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  • richardmitnick 11:53 am on July 29, 2017 Permalink | Reply
    Tags: 3D structure of human chromatin, , ChromEMT, , , Genomics, , ,   

    From Salk: “Salk scientists solve longstanding biological mystery of DNA organization” 

    Salk Institute bloc

    Salk Institute for Biological Studies

    July 27, 2017

    Stretched out, the DNA from all the cells in our body would reach Pluto. So how does each tiny cell pack a two-meter length of DNA into its nucleus, which is just one-thousandth of a millimeter across?

    The answer to this daunting biological riddle is central to understanding how the three-dimensional organization of DNA in the nucleus influences our biology, from how our genome orchestrates our cellular activity to how genes are passed from parents to children.

    Now, scientists at the Salk Institute and the University of California, San Diego, have for the first time provided an unprecedented view of the 3D structure of human chromatin—the combination of DNA and proteins—in the nucleus of living human cells.

    In the tour de force study, described in Science on July 27, 2017, the Salk researchers identified a novel DNA dye that, when paired with advanced microscopy in a combined technology called ChromEMT, allows highly detailed visualization of chromatin structure in cells in the resting and mitotic (dividing) stages. By revealing nuclear chromatin structure in living cells, the work may help rewrite the textbook model of DNA organization and even change how we approach treatments for disease.

    “One of the most intractable challenges in biology is to discover the higher-order structure of DNA in the nucleus and how is this linked to its functions in the genome,” says Salk Associate Professor Clodagh O’Shea, a Howard Hughes Medical Institute Faculty Scholar and senior author of the paper. “It is of eminent importance, for this is the biologically relevant structure of DNA that determines both gene function and activity.”

    2
    A new technique enables 3D visualization of chromatin (DNA plus associated proteins) structure and organization within a cell nucleus (purple, bottom left) by painting the chromatin with a metal cast and imaging it with electron microscopy (EM). The middle block shows the captured EM image data, the front block illustrates the chromatin organization from the EM data, and the rear block shows the contour lines of chromatin density from sparse (cyan and green) to dense (orange and red). Credit: Salk Institute.

    Ever since Francis Crick and James Watson determined the primary structure of DNA to be a double helix, scientists have wondered how DNA is further organized to allow its entire length to pack into the nucleus such that the cell’s copying machinery can access it at different points in the cell’s cycle of activity. X-rays and microscopy showed that the primary level of chromatin organization involves 147 bases of DNA spooling around proteins to form particles approximately 11 nanometers (nm) in diameter called nucleosomes. These nucleosome “beads on a string” are then thought to fold into discrete fibers of increasing diameter (30, 120, 320 nm etc.), until they form chromosomes. The problem is, no one has seen chromatin in these discrete intermediate sizes in cells that have not been broken apart and had their DNA harshly processed, so the textbook model of chromatin’s hierarchical higher-order organization in intact cells has remained unverified.

    To overcome the problem of visualizing chromatin in an intact nucleus, O’Shea’s team screened a number of candidate dyes, eventually finding one that could be precisely manipulated with light to undergo a complex series of chemical reactions that would essentially “paint” the surface of DNA with a metal so that its local structure and 3D polymer organization could be imaged in a living cell. The team partnered with UC San Diego professor and microscopy expert Mark Ellisman, one of the paper’s coauthors, to exploit an advanced form of electron microscopy that tilts samples in an electron beam enabling their 3D structure to be reconstructed. By combining their chromatin dye with electron-microscope tomography, they created ChromEMT.

    The team used ChromEMT to image and measure chromatin in resting human cells and during cell division when DNA is compacted into its most dense form—the 23 pairs of mitotic chromosomes that are the iconic image of the human genome. Surprisingly, they did not see any of the higher-order structures of the textbook model anywhere.

    3
    From left: Horng Ou and Clodagh O’Shea. Credit: Salk Institute.

    “The textbook model is a cartoon illustration for a reason,” says Horng Ou, a Salk research associate and the paper’s first author. “Chromatin that has been extracted from the nucleus and subjected to processing in vitro—in test tubes—may not look like chromatin in an intact cell, so it is tremendously important to be able to see it in vivo.”

    What O’Shea’s team saw, in both resting and dividing cells, was chromatin whose “beads on a string” did not form any higher-order structure like the theorized 30 or 120 or 320 nanometers. Instead, it formed a semi-flexible chain, which they painstakingly measured as varying continuously along its length between just 5 and 24 nanometers, bending and flexing to achieve different levels of compaction. This suggests that it is chromatin’s packing density, and not some higher-order structure, that determines which areas of the genome are active and which are suppressed.

    With their 3D microscopy reconstructions, the team was able to move through a 250 nm x 1000 nm x 1000 nm volume of chromatin’s twists and turns, and envision how a large molecule like RNA polymerase, which transcribes (copies) DNA, might be directed by chromatin’s variable packing density, like a video game aircraft flying through a series of canyons, to a particular spot in the genome. Besides potentially upending the textbook model of DNA organization, the team’s results suggest that controlling access to chromatin could be a useful approach to preventing, diagnosing and treating diseases such as cancer.

    “We show that chromatin does not need to form discrete higher-order structures to fit in the nucleus,” adds O’Shea. “It’s the packing density that could change and limit the accessibility of chromatin, providing a local and global structural basis through which different combinations of DNA sequences, nucleosome variations and modifications could be integrated in the nucleus to exquisitely fine-tune the functional activity and accessibility of our genomes.”

    Future work will examine whether chromatin’s structure is universal among cell types or even among organisms.

    Other authors included Sébastien Phan, Thomas Deerinck and Andrea Thor of the UC San Diego.

    The work was largely funded by the W. M. Keck Foundation, the NIH 4D Nucleome Roadmap Initiative and the Howard Hughes Medical Institute, with additional support from the William Scandling Trust, the Price Family Foundation and the Leona M. and Harry B. Helmsley Charitable Trust.

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    Every cure has a starting point. Like Dr. Jonas Salk when he conquered polio, Salk scientists are dedicated to innovative biological research. Exploring the molecular basis of diseases makes curing them more likely. In an outstanding and unique environment we gather the foremost scientific minds in the world and give them the freedom to work collaboratively and think creatively. For over 50 years this wide-ranging scientific inquiry has yielded life-changing discoveries impacting human health. We are home to Nobel Laureates and members of the National Academy of Sciences who train and mentor the next generation of international scientists. We lead biological research. We prize discovery. Salk is where cures begin.

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  • richardmitnick 10:26 am on July 20, 2017 Permalink | Reply
    Tags: , Elizabeth Davis spent 21 years trying to receive a correct diagnosis from doctors about her condition which prevented her toes from uncurling causing her to walk with crutches for the most of her life, , Genomics, GTPCH1 impairs her ability to produce dopa, , Mutagenesis, NCGENES project, They were able to treat it — with something as simple as a pill. A pill that has been on the market since 1988 used to treat patients with Parkinson’s disease,   

    From UNC: “The Cure Code” 

    University of North Carolina

    July 18th, 2017
    Alyssa LaFaro

    1
    Davis can now walk fully unsupported and live a relatively normal life thanks to a correct diagnosis from UNC researchers within the NCGENES project. No image credit.

    “Consider this: In 1969, if a disease-linked gene was found in humans, scientists had no simple means to understand the nature of the mutation, no mechanism to compare the altered gene to normal form, and no obvious method to reconstruct the gene mutation in a different organism to study its function. By 1979, that same gene could be shuttled into bacteria, spliced into a viral vector, delivered into the genome of a mammalian cell, cloned, sequenced, and compared to the normal form.” —Siddhartha Mukherjee, “The Gene: An Intimate History”

    “I can move my toes,” Elizabeth Davis says.

    Her 9-year-old son looks at her in awe. The two stand, wide-eyed in the middle of a Verizon Wireless store in Goldsboro, North Carolina. Davis leans hard against her crutches, staring at her feet. She looks up and smiles.

    At age 37 — for the first time in 31 years — Davis can uncurl her toes from a locked position, the symptom of a condition gone misdiagnosed for just as long. Three months later, she sheds her crutches, walking fully unsupported — something she hasn’t done since she was 14 years old.

