Children with significant congenital heart disease have a far better chance of surviving today than in decades past, thanks to major advances in surgery. But some infants who recover from repairs to their hearts later show the effects of delays in brain development, including impairments to cognitive, language and social functioning. Such impairments can affect how well these children do in school and in the workplace; they can even diminish their overall quality of life.
Epidemiological studies have given numbers to what doctors and families have long observed: The risk of neurodevelopmental delays is tenfold higher for children with moderate to severe congenital heart disease than for other children.
Over the years, those who study these phenomena have considered several possible reasons. Do the rigors of open-heart surgery so soon after birth play a role? Could heart defects limit nutrients and oxygen needed by the fetus? Or could spontaneous genetic mutations cause congenital problems that affect both the heart and the brain of a child?
Now, the “why” may have been answered by the efforts of the Pediatric Cardiovascular Genetics Consortium, led by a team of Harvard Medical School scientists. In a recent issue of Science the consortium reported exome sequence analyses of more than 1,200 children and their parents and showed that children with both congenital heart disease and neurodevelopmental delays share certain genetic mutations that thwart the normal development of both the heart and the brain.
“We’re homing in on a set of genes that have multiple different roles on multiple different tissues during development: heart tissue, brain tissue, other developing organs, limb tissue.” —Jason Homsy
Using a mathematical model created by co-authors Kaitlin Samocha and Mark Daly of the Analytical and Translational Genetics Unit at Massachusetts General Hospital, the team analyzed mutations in the protein-coding portion of the genomes of children with congenital heart disease that were not present in their parents’ genomes. They found that these children have more of these de novo mutations in genes that are highly expressed in the developing heart, compared to a control cohort of children without congenital heart disease.
The de novo mutations were also found to be more frequent in children with congenital heart disease plus another birth defect, either neurodevelopmental delay or more-subtle abnormalities of finger or ear shape. These findings bolster the case for shared genetic causes of the cardiac and extra-cardiac abnormalities rather than surgeries or environmental factors.
“We’re homing in on a set of genes that have multiple different roles in multiple different tissues during development: heart tissue, brain tissue, other developing organs, limb tissue,” said Jason Homsy, an HMS LaDue Fellow who trained at Mass General and co-lead author of the Science paper. “Our study shows a common genetic link for the development of these diseases.”
Potential for early testing
According to Homsy and co-senior author Christine Seidman, the HMS Thomas W. Smith Professor of Genetics and Medicine at Brigham and Women’s Hospital and a Howard Hughes Medical Institute investigator, these findings could lead to early testing that would help identify newborns with congenital heart disease who are at high risk of neurodevelopmental difficulties.
“We can pretty clearly tell the parents of children with congenital heart disease what’s going to happen after the heart surgery, but there’s always a big question: Will my kid learn well in school?” Seidman said. “If we could identify children at high risk for neurodevelopmental delays, they could receive increased surveillance and earlier interventions than occur now.”
The mutations primarily affected genes involved in three areas: morphogenesis, chromatin modification and transcriptional regulation. If any one of these processes is perturbed even slightly at a critical time in development, the heart is malformed; sometimes another developmental defect occurs, such as a missed connection in the brain.
“These genes are not just involved in shaping the heart,” Seidman said. “They are master regulators of organ development.”
One of the mutated genes is RBFOX2, which encodes a molecule that regulates RNA splicing. Although RBFOX2 has not been previously implicated in congenital heart disease, de novo mutations were identified in multiple affected children.
“There are still many unanswered questions, including why the same mutation can cause very different clinical manifestations,” Seidman said. Perhaps additional genetic variants in the multiple layers of transcriptional regulation allow compensation for some mutations but worsen the consequences of others. For now, Seidman said, knowing that a genetic mutation is present is different from knowing the outcome.
“It’s a long, long, long way down the road,” Seidman said, “but we’d like to believe that if you knew the steps by which these mutations perturbed the regulation of gene expression, there might even be ways to actually treat it.”
This work was supported by grants from the National Heart, Lung, and Blood Institute and the National Human Genome Research Institute of the National Institutes of Health, Howard Hughes Medical Institute, Simons Foundation for Autism Research, John S. LaDue Fellowship at HMS, Medical Scientist Training Program and National Research Science Award, Academy of Medical Sciences, British Heart Foundation, Wellcome Trust, Arthritis Research UK and the NIHR Cardiovascular Biomedical Research Unit at Royal Brompton and Harefield NHS Foundation Trust and Imperial College London, Leducq Foundation, Heart and Stroke Foundation of Ontario, Ted Rogers Centre for Heart Research, Kostin Family Innovation Fund, Aaron Stern Professorship at the University of Michigan, and Braylon’s Gift of Hope Fund.
WASHINGTON, D.C.—Genetics is not your destiny, George Church told an overflow audience gathered in a Congressional briefing room. This was one of many head-turning statements he and two other speakers made about advances in the rapidly progressing field of genetic technologies.
“You can change it,” Church said about the genome. “Certainly you need to know it first.”
In a lightning-quick 10 minutes, Church, the Robert Winthrop Professor of Genetics at Harvard Medical School, explained how gene editing can potentially eliminate malaria and allow transplants of pig organs into humans, among other innovations.
