Recent advances in high throughput genomic technologies, coupled with large patient cohorts and and an evolving culture of rapid data sharing have led to remarkable advances in the understanding of the genetics of autism spectrum disorders. To date, the lion’s share of this progress has been with regard to the contribution of rare and de novo mutations, both in DNA sequence and chromosomal structure. The ability now to reliably and systematically identify ASD risk genes and loci provides important initial insights into both the opportunities as well as the challenges the field now faces in moving from gene discovery to an actionable understanding of pathophysiological mechanisms underlying these complex common neurodevelopmental syndromes. The lecture will provide an overview of what is now known about the genomic architecture and specific risk mutations associated with ASD, address the particular challenges posed by the discovery of mutations that have large biological effect but low population allele frequency, and consider the role that whole genome sequencing will play in the near future in enhancing the understanding of the developmental aspects of ASD risk.
Category: Uncategorized
How a single gene contributes to autism and schizophrenia
Although it is known that psychiatric disorders have a strong genetic component, untangling the web of genes contributing to each disease is a daunting task. Scientists have found hundreds of genes that are mutated in patients with disorders such as autism, but each patient usually has only a handful of these variations.
To further complicate matters, some of these genes contribute to more than one disorder. One such gene, known as Shank3, has been linked to both autism and schizophrenia.
MIT neuroscientists have now shed some light on how a single gene can play a role in more than one disease. In a study appearing in the Dec. 10 online edition of Neuron, they revealed that two different mutations of the Shank3 gene produce some distinct molecular and behavioral effects in mice.
“This study gives a glimpse into the mechanism by which different mutations within the same gene can cause distinct defects in the brain, and may help to explain how they may contribute to different disorders,” says Guoping Feng, the James W. and Patricia Poitras Professor of Neuroscience at MIT, a member of MIT’s McGovern Institute for Brain Research, a member of the Stanley Center for Psychiatric Research at the Broad Institute, and the senior author of the study.
The findings also suggest that identifying the brain circuits affected by mutated genes linked to psychiatric disease could help scientists develop more personalized treatments for patients in the future, Feng says.
The paper’s lead authors are McGovern Institute research scientist Yang Zhou, graduate students Tobias Kaiser and Xiangyu Zhang, and research affiliate Patricia Monteiro.
Disrupted communication
The protein encoded by Shank3 is found in synapses — the junctions between neurons that allow them to communicate with each other. Shank3 is a scaffold protein, meaning it helps to organize hundreds of other proteins clustered on the postsynaptic cell membrane, which are required to coordinate the cell’s response to signals from the presynaptic cell.
In 2011, Feng and colleagues showed that by deleting Shank3 in mice they could induce two of the most common traits of autism — avoidance of social interaction, and compulsive, repetitive behavior. A year earlier, researchers at the University of Montreal identified a Shank3 mutation in patients suffering from schizophrenia, which is characterized by hallucinations, cognitive impairment, and abnormal social behavior.
Feng wanted to find out how these two different mutations in the Shank3 gene could play a role in such different disorders. To do that, he and his colleagues engineered mice with each of the two mutations: The schizophrenia-related mutation results in a truncated version of the Shank3 protein, while the autism-linked mutation leads to a total loss of the Shank3 protein.
Behaviorally, the mice shared many defects, including strong anxiety. However, the mice with the autism mutation had very strong compulsive behavior, manifested by excessive grooming, which was rarely seen in mice with the schizophrenia mutation.
In the mice with the schizophrenia mutation, the researchers saw a type of behavior known as social dominance. These mice trimmed the whiskers and facial hair of the genetically normal mice sharing their cages, to an extreme extent. This is a typical way for mice to display their social dominance, Feng says.
By activating the mutations in different parts of the brain and at different stages of development, the researchers found that the two mutations affected brain circuits in different ways. The autism mutation exerted its effects early in development, primarily in a part of the brain known as the striatum, which is involved in coordinating motor planning, motivation, and habitual behavior. Feng believes that disruption of synapses in the striatum contributes to the compulsive behavior seen in those mice.
In mice carrying the schizophrenia-associated mutation, early development was normal, suggesting that truncated Shank3 can adequately fill in for the normal version during this stage. However, later in life, the truncated version of Shank3 interfered with synaptic functions and connections in the brain’s cortex, where executive functions such as thought and planning occur. This suggests that different segments of the protein — including the stretch that is missing in the schizophrenia-linked mutation — may be crucial for different roles, Feng says.