    In 1975, the same year Davis was born, UNC microbiologists Clyde Hutchison and Marshall Edgell experienced a different kind of life-changing event. They’d been working rigorously to isolate DNA within the smallest-known virus at the time, Phi-X174. More than anything, they wanted to understand how to read the genetic code. Then, later that year and across the pond at St. John’s College in Cambridge, Fred Sanger figured it out. The British biochemist became the first person to develop a relatively rapid method for sequencing DNA, a discovery that won him a Nobel Prize in Chemistry — for the second time.

    In response to Sanger’s discovery, Hutchison took a sabbatical and headed to England to work in his lab. During his first year there, he helped uncover the entire sequence of Phi-X174 — the first time this had been done for any organism. While there, he realized the new ability to read DNA could help him and Edgell solve a different problem they’d been having back in North Carolina: fusing two pieces of DNA code together to create an entirely different sequence.

    After returning to Chapel Hill, Hutchison continued his work with Edgell and also Michael Smith, a researcher at the University of British Columbia who he met while working in Sanger’s lab. Together, the trio successfully fused two differing DNA strands using a more flexible approach to site-directed mutagenesis — a technique that makes gene therapy possible today. They published their results in 1978. Smith would go on to receive the Nobel Prize for this work in 1992.

    —-

    The scientific breakthroughs of the 1970s changed the field of genetics forever. In 1980, Sanger received the Nobel Prize for Chemistry for his contributions, along with Walter Gilbert (Harvard), who discovered that individual modules from different genes mix and match to construct entirely new genes; and Paul Berg (Stanford), who developed a technique for splicing recombinant DNA.

    Meanwhile, researchers in Chapel Hill continued to chip away at the mysteries of the gene. Oliver Smithies, who came to UNC in 1987, would later win the Nobel Prize for his work in gene targeting using mouse models. That same year, UNC cancer geneticist Michael Swift and team discover the AT gene, which predisposes women to breast cancer; and George McCoy becomes the first clinical trial participant in the world to receive the genetically engineered Factor VIII gene to treat his hemophilia at the then UNC-Thrombosis and Hemostasis Center.

    Genetics was changing the world. And this was only the beginning.

    An unsolved mystery

    One year after Sanger won the Nobel Prize, Elizabeth Davis turned 6. She soon began walking on her toes, which had suddenly, one day, curled under in pain, making it nearly impossible for her to stride with feet flat on the ground. Her knees knocked together as she struggled to move with the swift pace characteristic of a child her age. Davis continued to walk on her toes for years.

    “I would even brace the school walls when walking down the hallway,” she says. Eventually, the pain became unbearable. By the time she was 12, she’d resigned herself to crutches.

    Doctors believed Davis’ condition could be treated with foot surgery, misdiagnosing her condition for years. By age 14, she had already undergone three procedures — two to lengthen her Achilles tendons and an experimental bone fusion. But each surgery offered little to no relief, and walking only grew more painful for Davis, both physically and emotionally. As her condition worsened, her classmates became cruel — so much so that she dropped out of high school when she was just 16.

    By age 20, Davis grew restless. “The pain was constant,” she remembers. “I could hardly move my legs — they just felt weak. I would drag them behind me as I used my crutches. I couldn’t even lift them.” Doctors suggested she undergo a third Achilles tendon lengthening surgery, the result of which minimally improved her condition.

    “By that age, I just wanted more,” Davis says. “I just wanted to do things, to go places. I wanted the surgery to work. But it didn’t. And the pain continued.”

    It would be another 17 years before doctors realized the problem was hidden in her genome.

    The birth of a department

    In 1990, the start of the Human Genome Project — an international research program to map out the 20,000 genes that define human beings — further fueled new discoveries in the field of genetics. So when Jeff Houpt, then-UNC School of Medicine dean, formed a research advisory committee in 1997 and asked his faculty what the number-one research program the university needed to focus on, they responded: genetics and genome sciences.

    Great minds think alike. At the same time, the College of Arts and Sciences was also hosting its own committee that vied to develop a genetics department. “At this point, I had a vision for a pan-university program,” Houpt shares. “This wasn’t just going to be a program of the medical school.”

    Along with the College, the schools of public health, dentistry, pharmacy, nursing, and information and library science all wanted in, offering financial assistance to the program. Then-Provost Robert Shelton and Chancellor James Moeser both signed off on it as well. “What we wanted from Shelton and Moeser was more money and more positions,” Houpt remembers. “And they agreed to that.”

    By 2000, a hiring committee was ready to interview candidates to chair the new department and genomics center. Terry Magnuson quickly emerged as the lead candidate. He and his team had spent the past 16 years researching developmental abnormalities using genetics and mouse models, successfully changing the genetic background of a mutated gene.

    “It was obvious he was going to have a following,” Houpt remembers. “People were going to listen to him because he’s a good scientist. But more than that, it was pretty clear that Terry was interested in building a program, and this university-wide effort appealed to him.”

    Unanswered pain

    By the time she reached her 30s, Davis’ condition had spread to her arms. She underwent multiple MRIs, nerve and muscle testing, and a spinal tap. She even endured a fifth, unsuccessful surgery on her feet. Physicians misdiagnosed her yet again. A few believed she suffered from hereditary spastic paraplegia, a genetic condition that causes weakness in the legs and hips. Another told her she had cerebral palsy. “But I didn’t want to believe him,” she says — and it’s a good thing she didn’t.

    As Davis continued her search for answers, walking grew more and more painful. “I was always in pain,” she admits. “But some weeks were really, really bad — to the point where I couldn’t even move.” She finally succumbed to the assistance of a wheelchair. “I hated it so much. I barely went anywhere.” And when she did, she needed help.

    Her mother assisted her regularly with everyday tasks like grocery shopping. Her youngest son, Alex, learned to expertly navigate her around high school gyms, baseball fields, and the local YMCA pool so she could watch her other son, Myles, compete in the plethora of sports he participated in.

    “Myles really experienced the worst of it,” Davis says. “I remember one time, in particular. I was taking a shower and knew I was about to fall. I called for him and he came running. He was always there to pick me back up.”

    Sequences and algorithms

    After the Human Genome Project published its results in 2004, genomic sequencing became an option for people with undiagnosed diseases. But analyzing and understanding the 3 billion base pairs that make up a person’s genetic identity was an expensive process. As time progressed and technology improved, though, the technique became more manageable for both physicians and patients.

    Using these new genomic technologies for outpatient care intrigued UNC geneticists James Evans and Jonathan Berg. In 2009, after gathering enough preliminary data, the NIH granted the team the funds to start the North Carolina Clinical Genomic Evaluation by NextGen Exome Sequencing (NCGENES), which uses whole exome sequencing (WES) to uncover the root cause of undiagnosed diseases. Using just two tablespoons of blood, WES tests 1 percent of the genome — a feat that is both miraculous and controversial, creating a whole new wave of ethical questions.

    Simply put: “Some people want information that other people don’t,” Evans explains. Most people want to know about genetic disorders that have treatment options, but when it comes to those that don’t, they’d rather not hear it. “Navigating those different viewpoints can be a challenge,” he says. Privacy and confidentiality also present problems within the insurance world. Although protections exist in the realm of medical insurance, major genetic predispositions could have large implications for life, disability, and long-term care insurance.

    Today, upward of 50 researchers from across Carolina participate in NCGENES to study everything from the protection of data to the delivery of results. More than 750 people with undiagnosed diseases have undergone testing.

    NCGENES wouldn’t exist without the technical infrastructure that tracks, categorizes, and helps analyze genetic material as it makes its way through multiple laboratories — all of which is provided by UNC’s Renaissance Computing Institute (RENCI). A developer of data science cyberinfrastructure, RENCI provides the software programming that helps the team at NCGENES analyze genomes more effectively.

    “You need new computer algorithms to solve new science problems,” RENCI Director Stan Ahalt says. “It takes a multidisciplinary team to understand science problems like genetics — and computer code to make that process go fast.”

    A transformative experience

    By 2013, Davis was in desperate need of a new algorithm. Thankfully, that year, she was referred to Jane Fan, a pediatric neurologist at UNC. After studying Davis’ file, Fan felt sure that the doctors who tried to diagnose her condition failed, making her the perfect candidate for NCGENES.