Change was the theme of a briefing designed to shed light on the frontiers of genetic technologies. Church was joined by Diana Bianchi, the Natalie V. Zucker Professor at Tufts University School of Medicine, and Jennifer Doudna, professor of molecular and cell biology and of chemistry at the University of California, Berkeley.
“This is a technology that seemed at first like science fiction: a technology for rewriting DNA itself. We can sequence DNA, we can find mutations. What if we had a way to fix those mutations? That technology is not science fiction; it is a reality.”— Jennifer Doudna, University of California, Berkeley
The Nov. 17 briefing was presented by the Personal Genetics Education Project (pgEd), a multifaceted effort based in the Department of Genetics at HMS. The mission of pgED is to educate individuals about advances in the field through school programs, libraries, religious institutions and other public forums, and to accelerate public awareness of genetics issues by advising the entertainment industry. It also seeks to engage lawmakers—the “eyes and ears of the nation”—in discussions.
pgEd takes no position on policy, preferring to educate from a neutral position so that its audience can make better-informed decisions.
Each speaker was passionate about the need to inform and educate the public about genetics and the power of new technologies to affect health and disease.
Bianchi is a leading medical geneticist in the burgeoning field of prenatal diagnosis. Doudna and her colleague Emmanuelle Charpentier, now at the Max Planck Institute for Infection Biology, are credited with translating an ancient mechanism in bacterial immunity into a powerful tool for editing genomes. Church optimized the gene-editing technology, now known by the acronym CRISPR, by developing what are known as gene drives, that is, molecular tools that can “drive” a DNA sequence to be inherited by all of an organism’s offspring.
Doudna described the technology she and others have advanced as a genome-engineering revolution.
“This is a technology that seemed at first like science fiction; a technology for rewriting DNA itself,” she said. “We can sequence DNA, we can find mutations. What if we had a way to fix those mutations? That technology is not science fiction; it is a reality.”
The possible therapeutic applications are promising, she said, but delivery into cells and tissues remains a challenge.
Doudna also pointed out the fundamental difference between gene editing that affects only one person versus gene editing in the germ line that is passed on to future generations.
“When do we apply that? And for what kind of afflictions?” she asked. “I’m very interested myself in trying to stimulate public conversation over the implications of this technology.”
Bianchi, who both sees patients and conducts research in medical genetics, is already using genomic sequencing in her practice. Non-invasive prenatal testing takes advantage of the fact that small amounts of DNA fragments from the placenta circulate within the blood of pregnant women. Cell-free fetal DNA can be analyzed to screen for extra or missing copies of chromosomes that might indicate Down syndrome or other inherited syndromes.
Since 2011, the DNA screening tests have been done on 2 million women. The DNA tests have both a high sensitivity and specificity and most important, Bianchi said, a higher positive predictive value than the current noninvasive screening tests that measure blood proteins and fetal ultrasound markers.
“Genomic testing has really changed prenatal medicine,” Bianchi said. “We think the future is linked to treatment.”
Bianchi’s lab is testing in mice whether administering therapy during pregnancy improves fetal brain development. Treatment in pregnant women could start as early as 12 weeks following a confirmed diagnosis of fetal Down syndrome and continue until delivery at 40 weeks.
If such a strategy works, it would give pregnant women three options to consider: continuing the pregnancy aware of a high risk for a genetic abnormality, terminating the pregnancy or treating the condition before birth.
Church explained how gene drives could make mosquitoes immune to malaria and therefore unable to transmit the disease to humans.
“That would have a huge impact. Like smallpox, once it’s gone, it’s gone,” he said.
Gene editing could also eliminate the viral components of the pig genome, thereby making organ transplants into humans more feasible. A recent paper showed that 62 genes could be altered at once, demonstrating the power of the technology.
While such projects are tantalizing, concern is also growing among scientists, bioethicists, policymakers and others over how gene-editing technologies, such as CRISPR, will be used, the speakers said, underscoring the importance of public discussion.
The briefing on Capitol Hill was sponsored by U.S. Rep. Louise Slaughter, D-N.Y., and Sen. Elizabeth Warren, D-Mass. More than 160 people, half of them Congressional staffers, heard where the science of genetic technologies is today, where it may be going and what concerns exist over its benefits and risks.
“It’s all about scalability,” said Ting Wu, HMS professor of genetics and pgEd’s director and co-founder. She hoped to send people out of the room astounded by the science and eager to spark discussion among policymakers, thus amplifying pgEd’s reach.
After the briefing, the speakers and the pgEd team met with the White House Office of Science and Technology, where they expressed hope that all segments of society could become part of the discussion.
“We think it is critical to engage across all communities, particularly the disenfranchised,” Marnie Gelbart, pgEd director of programs, said later. “This piece is as important as the technologies themselves, for safely and fairly integrating the technologies into society.”
The briefing was the fourth presented by pgEd. The first briefing highlighted the science of genomics, personalized medicine and genetic engineering, as well as ways to reach out to the public. The second focused on two topics: the role of genetics research in the unfolding Ebola outbreak in West Africa and the issues addressed by the Genetic Information Nondiscrimination Act, also known as GINA. The third centered on law enforcement and highlighted research on how microbial genomics is being used to improve health and increase public safety. For the fourth, pgEd wanted to bring attention to the excitement at the forefront of genetics and the questions being raised about how we as a society use these new technologies.
pgEd is supported by the HMS Department of Genetics and private funding from Sigma-Aldrich, Autodesk, Genentech, IDT (targeted specifically for GETed conferences and Map-Ed) and an anonymous donor.
