Chaperone enzyme provides new target for cancer treatments

Chaperone enzyme provides new target for cancer treatments

UNC scientists who study how cells repair damage from environmental factors like sunlight and cigarette smoke have discovered how a "chaperone" enzyme plays a key role in cells' ability to tolerate the DNA damage that leads to cancer and other diseases.

The enzyme, known as Rad18, detects a protein called DNA polymerase eta (Pol eta) and accompanies it to the sites of sunlight-induced DNA damage, enabling accurate repair. When Pol eta is not present, alternative error-prone polymerases take its place – a process that leads to DNA mutations often found in cancer cells.

In one known example, faulty DNA repair due to Pol eta- deficiency is responsible for the genetic disease xeroderma pigmentosum-variant, which makes patients extremely susceptible to skin cancers caused by exposure to sunlight. However, scientists did not know how the cells selected the correct DNA Polymerase for error-free repair of each type of DNA damage.

"We found that the mechanism that promotes the 'chaperone' enzyme to recruit Pol eta to sites of DNA damage is managed by another signaling protein termed 'Cdc7' which we know is essential to normal regulation of the cellular lifecycle," said lead author Cyrus Vaziri, PhD, who is an associate professor of pathology and laboratory medicine and member of UNC Lineberger Comprehensive Cancer Center. Thus cells employ Cdc7 to ensure accurate DNA repair during the stage of their lifecycle that is most vulnerable to cancer-causing mutations.

The study was published in November in the Journal of Cell Biology.

According to Vaziri, the dual role that Cdc7 plays in the cell lifecycle and DNA repair offers a promising target for potential cancer therapies.

"We know that cancer cells have high levels of Cdc7 activity and can evade some DNA-damaging therapies such as cis-Platinum through Rad18 and Pol eta activity. We may be able to target this pathway in platinum-resistant tumors to prevent DNA repair and enhance cancer cell killing by platinating agents," he said.

Global view of blood cell development reveals new and complex circuitry

Global view of blood cell development reveals new and complex circuitry

A small pool of stem cells replenishes the human body with about 200 billion new blood cells daily. But the elaborate circuitry that determines if a cell will develop into a T cell, red blood cell, or one of the nine or more other blood cell types remains largely unknown. A research team led by scientists from the Broad Institute and Brigham and Women's Hospital has taken a systematic approach to help decipher this circuitry, compiling a comprehensive catalog of the factors that determine a blood cell's fate. Their work appears in the January 21 issue of Cell.

The researchers found that blood cells are directed by a multitude of transcription factors, proteins that turn on and off genes. While many previous studies have focused on individual transcription factors or types of blood cells, this study examined the expression and regulation of all transcription factors throughout blood development. The findings point to densely, interconnected circuits that control this process, suggesting that the wiring for blood cell fate is far more complex than previously thought.

"One assumption in the field had been that there are a small number of transcription factors that orchestrate this process," said Aviv Regev, a Broad Institute core member and co-senior corresponding author of the study. "Some people have always thought there would be a lot of factors and that it would just take time to find them. It turns out there are more masters than we would have thought."

The researchers looked globally at how the expression of all 20,000 or so genes in the genome change as blood stem cells become specialized cell types (a process known as differentiation). They discovered that while a small fraction of genes are uniquely expressed in a single type of cell, other genes are more broadly expressed — present in a variety of cell types but at varying levels. Some of these genes are turned on in the blood stem cells and switched off at certain points in development while others are reused in several parallel developmental branches. The researchers found about 80 of these patterns of variable genes, called modules. Each kind of specialized cell has a unique profile, or combination, of these modules.

Looking at the genes modulated in the course of healthy cell development could give researchers clues about what events lead to blood cancers, such as leukemia, a disease where differentiation has gone wrong.

"When you look at leukemia cells beneath a microscope, they have a lack of differentiation and they look abnormal," said Broad associate member Ben Ebert, an associate physician of hematology at Brigham and Women's Hospital and a senior corresponding author of the study. "They've ended up in a place that doesn't exist in normal development." Now that the researchers have a clearer picture of the modules that normal cells exhibit, they can apply this knowledge to help identify the similarities and critical changes in leukemia cells' profiles.

"Leukemia cells have the same set of building blocks as normal blood cells – some, they keep the right way so a piece of the profile is right, and a piece of the profile is wrong," said Regev, who is also an assistant professor in the department of biology at MIT and an Early Career Scientist at Howard Hughes Medical Institute.