    Four tubes of blood, 100,000 possible genetic locations, and just over six months later, Fan called Davis. A single gene mutation called GTPCH1 impairs her ability to produce dopa, an amino acid crucial for nervous system function. “I had to hear it in person before I believed it,” Davis admits. “I had been misdiagnosed many times before.”

    Not only were UNC geneticist James Evans and his NCGENES team finally able to accurately diagnose Davis, but they were able to treat it — with something as simple as a pill. A pill that has been on the market since 1988, used to treat patients with Parkinson’s disease.

    And just like that, Davis ‘life was changed forever by genome sequencing.

    Three days after she took one-quarter of a pill, movement returned to her toes while standing in the middle of a Verizon Wireless store in Goldsboro. She began to cry.

    Top-five in the country

    UNC’s genetics department has ranked in the top-five programs for NIH funding across the nation every year since 2012 (and top-10 each year since 2006). “I think we’ve built one of the best genetics departments in the country,” Magnuson says. In 2016 alone, genetics department faculty brought $38 million to Carolina.

    Houpt agrees with Magnuson’s sentiment. “The genetics department is a great example of how universities should run,” he says. “People need to put aside their own interests and see what’s needed. Terry is a leader who’s made each school involved feel like it’s their program and not just a medical school program – which is why he’s now the vice chancellor for research.”

    Today, more than 80 faculty members from across campus conduct world-recognized genetics research in multiple disciplines.

    Ned Sharpless, for example, focuses on cancer. Most recently, the director of the UNC Lineberger Comprehensive Cancer Center lead a study that paired UNCseq — a genetic sequencing protocol that produces volumes of genetic information from a patient’s tumor — with IBM Watson’s ability to quickly pull information from millions of medical papers. A procedure much too intense and time-consuming for the human mind, this data analysis can help physicians make more informed decisions about patient care.

    Another member of Carolina’s Cancer Genetics Program, Charles Perou uses genomics to characterize the diversity of breast cancer tumors — research that helps doctors guarantee patients more individualized care. In 2011, he cofounded GeneCentric, which uses personalized molecular diagnostic assays and targeted drug development to treat cancer.

    In 2015, geneticist Aravind Asokan started StrideBio with University of Florida biochemist Mavis Agbandje-McKenna. The gene therapy company develops novel adeno-associated viral (AAV) vector technologies for treating rare diseases. Although still in its infancy, the company has already partnered with CRISPR Therapeutics and received an initial investment from Hatteras Venture Partners. Asokan has spent nearly a decade studying AAV — and even helped to, previously, cofound Bamboo Therapeutics, acquired by Pfizer for $645 million just last year.

    In 2016, current genetics department Chair Fernando Pardo-Manuel de Villena challenged both Darwin’s theory of natural selection and Mendel’s law of segregation through researching a mouse gene called R2d2. In doing so, he found that a selfish gene can become fixed in a population of organisms while, at the same time, being detrimental to “reproductive fitness” — a discovery that shows the swiftness at which the genome can change, creating implications for an array of fields from basic biology to agriculture and human health.

    A former student of Oliver Smithies, Beverly Koller uses gene targeting in mice to better understand diseases like cystic fibrosis, asthma, and arthritis — research that will ultimately lead to better treatments. Similarly, Mark Heise observes mice to study diseases caused by viruses including infectious arthritis and encephalitis (inflammation of the brain). Both researchers are part of the Collaborative Cross project, a large panel of inbred mouse strains that help map genetic traits — a resource that is UNC lead, according to Magnuson.

    Genetics research stems far beyond the UNC School of Medicine. In 2009, for example, chemist Kevin Weeks and his research team decoded the HIV genome, advancing the development of new therapies and treatments. UNC sociologist Gail Henderson runs the Center for Genomics and Society, which provides research and training on ethical, legal, and social implications of genomic research. In 2015, UNC Eshelman School of Pharmacy Dean Bob Blouin helped the school become the first U.S. hub to join the international Structural Genomics Consortium — focused on discovering selective, small molecules and protein kinases to help speed the creation of new medicines for patients.

    From crutches to a 5K

    After just three months of treatment, Davis walked fully unsupported for the first time since she was 6 years old. She’s since traversed Hershey Park in Pennsylvania, strolled around the World Trade Center in New York, and regularly participated in yoga and spin classes. This past May, she walked her first 5K. “I have crazy endurance,” she says. “When your body feels good, you just want to keep on going.”

    Perhaps, more importantly, Davis is able attend Alex’s sports games without assistance. “When I used to walk into the gym on crutches to watch my oldest son play basketball, everyone would look at my crutches and my legs,” she says. “Now, when I go watch my youngest son play, I have so much more confidence walking in to the gym. People see me.”

    See the full article here .

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    Carolina’s vibrant people and programs attest to the University’s long-standing place among leaders in higher education since it was chartered in 1789 and opened its doors for students in 1795 as the nation’s first public university. Situated in the beautiful college town of Chapel Hill, N.C., UNC has earned a reputation as one of the best universities in the world. Carolina prides itself on a strong, diverse student body, academic opportunities not found anywhere else, and a value unmatched by any public university in the nation.

     
  • richardmitnick 8:47 am on July 17, 2017 Permalink | Reply
    Tags: , CRISPR-Cas3, , , Genomics, ,   

    From HMS: “Bringing CRISPR into Focus” 

    Harvard University

    Harvard University

    Harvard Medical School

    Harvard Medical School

    June 29, 2017
    KEVIN JIANG

    CRISPR-Cas3 is a subtype of the CRISPR-Cas system, a widely adopted molecular tool for precision gene editing in biomedical research. Aspects of its mechanism of action, however, particularly how it searches for its DNA targets, were unclear, and concerns about unintended off-target effects have raised questions about the safety of CRISPR-Cas for treating human diseases.

    Harvard Medical School and Cornell University scientists have now generated near-atomic resolution snapshots of CRISPR that reveal key steps in its mechanism of action. The findings, published in Cell on June 29, provide the structural data necessary for efforts to improve the efficiency and accuracy of CRISPR for biomedical applications.

    Through cryo-electron microscopy, the researchers describe for the first time the exact chain of events as the CRISPR complex loads target DNA and prepares it for cutting by the Cas3 enzyme. These structures reveal a process with multiple layers of error detection—a molecular redundancy that prevents unintended genomic damage, the researchers say.

    High-resolution details of these structures shed light on ways to ensure accuracy and avert off-target effects when using CRISPR for gene editing.

    “To solve problems of specificity, we need to understand every step of CRISPR complex formation,” said Maofu Liao, assistant professor of cell biology at Harvard Medical School and co-senior author of the study. “Our study now shows the precise mechanism for how invading DNA is captured by CRISPR, from initial recognition of target DNA and through a process of conformational changes that make DNA accessible for final cleavage by Cas3.”

    Target search

    Discovered less than a decade ago, CRISPR-Cas is an adaptive defense mechanism that bacteria use to fend off viral invaders. This process involves bacteria capturing snippets of viral DNA, which are then integrated into its genome and which produce short RNA sequences known as crRNA (CRISPR RNA). These crRNA snippets are used to spot “enemy” presence.

    Acting like a barcode, crRNA is loaded onto members of the CRISPR family of enzymes, which perform the function of sentries that roam the bacteria and monitor for foreign code. If these riboprotein complexes encounter genetic material that matches its crRNA, they chop up that DNA to render it harmless. CRISPR-Cas subtypes, notably Cas9, can be programmed with synthetic RNA in order to cut genomes at precise locations, allowing researchers to edit genes with unprecedented ease.

    To better understand how CRISPR-Cas functions, Liao partnered with Ailong Ke of Cornell University. Their teams focused on type 1 CRISPR, the most common subtype in bacteria, which utilizes a riboprotein complex known as CRISPR Cascade for DNA capture and the enzyme Cas3 for cutting foreign DNA.

    Through a combination of biochemical techniques and cryo-electron microscopy, they reconstituted stable Cascade in different functional states, and further generated snapshots of Cascade as it captured and processed DNA at a resolution of up to 3.3 angstroms—or roughly three times the diameter of a carbon atom.