It used to be enough to call a serotonergic neuron a serotonergic neuron.
These brain cells make the neurotransmitter serotonin, which helps regulate mood, appetite, breathing rate, body temperature and more.
Recently, however, scientists have begun to learn that these neurons differ from one another—and that the differences likely matter in dysfunction and disease.
Last year, a team led by Harvard Medical School genetics professor Susan Dymecki defined a subgroup of serotonergic neurons in mice by showing that those cells specifically, among all serotonergic neurons, were responsible for increasing the breathing rate when too much carbon dioxide builds up in the body.
Now, Dymecki and colleagues have taken a first stab at systematically characterizing serotonergic neurons at the molecular level and defining a full set of subtypes, again in mice.
The researchers report in Neuron that serotonergic neurons come in at least six major molecular subtypes defined by distinct expression patterns of hundreds of genes. In many cases, the subtypes modulate different behaviors in the body.
By conducting a cross-disciplinary series of experiments, the researchers found that the subtypes also vary in their developmental lineage, anatomical distribution, combinations of receptors on the cell surface and electrical firing properties.
“This work reveals how diverse serotonin neurons are at the molecular level, which may help to explain how, collectively, they are able to perform so many distinct functions,” said Benjamin Okaty, a postdoctoral researcher in the Dymecki lab and co-first author of the paper.
“To have the list of molecular players that make each of these subtypes different from one another gives us an important handle on learning more about what that cell type does and how we can manipulate only that subtype,” said Dymecki. “It holds enormous therapeutic potential.”
“This is an ancient neurotransmitter system that’s implicated in many different diseases, and it’s starting to be cracked open,” said Morgan Freret, a graduate student in the Dymecki lab and co-first author of the paper. “We can now ask questions in a more systematic way about which serotonergic cells and molecules are important in, for example, pain, sleep apnea or anxiety.”
Crucially, the team also showed that a serotonergic neuron’s gene expression and function depend not only on its location in the adult brain stem, but also on its cellular ancestor in the developing brain.
“Earlier work had shown that you could explore the relationship between a mature neuronal system and the different developmental lineages that gave rise to it, but we had no idea whether it was meaningful,” said Dymecki. “We show that the molecular phenotypes of these neurons track quite tightly to their developmental origin, with anatomy making some interesting contributions as well.”
While the work was done in mice, Dymecki is optimistic that it will be replicated in humans because the serotonergic neuronal system is in a highly conserved region of the brain, meaning it tends to remain consistent across vertebrate species.
Because of this, researchers can look for the same molecular signatures in human tissue and begin to tease apart whether particular subtypes of serotonergic neurons are involved in conditions such as sudden infant death syndrome (SIDS) or autism.
Such research could ultimately reveal previously unknown contributions of the serotonergic neuronal system to disease, inform the development of biomarkers or lead to more targeted therapies.
The team’s findings could also inform stem cell research. “Which subtype of serotonergic neuron are we getting when we use current stem cell protocols?” asked Dymecki. “Can we drive the development of different subtypes? Can we watch how gene expression patterns change over time during development for each subtype?”
Finally, the study provides an example of a highly integrative approach to understanding brain function at multiple scales, “linking genes and gene networks to the properties of single neurons and populations of neuron subtypes, all the way up to the level of animal behaviors,” said Okaty. “I think it’s a useful template going forward. Imagine what we’d learn by applying this approach to all the neurotransmitter systems in the brain.”
This research was supported by funding from the National Institutes of Health (R01 DA034022, P01 HD036379, T32 GM007753, R21 MH083613, R21 DA023643), the American SIDS Institute, a Harvard Stem Cell Institute seed grant, a NARSAD Distinguished Investigator Grant from the Brain and Behavior Foundation, and Harvard’s Blavatnik Biomedical Accelerator, which provides resources to develop early-stage biomedical technologies toward clinical applications. Harvard’s Office of Technology Development has filed a patent application on the technology.
Four tiny segments of RNA appear to play critical roles in controlling cholesterol and triglyceride metabolism. In their report in Nature Medicine, Harvard Medical School researchers at Massachusetts General Hospital describe how these microRNAs could reduce the expression of proteins playing key roles in generating beneficial HDL cholesterol, disposing of artery-clogging LDL cholesterol, controlling triglyceride levels and other risk factors for cardiovascular disease.
“While we and others have recently identified microRNAs that control cholesterol and fat metabolism and trafficking, no studies to date have systematically looked at all non-coding factors such as microRNAs in genetic studies of human diseases and other traits,” said Anders Näär, HMS professor of cell biology and corresponding author of the current study.
“Using human genetic data from almost 190,000 individuals, we have linked 69 microRNAs to increased genetic risk for abnormal cholesterol and triglyceride levels, and showed that four of these act to control proteins we know are involved in those metabolic activities,” Näär said.
“We hope these findings will lead to new, more effective ways of treating or even preventing cardiovascular disease and other metabolic disorders.”— Anders Näär
Less than 2 percent of human DNA represents genes that code for the production of proteins. While it was originally hypothesized that the other 98 percent had no function—leading to the term “junk DNA”—it has now become apparent that these DNA sequences play essential roles in determining how, when and where protein-coding DNA is expressed.
