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Science Feature Articles</p>

Living with the Quick Fix

English
 
(l-r) Omer Ziv and Prof. Zvi Livneh
 
Our DNA is thought to get damaged at the staggering rate of 50,000 times a day, mostly from byproducts of our metabolism and such external agents as sunlight and tobacco smoke. If left unmended, this damage can eventually lead to cancer, immunodeficiency, premature aging and neurodegeneration. Fortunately, organisms have evolved a whole array of DNA repair mechanisms – different kinds for every type of damage.
 
Among them are various “quick fix” mechanisms; these have been the focus of research in Prof. Zvi Livneh’s lab in the Weizmann Institute’s Biological Chemistry Department. In a new study published in Nature Communications, Livneh and his colleagues have now revealed how one of these mechanisms, called translesion DNA synthesis (TLS), is regulated in mammalian cells. These findings may have potential applications for cancer prevention and therapy, especially for certain cancers that have mutations in the TLS genes.
 

Bridge over troubled DNA

 
“Fixes” may come at a price. The methodical routine that accurately restores the damaged DNA to its original code is time- and energy-consuming. So the cell may turn to “fast,” but error-prone methods, which, in the case of TLS, merely tolerate the damage, “bridging” the damaged section and allowing DNA replication to bypass the hurdle and continue on its way. The cost of this fast mechanism lies in the possible introduction of a wrong DNA code – a gamble that carries risks of the onset of disease and even death. In previous research, Livneh’s lab discovered that at the heart of the TLS mechanism is a group of DNA polymerases – enzymes that assemble the DNA strand. With no fewer than 10 of these polymerases, all “sloppy” workers, one might expect a host of problems, yet, surprisingly, the error rate is relatively low. The research of Livneh and others offered an explanation: Each of these polymerases is finely tuned to deal with a specific type of DNA damage, thus lowering the chance of a mistake. The question now was: What keeps all these polymerases in check – ensuring their action at the right place and time?
 

Unique genes revealed

 
To identify the genes that regulate TLS, Livneh and PhD student Omer Ziv, in collaboration with Prof. Eytan Domany and former PhD student Amit Zeisel of the Physics of Complex Systems Department, developed a novel two-stage screening approach: First, they took human cells with a specific defect – in the repair of UV-induced DNA damage. One by one, they “turned off” 1000 different genes, winnowing out those cells that had either reduced or increased survival rates after being exposed to damaging UV light. These cells then went through a second round of more refined screening developed by the team; this time, to determine whether the cells’ survival was specifically dependent on TLS. Of the 1000 genes screened, the scientists discovered 17 new ones involved in TLS, six of which appear to be unique to mammals.
 
NPM1 interacts with DNA polymerase eta and regulates polymerase eta-promoted TLS. Assay of polymerase eta and NPM1 in unirradiated (l) and ultraviolet irradiated cells ((c) after one hour, (r) after 18 hours). Blue: DNA in the nucleus; green: polymerase eta–NPM1 interaction
 
 
 

New regulatory system, old cancer mystery

 
The scientists chose to further investigate one of the novel TLS genes they identified; this gene encodes a multifunctional protein, nucleophosmin (NPM1), which is involved in the biogenesis of ribosomes and cell proliferation, among its many roles. The team discovered that NPM1 regulates TLS by physically interacting with one of the “sloppy” polymerases – called DNA polymerase eta – in the nucleus of the cell. NPM1 binds to polymerase eta as long as there is no damage, thus “locking” it away to protect it against degradation while maintaining a functional pool for fast action when needed. In the meantime, the polymerase is prevented from acting on any intact DNA and introducing needless errors. When damage does occur, i.e., following exposure to UV radiation, NPM1 releases its hold on the polymerase. The team traced the effect experimentally: NPM1 deficiency resulted in decreased DNA polymerase eta levels, leading to defective TLS.
 
Domany: “NPM1 is essentially the ‘guardian’ of DNA polymerase eta.” The scientists think that the multiple new TLS genes uncovered in this research might play a similar role for the other “sloppy” polymerases, thus providing a tightly controlled system that works in harmony, with minimal error.
 
For a specific cancer – acute myeloid leukemia (AML) – these findings may provide an explanation to a longstanding mystery: NPM1 has been found to be mutated in approximately 30% of AML patients; and patients harboring such mutations puzzlingly tend to have a better response to chemotherapy. Studies have shown that TLS is involved in resistance to chemotherapy, which damages DNA. The Weizmann team’s research suggests that AML cells carrying the mutated NPM1 gene have lower TLS rates, as their DNA polymerase eta degrades without its “guardian,” and they thus are more easily killed by the treatment.
 
Livneh: “Plans are currently under way to test these observations in AML patients, and if evidence supports these findings, it could indicate that NPM1 and DNA polymerase eta, and in particular their interaction, could be potential targets for AML drugs.”
 
Also participating in the study were Nataly Mirlas-Neisberg, Dr. Umakanta Swain, Dr. Reinat Nevo and Nir Ben-Chetrit of the Weizmann Institute, as well as Prof. Brunangelo Falini, Dr. Maria Paola Martelli and Roberta Rossi of the University of Perugia, Italy; Prof. Nicholas Geacintov of New York University, USA; Prof. Thomas Carell and Stefan Schiesser of Ludwig Maximilians University, Germany; and Prof. Christine E. Canman of the University of Michigan, USA.
 
Prof. Eytan Domany’s research is supported by the Leir Charitable Foundations; Mordechai Segal, Israel; and the Louis and Fannie Tolz Collaborative Research Project. Prof. Domany is the incumbent of the Henry J. Leir Professorial Chair.

Prof. Zvi Livneh’s research is supported by the Y. Leon Benoziyo Institute for Molecular Medicine, which he heads; the Dr. Erhard, Emmi and Fred Loewinsohn Center for Pediatric Health, which he heads; the Leona M. and Harry B. Helmsley Charitable Trust; the David M. Polen Charitable Trust; the Mike and Valeria Rosenbloom through the Mike Rosenbloom Foundation; and the Sergio Lombroso Award for Cancer Research. Prof. Livneh is the incumbent of the Maxwell Ellis Professorial Chair of Biomedical Research.