    1
    A sample cryo-electron microscope image of CRISPR molecules(left). The research team combined hundreds of thousands of particles into 2D averages (right), before turning them into 3D projections. Image: Xiao et al.

    Seeing is believing

    In CRISPR-Cas3, crRNA is loaded onto CRISPR Cascade, which searches for a very short DNA sequence known as PAM that indicates the presence of foreign viral DNA.

    Liao, Ke and their colleagues discovered that as Cascade detects PAM, it bends DNA at a sharp angle, forcing a small portion of the DNA to unwind. This allows an 11-nucleotide stretch of crRNA to bind with one strand of target DNA, forming a “seed bubble.”

    The seed bubble acts as a fail-safe mechanism to check whether the target DNA matches the crRNA. If they match correctly, the bubble is enlarged and the remainder of the crRNA binds with its corresponding target DNA, forming what is known as an “R-loop” structure.

    Once the R-loop is completely formed, the CRISPR Cascade complex undergoes a conformational change that locks the DNA into place. It also creates a bulge in the second, non-target strand of DNA, which is run through a separate location on the Cascade complex.

    Only when a full R-loop state is formed does the Cas3 enzyme bind and cut the DNA at the bulge created in the non-target DNA strand.

    The findings reveal an elaborate redundancy to ensure precision and avoid mistakenly chopping up the bacteria’s own DNA.

    2
    CRISPR forms a “seed bubble” state, which acts as an initial fail-safe mechanism to ensure that CRISPR RNA matches its target DNA. Image: Liao Lab/HMS.

    “To apply CRISPR in human medicine, we must be sure the system is accurate and that it does not target the wrong genes,” said Ke, who is co-senior author of the study. “Our argument is that the CRISPR-Cas3 subtype has evolved to be a precise system that carries the potential to be a more accurate system to use for gene editing. If there is mistargeting, we know how to manipulate the system because we know the steps involved and where we might need to intervene.”

    Setting the sights

    Structures of CRISPR Cascade without target DNA and in its post-R-loop conformational states have been described, but this study is the first to reveal the full sequence of events from seed bubble formation to R-loop formation at high resolution.

    In contrast to the scalpel-like Cas9, CRISPR-Cas3 acts like a shredder that chews DNA up beyond repair. While CRISPR-Cas3 has, thus far, limited utility for precision gene editing, it is being developed as a tool to combat antibiotic-resistant strains of bacteria. A better understanding of its mechanisms may broaden the range of potential applications for CRISPR-Cas3.

    In addition, all CRISPR-Cas subtypes utilize some version of an R-loop formation to detect and prepare target DNA for cleavage. The improved structural understanding of this process can now enable researchers to work toward modifying multiple types of CRISPR-Cas systems to improve their accuracy and reduce the chance of off-target effects in biomedical applications.

    “Scientists hypothesized that these states existed but they were lacking the visual proof of their existence,” said co-first author Min Luo, postdoctoral fellow in the Liao lab at HMS. “The main obstacles came from stable biochemical reconstitution of these states and high-resolution structural visualization. Now, seeing really is believing.”

    “We’ve found that these steps must occur in a precise order,” Luo said. “Evolutionarily, this mechanism is very stringent and has triple redundancy, to ensure that this complex degrades only invading DNA.”

    Additional authors on the study include Yibei Xiao, Robert P. Hayes, Jonathan Kim, Sherwin Ng, and Fang Ding.

    This work is supported by National Institutes of Health grants GM 118174 and GM102543.

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    Harvard is the oldest institution of higher education in the United States, established in 1636 by vote of the Great and General Court of the Massachusetts Bay Colony. It was named after the College’s first benefactor, the young minister John Harvard of Charlestown, who upon his death in 1638 left his library and half his estate to the institution. A statue of John Harvard stands today in front of University Hall in Harvard Yard, and is perhaps the University’s best known landmark.

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  • richardmitnick 1:21 pm on July 8, 2017 Permalink | Reply
    Tags: , , , , , , Genomics, , , UCSD Comet supercomputer   

    From Science Node: “Cracking the CRISPR clock” 

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    Science Node

    05 Jul, 2017
    Jan Zverina

    SDSC Dell Comet supercomputer

    Capturing the motion of gyrating proteins at time intervals up to one thousand times greater than previous efforts, a team led by University of California, San Diego (UCSD) researchers has identified the myriad structural changes that activate and drive CRISPR-Cas9, the innovative gene-splicing technology that’s transforming the field of genetic engineering.

    By shedding light on the biophysical details governing the mechanics of CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats) activity, the study provides a fundamental framework for designing a more efficient and accurate genome-splicing technology that doesn’t yield ‘off-target’ DNA breaks currently frustrating the potential of the CRISPR-Cas9- system, particularly for clinical uses.


    Shake and bake. Gaussian accelerated molecular dynamics simulations and state-of-the-art supercomputing resources reveal the conformational change of the HNH domain (green) from its inactive to active state. Courtesy Giulia Palermo, McCammon Lab, UC San Diego.

    “Although the CRISPR-Cas9 system is rapidly revolutionizing life sciences toward a facile genome editing technology, structural and mechanistic details underlying its function have remained unknown,” says Giulia Palermo, a postdoctoral scholar with the UC San Diego Department of Pharmacology and lead author of the study [PNAS].

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    Science Node is an international weekly online publication that covers distributed computing and the research it enables.

    “We report on all aspects of distributed computing technology, such as grids and clouds. We also regularly feature articles on distributed computing-enabled research in a large variety of disciplines, including physics, biology, sociology, earth sciences, archaeology, medicine, disaster management, crime, and art. (Note that we do not cover stories that are purely about commercial technology.)

    In its current incarnation, Science Node is also an online destination where you can host a profile and blog, and find and disseminate announcements and information about events, deadlines, and jobs. In the near future it will also be a place where you can network with colleagues.

    You can read Science Node via our homepage, RSS, or email. For the complete iSGTW experience, sign up for an account or log in with OpenID and manage your email subscription from your account preferences. If you do not wish to access the website’s features, you can just subscribe to the weekly email.”

     
  • richardmitnick 2:36 pm on July 5, 2017 Permalink | Reply
    Tags: A Whole-Genome Sequenced Rice Mutant Resource for the Study of Biofuel Feedstocks, , Fast-neutron irradiation causes different types of mutations, , Genomics, Kitaake: a model rice variety with a short life cycle,   

    From LBNL: “A Whole-Genome Sequenced Rice Mutant Resource for the Study of Biofuel Feedstocks” 

    Berkeley Logo

    Berkeley Lab

    July 5, 2017
    Sarah Yang
    scyang@lbl.gov
    (510) 486-4575

    JBEI researchers create open-access web portal to accelerate functional genetic research in plants.

    1
    Genome-wide distribution of fast-neutron-induced mutations in the Kitaake rice mutant population (green). Even distribution of mutations is important to achieve saturation of the genome. Colored lines (center) represent translocations of DNA fragments from one chromosome to another. (Credit: Guotian Li and Rashmi Jain/Berkeley Lab).

    Rice is a staple food for over half of the world’s population and a model for studies of candidate bioenergy grasses such as sorghum, switchgrass, and Miscanthus. To optimize crops for biofuel production, scientists are seeking to identify genes that control key traits such as yield, resistance to disease, and water use efficiency.

    Populations of mutant plants, each one having one or more genes altered, are an important tool for elucidating gene function. With whole-genome sequencing at the single nucleotide level, researchers can infer the functions of the genes by observing the gain or loss of particular traits. But the utility of existing rice mutant collections has been limited by several factors, including the cultivars’ relatively long six-month life cycle and the lack of sequence information for most of the mutant lines.

    In a paper published in The Plant Cell, a team led by Pamela Ronald, a professor in the Genome Center and the Department of Plant Pathology at UC Davis and director of Grass Genetics at the Department of Energy’s (DOE’s) Joint BioEnergy Institute (JBEI), with collaborators from UC Davis and the DOE Joint Genome Institute (JGI), reported the first whole-genome sequenced fast-neutron induced mutant population of Kitaake, a model rice variety with a short life cycle.

    Kitaake (Oryza sativa L. ssp. japonica) completes its life cycle in just nine weeks and is not sensitive to photoperiod changes. This novel collection will accelerate functional genetic research in rice and other monocots, a type of flowering plant species that includes grasses.