One such control mechanism is through single-stranded microRNAs, which block the expression of protein-coding genes by binding to messenger RNAs and preventing their translation into protein. In previous studies, Näär and his colleagues found that a microRNA called miR-33 suppresses production of beneficial HDL cholesterol and that antisense blocking of miR-33 increased HDL levels in an animal model.
The current study began with analysis of genome-wide association studies involving more than 188,000 people. The researchers identified 69 microRNAs located near gene variants previously associated with lipid abnormalities.
Using a tool that predicts the targets of microRNAs based on matches between their nucleotide sequences and those of protein-coding genes and a database of identified gene functions, the researchers arrived at four microRNAs that appear to control genes involved in cholesterol and triglyceride levels and in other metabolic functions, such as glucose metabolism.
Two of these—miR-128-1 and miR-148a—were found to control the expression of proteins essential to the regulation of cholesterol/lipid levels in cells and in animal models; miR-128-1 was also found to regulate fatty liver deposits, insulin signaling and maintenance of blood sugar levels.
“We are following up these findings with studies to address whether antisense blocking of these microRNAs could decrease atherosclerosis, cardiovascular disease and inflammatory fatty liver diseases in animals,” Näär said. “We hope these findings will lead to new, more effective ways of treating or even preventing cardiovascular disease and other metabolic disorders.”
Support for the study includes National Institutes of Health grants R21DK084459, R01DK094184, R37DK048873, R01DK056626, K24DK078772, R01HL107953 and R01HL106063.
Adapted from a Mass General news release.
Brain metastases are a devastating complication of cancer, leading to the death of more than half of patients whose cancer spreads to the brain. A new study finds that while brain metastases share some genetic characteristics with the primary tumors from which they originated, they also carry unique genetic mutations.
The diverging evolutionary pathways of the metastatic and the primary tumors may change sensitivities to targeted therapy drugs, an international collaboration led by Harvard Medical School scientists at Massachusetts General Hospital report in Cancer Discovery.
“Our study demonstrates that while brain metastases and primary tumors share a common ancestry, they continue to evolve separately,” said Priscilla Brastianos, HMS instructor in medicine at Mass General and co-lead author of the paper.
“This is tremendously important, as we demonstrate that brain metastases may have clinically significant mutations that have not been detected either in the primary tumor biopsy or in metastases from other parts of the body,” she said. “We also showed that multiple brain metastases from the same patient share nearly all clinically significant mutations.”
Brain metastases commonly develop from melanoma, lung cancer or breast cancer and can appear despite the primary tumor’s being well controlled by drugs that target mutations driving its growth. Once brain metastases develop, patients usually die within a matter of months, and patients with brain metastases are typically excluded from most clinical trials.
“While brain metastases and primary tumors share a common ancestry, they continue to evolve separately.” —Priscilla Brastianos
In treating cancers known to be driven by targetable gene mutations, treatment planning is usually based on genetic analysis of tissue from the primary tumor. Because treatment of brain metastases often involves removal of the metastasis, samples of that tumor are often available for analysis. The current study was designed to investigate whether the genetic profiles of brain metastases are identical to those of the primary tumors.
The research team conducted whole-exome gene sequencing on three tissue samples—primary tumor, brain metastasis and normal tissue—from each of 86 patients with lung, breast or kidney cancers. The exome is the tiny portion of the genome that encodes proteins.
In each instance, while the investigators found that the primary tumor and the metastasis shared some mutations, the brain metastases had new mutations that were not related to those of the primary tumors. In four of the 86 patients, the brain metastases actually appeared to have originated from additional primary tumors.
The new mutations detected in the metastases often signaled potential sensitivity to targeted therapy drugs that would not have been effective against the primary tumors. Overall, more than half of the patients appeared to have clinically targetable new mutations in their brain metastases.
Analysis of multiple brain metastases samples from the same patient showed that nearly all of the significant mutations appeared in all of the brain metastases. In contrast, metastases from other parts of the body differed significantly from the brain metastases.
“It has been unclear whether brain metastases from well-controlled primary tumors develop because the chemotherapy drugs don’t cross the blood-brain barrier or because of different genetic mutations in the metastasis,” said Brastianos. “Our data suggest that genetic differences may contribute to the formation and treatment resistance of brain metastases.”
The clinical impact of directly targeting brain-metastasis specific mutations needs to be evaluated more fully, she said, and it is something the scientists are now investigating.
“We believe that routinely looking for clinically significant alterations in brain metastases may open the door to new therapeutic options for these patients,” Brastianos said.
Support for the study includes National Institutes of Health grants U54 HG003067, 5U24 CA143687 and U54 CA143798, and grants from the Brain Science Foundation, Susan G. Komen for the Cure, Terri Brodeur Breast Cancer Foundation, Conquer Cancer Foundation, the American Brain Tumor Association, Breast Cancer Research Foundation and the Mary Kay Foundation.
Adapted from a Mass General news release.
Stephen J. Elledge, the Gregor Mendel Professor of Genetics and of Medicine at HMS, and professor of medicine at Brigham and Women’s Hospital, is a co-recipient, with Evelyn Witkin of Rutgers University, of the 2015 Albert Lasker Basic Medical Research Award.
The award, widely considered to be among the most respected in biomedicine, will be presented on Friday, Sept. 18, in New York City.
