mechanistically, LIF was found to inhibit the proliferation of brain cells known as astrocytes as well as inhibit their production of a factor known to promote blood vessel growth, VEGF.  It therefore seems that LIF is an important part of the communication between tissues and developing blood vessels, meaning that LIF and the signaling pathway it triggers might serve as a target for new treatment approaches for preventing diabetic retinopathy and other diseases that are associated with uncontrolled blood vessel growth, such as cancer.

Montana State University scientists in the Department of Chemistry and Biochemistry published new research this week that could one day affect the lives of millions around the world who suffer from blood iron disorders.
Martin Lawrence (left) and Anoop Sendamarai (MSU photo by Kelly Gorham)
The paper, which will appear in the Proceedings of the National Academy of Sciences, details the work of Associate Professor Martin Lawrence and doctoral candidate Anoop Sendamarai. The pair have spent the past two years studying Steap3, a protein involved in regulating the body’s absorption of iron.

The results of their studies - the first three-dimensional maps of the atoms that make up Steap3 - could allow pharmaceutical companies to someday design drugs to regulate iron levels in the blood.

“Iron is essential,” Lawrence said. “You can’t live without it, but it’s a double-edged sword. Too much of a good thing can kill you.”

Iron serves several important functions in the bloodstream. It carries oxygen, transports electrons within cells and plays an important role in enzyme systems.

Iron irregularities are some of the most common blood disorders in the world. According to the World Health Organization, iron deficiency, which can lead to anemia, affects more than a billion people around the world and can cause developmental and immune system problems.

Conversely, having too much iron, a condition called hemochromatosis, can also hurt the body by releasing destructive free radicals, Lawrence said. Hemochromatosis affects about one in every 300 people and is most common in people of northern European ancestry. Left untreated, it can lead to early death, often by age 50.

“We’re struck by how many people have too much or too little iron,” Lawrence said.

To understand Steap3’s role in transporting and maintaining balanced levels of iron, Lawrence and Sendamarai first had find and purify samples of the protein and then turn those samples into crystals.

Lawrence said the result of the crystallization process, if done correctly, is analogous to the rigid structure of a brick wall. If done incorrectly, it more closely resembles a pile of bricks.

“It’s kind of a black art really more than a science,” Lawrence said. “You can’t always predict the kind of witch’s brew that needs to be around to get it to crystallize.”

He said only a handful of labs in the country are crystallizing iron transport proteins like Steap3, a fact that places MSU on the same shelf as places like Harvard Medical School.

Once crystallized, the samples are shot with a powerful X-ray beam. Electrons in the sample diffract the X-rays, creating patterns on a digital sensor. The technique, called X-ray crystallography, has been used since the 1950s to de-termine the structure of different substances.

In their basement lab in the campus’s New Chemistry Building, Lawrence and Sendamarai then examined the diffraction patterns created by Steap3.

“It’s kind of like a contour map,” Sendamarai said. “Whenever we see the peaks, we know there are atoms.”

Working backward, they can mathematically determine the position of atoms in the protein and display them in three dimensions.

The computer-drawn result, a three-dimensional image that resembles tangled ribbons and strings, is an picture of what the atoms of Steap3 look like.

Sendamarai said having that picture, which depicts all the nooks and crannies on the protein’s surface, could allow drug companies to design drugs to fit those spots like puzzle pieces.

If a future drug fits those nooks just right, it could help treat hemochromatosis. From there, Sendamarai said it would be conceivable to work backward and possibly treat iron deficiencies or anemia.

Lawrence said that Steap3 is only one in a family of proteins that affect iron transport. This summer, in addition to continuing to study Steap3, Lawrence and Sendamarai hope to learn whether the lab will receive a grant from the National Institutes of Health to work on other iron transport proteins.

“It’s a critical step towards toward learning to modulate iron levels in patients with too much or too little iron,” Sendamarai said. “But, there are a lot of question marks left in iron transport. It’s a big field.”

Biologists from Austria and Singapore developed a technique that adds a new twist on the relationship between biology and art. In an article recently published online in The FASEB Journal (http://www.fasebj.org) and scheduled for the August 2008 print issue, these researchers describe how they were able to coat—or paint—viruses with proteins. This breakthrough should give a much-needed boost to the efficiency of some forms of gene therapy, help track and treat viral disease and evolution, improve the efficiency of vaccines, and ultimately allow health care professionals track the movement of viral infections within the body. Specifically, the new method should make it easier to track and treat infectious diseases such as HIV/AIDS, influenza, hepatitis C, and dengue fever. And because viruses can also be used to introduce biotechnology drugs and replacement genes, and act as vaccines, this research should lead to new treatments for cancer, cardiovascular, metabolic and inherited disorders.

“This technology should provide a new tool for the treatment of many diseases,” said Brian Salmons, one of scientists who co-authored the study. “Even if you are working with a virus that is unknown or poorly characterized, it is still possible to modify or paint it. This is very interesting for emerging diseases.”

In the article, Salmons and colleagues explain how they mixed purified proteins (glycosylphophatidylinositol anchor proteins) with lipid membranes to make it possible to bind these proteins to the outer “skin” (the lipid envelope) of viruses. Even with the new paint job, the viruses remained infectious. While the experiment only involved one type of protein and two types of viral vectors, Salmons says the technique could be expanded and used to apply “paint” made up of other proteins, dyes, and a variety of unique markers.

“Biology and art converge daily: people paint their nails, color their hair, and tattoo their skin,” said Gerald Weissmann, M.D., Editor-in-Chief of The FASEB Journal. “Now this convergence has entered a new dimension as painted viruses permit scientists to track, cure and prevent disease.”