Research Area: Genetics

Francisco J. Sánchez-Rivera

Education

  • PhD, 2016, Biology, MIT
  • BS, 2008, Microbiology, University of Puerto Rico at Mayagüez

Research Summary

The overarching goal of the Sánchez-Rivera laboratory is to elucidate the cellular and molecular mechanisms by which genetic variation shapes normal physiology and disease, particularly in the context of cancer. To do so, we develop and apply genome engineering technologies, genetically-engineered mouse models (GEMMs), and single cell lineage tracing and omics approaches to obtain comprehensive biological pictures of disease evolution at single cell resolution. By doing so, we hope to produce actionable discoveries that could pave the way for better therapeutic strategies to treat cancer and other diseases.

Awards

  • V Foundation Award, 2022
  • Hanna H. Gray Fellowship, Howard Hughes Medical Institute, 2018-2026
  • GMTEC Postdoctoral Researcher Innovation Grant, Memorial Sloan Kettering Cancer Center, 2020-2022
  • 100 inspiring Hispanic/Latinx scientists in America, Cell Mentor/Cell Press, 2020
The proteins that package DNA to fit inside cells have another role: tuning gene expression
Raleigh McElvery
May 19, 2021

The DNA inside a single human cell is several meters long — yet it must be condensed to fit inside a space one-tenth the diameter of a hair. That’s like stretching a string from Philadelphia, Pennsylvania to Washington, D.C., and then trying to stuff it into a soccer ball. Imagine then organizing all of this information for each of the body’s 3 trillion cells! The DNA is condensed by proteins called histones that create a spool around which the DNA can wrap itself. How tightly the DNA is wound determines whether it is accessible enough for other proteins to bind to and copy into RNA, toggling gene expression levels up or down.

One specialized type of histone, H2A.Z, is ubiquitous and essential among multicellular organisms. But there have been conflicting reports about how it affects gene expression, especially during embryonic development.

Several years ago, Laurie Boyer’s lab at MIT was the first to show that H2A.Z wraps the DNA located around the start sites of most genes, where the molecular machine RNA polymerase II (RNAPII) binds to copy the DNA into RNA. Boyer’s team demonstrated that removing H2A.Z prevented embryonic cells from turning on genes that are important for forming organs and tissues. But scientists still weren’t sure how H2A.Z exerted its effects.

Now, in a recent Nature Structural and Molecular Biology study, a team from the Boyer lab, led by former postdoc Constantine Mylonas, has revealed how H2A.Z regulates the ability of RNAPII to properly transcribe DNA into the messages that specify all cell types in the body. The researchers found that in embryonic stem cells, H2A.Z serves as a “yellow traffic light,” signaling RNAPII to slow the process of transcribing DNA into RNA. Although there are other proteins that also contribute to RNAPII pausing, H2A.Z establishes a second barrier to transcription that allows gene expression to be tuned in response to developmental signals.

“H2A.Z appears to regulate how fast RNAPII begins to transcribe DNA, and this allows the cell time to respond to important cues that ultimately direct a stem cell to become a brain or heart cell, for example,” says Boyer, a professor of biology and biological engineering. “This connection was a critical missing piece of the puzzle, and explains why H2A.Z is essential for development across all multicellular organisms.”

Illustration of molecules
As RNAPII starts to transcribe a gene, it encounters a cluster of eight histones (a “nucleosome”) including H2A.z, which slows its progression — allowing for tuning of gene expression in response to developmental signals.

According to Boyer, H2A.Z’s role in gene expression has been difficult to pin down because previous approaches only provided static snapshots of how proteins interact with DNA days after loss of the histone. Boyer’s team overcame this shortcoming by leveraging a system that allowed for targeted degradation of H2A.Z within hours. They combined this technique with high-resolution genomic approaches and live cell imaging of RNAPII dynamics using super-resolution microscopy. With help from Ibrahim Cissé’s lab, they were able to visualize RNAPII dynamics in real time at the single molecule level in embryonic stem cells. Upon loss of H2A.Z, they found a remarkable increase in RNAPII movement in the cells, consistent with their genomic results showing a faster release of RNAPII and an increase in transcription in the absence of H2A.Z.

Next, the researchers plan to determine precisely how H2A.Z is targeted to the start sites of genes and how it forms a barrier to RNAPII passage.

Boyer says pinpointing the way histone variants like H2A.Z control gene expression is fundamental to understanding how developmental decisions are made, and will help researchers understand why misregulation of H2A.Z has been linked to diseases such as cancer.