The new paper represents an important first step in understanding how different mutations in the same gene can lead to different diseases, says Joshua Gordon, an associate professor of psychiatry at Columbia University.
“The key is to identify how the different mutations alter brain function in different ways, as done here,” says Gordon, who was not involved in the research. “Autism strikes early in childhood, while schizophrenia typically arises in adolescence or early adulthood. The finding that the autism-associated mutation has effects at a younger age than the schizophrenia-associated mutation is particularly intriguing in this context.”
Modeling disease
Although only a small percentage of autism patients have mutations in Shank3, many other variant synaptic proteins have been associated with the disorder. Future studies should help to reveal more about the role of the many genes and mutations that contribute to autism and other disorders, Feng says. Shank3 alone has at least 40 identified mutations, he says.
“We cannot consider them all to be the same,” he says. “To really model these diseases, precisely mimicking each human mutation is critical.”
Understanding exactly how these mutations influence brain circuits should help researchers develop drugs that target those circuits and match them with the patients who would benefit most, Feng says, adding that a tremendous amount of work needs to be done to get to that point.
His lab is now investigating what happens in the earliest stages of the development of mice with the autism-related Shank3 mutation, and whether any of those effects can be reversed either during development or later in life.
The research was funded by the Simons Center for the Social Brain at MIT, the Stanley Center for Psychiatric Research at the Broad Institute of MIT and Harvard, the Poitras Center for Affective Disorders Research at MIT, and National Institute of Mental Health.
Stanley Center & Poitras Center Joint Translational Neuroscience Seminar Spring 2016 Series
Sounds of the McGovern Institute
Stanley Center & Poitras Center Translational Neuroscience Joint Seminar: Amy Arnsten, PhD
Stanley Center & Poitras Center Translational Neuroscience Joint Seminar
Speaker: Amy Arnsten, Yale University
December 1, 2015
The newly evolved circuits of the primate dorsolateral prefrontal cortex (dlPFC) generate the mental representations needed for working memory, the foundation of abstract thought. These layer III dlPFC pyramidal cell microcircuits are a focus of pathology in cognitive disorders such as schizophrenia and Alzheimer’s Disease. Research in the Arnsten lab has found that these circuits are uniquely regulated at the molecular level in ways that facilitate mental flexibility but make them particularly vulnerable to atrophy and degeneration. For example, in contrast to the primary visual cortex where calcium-cAMP signaling strengthens connections and increases neuronal firing, increased calcium-cAMP signaling in layer III of dlPFC weakens connections and decreases neuronal firing by opening K+ channels near the synapse. Understanding these unique properties has led to the development of treatments for dlPFC cognitive disorders in humans, e.g. Intuniv™, illustrating the importance of translational research.
MIT, Broad scientists overcome key CRISPR-Cas9 genome editing hurdle
Researchers at the Broad Institute of MIT and Harvard and the McGovern Institute for Brain Research at MIT have engineered changes to the revolutionary CRISPR-Cas9 genome editing system that significantly cut down on “off-target” editing errors. The refined technique addresses one of the major technical issues in the use of genome editing.
The CRISPR-Cas9 system works by making a precisely targeted modification in a cell’s DNA. The protein Cas9 alters the DNA at a location that is specified by a short RNA whose sequence matches that of the target site. While Cas9 is known to be highly efficient at cutting its target site, a major drawback of the system has been that, once inside a cell, it can bind to and cut additional sites that are not targeted. This has the potential to produce undesired edits that can alter gene expression or knock a gene out entirely, which might lead to the development of cancer or other problems. In a paper published today in Science, Feng Zhang and his colleagues report that changing three of the approximately 1,400 amino acids that make up the Cas9 enzyme from S. pyogenes dramatically reduced “off-target editing” to undetectable levels in the specific cases examined.
Zhang and his colleagues used knowledge about the structure of the Cas9 protein to decrease off-target cutting. DNA, which is negatively charged, binds to a groove in the Cas9 protein that is positively charged. Knowing the structure, the scientists were able to predict that replacing some of the positively charged amino acids with neutral ones would decrease the binding of “off target” sequences much more than “on target” sequences.
After experimenting with various possible changes, Zhang’s team found that mutations in three amino acids dramatically reduced “off-target” cuts. For the guide RNAs tested, “off-target” cutting was so low as to be undetectable.