Elledge and Witkin are being honored for their seminal discoveries that have illuminated the DNA damage response, a cellular pathway that senses when DNA is altered and sets in a motion a series of responses to protect the cell. This pathway is critical to a better understanding of many diseases and conditions, such as cancer.
"Steve is an amazing scientist, mentor and colleague," said Jeffrey S. Flier, dean of Harvard Medical School.
"His insights into the basic mechanisms of the DNA damage response have profoundly enriched our understanding not only of the fundamental genetics of all cellular life, but also of how we conceptualize many diseases and conditions, especially cancer. This distinction is richly deserved, and I am delighted that Steve is being honored for this extraordinary body of work," Flier said.
"We are extremely proud of Steve, who is truly deserving of this recognition,” said Elizabeth G. Nabel, MD, president of Brigham and Women's Health Care.
“Courageous and insatiably inquisitive, he represents the best of Brigham and Women’s and our mission of driving innovation in basic science to improve human health. As a devoted mentor, Steve is deeply committed to guiding the careers of young investigators, ensuring that the next generation of scientists is filled with curious, passionate and talented researchers," she added.
Elledge often describes the process by which a cell duplicates itself as akin to the duplication of a small city. It is a vastly complex process that requires many levels of intricate coordination. Each cell contains a detailed blueprint for this entire process: DNA.
But not every duplication results in a perfect copy. That is because each time a cell makes a copy of itself, DNA is vulnerable to damage, not only from faulty cellular processes, but also from such entities as environmental chemicals. As DNA damage accumulates, it profoundly complicates a cell’s ability to make a faithful copy of itself. This can lead to serious illnesses, birth defects, cancer and other health problems.
Witkin discovered how bacteria respond to DNA damage, detailing the response to UV radiation. Elledge uncovered a DNA-damage response pathway that operates in more complex organisms, including humans.
Over the years, Elledge and his colleagues elucidated a signaling network that informs a cell when DNA sustains an injury.
Called the DNA damage response, this network senses the problem and sends a signal to the rest of the cell so it can properly repair itself, otherwise severe mutations can occur. As a result, this pathway helps keep the genome stable and suppresses adverse events such as tumor development.
When individuals are born with mutations in this pathway, they often have severe developmental defects. If the pathway is interfered with later in life, cancer can result.
In addition to the award in basic medical research, the Lasker Foundation is also presenting awards to individuals in clinical research and in public service.
According to Claire Pomeroy, president of the Albert and Mary Lasker Foundation, this year’s recipients “remind us all that investing in biological sciences and medical research is crucial for our future.”
Joseph L. Goldstein of the University of Texas Southwestern Medical Center and chair of the Lasker Medical Research Awards Jury, added, “The 2015 Lasker winners had bold ideas and pursued novel questions that they tested through fearless experimentation.”
When mitochondria—the cell’s power plant—are sick or damaged, they must be cleared away so the cell can survive.
In the brain, this mitochondrial quality control pathway is so critical that neurodegenerative disease can result if bad mitochondria accumulate in neurons.
Now, Harvard Medical School researchers have mechanistically connected this pathway—which is already linked to Parkinson’s disease—with proteins that are mutated in amyotrophic lateral sclerosis, the motor-neuron disease also known as Lou Gehrig’s disease. Their findings are described in Molecular Cell.
In healthy cells, damaged mitochondria are broken down and disposed of by a process called selective autophagy, a term stemming from Greek roots that translates as “self-eating.” How the mitochondria get tagged for disposal has been the subject of research in many labs around the world in recent years.
Proteins linked to Parkinson’s
Mitochondrial damage triggers activation of two proteins—PARKIN and PINK1—that tag the mitochondria surface with chains of ubiquitin, molecules that signal the cell to get rid of the defective mitochondria. These proteins have been known for more than a decade to be faulty in early-onset familial forms of Parkinson’s disease, but only recently have their roles in mitochondrial quality control been elucidated.
Most recently, scientists have been studying how cells recognize these disposal signals and which proteins are involved further downstream in the disposal process.
Initial key insights into the downstream mechanism were suggested several years ago by the finding that the autophagy cargo receptor protein OPTN functions together with the TBK1 protein kinase in the removal of pathogenic bacteria from cells via autophagy. Interestingly, both OPTN and very recently TBK1 were also found to be mutated in ALS, but how these proteins contribute to this neurodegenerative disease has remained poorly understood.
Now scientists led by Wade Harper have mechanistically connected these two sets of proteins and have described a multistep mitochondrial quality control pathway. Upstream proteins—PARKIN and PINK1—function early in the mitochondrial disposal process by assembling ubiquitin chains on damaged mitochondria while the downstream proteins—OPTN and TBK1—bind to these ubiquitin chains to target the damaged mitochondria to the autophagy machinery.
Importantly, binding of the OPTN-TBK1 complex to these ubiquitin chains promotes TBK1 activation and further activation of OPTN’s ubiquitin-binding activity, establishing a self-reinforced feed-forward mechanism that is critical for the ultimate delivery of mitochondria to the autophagosome.
Connection to ALS
By using state-of-the-art quantitative mass spectrometry coupled with gene-editing and cell-imaging tools, the new study provides a detailed picture of how these two pathways work together and suggests how they may link the two neurodegenerative diseases.