The research team included co-first author Noa Novershtern from the School of Computer Science at the Hebrew University of Jerusalem, co-first author Aravind Subramanian in Todd Golub's laboratory at the Broad, and Lee Lawton and other collaborators in Richard Young's laboratory at the Whitehead Institute. All of their results will be made publicly available online through a database known as the Differentiation Map Portal (or D-Map). Ebert, Regev and their colleagues intend for D-Map to be a starting point for other researchers, empowering their investigations into the biology of blood cells as well as leukemia and other human diseases.

"Already, many people are asking for the data. Other groups can now combine their data with ours to ask new questions," said Novershtern. "What's also exciting is that people can see the power of computational models, tools that can be used to find new biological insights from the data."

Scientists find measles' natural nemesis

Scientists find measles' natural nemesis

Scientists at The Scripps Research Institute have found that a known enzyme in cells protects against measles virus, likely by altering the virus's genetic material, RNA. Cells lacking the enzyme become highly vulnerable to the virus's destructive effects. The enzyme also protects against several other respiratory viruses, including influenza A.

"We believe that host cells use this RNA-editing enzyme to slow these viruses' ability to replicate," said Michael B. A. Oldstone, the study's senior author and a professor at Scripps Research's La Jolla, California campus. The study's first authors are Simone V. Ward, a senior research associate in the Scripps Research Oldstone laboratory, and Cyril X. George of the University of California, Santa Barbara.

The finding represents a significant improvement in the understanding of measles infections, which still kill about 150,000 children and adults around the world every year. The paper, which was published recently in Proceedings of the National Academy of Sciences, has prompted commentaries in the journals Nature Reviews Microbiology and Viruses.

The focus of the study was the enzyme ADAR1 ("adenosine deaminase acting on RNA, 1"), which is known to be produced in high amounts in measles-infected cells. ADAR1 has been suspected as a "restriction factor" that inhibits viral replication.

ADAR1's role against measles has been difficult to nail down, however. In mice genetically engineered to be infectable by measles – a virus that normally infects only humans – ADAR1 is required for embryonic development, as in all mice. Thus the standard "gene knockout" technique, which would enable scientists to see how measles infections proceed without ADAR1, hasn't been feasible.

In this study, Ward, George, and Oldstone, and their colleagues knocked out only one of the two forms of ADAR1 produced in cells. This form, p150, is the one produced in response to infections. For reasons that still aren't clear, mouse embryos cannot grow for long without p150, so the researchers used a standard technique to "immortalize" these p150-knockout embryonic cells—ensuring their continuous supply—and in this way created a useful cell model.

When infected by measles virus, the p150-knockout cells succumbed quickly compared to immortalized control cells that produced p150 normally. "When I looked at the cells only 21 hours after infection, the p150-knockout cells already showed the signs of cell damage typical of measles infection," said Ward. "But the control cells looked exactly like uninfected cells."

In further tests, Ward found p150 also provided significant protection for cells against Newcastle disease virus, Sendai virus, canine distemper virus, and influenza A virus – which are all respiratory viruses like measles, and all members of the paramyxovirus or orthomyxovirus families.

Nine years ago, it was reported that a cellular enzyme known as APOBEC3G protects cells from DNA-based viruses such as HIV, by mutating viral genes. "We're now showing that an analogous gene-editing enzyme also seems to exist for RNA viruses," said Oldstone.

With the new cell model, and advanced "conditional knockout" techniques that allow genes to be disrupted in specific organs in adult mice, Ward, Oldstone, and their colleagues now hope to study ADAR1-p150's role in more detail.

One key issue to be resolved is the enzyme's role during brain infections. Measles virus usually results in a relatively mild illness lasting only a week or two, but in rare cases it spreads to the brain and becomes a persistent, always fatal infection known as subacute sclerosing panencephalitis (SSPE). In such cases, the virus doesn't have to spread via cell-to-cell contact, thus exposing itself to the immune system; it can spread more stealthily along the axons and dendrites that connect neurons.

"What we hope to show with our ongoing work is that host neurons are using ADAR1 to slow down this process, turning it into a gradual neurological disease," said Oldstone. ADAR1 might also be exacerbating the neurological symptoms of SSPE, he adds, because its enzymatic activity is known to affect the production of the important neurotransmitter receptors for serotonin and glutamate: "It's an enzyme that has multiple roles," Oldstone concluded.