 
 
 


 

NPM1 interacts with DNA polymerase eta and regulates polymerase eta-promoted TLS. Assay of polymerase eta and NPM1 in unirradiated (l) and ultraviolet irradiated cells ((c) after one hour, (r) after 18 hours). Blue: DNA in the nucleus; green: polymerase eta–NPM1 interaction
Space & Physics
English
  • Biological Chemistry
  • Cancer
  • DNA damage and repair›DNA repair
  • Eytan Domany
  • Leukemia
  • Physics of Complex Systems
  • Zvi Livneh

Use It or Lose It

English

 

muscle
         A 3-D computer model of a fruit fly muscle fiber with multiple nuclei

 

Everyone knows that exercise can build muscle; without exercise, muscles quickly lose their strength and volume. These facts have been common knowledge for so long, it might come as a surprise to find that very little is known about the molecular mechanisms behind them. How indeed does physical activity cause muscles to strengthen?
Talila Volk
Prof. Talila Volk
Weizmann Institute scientists have now proposed an explanation. Research conducted by the team of Prof. Talila Volk in the Molecular Genetics Department suggests that the long, cylinder-shaped muscle cells – the muscle fibers – contain proteins that function as biological “mechanosensors.” These spring-like, elastic proteins are connected to the cellular supporting structure – the cytoskeleton – on the one side, and on the other side, to the cell’s nucleus. Muscle contractions exert mechanical pressure on the cytoskeleton, which, in turn, puts pressure on these sensor proteins, causing them to transmit a signal to the nucleus.

This signal presumably alters the internal architecture of the chromosomes within the nucleus, changing gene expression – that is, the activity of genes – in the affected portion of the DNA. As a result, certain genes become activated, prompting the release of proteins that make up the filaments responsible for the contraction of the muscle fiber. In addition to contraction, the proteins fortify existing filaments and help produce new ones, building up muscle mass.

In this manner, exercise triggers a strengthening of the muscle. But because muscle-building proteins have a high turnover, the signal for their production must be periodically repeated. If, in the absence of exercise, the muscle fails to contract for a while, the mechanosensors no longer send their signals to the nucleus, ultimately leading to a loss of muscle mass.
 
muscle fibers
Muscle fibers of a fruit fly viewed under a confocal microscope: a normal fiber has normally-shaped, properly distributed nuclei (A), whereas the nuclei of fibers with mutated MSP-300 proteins are distorted and distributed abnormally (B, C and D)

 

 
Evidence for this explanation comes from a series of studies that Volk and her team – including research students Hadas Elhanany-Tamir and Miri Shnayder, and postdoctoral fellow Dr. Shuoshuo Wang – conducted in fruit flies. As reported in the Journal of Cell Biology, the scientists have identified a fruit fly protein, called MSP-300, whose shape and mechanical properties make it perfectly suited to serving as a mechanosensor. MSP-300 forms a ring around the nucleus, with numerous radial extensions connected to the cytoskeleton. It has elastic properties, as would be required from a mechanosensor; but on the other hand, it operates in close cooperation with two other proteins that help create rigid scaffolding around the nucleus, protecting it from muscle contractions. By introducing mutations into MSP-300, the scientists showed that it is indeed essential for maintaining muscle mass. The effect of the mutations on the fly’s muscles was devastating: The muscles fibers thinned out and their nuclei were deformed and clumped together abnormally. As a result, the muscles didn’t work properly, so the larvae couldn’t crawl and adult flies couldn’t fly.
 
mechanosensor protein
The MSP-300 protein, viewed here under a confocal microscope, forms a ring (red in the left image, green on the right) around the muscle cell nucleus, with extensions to the cytoskeleton

 

 
Because human muscle cells contain proteins equivalent to MSP-300, suggesting they may also function as mechanosensors, these findings can shed new light on the connection between physical activity and a healthy build-up of muscle in humans. A better understanding of this connection may in the future lead to improved ways to prevent muscle loss resulting from aging, forced inactivity as in paralysis, or such disorders as muscular dystrophy.
 
Prof. Talila Volk's research is supported by Erica A. Drake and Robert Drake. Prof. Volk is the incumbent of the Sir Ernst B. Chain Professorial Chair.
 
Life Sciences
English
  • Cell
  • Molecular Genetics
  • Muscle tissue
  • Protein
  • Sensor
  • Talila Volk

“Big Eaters” Get a Makeover

English

When in the 1880s Russian zoologist Ilya Mechnikov first described large, voracious cells that devour bacteria and cellular debris – findings for which he later received a Nobel Prize – he called them macrophages, or “big eaters.”  Although macrophages have since been assigned numerous additional responsibilities, both in immunity and in tissue maintenance, it is the “big-eater” label that has stuck for nearly 130 years.  But recent research at the Weizmann Institute, which recently appeared in Cell, is putting a new face on the macrophage. In fact, not only one face but many: These studies show that macrophages take on different functions and appearances depending on which tissue they call home.  Prof. Steffen Jung of the Immunology Department says: “If we can learn the tricks they use to change their functions, we might be able to harness their activities to control disease.”

In the gut: Macrophages visualised as green cells in reporter mice
 
The tasks that the macrophages carry out in the various organs can be highly specific. In the nervous system, for example, they play a role in shaping nerve cells, while in the spleen they dine on well-aged red blood cells. Jung, who has been studying macrophages for a decade, recently revealed that, unlike most immune cells, many macrophages are long-lived cells that arise in the embryo, developing alongside their host organs. These findings raised new questions about macrophage development. For example, how do they get trained to serve each particular organ and tissue?

To begin to answer these questions, Jung joined up with departmental colleague Dr. Ido Amit, who investigates how regulatory regions in the genome control immunity. There are only a handful of macrophages in any one organ, but Amit and his team have developed cutting-edge techniques, including advanced robotics and next-generation sequencing technologies, to study such rare, specialized cells in their natural state. The scientists profiled these cells down to the molecular level, using the data to explain the unique ways their genes are expressed and regulated.
 