“This is the first time we could mechanistically show how ubiquitination of damaged mitochondria promotes their clearance, and suggests that mitophagy defects potentially contribute to ALS as well as Parkinson’s disease,” said Harper, who is Bert and Natalie Vallee Professor of Molecular Pathology and chair of the HMS Department of Cell Biology.
Alban Ordureau, a postdoctoral fellow in the Harper lab and a co-author of the paper, had previously studied how PARKIN and PINK1 work together biochemically to specifically tag damaged mitochondria with ubiquitin.
“The story here focuses on how, once mitochondria are ubiquitinated, they are recognized by the cell and gotten rid of by proteins implicated in two distinct neurodegenerative diseases,” said Ordureau.
Jin-Mi Heo, postdoctoral fellow in the Harper lab and lead author of the paper, discovered that TBK1 becomes activated when mitochondria are damaged via ubiquitin chain binding by OPTN.
Stress and sensitivity
“If there is stress happening in the same cell in one pathway, that might cause defects in other pathways,” Heo said. “That could be why we see similar characteristics in different diseases with different origins.”
In Parkinson’s disease, neurons that make the neurotransmitter dopamine are defective while in ALS, the motor neurons do not perform as they should.
“There are some very complex relationships between different mutations that are going to lead to disease and which cell types they act in,” Harper said. “Depending on the cell type, you get one or the other disease.”
One idea is that individual types of neurons are sensitive to toxic proteins or toxic organelles—such as damaged mitochondria—in different ways. Motor neurons, for example, are much longer than most neurons in the brain, so they may be more sensitive to the accumulation of certain types of toxic proteins or organelles than other types of neurons, Harper said. The mutations that are present in the individual diseases may reflect sensitivities to different types of autophagic cargo; the result is motor neuron disease in ALS or dopaminergic disease in Parkinson’s.
“The surprising thing is that the Parkinson’s genes are functioning upstream of a pathway that’s mutated downstream in motor neuron disease,” Harper said. “So there is a genetic sensitivity within the pathway that must be different in different cells.”
It may turn out that this is a general mechanism that cells use to get rid of a variety of damaged material in different kinds of neurons.
“We’re starting to work that out,” Harper said.
This study was supported by National Institutes of Health grant R37 NS083524, Biogen and the Edward R. and Anne G. Lefler Center for the Study of Neurodegenerative Disorders at HMS. Harper is a consultant for Millennium: the Takeda Oncology Company and Biogen.
Harvard Medical School and the Qatar Biomedical Research Institute, with a commitment from the Shenzhen-HMS Initiative in International Education in China, are working together to make history with a joint global initiative designed to disseminate knowledge of cancer biology, prevention and treatment.
A new 12-month program aimed at international clinicians and scientists who specialize in cancer, Cancer Biology and Therapeutics will break new ground as the first HMS global program to combine basic and applied science in a blended-learning format, providing insights from leading researchers, oncologists and therapeutic developers both online and in person.
“This collaborative initiative will benefit Qatar Biomedical Research Institute, the Shenzhen-HMS Initiative in International Education and Harvard Medical School through the exchange of ideas and will develop a regional and global network of future leaders in cancer research and treatment,” said Ajay Singh, HMS associate dean for global education and continuing education.
HMS-CBT, as the program is known, highlights the global nature of the HMS mission, said George Demetri, co-director of the Harvard Ludwig Center, HMS professor of medicine at Dana-Farber Cancer Institute and a co-director of HMS-CBT.
“We are very focused on the terrific talent we’re pleased to have in the United States, but many brilliant people outside the United States are not able to come here for their training,” Demetri said.
“The more we can share the expanding body of knowledge about cancer biology and therapeutics with them, I think the better it will be for patients and for the field,” he said.
Combining the study of cancer biology with the study of cancer therapeutics is intentional and necessary, Demetri said.
“We have had an explosion in knowledge relating to cancer—the diversity of cancers, the fact that cancer is not one disease but probably thousands of different diseases that we can now parse very finely in the new world of precision cancer medicine,” he said.
“How do we develop therapeutics from that knowledge? How do we apply those therapeutics? That has so much depth and robustness we can address in this program,” Demetri added.
Emphasis on interactivity
Through a combination of three in-person, three-day workshops in cities on three continents—in Doha (Qatar), London and Boston—the program emphasizes interactivity between students and Harvard faculty and between groups of students who will organize themselves via internet-connected teams around projects and ideas.
The program, which begins Oct. 24 with a workshop in Doha, will be multi-faceted, reflecting both the evolving science of cancer and the interests of its students. The first workshop will feature an introductory session on cancer biology, followed by sessions focused on links between infectious diseases, viruses and cancer.
The second workshop, starting March 18, 2016, in London, will focus on targeted therapies in cancer.
The third workshop and capstone event for the program, beginning Oct. 26, 2016, in Boston, will feature the latest developments in cancer immunotherapy, a dynamic field that is changing so rapidly that the organizers predict topics will evolve over the coming year.
Students will also be asked about what they are particularly interested in learning.
HMS-CBT program co-director Ed Harlow, the Virginia and D.K. Ludwig Professor of Cancer Research and Teaching in the HMS Department of Biological Chemistry and Molecular Pharmacology, is well known for Provocative Questions, a program he initiated in 2012 at the National Cancer Institute with former director Harold Varmus. Harlow remains special assistant to the current NCI director.