    “Some of the most popular rice varieties people use right now only have two generations per year. Kitaake has up to four, which really speeds up functional genomics work,” said Guotian Li, a project scientist at Lawrence Berkeley National Laboratory (Berkeley Lab) and deputy director of Grass Genetics at JBEI.

    In a previously published pilot study [Molecular Plant], Li, Mawsheng Chern, and Rashmi Jain, co-first authors on The Plant Cell paper, demonstrated that fast-neutron irradiation produced abundant and diverse mutations in Kitaake, including single base substitutions, deletions, insertions, inversions, translocations, and duplications. Other techniques that have been used to generate rice mutant populations, such as the insertion of gene and chromosome segments and the use of gene editing tools like CRISPR-Cas9, generally produce a single type of mutation, Li noted.

    “Fast-neutron irradiation causes different types of mutations and gives different alleles of genes so we really can get something that’s not achievable from other collections,” he said.

    Whole-genome sequencing of this mutant population – 1,504 lines in total with 45-fold coverage – allowed the researchers to pinpoint each mutation at a single-nucleotide resolution. They identified 91,513 mutations affecting 32,307 genes, 58 percent of all genes in the roughly 389-megabase rice genome. A high proportion of these were loss-of-function mutations.

    Using this mutant collection, the Grass Genetics group identified an inversion affecting a single gene as the causative mutation for the short-grain phenotype in one mutant line with a population containing just 50 plants. In contrast, researchers needed more than 16,000 plants to identify the same gene using the conventional approach.

    “This comparison clearly demonstrates the power of the sequenced mutant population for rapid genetic analysis,” said Ronald.

    This high-density, high-resolution catalog of mutations, developed with JGI’s help, provides researchers opportunities to discover novel genes and functional elements controlling diverse biological pathways. To facilitate open access to this resource, the Grass Genetics group has established a web portal called KitBase, which allows users to find information related to the mutant collection, including sequence, mutation and phenotypic data for each rice line. Additional information about the database can be found through JGI.

    Additional Berkeley Lab scientists who contributed to this work include co-first authors Rashmi Jain and Mawsheng Chern; Tong Wei and Deling Ruan, both affiliated with JBEI’s Feedstocks Division and with Berkeley Lab’s Environmental Genomics and Systems Biology Division; Nikki Pham and Kyle Jones of JBEI’s Feedstocks Division; and Joel Martin, Wendy Schackwitz, Anna Lipzen, Diane Bauer, Yi Peng, and Kerrie Barry of the JGI.

    Support for the research at JBEI, a DOE Bioenergy Research Center, and JGI, a DOE Office of Science User Facility, was provided by DOE’s Office of Science. Additional support was provide by the National Institutes of Health and the National Science Foundation.

    See the full article here .

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  • richardmitnick 12:20 pm on June 5, 2017 Permalink | Reply
    Tags: and personalized therapies, , DiscovEHR, , Genomics, Harvard Graduate School of Arts and Sciences, Learning to read your DNA, Population sequencing lands a knockout punch, Precision medicine, Scaling up genetic sequencing studies, The future: population sequencing   

    From Harvard: “Strength in Numbers: genetic sequencing of large populations is shaping the future of medicine” 

    Harvard University

    Harvard University

    June 5, 2017
    Ryan L. Collins
    Figures by Brad Wierbowski

    Thanks to modern genetics, “precision medicine” is slowly becoming a reality: doctors can perform genetic tests to determine your risk for dozens of diseases, like stroke or liver disease, and can prescribe treatments or therapies tailored to your individual genetic makeup. Yet before doctors can provide you with precision medicine in practice, they first need to understand the genes of tens of thousands of other people. Excitingly, recent breakthroughs in genetics research have made it possible to study the genes of whole populations at once, and the lessons we are learning from those studies are rapidly changing our approach to diagnosing and treating disease.

    Learning to read your DNA

    You can picture your genome as a book with 3.1 billion letters, known as nucleotides, that encode a list of the ~20,000 different molecular parts, or proteins, that comprise every cell in your body. Much like a book with 3.1 billion letters, your genome isn’t exactly a light read: for example, the first time a human genome was ever read (“sequenced”) in its entirety required the combined efforts of more than 200 scientists, a timespan of 12 years (finally finishing in 2003), and a staggering total cost of $2.7 billion.

    In the subsequent 14 years, massive advances in sequencing technologies [NIH] [National Human Genome Research Institute] have transformed scientists into genetic speed-readers, with cutting-edge sequencing methods able to process your entire genome in under two days for a mere ~$1,500. Additional technologies have improved our efficiency by allowing targeted sequencing of the bits of your genome that spell out the blueprints for all human proteins, known as the exome. Surprisingly, the exome takes up barely more than ~1% of all of the nucleotides in your whole genome, which means exome sequencing is both faster and cheaper than genome sequencing (see Figure 1 for a comparison of exome vs genome sequencing). While the other ~99% of your DNA plays various “helper” functions in your cells, it does not code for proteins. Since proteins are the most important cellular building blocks, and thus the most important determinants of disease, exome sequencing is a great way to hone in on the most critical sections of your DNA.

    2
    CDC

    Scaling up genetic sequencing studies

    The development of these groundbreaking sequencing technologies has opened countless promising research avenues. Not least among these is an effort known as “population sequencing,” or the process of sequencing the exomes or genomes of entire human communities, illustrated in Figure 2. Early examples of population sequencing, including the 1000 Genomes Project or the Exome Sequencing Project, combined genetic data from thousands of volunteers into vast datasets bursting with new knowledge about human biology.

    3
    Figure 2: Overview of population sequencing. Studying the genomes of many thousands of people at once, known as population sequencing, is an exciting area of research and is producing countless new insights into human biology and medicine. Typically, a population sequencing experiment will involve sequencing the exomes or genomes of large groups of individuals with and without a trait, such as a disease like cancer, then comparing the differences in the genes of the two groups. Once researchers identify DNA changes associated with the trait of interest, they can then use that information to answer many important questions that help guide drug development, clinical practice, and therapeutic selection.

    The initial successes of these population sequencing projects triggered a tidal wave of similar studies. Dozens of research groups rushed to apply similar methods, and the results came pouring in. For example, one 2014 study [Nature Genetics] sequenced the genomes of several thousand people from Iceland, identifying specific genes that may predispose Icelanders to early-onset heart disease and liver disease. As a second example, multiple studies [Nature] have used exome and genome sequencing in children to pinpoint over 60 genes strongly linked to autism. The list of these sequencing success stories is already lengthy, and continues to grow every month. Even more importantly, sequencing studies like these produce the information doctors and researchers need to screen your genome and provide a more complete picture of how your genes influence your individual health.

    Data sharing: ExACtly what the doctor ordered

    As of 2017, well over one million human exomes and genomes have been sequenced worldwide. In the realm of human genetics, bigger datasets are almost always more informative, so analyzing all of these data together seems like an obvious choice. These population-scale studies depend on volunteers like you to contribute their DNA, but unfortunately this process isn’t always straightforward. Genetic data sharing, even if performed strictly in a research context where no personal health information is transferred, is still fraught with ethical, legal, practical, and bureaucratic hurdles.

    In 2014, a large international alliance of researchers, led by Daniel MacArthur at The Broad Institute of M.I.T. and Harvard, set out to tackle these obstacles. They formed a collaborative group known as the Exome Aggregation Consortium (abbreviated “ExAC”), and combined exome sequences from over 60,000 healthy individuals from more than two-dozen independent studies conducted around the world to build a dataset nearly ten times larger than any other ever assembled. Their results, reported in the journal Nature, outlined the most comprehensive atlas of human genetic diversity to date, including individuals from nearly all major global populations and uncovering nearly five-and-a-half million genetic changes, known as mutations, never seen in any previous studies. This detailed mutation map immediately changed the landscape of human genetics research: in the short span since the team publicly released a draft of their results in late 2015, hundreds of scientific groups around the world have used the ExAC dataset, with over 600 peer-reviewed scientific publications citing ExAC in the last two years alone.