(l-r) Prof. Steffen Jung, Dr. Deborah Winter, Dr. Ido Amit and Dr. Ronnie Blecher-Gonen
 

 

        
 
The team, which also included Dr. Deborah Winter, Dr. Ronnie Blecher-Gonen, Eyal David and Dr. Hadas Keren-Shaul, all of the Weizmann Institute’s Immunology Department, and Yonit Lavin and Prof. Miriam Merad of the Icahn School of Medicine at Mount Sinai, New York, examined macrophages in seven different organs – from brain to lungs to intestines. They succeeded in identifying a unique signature for each of these populations, based on their genomic profiles and regulatory landscapes – a sort of textured map of their convoluted genetic activities. In particular, the team noted that each macrophage population used a distinct set of regulatory elements, or “enhancers,” for turning “on” or “off” the expression of certain genes.

Next the researchers switched these cells between organs. To their surprise, the macrophages began to change their signatures, taking on new profiles to fit their new surroundings. When the cells grew from immature cells, the revamping process was nearly complete – over 90%. But even when the experiment was repeated with fully differentiated cells, there were substantial changes in the macrophages’ personality profiles. Specifically, macrophages that were moved from the peritoneum to the lungs started to look and act like functional lung macrophages.
 
 
Macrophages in the brain shown in green
 
These findings, Amit and Jung believe, could change quite a few preconceived notions about macrophages. For one, these cells turn out to be incredibly plastic: “It’s a sort of ‘nature versus nurture’ issue,” says Winter, who led the study together with Blecher-Gonen. “Our findings suggest that nurture plays a much stronger role in shaping these cells’ identities than anyone had thought.” And the ways that the cells take their cues from their surroundings support Jung’s earlier findings: Macrophages arise from common precursor cells in the embryo and specialize after receiving signals from their host tissue, thus becoming an integral part of their surroundings. This goes beyond mere genetics, say the researchers: Understanding how the fate of such cells as macrophages is determined could help decipher the molecular mechanisms of diseases, including immunological disorders, anemia, leukemia and more.
 
The study was conducted in mice, but the ultimate goal is to apply the findings to humans. “In the future,” says Blecher-Gonen, “we want to learn how to retrain the macrophages ourselves. Then we could use them to treat diseases in which the patient’s macrophages are faulty or inadequate.” As an example, the team envisions creating lung macrophages that could clean up the thick secretions blocking the lungs of cystic fibrosis patients, or designer gut macrophages that could be used to treat irritable bowel disease. Amit: “Since most disease-causing mutations are located in regulatory regions, further studies could shed new light on the pathways involved in such diseases as inflammatory bowel disease and multiple sclerosis, and perhaps lead to the more precise treatment of patients.”
 
 
 
 
Ido Amit's research is supported by the M.D. Moross Institute for Cancer Research; the J&R Center for Scientific Research; the Jeanne and Joseph Nissim Foundation for Life Sciences Research; the Abramson Family Center for Young Scientists; the Wolfson Family Charitable Trust; the Abisch Frenkel Foundation for the Promotion of Life Sciences; the Leona M. and Harry B. Helmsley Charitable Trust; Sam Revusky, Canada; the Florence Blau, Morris Blau and Rose Peterson Fund; the estate of Ernst and Anni Deutsch; the estate of Irwin Mandel; and the estate of David Levinson. Dr. Amit is the incumbent of the Alan and Laraine Fischer Career Development Chair.
 
Prof. Steffen Jung's research is supported by the Leir Charitable Foundations; the Leona M. and Harry B. Helmsley Charitable Trust; the Maurice and Vivienne Wohl Biology Endowment; the Adelis Foundation; Lord David Alliance, CBE; the Wolfson Family Charitable Trust; the estate of Olga Klein Astrachan; and the European Research Council.


 
 
Macrophages in the brain shown in green
Life Sciences
English
  • Ido Amit
  • Immune cells
  • Immunology
  • Macrophage
  • Steffen Jung

Cellular Signature Style

English
To divide or not to divide: This is one of the most crucial decisions in the life of a cell. A new Weizmann Institute of Science study, conducted in collaboration with Danish researchers, has revealed a previously unknown mechanism that is involved in the control of such decisions in both healthy and cancerous cells. The findings, reported recently in Cell, provide an entirely new feature by which dividing and non-dividing cells can be told apart – a discovery that may, in the future, help advance the diagnosis, prognosis and perhaps even therapy of cancer.

The study focused on the presence of “synonyms” in the language of DNA: different three-letter portions of the genetic code that encode the same amino acid, a building block of proteins. For example, the two synonyms, GAA and GAG, differ chemically but produce the same amino acid. When the genetic code is translated into proteins – that is, when amino acids are manufactured on the basis of a particular DNA sequence – each synonym has its own messenger molecule, called transfer RNA (tRNA), which enables the translation. A tRNA is a two-headed molecular machine that on one end “reads” different synonyms of the code and on the other supplies the required amino acid for the synthesized protein.
 
Dr. Hila Gingold and Prof. Yitzhak Pilpel
 
Until now, the existence of more than 60 genetic three-letter synonyms, each having a corresponding tRNA, was not thought to have a major bearing on cell division. The ratios of different tRNAs were thought to be constant throughout the life of each cell, and in any event deviations or fluctuations in these ratios were not known to have particular biological significance.

Prof. Yitzhak Pilpel of the Weizmann Institute’s Molecular Genetics Department decided to look into the potential role played by the relative amounts of different tRNAs in the cell when he learned that Danish researchers had compiled an enormous database of genomic information from more than 400 people, including some 300 cancer patients, which contained a large volume of tRNA data. Pilpel’s intern Dr. Hila Gingold used DNA chip technology and mathematical algorithms to analyze the data, working together with Dr. Disa Tehler and other members from the team of Prof. Anders Lund of the University of Copenhagen, as well as other researchers from Denmark, the Netherlands, Switzerland and the United States.
 

Division signs


The results were so striking they surprised even the study authors themselves. The analysis revealed that two distinct subsets of tRNAs are present in the cell depending on whether it is dividing or not. For example, dividing cells tend to have large amounts of the tRNA that translates the GAG synonym, whereas non-dividing cells have a preference for the tRNA associated with the GAA synonym. As a result, dividing and non-dividing cells have discernible tRNA signatures – that is, characteristic amounts of all the different tRNAs.
 