The newly-engineered enzyme, which the team calls “enhanced” S. pyogenes Cas9, or eSpCas9, will be useful for genome editing applications that require a high level of specificity. The Zhang lab is immediately making the eSpCas9 enzyme available for researchers worldwide. The team believes the same charge-changing approach will work with other recently described RNA-guided DNA targeting enzymes, including Cpf1, C2C1, and C2C3, which Zhang and his collaborators reported on earlier this year.
The prospect of rapid and efficient genome editing raises many ethical and societal concerns, says Zhang, who is speaking this morning at the International Summit on Gene Editing in Washington, DC. “Many of the safety concerns are related to off-target effects,” he said. “We hope the development of eSpCas9 will help address some of those concerns, but we certainly don’t see this as a magic bullet. The field is advancing at a rapid pace, and there is still a lot to learn before we can consider applying this technology for clinical use.”
Singing in the brain
Male zebra finches, small songbirds native to central Australia, learn their songs by copying what they hear from their fathers. These songs, often used as mating calls, develop early in life as juvenile birds experiment with mimicking the sounds they hear.
MIT neuroscientists have now uncovered the brain activity that supports this learning process. Sequences of neural activity that encode the birds’ first song syllable are duplicated and altered slightly, allowing the birds to produce several variations on the original syllable. Eventually these syllables are strung together into the bird’s signature song, which remains constant for life.
“The advantage here is that in order to learn new syllables, you don’t have to learn them from scratch. You can reuse what you’ve learned and modify it slightly. We think it’s an efficient way to learn various types of syllables,” says Tatsuo Okubo, a former MIT graduate student and lead author of the study, which appears in the Nov. 30 online edition of Nature.
Okubo and his colleagues believe that this type of neural sequence duplication may also underlie other types of motor learning. For example, the sequence used to swing a tennis racket might be repurposed for a similar motion such as playing Ping-Pong. “This seems like a way that sequences might be learned and reused for anything that involves timing,” says Emily Mackevicius, an MIT graduate student who is also an author of the paper.
The paper’s senior author is Michale Fee, a professor of brain and cognitive sciences at MIT and a member of the McGovern Institute for Brain Research.
Bursting into song
Previous studies from Fee’s lab have found that a part of the brain’s cortex known as the HVC is critical for song production.
Typically, each song lasts for about one second and consists of multiple syllables. Fee’s lab has found that in adult birds, individual HVC neurons show a very brief burst of activity — about 10 milliseconds or less — at one moment during the song. Different sets of neurons are active at different times, and collectively the song is represented by this sequence of bursts.
In the new Nature study, the researchers wanted to figure out how those neural patterns develop in newly hatched zebra finches. To do that, they recorded electrical activity in HVC neurons for up to three months after the birds hatched.
When zebra finches begin to sing, about 30 days after hatching, they produce only nonsense syllables known as subsong, similar to the babble of human babies. At first, the duration of these syllables is highly variable, but after a week or so they turn into more consistent sounds called protosyllables, which last about 100 milliseconds. Each bird learns one protosyllable that forms a scaffold for subsequent syllables.
The researchers found that within the HVC, neurons fire in a sequence of short bursts corresponding to the first protosyllable that each bird learns. Most of the neurons in the HVC participate in this original sequence, but as time goes by, some of these neurons are extracted from the original sequence and produce a new, very similar sequence. This chain of neural sequences can be repurposed to produce different syllables.
“From that short sequence it splits into new sequences for the next new syllables,” Mackevicius says. “It starts with that short chain that has a lot of redundancy in it, and splits off some neurons for syllable A and some neurons for syllable B.”
This splitting of neural sequences happens repeatedly until the birds can produce between three and seven different syllables, the researchers found. This entire process takes about two months, at which point each bird has settled on its final song.
Evolution by duplication
The researchers note that this process is similar to what is believed to drive the production of new genes and traits during evolution.
“If you duplicate a gene, then you could have separate mutations in both copies of the gene and they could eventually do different functions,” Okubo says. “It’s similar with motor programs. You can duplicate the sequence and then independently modify the two daughter motor programs so that they can now each do slightly different things.”
Mackevicius is now studying how input from sound-processing parts of the brain to the HVC contributes to the formation of these neural sequences.
Mother and Child
“The Mother and Child is a powerful symbol of love and innocence, beauty and fertility. Although these maternal values, and the women who embody them, may be venerated, they are usually viewed in opposition to other values: inquiry and intellect, progress and power. But I am a neuroscientist, and I worked to create this image; and I am also the mother in it, curled up inside the tube with my infant son.” — Rebecca Saxe
This image appeared in the 2015 issue of Smithsonian Magazine.