How bacteria keep us healthy

How bacteria keep us healthy

Enlarge

Joerg Graf, associate professor of molecular and cell biology, with medicinal leeches at his lab. His research on bacteria in the gut uses the medicinal leech as a model system. Photo by Peter Morenus

(PhysOrg.com) -- Joerg Graf is studying medicinal leeches for clues about how changes in diet affect microorganisms in the digestive tract.

Many people think of bacteria as disease-causing agents that should be avoided for fear of getting sick. But molecular and cell biologist Joerg Graf points out that we can’t live without them.

He says there are actually more bacteria on and in humans than there are human cells. “If you took a person and removed all the human cells, you would still see the outline of a human body,” made up of bacterial cells, says Graf, an associate professor of molecular and cell biology in the College of Liberal Arts and Sciences.

Bacteria are on our skin, in our mouth and throat, in our intestines, nose, and virtually every nook and cranny of our bodies that is connected with the outside – and almost of all them are not bad for us.

Graf recently received a $1.6 million grant from the National Institutes of Health (NIH) to study bacteria that live in digestive tracts. The project is a collaboration with Pieter Visscher, a professor of marine sciences, and Hillary Morrison, a researcher at the Marine Biological Laboratory at Woods Hole. Their work will help scientists understand the vital role that bacteria play in our everyday health.

Many bacteria are essential to the normal functioning of physiological processes, including digestion and immune responses. The gut microbiome consists of all of the bacteria in the human gut and, for example, digests food that humans otherwise can’t, such as some plant material, as well as providing nutrients in forms that humans can use. Humans acquire these bacteria from their mothers as they pass through the birth canal and from the outside world through the course of their childhood. This is why, explains Graf, people in different parts of the world have different combinations of gut bacteria.

Enlarge

Joerg Graf displays a leech. Photo by Peter Morenus

Graf and his colleagues are interested in how relationships between bacteria and their hosts have evolved, and how they change over an organism’s lifetime.

“We don’t understand what drives the changes in the gut microbiome,” he says, adding that it’s difficult to study the human gut because there are hundreds of different kinds of bacteria present.

Instead Graf and his colleagues work on a model system. “By having a simple system, we can understand it better using molecular tools.”

The model system they use is the medicinal leech, because the microbiome of its digestive tract is normally dominated by only two types of bacteria. These species help to degrade blood that the leech consumes, providing nutrients to the leech. With only two species to observe, Graf can study how this bacterial community changes with dietary shifts in the organism.

Like humans, leeches undergo a shift in their diet at a young age: while humans spend the first stages of their life drinking only milk, leeches also spend their first few days living only on protein provided by their mother. After feeding the leech its first actual blood meal, the scientists will observe the changes in the leeches’ gut bacteria by looking at the RNA they make, which will in turn produce different proteins that allow the bacteria to degrade the food and provide nutrients to the leech. To perform this work, they will use a gas chromatography-mass spectrometer that was purchased by Kenneth Noll, a professor of molecular and cell biology, using funds from the Provost’s Intermediate Research Equipment Competition.

“The function of these proteins is related to the bacterial species and can determine how diverse the functions are within one species,” and how the function changes as the microbial community changes, says Graf.

One of Graf’s hypotheses is that one of the bacteria uses a “molecular syringe” to inject a toxin that prevents the leech’s natural immune cells from attacking it. This molecular syringe is also a virulence factor that is required to cause disease in other animals, but not to the leech. In the leech, it simply signals that the bacteria are not enemies, and should be left alone.

Additionally, Graf is interested to find out whether these bacteria rely on each other for nutrients. Like the leech itself, one of the bacteria might rely on the other to converting food into a form that it can use, allowing it to grow faster.

All of these questions will help Graf and his colleagues determine how physiology changes when the gut microbiome changes, which could lead to insights about our dependence on the bacteria themselves. For example, people with inflammatory bowel disease have a different microbial community than those without, says Graf. Learning about the changes in bacterial gut communities and its contribution to the host animal could give clues about other diseases.

Graf is also involved with the Human Microbiome Project, which aims to characterize human microbial communities, including that of the gut, and to analyze their role in health and disease. Hundreds of scientists across the U.S. are working on this NIH-funded initiative.

Graf says that for him, being involved with these projects is like working to solve a puzzle.

“There are many different ways to do science,” he says. “We’re at a very exciting time in biology, with new techniques to sequence millions of DNA and RNA molecules. These techniques are proving very powerful.”