Algorithm-generated comparisons among the genomes of cells from 450 tissue samples: The analysis revealed a striking similarity (red colors, upper right rectangle) in tRNA signatures among cancerous cells and healthy dividing cells, as well as a degree of similarity among the non-dividing cells (red colors, lower left rectangle), whereas no such similarity (blue) was found when dividing cells were compared with non-dividing ones
 

 

 
In other words, by measuring the relative amounts of tRNAs it’s possible to tell whether the cell is dividing, as is the case with cancerous cells, embryonic stem cells and other stem cells, or whether it’s a healthy cell that has stopped dividing after having specialized into a particular tissue type, such as a bone, blood or skin. In fact, all cancerous cells in the study were found to have characteristic tRNA signatures, which could be further classified by cancer type.
 

Changes in behavior

 
When the scientists examined the connection between tRNAs and different types of genes, they found, unsurprisingly, that the tRNAs more often present in dividing cells tended to be involved in the translation of genes that govern more “individualistic” cellular behavior – for example, rapid growth that doesn’t take neighboring cells into consideration – a feature typical of cancer. On the other hand, tRNAs present in non-dividing cells were associated with genes responsible for more “group-oriented” behaviors, such as cellular adhesion and communication, typical of healthy tissue.  

The scientists think the two tRNA signatures could have evolved as one of nature’s many ways of trying to block cancer. These signatures represent a mechanism that may add to cellular stability by keeping the two states, dividing and non-dividing, separate. When a normal non-dividing cell is on its way to becoming cancerous, it needs to alter its tRNA repertoire, a task that presents yet another hurdle it needs to overcome before it can start dividing uncontrollably: until this repertoire is changed, the translation of genes with cancerous potential may be less efficient.

Whatever their evolutionary significance, the analysis of tRNA signatures may in the future be developed into a tool for detecting or diagnosing cancer or for making a clinical prognosis of cancer development. An improved ability to distinguish between cancerous and healthy cells may also facilitate the design of therapies that will selectively target the tumor while sparing healthy tissue.
 
Prof. Yitzhak Pilpel's research is supported by the Sharon Zuckerman Laboratory for Research in Systems Biology; and the Braginsky Center for the Interface between Science and Humanities. Prof. Pilpel is the incumbent of the Ben May Professorial Chair.
 
Algorithm-generated comparisons among the genomes of cells from 450 tissue samples: The analysis revealed a striking similarity (red colors, upper right rectangle) in tRNA signatures among cancerous cells and healthy dividing cells, as well as a degree of similarity among the non-dividing cells (red colors, lower left rectangle), whereas no such similarity (blue) was found when dividing cells were compared with non-dividing ones
Life Sciences
English
  • Algorithm
  • Amino acids
  • Cancer genetics
  • Gene code
  • Molecular Genetics
  • Stem cell
  • Yitzhak Pilpel

The Time-Bomb Fungus

English
Stages in the invasion of the fungus Colletotrichum gloeosporioides: 1)germination
 
The frustration is familiar: You buy a perfect, slightly hard avocado, mango or tomato, but by the time it ripens sufficiently to be eaten, it has developed soft rotten spots that force you to discard at least part of the fruit and wonder if you need to change your greengrocer. A new study conducted by Weizmann Institute scientists, in collaboration with Israel’s Agricultural Research Organization at the Volcani Center, reveals in great detail the stages of a fierce, protracted battle waged by the fruit to try to prevent the delayed rotting. The findings point to future preventive treatments.
 
 
 
 
 
The story begins long before the fruit makes its way to your greengrocer. The new study, recently reported in the New Phytologist, has shown that a widespread fungus, Colletotrichum gloeosporioides, which infects close to 500 common plants and crops, starts its insidious invasion while the unripe fruit is still hanging on the tree. Once the fungus’s spore-like particles land on the fruit’s surface, they germinate. Within hours, each fungal particle grows a structure called an appressorium. This, in turn, releases a spear-like extension that pierces the fruit’s waxy surface and penetrates an unsuspecting cell, turning it into a “zombie” – that is, causing this cell to suspend most of its vital processes and devote itself to supporting the parasitic fungus. At this stage, the fungus switches into a dormant state that can last for months. To achieve such dormancy, the fungus structurally rearranges its chromosomes, shutting down hundreds of genes to reduce its own metabolism to the minimum required for maintenance.
2) penetration
 
Despite the furtiveness of the fungal attack, already at the initial, appressorial stage the plant senses the invasion by means of special cellular receptors and mounts a massive counterattack, activating thousands of genes, some of them encoding for antifungal compounds. These compounds are aimed at damaging the fungus and slowing its growth without affecting the plant’s own cells.
 
A fungus that survives the counterattack bides its time, encased within the fruit, and waits for ripening, a point at which the fruit’s defensive chemicals disappear. That is precisely when the fungus goes off like a time bomb.  Emerging from the dormant state, it starts releasing an array of digestive enzymes that rapidly kill the surrounding plant cells. Using these dead cells as a new source of nutrients, it begins to multiply rapidly, causing the death of even more fruit tissue and creating spots of soft rot on what was the greengrocers' prized fruit.
(l-r) Drs. Dana Ment and Gilgi Friedlander, Prov. Dov Prusky, Dr. Noam Alkan and Prof. Robert Fluhr
 
 
 
The research was performed in tomatoes, a convenient plant because its genome is known and because its fruit is available year-round, but the findings are applicable to other plant species infected by Colletotrichum gloeosporioides. After sequencing the entire genome of the fungus, the scientists performed in-depth analyses, determining which genes were activated or shut down at different points of the infection and thus which enzymes and other substances were released, in the fungus and in the plant simultaneously, at each stage of the fungal infection.
 
 
 
 
 
The study was performed in the laboratory of Prof. Robert Fluhr of the Weizmann Institute’s Plant Sciences Department; and by Dr. Noam Alkan of the Weizmann Institute and of the Agricultural Research Organization, with Prof. Dov Prusky and Dr. Dana Ment of the Agricultural Research Organization; and Dr. Gilgi Friedlander of the Weizmann Institute’s Nancy and Stephen Grand Israel National Center for Personalized Medicine.