Harlow hopes to spark the same impulse with students at the first HMS-CBT workshop by inviting them to ask questions for which there may not be one simple answer.
“This sounds interesting to me, this idea that an international group of scientists will be talking to one another about this project they’ve got to work on, and how they’ll do it across global time zones and cultures,” Harlow said.
Demetri calls this adaptable form of education “bespoke.” While a library of lectures from Harvard faculty is being assembled, webinars and interaction among students and with professors will supplement them, addressing the needs and interests of the students as they emerge.
The program directors are committed to interactivity, which they say will sprout at the workshops and flower during the year through peer-to-peer projects.
“The course will create opportunities for the students to network,” said Peter Howley, the Shattuck Professor of Pathological Anatomy in the HMS Department of Microbiology and Immunobiology and an HMS-CBT co-director.
“One could imagine wonderful and creative outcomes from such learning collaborations. The course faculty will give them assignments and will be around to help the students refine and work on them, but the actual learning is going to be among themselves,” he said.
In the process, the program directors will actively assess the course to learn what succeeds, and, along with the students, will work to build upon the experiences for future offerings of this course.
”We want to provide an educational experience that’s useful for all the participants,” Demetri said.
“It is really going to be an exciting year.”
Like many other conditions, obesity is caused by an interplay between genetic and environmental factors. While efforts to combat the obesity epidemic will need to include changes in diet and exercise, insights into the genes involved may also help with prevention and treatment.
Now a research team led by Harvard Medical School investigators at Beth Israel Deaconess Medical Center and MIT reveals the mechanistic explanation behind the strongest genetic association with obesity.
The findings, published in the New England Journal of Medicine, uncover a genetic circuit that controls whether our bodies burn or store fat. Manipulating that genetic circuit may offer a new approach for obesity treatments.
The strongest genetic association with obesity lies within an unexpressed region of the FTO gene and contains 89 common variants in a region of 47,000 nucelotides.
The obesity-risk version of the region that predisposes individuals to increased body weight is found in 44 percent of individuals in European populations, but its mechanistic basis has until now remained unknown, despite extensive investigation.
Assessing epigenetic modifications
To identify the cell types in which the FTO obesity-risk region might exert its effects, the researchers analyzed information from the Roadmap Epigenomics project, which assesses chemical or “epigenetic” modifications within chromosomes that switch genes on or off.
The project’s data revealed that the strongest epigenetic signal was found in “preadipocyte” cells, the progenitor cells that go on to become fat cells.
“Previous studies attempted to uncover a link between FTO and the regulation of appetite or propensity to exercise controlled by the brain,” said the study’s lead and corresponding author Melina Claussnitzer, an HMS instructor in medicine and an investigator in the Division of Gerontology at Beth Israel Deaconess and Hebrew SeniorLife, a visiting professor at MIT’s Computer Science and Artificial Intelligence Laboratory (CSAIL), and member of the Broad Institute.
“But an unbiased look across more than one hundred human tissues and cell types indicated that the obesity-associated region acts primarily in adipocyte progenitor cells—not the brain,” she said.
The researchers collected fat, or adipose, tissue samples from individuals who carried the genetic obesity risk variant and compared it with tissue samples of individuals who did not carry the variant; they found increased expression of two distant genes, IRX3 and IRX5, indicating that these genes are under genetic control by the obesity risk variant.
“Despite years of investigating the FTO obesity region, no substantial expression differences were found between obesity-risk and non-risk individuals in brain or other tissue types, making it difficult to trace its mechanism of action,” said Manolis Kellis, professor at MIT’s CSAIL.
“We found a strong difference for both IRX3 and IRX5 in preadipocytes, revealing the target genes, cell type and developmental stage where the genetic variant acts, thus enabling us to begin dissecting its mechanism of action,” said Kellis.
Manipulating the pathways
Elevated expression of these genes resulted in a shift from energy-burning beige fat cells to energy-storing white fat cells. The researchers showed that they could manipulate this new pathway to reverse the signatures of obesity.
“By altering the expression of either gene in human preadipoctyes, we could alter adipocyte metabolism between energy storage and energy dissipation, providing a direct link between IRX3 and IRX5 expression and energy balance,” said Kellis.
To evaluate the effect of IRX3 inhibition on whole-body energy metabolism and body weight, the team inhibited the corresponding gene in the fat cells of mice. The animals’ metabolism increased and they lost weight, even though their physical activity and appetite were unchanged.
“The results at the organism level were dramatic,” said Claussnitzer.
“These mice were 50 percent thinner than the control mice, and they did not gain any weight on a high-fat diet. Instead they dissipated more energy, even in their sleep, suggesting a dramatic shift in their global metabolism. The circuitry underlying the FTO region functions like a master regulatory switch between energy storage and energy dissipation,” she said.
The researchers then sought to connect these differences in metabolism and gene expression to the genetic differences between lean and obese people within the FTO gene.
They predicted that the specific T-to-C single-nucleotide alteration within FTO is responsible for the obesity association by repressing an evolutionarily conserved gene regulator called ARID5B.
Loss of repression turns on IRX3 and IRX5 during early adipocyte differentiation, leading to a shift from beige adipocyte functions and thermogenesis, or energy burning, to white adipocyte lipid accumulation.