Cell death pathway linked to mitochondrial fusion

Cell death pathway linked to mitochondrial fusion

New research led by UC Davis scientists provides insight into why some body organs are more susceptible to cell death than others and could eventually lead to advances in treating or preventing heart attack or stroke.

In a paper published Jan. 21 in the journal Molecular Cell, the UC Davis team and their collaborators at the National Institutes of Health and Johns Hopkins University report that Bax, a factor known to promote cell death, is also involved in regulating the behavior of mitochondria, the structures that provide energy inside living cells.

Mitochondria constantly split and fuse. The proteins that control the splitting of mitochondria also promote a process called apoptosis, or programmed cell death. In contrast, the proteins that control mitochondrial fusion help protect against cell death. Cell death can happen when cells are starved of oxygen, for example during a heart attack or stroke.

Yeast have a single protein that controls outer membrane fusion, but both human and mouse cells have two proteins, called MFN1 and MFN2, which control outer membrane fusion. Using mitochondria from cells derived from genetically modified "knockout" mice, Suzanne Hoppins, a postdoctoral researcher at UC Davis, and Jodi Nunnari, a professor of molecular cell biology, studied how these two proteins work together and the role specific genes play in that process.

The research team discovered that these proteins combine with themselves or each other to form a tether between two mitochondria, leading to fusion. All three combinations -- MFN1/MFN1, MFN1/MFN2 and MFN2/MFN2 -- can promote membrane fusion, but the combination of MFN1/MFN2 is by far the most efficient, Hoppins said.

Hoppins also found that a soluble form of Bax, a protein that triggers apoptosis, can also stimulate mitochondria to fuse. It acts only through the MFN2/MFN2 combination, she found.

The form of Bax that promotes mitochondrial fusion is different from the type that leads to cell death, Nunnari said. Bax leads to cell death when it inserts itself in the mitochondrial membrane. In its soluble, free-floating form, it causes mitochondria to fuse instead.

MFN1 and MFN2 are found in different amounts in different body organs. MFN2 is more abundant in the brain and heart -- tissues where cell death can have disastrous consequences.

The paper shows how MFN2 could act to protect the brain or heart from cell death, by using Bax in a different form, Nunnari said.

"This shows that the fusion machine is both positively and negatively regulated in cells and opens doors to finding the regulatory mechanisms and discovering ways to increase or decrease the sensitivity of cells to apoptosis," Hoppins said. That could lead to new drugs that save cells, for heart disease and stroke, or that kill cells, for cancer.

The genius of bacteria: Scientists develop IQ test to assess and outsmart bacteria's 'social intelligence'

The genius of bacteria: Scientists develop IQ test to assess and outsmart bacteria's 'social intelligence'

Enlarge

This is a "smart community" of Paenibacillus vortex bacteria. Credit: Prof. Eshel Ben-Jacob, Tel Aviv University

IQ scores are used to assess the intelligence of human beings. Now Tel Aviv University has developed a "Social-IQ score" for bacteria ? and it may lead to new antibiotics and powerful bacteria-based "green" pesticides for the agricultural industry.

An international team led by Prof. Eshel Ben-Jacob of Tel Aviv University's Department of Physics and Astronomy and his research student Alexandra Sirota-Madi says that their results deepen science's knowledge of the social capabilities of bacteria, one of the most prolific and important organisms on earth. "Bacteria are our worst enemies but they can also be our best friends. To better exploit their capabilities and to outsmart pathogenic bacteria, we must realize their social intelligence," says Prof. Ben-Jacob.

The international team was first to sequence the genome of pattern-forming bacteria, the Paenibacillus vortex (Vortex) discovered two decades ago by Prof. Ben-Jacob and his collaborators. While sequencing the genome, the team developed the first "Bacteria Social-IQ Score" and found that Vortex and two other Paenibacillus strains have the world's highest Social-IQ scores among all 500 sequenced bacteria. The research was recently published in the journal BMC Genomics.

Highly evolved communities

The impact of the team's research is three-fold. First, it shows just how "smart" bacteria can really be –– a new paradigm that has just begun to be recognised by the science community today. Second, it demonstrates bacteria's high level of social intelligence –– how bacteria work together to communicate and grow. And finally, the work points out some potentially significant applications in medicine and agriculture.