    The effects of ExAC on human genetics research have been profound. For instance, specific mutations in important, disease-causing genes might not always result in disease for certain people; researchers have now used the ExAC dataset to decipher why this might happen for a peculiar gene, known as PRNP, that causes multiple neurological disorders, such as fatal familial insomnia. A different 2016 study [Nature] performed exome sequencing on 14,133 individuals from northern Europe to identify “ultra-rare” mutations—genetic changes never seen in any of the 60,708 ExAC participants—and showed the number of these ultra-rare mutations in genes important for brain development can partially predict how many years an otherwise healthy individual is likely to stay in school. These and similar discoveries are already having a palpable impact in translational research and clinical medicine, and these advances wouldn’t have been possible without population-scale resources like ExAC.

    Combining genetics with medical records to make DiscovEHRies

    Like ExAC, a recent collaboration between Regeneron Pharmaceuticals, Inc., and Geisinger Health System, dubbed DiscovEHR, combined the exome sequences and full medical records of over 50,000 volunteers recruited at one of Geisinger’s clinics in Pennsylvania. As depicted in Figure 3, this fusion of genetic and medical data proved to be an even more powerful approach than analyzing just the genetic data alone. The DiscovEHR study, published in 2016 in the journal Science, compared medical data between patients with and without mutations in certain genes, and found that patients with mutations that disabled a small group of specific genes had lower cholesterol levels, which lowered their risk of serious heart disease. Current pharmaceutical strategies involve identifying such genes as “targets” for drug development, aimed at recreating the lower cholesterol levels caused by the mutation that disabled the gene in patients, with the ultimate goal of providing those drugs to individuals at high risk for heart disease but who lack these rare, protective gene mutations.

    4
    Figure 3: Combining population sequencing with health records can identify new drug targets. A recent sequencing study of over 50,000 individuals, known as DiscovEHR, demonstrated the ability for population sequencing to generate new medical knowledge that can directly inform drug development and patient treatment choices. For example, the DiscovEHR study found genes that caused individuals to have lower blood cholesterol levels when inactivated by rare mutations, resulting in a reduced risk for major heart disease.

    Population sequencing lands a knockout punch

    Except for genes on the sex chromosomes (X & Y), there are two copies of every gene in your genome, one inherited from your father and one from your mother. Gene-inactivating mutations are generally rare events, so it is extremely uncommon for a single individual to inherit disabled copies of the same gene from both parents. When this does occur, it’s called a “gene knockout,” and means that individual lacks the ability to produce any of the protein encoded by that gene. Not surprisingly, gene knockouts are the known cause for hundreds (if not thousands) of rare diseases.

    In some cultures, marriages between first-cousins is commonplace. Since first-cousins are genetically closely related, their children are at a much higher risk of inheriting gene knockouts, making those children ideal individuals to study the effects of gene knockouts in humans (such studies are usually conducted in mice or other “model organisms”). By studying populations of children from first-cousin marriages, Last month, a team of researchers led by Sekar Kathiresan at the Massachusetts General Hospital reported in Nature on a population sequencing study of over 10,000 individuals from Pakistan, known as the PROMIS study, where the rate of first-cousin marriages is particularly high. Remarkably, the team found that at least 7% of all known protein-coding genes were knocked out in at least one individual without resulting in any obvious medical issues, meaning these genes might represent safe drug targets with little side-effect risks, as shown in Figure 4. Conversely, the PROMIS study also reported on a subset of individuals who were knockouts for current drug target genes that are thought to protect against heart disease, but those individuals developed heart disease at the same rate as the general population. Whatever the conclusion, this study drives home the point that population sequencing can inform—and, in some cases, correct—drug development and prescription of clinical treatments.

    5
    Figure 4: Rare gene knockouts can teach us why some drugs work and others don’t. Gene knockouts, or the situation where an individual inherits inactivated copies of a gene from both parents, are the known cause of hundreds of diseases because that individual is unable to produce any of the protein encoded by the knocked-out gene. However, not all gene knockouts cause disease. By studying healthy individuals with gene knockouts, scientists can learn about genes that might be safely knocked out by drugs, allowing for development of new targeted therapies. Conversely, scientists can also find genes that are targets of existing drugs yet, when knocked out in individuals in the general population, doesn’t result in the expected health benefit.

    The future: population sequencing, precision medicine, and personalized therapies

    Population sequencing is changing the way companies design drugs and doctors diagnose diseases and choose therapies. Landmark large-scale studies like ExAC and DiscovEHR have proven that pooling genetic data across tens of thousands of individuals dramatically improves researchers’ abilities to make new discoveries about the causes of disease. Cataloguing healthy individuals with rare gene knockouts, like the PROMIS study, can advance our understanding of human physiology, produce new drug targets, and shed new light onto why drugs might fail in certain patients. Yet numerous issues continue to hinder advances in medical genetics: we still know very little about the genetics of disease in people of African or Asian ancestry, genetic data remains difficult to share between research groups, and we have still only sequenced less than 0.01% of all people on earth.

    Even despite these challenges, our knowledge of the genome has already revolutionized medicine. Modern clinics can now perform dozens of genetic tests to evaluate your risk for cancer, Alzheimer’s disease, and heart attacks, or can tell you the odds your future children might have autism, epilepsy, or physical birth defects. For certain diseases, especially cancer, having your genome sequenced helps doctors select drugs designed specifically for your genetic makeup, making sure you get the most effective personalized treatment possible.

    Just like the famous Greek philosopher Socrates once quipped, “to know thyself is the beginning of wisdom.” Today’s geneticists and doctors would probably agree with him: genetic testing can teach you more about yourself, and in the process, help you live a longer, happier, healthier life.

    See the full article here .

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    Harvard University campus

    Harvard is the oldest institution of higher education in the United States, established in 1636 by vote of the Great and General Court of the Massachusetts Bay Colony. It was named after the College’s first benefactor, the young minister John Harvard of Charlestown, who upon his death in 1638 left his library and half his estate to the institution. A statue of John Harvard stands today in front of University Hall in Harvard Yard, and is perhaps the University’s best known landmark.

    Harvard University has 12 degree-granting Schools in addition to the Radcliffe Institute for Advanced Study. The University has grown from nine students with a single master to an enrollment of more than 20,000 degree candidates including undergraduate, graduate, and professional students. There are more than 360,000 living alumni in the U.S. and over 190 other countries.

     
  • richardmitnick 8:35 am on March 10, 2017 Permalink | Reply
    Tags: , Genomics, Hyak and Lolo supercomputing, Seeking to unravel DNA, U Wahington   

    From U Washington: “Seeking to unravel DNA” 

    U Washington

    University of Washington

    3.10.17
    No writer credit

    1
    Tim Durham, a graduate student in the William Stafford Noble Lab in the UW Department of Genome Sciences, adopted cloud computing to do his research.

    When Timothy Durham looks at the human genome, he sees an encyclopedia of precise instructions that tell approximately 31 trillion cells in the human body how to do their jobs.

    Figuring out how cells read and interpret these instructions—and how they can misread them—could help researchers unravel the mysteries of what leads to disease and point to cures. This is a complicated ongoing work being performed by thousands of researchers across the globe.

    Over the past decade, their efforts have produced large amounts of rich data. So when Durham, a graduate student and researcher in the William Stafford Noble Lab in the UW Department of Genome Sciences, decided to join the research, he found that a desktop computer and small department servers would not be up to the task.

    That’s why he turned to University of Washington Information Technology’s Research Computing experts, who recommended a cloud computing solution to do his work. The cloud, Durham said, provided him with virtually unlimited resources for computation, storage, networking and data management, the sort of tools he needed to build a complex three-dimensional model that would capture the state of the genome in different cell types. The model, he hopes, will help other researchers advance the field of genomics.

    Interpreting the human genome has been a tremendous challenge. It is like looking at a cookbook written in a foreign language with its own unique rules of grammar. In this cookbook, Durham said, genes are like “recipes” that cells use to construct the machinery they need to function

    “Now, we are starting to learn the language and the grammar of the genome, which is like learning to read the recipes and to understand which ones work well together and how the cell decides what to make,” he said.

    The ultimate goal is to be able to understand how the genome is used in different types of cells in the body to answer questions such as, “Which genes are important to the function of skin cells versus liver cells?”