“We could narrow down a genetic region spanning 47,000 nucleotides to reveal a single-nucleotide alteration, and explain precisely how it leads to loss of repressor binding, activation of a regulatory region, gain of distal gene expression, a change in adipocyte metabolism, and ultimately, obesity at the organism level,” explained Claussnitzer.
“This can serve as a model for understanding the mechanistic basis of other non-coding variants in other diseases and traits. Noncoding variants make up more than 90 percent of top-scoring variants that have emerged from genome-wide association studies, which find association between genetic variants and disease risk, she said.”
Using the genome editing technique known as CRISPR/Cas9, the team found that switching the risk variant to the protective variant in preadipocytes turned off IRX3 and IRX5 and restored thermogenesis, while the reverse change turned on IRX3 and IRX5 and turned off thermogenesis.
“Bidirectional genome editing of the causal nucleotide variant allowed us to demonstrate that a single nucleotide is responsible for flipping this metabolic switch between obese and lean individuals,” said Claussnitzer.
“This is the first time that causality has been demonstrated for a genetic variant in a distal non-coding region, but we hope it will be the first of many such studies to come, now that genome editing is becoming broadly adopted,” she added.
This study was supported, in part, by grants from National Institutes of Health (R01HG004037, R01GM113708 and RC1HG005334).
An international research collaboration led by Harvard Medical School investigators at Massachusetts General Hospital has identified the first gene whose mutations cause the common form of mitral valve prolapse, a heart valve disorder that affects almost 2.5 percent of the U. S. population.
In a paper published in Nature, the team reports finding mutations in a gene called DCHS1 in affected members of three families in which mitral valve prolapse is inherited.
“This work provides insights into the pathways regulating valve growth and development and implicates a previously unrecognized basis for the long-term structural integrity of the mitral valve,” said senior author Susan Slaugenhaupt, HMS professor of neurology.
She is also scientific director of the Mass General Research Institute, an investigator at the Mass General Center for Human Genetic Research and one of the lead scientists in the collaborative group that conducted the research.
“This finding can teach us how to prevent this inborn disease from manifesting as an illness in people who inherit mutated forms of this gene,” said Robert Levine, HMS professor of medicine at Mass General and co-senior author of the Nature paper. “Understanding how defects in this gene cause errors in early valve formation can point to ways we can prevent the progression of this condition to keep the valve and the heart healthy and help the patient avoid complications.”
One of four valves controlling the flow of blood through the heart, the mitral valve lies between the left atrium and the left ventricle, which handle oxygenated blood returning from the lungs. The valve consists of two leaflets that open to let blood pass through and close to keep it from moving backwards.
In mitral valve prolapse, the leaflets become thickened, elongated and floppy, preventing the valve from closing completely and allowing blood to leak backward in a process called regurgitation. Patients with serious mitral valve prolapse can develop shortness of breath, cardiac arrhythmia, heart failure or an infection of the heart valves. Mitral valve prolapse is the most common reason for mitral valve surgery.
While mitral valve prolapse can accompany connective tissue disorders such as Marfan syndrome, its most common form runs in families without such syndromes. A specific genetic cause of familial mitral valve prolapse had not previously been identified, and the Mass General research team’s first step was to link the occurrence of mitral valve prolapse in one large family to a genetic risk factor located on chromosome 11.
Their work depended on advanced diagnostics developed by Levine and colleagues in the Mass General Cardiac Ultrasound Laboratory, which in turn relied on their discovery of the three-dimensional shape of the mitral valve.
In the current study, detailed DNA analysis of the affected members of that family identified two rare mutations in DCHS1, a chromosome 11 gene previously studied in fruit flies. Experiments in zebrafish led by David Milan, HMS assistant professor of medicine at Mass General and co-senior author of the paper, revealed that inactivation of an analogous gene led to significant defects in the development of the heart at the site corresponding to the mitral valve. These defects could be prevented by the introduction of the normal copy of the human DCHS1 gene but not the mutated version.
Based on those findings, the Mass General team sought to collaborate with other groups studying mitral valve prolapse in order to determine whether the same mutations had a role in other families with the condition. A major grant from the Leducq Foundation enabled the formation of a network consisting of 11 centers in the U.S. and four European countries. Analysis of DNA from those centers identified two French families in which mitral valve prolapse was caused by another DCHS1 mutation.
Further experiments in cells indicated that mutations associated with mitral valve prolapse significantly reduced the expression of the DCHS1 protein, which helps organize how cells are patterned into tissues.
Co-senior author Russell Norris and his team at the Medical University of South Carolina analyzed the development of mice in which one copy of the DCHS1 gene was mutated, as it is in affected family members. Their findings revealed that the gene plays a critical role in the proper formation of the mitral valve, the first evidence for the gene’s role in cardiac development, and that the mutation led to mitral valve changes resembling the human disease.
“This discovery required the cooperation of multiple disciplines and teams—ranging from clinical cardiology and ultrasound diagnostics to classical genetics, screening of potential mutations in zebrafish and functional studies in our mouse models,” said Levine.
Milan added, “As a follow-up, this same international network has been seeking other genes that cause mitral valve prolapse across the population, which should point us to common pathways that could be targets of therapies designed to prevent progression into symptomatic disease.”
In addition to a grant from the Leducq Foundation, support for the study includes grants from the Doris Duke Medical Foundation and several grants from the National Heart, Lung, and Blood Institute.
Adapted from a Mass General news release.