On the basis of these findings, it should be possible to devise methods for fighting the time-bomb fungus: by developing tools to screen infected fruit, breeding plants with a heightened resistance to the infection or designing chemicals that affect the growth of the fungus but not plant growth or human health. Until then, as you cut out that rotten spot, you can at least marvel at this scene of a grand battle waged between the two kingdoms, fungal and fruit.
 

 

3) dormancy and 4) explosive growth
  
 

Prof.  Robert Fluhr's research is supported by the Angel Faivovich Foundation for Ecological Research; Lord David Alliance, CBE; and the estate of David Levinson. Prof. Fluhr is the incumbent of the Sir Siegmund Warburg Professorial Chair of Agricultural Molecular Biology.

 

 
3) dormancy and 4) explosive growth
Environment
English
  • Plant Genetics
  • Plant and Environmental Sciences
  • Plant disease
  • Robert Fluhr
  • Tomato

Bone: Heal Thyself

English
 
Bones break; it happens. It also happens that bones are one of the few organs that are able to regenerate. Those unfortunate enough ever to have experienced a broken bone know the procedure only too well: A physician physically “yanks” the bone – or, in the case of severe fractures, operates – to realign it with its other half, applies a cast to keep the two firmly in place, and within about one month, the bone halves reunite.
 
(l-r) Prof. Eli Zelzer and Chagai Rot
 
Though the first recorded use of rigid bone setting dates as far back as ancient Egypt, Prof. Elazar Zelzer and PhD student Chagai Rot of the Weizmann Institute’s Molecular Genetics Department now suggest, in a paper published in Developmental Cell, that constant movement of the bones could actually result in effective healing.

“In terms of evolution, the need for medical intervention just doesn’t make sense,” says Zelzer. “It’s a paradox: Evolution ‘invested’ great effort in affording bones with regenerative capabilities, and an intact skeleton is crucial for functioning and survival. Yet we are unaware of any natural mechanism able to align bones when they break.”

The medical literature has indicated the existence of such a natural system for some time. Some physicians are also aware of the phenomenon: They sometimes send very young children home with just a bandage wrapped around the limb. Even in cases of severe fracture, after some time the bones have often completely aligned and healed on their own. Until now, doctors and researchers have assumed that the bones initially rejoin at an angle and then are sculpted through a process of bone remodeling as the bone heals, mature bone being removed from one side and new bone being formed on the other to achieve the correct alignment.
 


To investigate the process, the Weizmann scientists allowed young mice with fractured bones to move around freely without any intervention, X-raying their bones on a daily basis. To their surprise, not only did the bones realign naturally, but this occurred within just a few days. This fast turnaround occurred while the bones were still separated, leading the scientists to believe that it is something other than the process of remodeling that brings about bone alignment.
 

Natural healing process of fractured bones. (A) Healing begins with a collection of blood and inflammation at the fracture site. (B) Soft callus (purple) is formed, which develops into the bidirectional growth plate at the concave side of the fracture site. (C) The growth plate drives bone growth in opposite directions. The result is a jack-like mechanical effect that moves the fragments toward straightening (red arrows). (D) New bone tissue is formed (orange). (E) The shape of the bone is fine-tuned by remodeling
 

 

 
Further analysis of the healing bones in the active mice revealed yet another surprise: New tissue similar to growth plates – an area at either end of growing bones from which new bone tissue is produced – had formed, but on the concave side of the fracture. The researchers observed that bone tissue is produced from both sides of the plate, acting like a “mechanical jack” to generate opposing forces that straighten the two bone fragments. Only once they are precisely realigned do the bone halves proceed with the modeling process to reunite and reshape.

Acting on previous research in Zelzer’s lab suggesting that muscle contraction may also play a role in the process, the scientists injected the mice with Botox to paralyze the muscles. They found that although the fractured bone had reunited, the halves were not aligned properly, remaining at an angle. The reason, they found, was that in the absence of muscle contraction, the new growth plate didn’t form.

The fact that this natural mechanism was found to be less effective in adults suggests that this newly discovered paradigm has helped solve the longstanding mystery of why fractures heal so much faster in the young. Rot: “In terms of evolution, a rapid and efficient fracture-healing process may be more important in the young, to ensure their ability to reproduce; while there is less ‘survival’ advantage for adults who have already passed on their genes to their offspring.”

The scientists suggest that a better understanding of spontaneous realignment in fracture healing may provide a new line of thinking – even in older children and adults – and help physicians reevaluate current bone-setting procedures.
 
Prof. Elazar Zelzer's research is supported by the Jeanne and Joseph Nissim Foundation for Life Sciences Research; the Irving and Dorothy Rom Charitable Trust; and the estate of David Levinson.
Natural healing process of fractured bones. (A) Healing begins with a collection of blood and inflammation at the fracture site. (B) Soft callus (purple) is formed, which develops into the bidirectional growth plate at the concave side of the fracture site. (C) The growth plate drives bone growth in opposite directions. The result is a jack-like mechanical effect that moves the fragments toward straightening (red arrows). (D) New bone tissue is formed (orange). (E) The shape of the bone is fine-tuned by rem
Life Sciences
English
  • Bone formation
  • Bone formation
  • Elazar Zelzer
  • Molecular Genetics
  • Musculoskeletal system

Sending a Mixed Message

English
 
In Alice in Wonderland, one side of the mushroom made her shrink, while the other side of the same mushroom made her grow. Alice may have been confused by this, but a similar paradox is often evident in biology: The same molecule can have completely opposite effects on the cells that receive its signals. These signaling molecules are secreted and consumed by cells in order to communicate with each other and coordinate actions. Thus, for example, a certain molecule can signal enhanced cell division to increase cell numbers and, at the same time, enhance the probability of cell death. Is there a reason why nature chose this counterintuitive strategy? Can one molecule somehow do both jobs better than two?
 