    And in the same way that a cook doesn’t make every recipe in a cookbook when planning a meal, specific kinds of cells only care about certain subsets of genes when they are doing their work.

    “If we can understand how cells pick the genes they need out of all 20,000 genes in the genome cookbook, it will have a profound impact on the way we understand human biology and disease,” Durham said.

    Noble’s Lab is a perfect place for Durham’s work. The lab develops and applies computational techniques for modeling and understanding biological processes at the molecular level. Machine learning, a subfield of computer science focused on the study and construction of algorithms that can learn from and make predictions on data, is an important area for research, and Durham relied on its principles to develop his model.

    “I am developing a model that captures the state of the genome across 127 different cell types. The full data set is more than 2 TB, which is more than the memory capacity of our entire lab cluster,” Durham said.

    UW-IT set up Durham with Microsoft Azure and Amazon Web Services, which offer cloud services to the University of Washington. To help fund this, Durham applied for awards from Amazon’s Cloud Credits for Research program and from Microsoft’s Azure for Research program, and was granted $30,000 in cloud research credits, an extremely valuable contribution that helped accelerate his work.

    “Research funding is not easy to come by, so the credit program is really valuable,” Durham said. “It helps you through the initial learning curve involved in moving to the cloud by removing some of the risk of adopting a new technology and allowing you an extended trial period in which you can really dive deep to see how well it works for your application,” Durham said.

    Before Durham moved to the cloud, he was using lab servers, and even one of his smallest processing runs would take up to two full weeks to complete, said Rob Fatland, a UW-IT Research Computing Director who offers consulting and support to researchers looking at cloud computing solutions or other innovative tools offered at the UW, such as Hyak, the University’s shared cluster supercomputer.

    3
    U Washington Hyak and Lolo

    2
    Rob Fatland, UW-IT Research Computing Director, helps researchers navigate the cloud.

    “When he was using the department servers for his work, no one else could use them,” Fatland said. “In the cloud, he reduced processing time to hours without the restrictions that come with shared resources.”

    Large-scale cloud computing for research is relatively new to the University, but it is quickly establishing itself as a valuable tool, Fatland said. When talking to researchers, he discusses security, management and cost to operate in the cloud.

    Fatland said many researchers who have switched to the cloud have found that it is more cost effective for many types of computing, with costs decreasing over time. It is also extremely secure, so they don’t have to worry about losing their work. And it offers an elastic environment, easily allowing researchers to scale up their work instantly.

    “It’s an empowering technology,” Fatland said.

    That has been the case for Durham, whose goal for his three-dimensional model is to predict what parts of the genome are most important in a particular cell type, such as a liver or a heart cell.

    “It is challenging to train one of these computing models,” he said. “You have to do a lot of fine tuning and it takes a lot of computing time to optimize it, a lot of trial and error.” But with the cloud, he doesn’t have to wait for anyone to get done with their work. It is always available when he needs it.

    “In the end, if we can predict the most relevant portions of the genome in a particular cell type, this can help us zero in on specific regions of the genome that might, for example, harbor mutations that can contribute to disease,” he said.

    See the full article here .

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    The University of Washington is one of the world’s preeminent public universities. Our impact on individuals, on our region, and on the world is profound — whether we are launching young people into a boundless future or confronting the grand challenges of our time through undaunted research and scholarship. Ranked number 10 in the world in Shanghai Jiao Tong University rankings and educating more than 54,000 students annually, our students and faculty work together to turn ideas into impact and in the process transform lives and our world. For more about our impact on the world, every day.

    So what defines us — the students, faculty and community members at the University of Washington? Above all, it’s our belief in possibility and our unshakable optimism. It’s a connection to others, both near and far. It’s a hunger that pushes us to tackle challenges and pursue progress. It’s the conviction that together we can create a world of good. Join us on the journey.

     
  • richardmitnick 9:54 am on March 9, 2017 Permalink | Reply
    Tags: , Genomics, Personal Genome Project,   

    From Wyss: “Wyss Institute and Lumos Labs Launch Research Collaboration on Memory of High Performing Individuals” 

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    Wyss Institute bloc
    Wyss Institute

    March 9, 2017
    Eriona Hysolli

    Personal Genome Project will integrate brain training tests to help identify key memory genes towards understanding neurodegeneration.

    Researchers at the Wyss Institute for Biologically Inspired Engineering and Harvard Medical School (HMS)’s Personal Genome Project (PGP) announced today a new collaboration with Lumos Labs, makers of brain training program Lumosity. The PGP-Lumosity memory project aims to leverage the PGP’s and Lumos Labs’ unique resources and expertise to investigate the relationship between genetics and memory, attention and reaction speed.

    Wyss scientists plan to recruit 10,000 members from the PGP which started in 2005 in the laboratory of George Church, PhD, a founding Core Faculty member of the Wyss Institute and also Professor of Genetics at Harvard Medical School. Participants in the PGP publicly share their genome sequences, biospecimens and healthcare data for unrestricted research on genetic and environmental relationships to disease and wellness. Wyss Institute researchers will use a select set of cognitive tests from Lumos Labs’ NeuroCognitive Performance Test (NCPT), a brief, repeatable, accessible web-based alternative to traditional pencil-paper cognitive assessments to evaluate participant’s memory functions, including their ability to recall objects, memorize object patterns, and response times.

    Church’s research team at the Wyss Institute and HMS Postdoctoral Fellows Elaine Lim, Ph.D., and Rigel Chan, Ph.D., will correlate extremely high performance scores with naturally-occurring variations in the participants’ genomes. “Our goal is to get people who have remarkable memory traits and engage them in the PGP. If you are exceptional in any way, you should share it not hoard it,” said Church.

    To validate their findings, the team will take advantage of the Wyss Institute’s exceptional abilities to sequence, edit and visualize DNA, model neuronal development in 3D brain organoids ex vivo, and, ultimately, to test emerging hypotheses in experimental models of neurodegeneration.

    “The Wyss Institute’s extraordinary scientific program and the Personal Genome Project’s commitment to research that is both pioneering and responsible make them ideal collaborators,” said Bob Schafer, Ph.D., Director of Research at Lumos Labs. “Combining Lumosity’s potential as a research tool could help us learn more about how our online assessment can help power innovative, large-scale studies.”

    Drs. Church, Lim and Chan plan to begin recruitment for this study in early March.

    The PGP-Lumosity memory project is the latest in a long line of exciting research collaborations supported by each platform. Through their Human Cognition Project, Lumos Labs is currently working with independent researchers at over 60 different institutions and investigating a range of topics, including normal aging, certain clinical conditions and the relationship between exercise and Lumosity training. Existing collaborative projects available to PGP participants include stem cell banking with the New York Stem Cell Foundation, “Go Viral” real-time Cold & Flu surveillance, the biology of Circles with Harvard Medical School, Genetics of Perfect pitch with the Feinstein Institute for Medical Research, characterizing the human microbiome in collaboration with American Gut, and discounted whole genome sequencing strategies.

    With the PGP’s aim to serve as a portal that empowers the public to drive scientific discovery through their participation, this collaboration is a synergistic convergence of two uniquely positioned organizations that combine science with broad outreach.

    “What excites us about this project is opening up groundbreaking technologies developed at the Wyss Institute to explore the relationship between genetics and memory with possible implications for Alzheimer’s and other diseases,” said Wyss Institute Founding Director Donald Ingber, M.D., Ph.D., who is also the Judah Folkman Professor of Vascular Biology at Harvard Medical School and Boston Children’s Hospital, and Professor of Bioengineering at Harvard SEAS.

    For more information or to register in the study, please visit: https://wyss.harvard.edu/pgp-lumosity

    See the full article here .

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    The Wyss (pronounced “Veese”) Institute for Biologically Inspired Engineering uses Nature’s design principles to develop bioinspired materials and devices that will transform medicine and create a more sustainable world.

    Working as an alliance among Harvard’s Schools of Medicine, Engineering, and Arts & Sciences, and in partnership with Beth Israel Deaconess Medical Center, Boston Children’s Hospital, Brigham and Women’s Hospital, Dana Farber Cancer Institute, Massachusetts General Hospital, the University of Massachusetts Medical School, Spaulding Rehabilitation Hospital, Tufts University, and Boston University, the Institute crosses disciplinary and institutional barriers to engage in high-risk research that leads to transformative technological breakthroughs.