The researchers looked at genes which allow the bacteria to communicate and process information about their environment, making decisions and synthesizing agents for defensive and offensive purposes. This research shows that bacteria are not simple solitary organisms, or "low level" entities, as earlier believed ? they are highly social and evolved creatures. They consistently foil the medical community as they constantly develop strategies against the latest antibiotics. In the West, bacteria are one of the top three killers in hospitals today.

The recent study shows that everyday pathogenic bacteria are not so smart: their S-IQ score is just at the average level. But the social intelligence of the Vortex bacteria is at the "genius range": if compared to human IQ scores it is about 60 points higher than the average IQ at 100. Armed with this kind of information on the social intelligence of bacteria, researchers will be better able to outsmart them, says Prof. Ben-Jacob.

This information can also be directly applied in "green" agriculture or biological control, where bacteria's advanced offense strategies and toxic agents can be used to fight harmful bacteria, fungi and even higher organisms.

Tiny biotechnology factories

Bacteria are often found in soil, and live in symbiotic harmony with a plant's roots. They help the roots access nutrients, and in exchange the bacteria eat sugar from the roots.

For that reason, bacteria are now applied in agriculture to increase the productivity of plants and make them stronger against pests and disease. They can be used instead of fertilizer, and also against insects and fungi themselves. Knowing the Social-IQ score could help developers determine which bacteria are the most efficient.

"Thanks to the special capabilities of our bacteria strain, it can be used by researchers globally to further investigate the social intelligence of bacteria," says co-author Sirota-Madi. "When we can determine how smart they really are, we can use them as biotechnology factories and apply them optimally in agriculture."

Research shows when stem cell descendants lose their versatility

Research shows when stem cell descendants lose their versatility

Becoming hair. To create a new hair follicle (blue), the body taps stem cells from a reservoir called the niche (green). Researchers have determined at what point stem cells irreversibly become hair follicle cells, and have found that these descendent cells send signals back to the niche that regulate the stem cells' activity.

(PhysOrg.com) -- Stem cells are the incomparably versatile progenitors of every cell in our body. Some maintain this remarkable plasticity throughout the life of an animal, prepared to respond as needed to repair an injury, for instance. Others differentiate into specialized cells, regenerating tissue or facilitating some other process before dying. Now new research from Rockefeller University defines the point at which hair follicle stem cells abandon their trademark versatility, or “stemness,” having left their niche to make new hairs. It also shows how these fated stem cell descendants then regulate the activity of their forebears.

“We and others have been focusing on what mobilizes stem cells to make tissue,” says Elaine Fuchs, Rebecca C. Lancefield Professor and head of the Laboratory of Mammalian Cell Biology and Development. “However, it is just as important for stem cells to know when to stop the process, which is what we’ve found here.”

The researchers, led by Ya-Chieh Hsu, a postdoctoral associate in Fuchs’ lab, focused on mouse hair follicles, which undergo cyclical bouts of growth, destruction and rest, a process requiring the activation of stem cells. Stem cells are usually inactive, at rest in their niche, but when activated, they proliferate and leave that niche to make new hairs. In the new research, published last week by Cell, the researchers drilled down on this cycle, defining the point at which activated stem cells become irreversibly committed to becoming the specialized cells needed to grow hair.

Through gene expression analysis and experiments designed to test the cells’ function at different stages in the cycle, the researchers show that early stem cell descendents can retain their stemness and return back to their niche when hair growth stops. In fact, even after their proliferating descendants irreversibly lose their stemness, some can still find their way back to the niche, where they continue to serve two primary purposes: they hold the hairs tightly in place to prevent hair loss, and they release inhibitory signals that prevent the stem cells from activating too early.

“This study shows that committed stem cell descendents transmit inhibitory signals back to the stem cells and return them to a dormant state,” says Fuchs, who is also a Howard Hughes Medical Insititute investigator. “The finding gives us new insights into why our spurts of hair growth are followed by a resting period. For many tissues of the body, such negative feedback loops could provide the necessary signals to prevent tissue overgrowth.”

These findings is work represents a new concept in stem cell biology — that an irreversibly committed cell that is downstream in a stem cell lineage can become an essential regulator of stem cells, the researchers say. In other words, the children tell the parents how to behave. “In many systems, stem cells and their differentiated progeny coexist in close proximity,” Hsu says. “The ability of the progeny to regulate stem cell activity could be a general but previously unrecognized phenomenon which enables stem cells to know when to stop making tissue.”