Paradox
 
The communication between T cells – part of the immune system’s protection against invading pathogens – provides a good basis for gleaning insight: T cells secrete a signaling molecule, called IL-2, which has the split function of increasing T-cell proliferation and inducing T-cell death. In previous collaborative research, the groups of Dr. Nir Friedman of the Weizmann Institute’s Immunology Department and Prof. Uri Alon of the Molecular Cell Biology Department had developed a mathematical model predicting that the paradoxical mechanism of IL-2 enables the body to reach a state of homeostasis – that is, maintaining a relatively steady number of cells. “Balance in such a system is important in order to prevent too large a response on the one hand, which could prove wasteful or harmful, or too weak a response on the other hand,” says Friedman.

The scientists recently joined forces again to put their theoretical predictions to the experimental test. Dr. Shlomit Reich-Zeliger and former postdoctoral fellow Dr. Yaron Antebi in Friedman’s lab, together with then PhD student Yuval Hart in Alon’s group, cultured T cells in groups of different starting sizes and followed their numbers for a week. Their findings, recently published in Cell, support the notion that T cells do indeed reach homeostasis as a result of the paradoxical IL-2  signaling – maintaining an almost fixed number of cells regardless of their initial concentration. However, again as predicted, this is only true above an initial threshold level of cell numbers. Below that threshold, the T-cell population shrinks to extinction. Friedman: “This function might serve as a kind of ‘safety switch,’ possibly to prevent an exaggerated response to a harmless trigger mistakenly identified by just a few cells.”
Prof. Uri Alon
 
The experiments helped validate further predictions: The body’s homeostasis is dependent on IL-2; T cells “lose their balance” if they can’t produce this molecule. The researchers also confirmed that IL-2 levels rise and fall, with peak levels occurring, counterintuitively, at relatively low cell numbers – just above the threshold. At this level, the higher signal probably functions to boost T-cell proliferation. 
 
Still, why use a “two-in-one” molecule? To answer the question, the scientists decided to test other possible scenarios by plugging a two-molecule solution into their mathematical model. The results showed that two separate molecules, each with its own function, would make the system more sensitive to environmental perturbations that could throw it completely off balance. In contrast, having one molecule with two functions proved to be a more robust setup, enabling the system to better sense and compensate for additional signals in the surrounding environment – something like Alice eating bits from alternate sides of the mushroom until her senses told her she had reached the right height.
Dr. Nir Friedman
 
IL-2 is used to treat various diseases, but its use is hampered by possible serious side effects. A better understanding of its complex mechanism of action may therefore help toward designing more effective interventions in the future. More generally, these findings can also be applied to other biological systems whose molecules display similarly paradoxical signaling – for example glucose, which has contradictory effects on the insulin-producing cells in the pancreas.  

This study provides an important step toward gaining a better, more quantitative understanding of the complex intercellular responses between communicating cells. By modeling the population dynamics of cells in such complex systems as the human immune system, studies like this can reveal the paradoxical nature of our existence.

Also participating in the research were Dr. Irina Zaretsky of the Immunology Department and Dr. Avraham Mayo of the Molecular Cell Biology Department.
 
Prof. Uri Alon’s research is supported by the Jeanne and Joseph Nissim Foundation for Life Sciences Research; the Braginsky Center for the Interface Between Science and Humanities; and the European Research Council. Prof. Alon is the incumbent of the Abisch-Frenkel Professorial Chair.

Dr. Nir Friedman’s research is supported by the Nella and Leon Benoziyo Center for Neurological Diseases; the Clore Center for Biological Physics; the Henry Chanoch Krenter Institute for Biomedical Imaging and Genomics; the Victor Pastor Fund for Cellular Disease Research; the Abraham and Sonia Rochlin Foundation; the Adelis Foundation; the Norman E. Alexander Family Foundation; the Jeanne and Joseph Nissim Foundation for Life Sciences Research; the Crown Endowment Fund for Immunological Research; the estate of John Hunter; and the estate of Suzy Knoll. Dr. Friedman is the incumbent of the Pauline Recanati Career Development Chair.
 
 

 

 
Paradox Alice in Wonderland
Life Sciences
English
  • Immunology
  • Molecular Cell Biology
  • Nir Friedman
  • Signaling molecule
  • T cell
  • Uri Alon

Curiosity and Constraint

English
Measuring the response to novelty: A mouse repeatedly touches the object and pulls away (nose and whisker contacts are color-coded; d is the distance of the snout from the object)
 
 
 
Put a young child in a new playground and she may take a while to start playing – approaching the slide and then running back to Mom before finally stepping on. A new model suggests that it is not fear that makes her run back and forth, but simply the fact that her brain is telling her to stop and take in the new information – the height of the slide or how slippery it appears – before going any further. Drs. Goren Gordon and Ehud Fonio, and Prof. Ehud Ahissar, believe that this is a basic pattern in mammals that governs how we learn. The mathematical model they developed and tested in experiments suggests that our innate curiosity is tempered by mechanisms in our brains that curb our ability to absorb novelty.   

In Ahissar’s lab in the Institute's Neurobiology Department, researchers investigate how animals sense their surroundings. Previous research in which Fonio had participated showed that in a new situation, a mouse would approach an unfamiliar space, retreat to familiar surroundings, and then approach again. When Gordon, Fonio and Ahissar examined how mice used their whiskers to feel out a novel object, a similar pattern ensued: The whisker would touch the object, pull back, and then touch it again. Gradually, as the mouse became familiar with one part of its surroundings, it would begin to explore further, moving away from the known part. The pattern was so consistent, the researchers thought they could create a model to explain how a mouse – or another mammal – explores new surroundings.
 
The researchers based their model on the premise that novelty can be measured and that the amount of novelty could be a primary factor in shaping the way that a mouse – or its whisker – will move through an environment. This model successfully reproduced the results of the previous study, in which the movement of the mouse gradually became more complex through the addition of measurable degrees of freedom. For example, it began with movement along a wall, as opposed to traveling across the open space. Using data from the previous experiments and others for which such data were available, they were able to construct a model that required very few additional assumptions.

The model suggested that novelty, per se, was not the deciding factor, but rather how much the novelty varied within a given situation. Approaching and retreating appear to be a way to keep the amount of new information within a constant range. Like the wavering child in the playground, the mice would absorb a certain amount of new sensory input – the curve of a new wall, for example – retreat, and approach again once the novel information was already starting to become familiar.