     
  • richardmitnick 10:12 am on January 13, 2017 Permalink | Reply
    Tags: , Could affect future treatments for some types of infertility, Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research, Genomics, , Metabolic proteins relocate to jump-start an embryo’s genome, UCLA study finds   

    From UCLA: “Metabolic proteins relocate to jump-start an embryo’s genome, UCLA study finds” 

    UCLA bloc

    UCLA

    January 12, 2017
    Sarah C.P. Williams

    FINDINGS

    1
    No image caption. No image credit.

    To turn on its genome — the full set of genes inherited from each parent — a mammalian embryo needs to relocate a group of proteins, researchers at the Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA have discovered. The metabolic proteins, normally found in the energy-generating mitochondria of cells, move to the DNA-containing nuclei about two days after a mouse embryo is fertilized, according to the new study, led by senior author Utpal Banerjee.

    BACKGROUND

    Early in development, a mammalian embryo — or zygote — has all the materials it needs to grow and divide from genes and proteins that were contained in the egg cell. But after a few cell divisions, the zygote needs to activate its own genome. Researchers have never fully understood how this shift is made. They knew that certain metabolic compounds, such a pyruvate, were required, but had also observed that the mitochondria — which normally process pyruvate into energy — were small and inactive during this stage of development.

    METHOD

    Banerjee, a professor of molecular, cell, and developmental biology and co-director of the UCLA Broad Stem Cell Research Center, and colleagues confirmed that pyruvate was required for zygotes to activate their genomes by growing mouse zygotes in a culture dish lacking pyruvate. Then, in both mouse and human embryos, researchers used a number of methods to determine the location of proteins that process pyruvate through a metabolic program called the TCA cycle. Just before the embryos activated their genomes, the two-cell stage in mice, the TCA cycle proteins moved from the mitochondria to the nuclei of cells, the researchers discovered. While mouse cells grown in dishes lacking pyruvate normally stopped growing at the two-cell stage, the researchers could rescue these cells by adding a metabolic compound that’s produced by the TCA cycle. Repeating some of the experiments in human embryos, they confirmed that the metabolic proteins move from the mitochondria to the nucleus just as the genome is activated — at the six- to eight-cell stage for humans.

    IMPACT

    The importance of metabolic proteins to early embryonic development could affect future treatments for some types of infertility. In addition, the researchers hypothesize that some stem cells that have similar metabolic properties to early zygotes — including cancer stem cells — may relocate the TCA cycle proteins. Better understanding of the relocation could shed light on stem cell biology and alter cancer treatments.

    AUTHORS

    In addition to Banerjee, the first authors of the study are Raghavendra Nagaraj and Mark Sharpley; the co-authors are Daniel Braas, Fangtao Chi, Amander Clark, Rachel Kim and Yonggang Zhou, all of UCLA.

    JOURNAL

    The study was published in the journal Cell.

    FUNDING

    The study was funded by an NIH Director’s Pioneer Award (DP1DK098059-04) and by the UCLA Broad Stem Cell Research Center.

    See the full article here .

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  • richardmitnick 1:59 pm on December 17, 2016 Permalink | Reply
    Tags: , Boy in the bubble, , Genomics, ,   

    From UC Berkeley: “From a single genetic mutation, secrets of ‘boy in the bubble’ disease revealed” 

    UC Berkeley

    UC Berkeley

    December 15, 2016
    Brett Israel
    brett.israel@berkeley.edu

    UC Berkeley was part of an interdisciplinary, international research team that has identified the rare genetic mutation responsible for a unique case of “boy in the bubble” disease, known as severe combined immunodeficiency (SCID), a deadly immune system disorder. The researchers found that the cause was a mutated version of a gene called BCL11B, which also plays an unexpected role in the normal processes of immune system development.

    1
    World of his own: David Vetter (Photo: Courtesy Baylor College of Medicine Archives) http://i2.mirror.co.uk/incoming/article3196066.ece/ALTERNATES/s810/The-Boy-in-the-Bubble.jpg. Just a single case chosen at random from many.

    The discovery of this genetic mutation is the latest of several breakthroughs from this team, which has been accomplished by analyzing exomes — the roughly 2 percent of DNA that contains the instructions for building proteins — to identify the cause of mysterious immunological diseases in newborns.

    “This is a gene that had never been associated with SCID before, which required more advanced genome analysis techniques to discover,” said Berkeley computational biologist Steven Brenner, co-author of the study. “Moreover, unlike variants in every other known SCID gene, this mutation is dominant, which means you only need one copy of this mutation to disrupt multiple aspects of development.”

    The study was published Dec. 1 in the New England Journal of Medicine. The research article was accompanied by a perspective by Michael Lenardo, chief of the Molecular Development of the Immune System Section at the National Institute of Allergy and Infectious Diseases, commissioned by the journal. Lenardo wrote that the study is “an exciting example of recent achievements in the application of contemporary molecular genomics to clinical medicine, especially with regard to congenital diseases…This study reflects remarkable advances in molecular diagnosis.”

    The infant patient featured in the new study was identified through a population-based neonatal screening approach for SCID, which was developed in 2005 by Jennifer Puck, the study’s senior author and a UCSF professor of immunology and pediatrics. The screening indicated a severely compromised immune system, leaving the patient open to a likely fatal series of infections. However, UCSF doctors performed a bone marrow transplant, the standard of care for SCID, which provided the infant with a fully functional immune system.

    In addition to SCID, however, the infant was born with a constellation of abnormal features including craniofacial deformities, loose skin, excess body hair and neurological abnormalities, which suggested that a single rare genetic defect could underlie the patient’s disease.

    In part to determine whether the infant’s parents were carriers of a genetic mutation that could be passed on to future children, the research team set out to scan the genomes of both infant and parents for mutations that could be responsible for the disease. Researchers at UC Berkeley and UCSF built on their productive collaboration with researchers at Tata Consultancy Services to use next-generation exome sequencing to identify a single mutation present in the infant but not the parents — referred to as a de novo mutation — in the BCL11B gene, which had previously been associated primarily with lymphatic cancer. So finding the BLC11B mutation to be causative for SCID was a surprise.

    “We’re entering a new era of genomic medicine,” Puck said. “Our technology has progressed to the point that we can learn a great deal about a disease, and even learn important new facts about normal biology, from just a single patient. In this case we were able to unearth the potentially unique underlying genetic cause of one patient’s disease and come away with brand new understanding of how the immune system develops.”

    In order to understand the biological effects of the patient’s mutation, the researchers collaborated with the team of David Wiest at Fox Chase Cancer Center, in Philadelphia, to introduce the patient’s mutated form of BCL11B into zebrafish, whose immune systems are similar to those of humans. They found that the mutated form of BCL11B produced abnormalities in the zebrafish that mimicked those observed in the patient, including not only a disabled immune system but also similar craniofacial abnormalities. Blocking the mutated gene and replacing it with the normal human gene in embryonic zebrafish reversed all these symptoms, strongly suggesting that abnormal BCL11B was the cause of the symptoms seen in both zebrafish and the human patient.

    The normal BCL11B protein binds to DNA at sites across the genome to activate a wide variety of developmental genes in a precisely orchestrated sequence. Experiments revealed that the BCL11B gene mutation identified in the new study disrupts this protein’s ability to bind to DNA, thereby resulting in the wide array of immunological, neurological and craniofacial disruptions seen in both the human patient and in zebrafish.

    “In this case, however, a mutation in BCL11B turned the protein it produces into a monkey wrench that disrupted many different systems in the body,” Puck said.

    According to Puck, the findings illustrate the power of deeply studying rare diseases in individual patients: “We may never get another patient just like this one,” she said. “But as a result of studying this one case we were able to learn so much about a critical gene in a critical pathway that hadn’t been appreciated before.”

    The research was supported by the National Institutes of Health, Tata Consultancy Services, the Commonwealth of Pennsylvania, the M.D. Anderson Cancer Center, the Fox Chase Cancer Center, the Jeffrey Modell Foundation, the Lisa and Douglas Goldman Fund and the Michelle Platt-Ross Foundation.

    See the full article here .

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