Dr. Goren Gordon and Prof. Ehud Ahissar
 
To test the model, the researchers designed an experimental setup in which a family of mice was born and raised in a den, and then a gate was opened from the den to a new area in which the pups could freely explore and return to their familiar den. The researchers found that the model was able to predict how the mice would explore their new surroundings. It held true whether it was applied to locomotion or to the motion of whiskers in feeling out new objects. The initial movements explored the most novel features of the new environment. After those were learned, just as the model predicted, the animals moved further afield, exploring the still-unknown parts of their surroundings.

Gordon: “The mice were not given rewards for their behavior – for them, as for humans, satisfying curiosity is its own reward.” Fonio: “This behavioral pattern enables the mice to control the level of sensory stimulus to their brains by regulating the amount of novelty they are exposed to.” These limits to novelty and exploration may, of course, have another evolutionary advantage: While the urge to explore is necessary for animals that must seek out food, stopping to check out the surroundings a bit at a time could be a prudent survival strategy. In other words, curiosity may have killed the cat, but a whisker pulled back in time might save the mouse.
 
Does this model apply to humans? Gordon points out that when we learn a new subject, we often need time to think things over before going on to the next topic. Further research might reveal whether young children, for example babies just learning to crawl, explore their new surroundings in the same way. Even an adult entering a new situation might undergo a similar process.

In the future, a mathematical model of learning might prove useful for teachers and students, as well as for research into neurological issues involving the ability to absorb new information. In addition, this model might someday be used in the field of robotics: Robots that learn on their own, like mice, to explore a new setting might be able to function in situations that are too dangerous for humans, for example, the aftermath of an earthquake or a nuclear power plant accident.
 
Prof. Ehud Ahissar’s research is supported by the Murray H. and Meyer Grodetsky Center for Research of Higher Brain Functions; the Jeanne and Joseph Nissim Foundation for Life Sciences Research; the Kahn Family Research Center for Systems Biology of the Human Cell; Lord David Alliance, CBE; the Berlin Family Foundation; Jack and Lenore Lowenthal, Brooklyn, NY; Research in Memory of Irving Bieber, M.D. and Toby Bieber, M.D.; the Harris Foundation for Brain Research; and the Joseph D. Shane Fund for Neurosciences. Prof. Ahissar is the incumbent of the Helen Diller Family Professorial Chair in Neurobiology.





 
 


 

 
Measuring the response to novelty: A mouse repeatedly touches the object and pulls away (nose and whisker contacts are color-coded; d is the distance of the snout from the object)
Life Sciences
English
  • Ehud Ahissar
  • Mathematical model
  • Neurobiology
  • Sensory stimulus
  • Whiskers

Where Two Paths Meet

English

 

protein map
 
With modern maps, we can zoom out to a region’s entire road system or zoom in to distinguish the streets leading from one particular point to another. Somewhere in the middle of our zooming we might note the pattern of roads connecting neighboring cities that create interdependence between them. In the lab of Prof. Adi Kimchi of the Molecular Genetics Department, researchers creating such a map for two “cities” in the cell are coming to understand just how important the interactions created by such connecting roads might be for our body’s cells. Among other things, this arrangement may help prevent the growth of cancer.
 
The “cities” are molecular pathways that end in two different types of cell death. A pathway, in the language of molecular biology, is the set of proteins and the series of events they mobilize that lead to a particular end. Such pathways may comprise large networks of proteins and complex, multi-stage communications systems. Hundreds of proteins in each cell death pathway must talk to one another to opt for, initiate and orchestrate the process.
Prof. Adi Kimchi and Yuval Gilad
 
Cell death is so important to the body that it has at least three different pathways for accomplishing this: apoptosis, autophagy and necrosis. As opposed to the latter, which is a violent death generally due to wounds or infection, apoptosis is a considered and planned rapid suicide. It often comes into play during development, when the embryo is constantly being remodeled, or later, when damage to the DNA of a cell threatens to turn it cancerous. Autophagy, unlike necrosis or apoptosis, does not always lead to cell death: It can be a survival mechanism, shutting down parts of the cell’s functioning in stressful conditions and recycling nonessential proteins for emergency use. But when needed, this same mechanism can also recycle the cell out of existence.

Until recently, researchers had thought that apoptosis and autophagy were like independent walled cities, each with its own separate system of roads. But recent research in Kimchi’s lab reveals a more modern landscape, in which the two pathways veer into one another’s territory, interconnected by crisscrossing trails.

To map out the region where the two pathways meet, Kimchi and research student Yuval Gilad created an experimental system to check for interactions between the proteins in both pathways. Gilad did this by attaching pieces of a protein called luciferase – the molecule that makes bioluminescent plankton glow – to the pathway proteins. In the experiment, the luciferase emitted light when two proteins with matching halves met, thus reporting on a connection between these pairs. Each connection then added another line to the map. This new map was recently published in Cell Reports.
 
 
Mapping cell death: The protein-protein interactions discovered and mapped out in Prof. Adi Kimchi’s lab (red lines are newly-revealed interactions) divulge the wealth of interconnections between two major cell death pathways
 

 

 
 
All together, after testing thousands of possible pairs, Kimchi and Gilad identified 46 new interactions, “each with a story to tell,” says Kimchi. Next, they zoomed in on one particular location on the map, a protein known as DAP kinase 2 (DAPK2). DAP kinases, a family of proteins discovered nearly two decades ago in Kimchi’s lab, play a prominent role in apoptosis. But the six lines on the map leading to and from DAPK2 suggested that it is active in both pathways. Zooming in further, the researchers looked closely at one of its connections – an interaction with a protein known as 14-3-3. The protein 14-3-3 binds to a unique region on DAPK2, preventing it from assuming its active form. The discovery of this interaction revealed a new layer of regulation on DAPK2 activity. Since it’s activation can lead to cell death, says Kimchi, it will be interesting to look for treatments that activate DAPK2 in cancer cells by inhibiting its interaction with 14-3-3.
 
DAPK2 map
The new map reveals just how complex the business of cell death can be. The researchers believe that the interconnections between the pathways provide the cell with a backup in case one of them fails. This is especially important when a cell that could be cancerous needs to self-destruct; the growth of cancer usually involves a failure of the cell death pathways. Kimchi and Gilad say that this map and the method they used to create it could provide a valuable tool to both cancer researchers and drug developers. Not only has it revealed a rich new landscape of protein interactions, it can expose the stages at which these interactions occur. Since cancer cells tend to evade death, drug designers might attempt to target specific stages of the cell death process in order to encourage the cells to “commit suicide.” The new map can help highlight the better targets for drug screening and the reporters detected in the study can be used as tools for the identification of such drugs.

Prof. Adi Kimchi's research is supported by the Helen and Martin Kimmel Institute for Stem Cell Research; the EMET Prize sponsored by the A.M.N. Foundation for the Advancement of Science, Art and Culture in Israel; and the estate of Katherine and Ladislaus Braun.
 
 

 
 
 
 
Mapping cell death: The protein-protein interactions discovered and mapped out in Prof. Adi Kimchi’s lab (red lines are newly-revealed interactions) divulge the wealth of interconnections between two major cell death pathways
Life Sciences
English
  • Adi Kimchi
  • Apoptosis
  • Autophagy
  • Cancer
  • Molecular Genetics
  • Molecular pathways

Contacts

English

 

(l-r) Drs. Yael Elbaz-Alon and Maya Schuldiner
 
Every country has borders and – no less important – border crossings where goods and people can enter and leave. In the world of the cell, the cellular organelles act like little independent states: each conducting its specialized, vital processes within its borders while at the same time ensuring the flow of information and materials from one to the other. The recent discovery of a new “border crossing” between two organelles that were not even known to have “diplomatic relations” sheds new light on the ways that the organelles work together in a “federated state.”

The trans-border business of organelles can be conducted through couriers – secreted proteins that go from one to the other. But some types of passage require physical contact – to move materials from one to the other, for example. This takes place at specific contact points located around the border of the organelle. The first such contact point was identified over three decades ago, and new ones are still being discovered.
 
Dr. Maya Schuldiner of the Molecular Genetics Department says that every new discovery of this sort adds to the growing picture of the interrelations between organelles, the movement of information and materials within the cell, and obstacles that can cause disease.

In her postdoctoral research at the University of California, San Francisco, in the group of Prof. Jonathan Weissman, Schuldiner had worked with the team in Prof. Peter Walter’s lab that discovered a contact point between two of the cell’s main organelles. It connected the mitochondria – the cell’s power plants – with the endoplasmic reticulum in which, in addition to its function in readying proteins for secretion outside the cell, fats are produced. Together, these two organelles take up close to a third of the cell’s volume. They cling together at their contact points through “zipper proteins,” allowing fat molecules from the endoplasmic reticulum to pass into the mitochondrion, which uses them to build membranes.
 
The scientists surmised that if there were damage to the zipper proteins, the passage of fats, and thus the membrane structure of the mitochondria, would be impaired. But to their surprise, no such effect ensued. The conclusion was that the mitochondria had an as-yet-undiscovered contact point for obtaining fat.

Dr. Yael Elbaz-Alon, a postdoctoral fellow in Schuldiner’s lab, took up the challenge of finding that unknown contact point. Her starting theory was that if the flow of material (in this case fat) to the mitochondria remains constant when one of two available sources dries up, crossing points for the other one must double in order to keep up with demand. So the scientists looked at yeast cells – which have a somewhat manageable 6,200 proteins – and disabled these proteins one by one, looking for any case in which the known contact point, highlighted with a fluorescent marker for visibility, would double its activity. The lab’s robotic system enabled the researchers to conduct this test for all the proteins automatically.
 
In a normal cell (left) there is some physical contact between the vacuole (V) and the mitochondria (M). In the cell on the right, the contact points have increased significantly following damage to the passage from the endoplasmic reticulum, leading to “clumping” of vacuoles around a mitochondrion
 

 

 
This strategy turned up four proteins that, when damaged, cause the known contact points to multiply. The team then looked at the locations of these four proteins in the cell. And, indeed, one of them appeared on the mitochondria’s borders – but in an unexpected place. This protein formed a border crossing to another organelle – the vacuole (similar to the lysosome in humans). This is a cellular recycling plant that naturally contains large quantities of fat molecules. It appears that the mitochondria have two sources of fat – freshly produced fat from the endoplasmic reticulum or recycled fat from the vacuole – and two types of border crossings to match.

The research, which recently appeared in Developmental Cell, not only revealed a new border crossing, but a new border as well. The new contact point, which has been named vCLAMP, has evaded detection until now, says Schuldiner, because “in normal cells there are very few of them. Only when the previously-known border crossings were damaged did the numbers of the second swell to the point where they could be easily seen with an electron microscope. In fact, when these contact points were forced to take over, we also saw unusual clumps of mitochondria surrounded by vacuoles.”

Evidence for the existence of vCLAMP has been found in humans, meaning that this phenomenon is an important one that has been preserved throughout evolution. Schuldiner’s research group plans to study this phenomenon in depth to understand just how important it is.

Also participating in this research were Prof. Tony Futerman of the Biological Chemistry Department and Eden Rosenfeld-Gur, a research student in his group; Dr. Vera Shinder of the Electron Microscopy Unit; and Dr. Tamar Geiger of the Sackler Medical Faculty of Tel Aviv University.   
 
Dr. Maya Schuldiner's research is supported by the Foundation Adelis; the Georges Lustgarten Cancer Research Fund; the Dora Yoachimowicz Endowed Fund for Research; the Berlin Family Foundation; Roberto and Renata Ruhman, Brazil; the European Research Council; and Karen Siem, UK.
 
Prof. Anthony H. Futerman's research is supported by the Nella and Leon Benoziyo Center for Neurological Diseases, which he heads; the M.D. Moross Institute for Cancer Research; the Carolito Stiftung; and the Rosetrees Trust. Prof. Futerman is the incumbent of the Joseph Meyerhoff Professorial Chair of Biochemistry.

 
 


 

 

 
(l-r) Drs. Yael Elbaz-Alon and Maya Schuldiner
Space & Physics
English
  • Biological Chemistry
  • Endoplasmic reticulum
  • Maya Schuldiner
  • Mitochondria
  • Molecular Genetics
  • Robot›Robotic lab
  • Tony Futerman

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