“Emerging evidence indicates that DNA ‘packaging proteins’ like histones directly participate in how RNAPII can read and transcribe DNA,” she explains, “and that crucial connection wasn’t clear before.”

Image credits: courtesy of Laurie Boyer
Top image: Live cell super-resolution imaging showing RNAPII dynamics at a single molecule level in embryonic stem cells. The bright and colored clusters represent RNAPII molecules.

Citation:
“A dual role for H2A. Z. 1 in modulating the dynamics of RNA Polymerase II initiation and elongation.”
Nature Structural & Molecular Biology, online May 10, 2021, DOI: 10.1038/s41594-021-00589-3
Constantine Mylonas, Choongman Lee, Alexander L. Auld, Ibrahim I. Cisse, and Laurie A. Boyer.

Kristin Knouse

Education

  • PhD, 2017, MIT; MD, 2018, Harvard Medical School
  • Undergraduate: BS, 2010, Biology, Duke University

Research Summary

We aim to understand how tissues sense and respond to damage with the goal of developing novel treatments for diverse human diseases. We focus on the mammalian liver, which has the unique ability to completely regenerate itself, in order to identify the molecular requirements for effective organ repair. To this end, we innovate genetic, molecular, and cellular tools that allow us to investigate and modulate organ injury and regeneration directly within living organisms.

Awards

  • NIH Director’s Early Independence Award, 2018
  • Henry Asbury Christian Award, 2018
Olivia Corradin

Education

  • PhD, 2015, Case Western Reserve University
  • BS, 2010, Biochemistry, Marquette University

Research Summary

Our lab studies genetic and epigenetic variation that contributes to human disease by disrupting gene expression programs. We utilize biological insights into the mechanisms of gene regulation in order to determine the impact of disease-associated variants on cellular function. We aim to identify actionable insights into disease pathogenesis by studying the confluence of genetic and epigenetic risk factors of human diseases, including multiple sclerosis and opioid use disorder.

Awards

  • NIH Director’s Pioneer Award Program Avenir Award, 2017
Using CRISPR as a research tool to develop cancer treatments

KSQ Therapeutics uses technology created at MIT to study the role of every human gene in disease biology.

Zach Winn | MIT News Office
April 23, 2021

CRISPR’s potential to prevent or treat disease is widely recognized. But the gene-editing technology can also be used as a research tool to probe and understand diseases.

That’s the basic insight behind KSQ Therapeutics. The company uses CRISPR to alter genes across millions of cells. By observing the effect of turning on and off individual genes, KSQ can decipher their role in diseases like cancer. The company uses those insights to develop new treatments.

The approach allows KSQ to evaluate the function of every gene in the human genome. It was developed at MIT by co-founder Tim Wang PhD ’17 in the labs of professors Eric Lander and David Sabatini.

“Now we can look at every single gene, which you really couldn’t do before in a human cell system, and therefore there are new aspects of biology and disease to discover, and some of these have clinical value,” says Sabatini, who is also a co-founder.

KSQ’s product pipeline includes small-molecule drugs as well as cell therapies that target genetic vulnerabilities identified from their experiments with cancer and tumor cells. KSQ believes its CRISPR-based methodology gives it a more complete understanding of disease biology than other pharmaceutical companies and thus a better chance of developing effective treatments to cancer and other complex diseases.

A tool for discovery

KSQ’s scientific co-founders had been studying the function of genes for years before advances in CRISPR allowed them to precisely edit genomes about 10 years ago. They immediately recognized CRISPR’s potential to help them understand the role of genes in disease biology.

During his PhD work, Wang and his collaborators developed a way to use CRISPR at scale, knocking out individual genes across millions of cells. By observing the impact of those changes over time, the researchers could tease out the functionality of each gene. If a cell died, they knew the gene they knocked out was essential. In cancer cells, the researchers could add drugs and see if knocking out any of the genes affected drug resistance. More sophisticated screening methods taught the researchers how different genes inhibit or drive tumor growth.

“It’s a tool for discovering human biology at scale that was not possible before CRISPR,” says KSQ co-founder Jonathan Weissman, a professor of biology at MIT and a member of the Whitehead Institute. “You can search for genes or mechanisms that can modulate essentially any disease process.”

Wang credits Sabatini with spearheading the commercialization efforts, speaking with investors, and working with MIT’s Technology Licensing Office. Wang also says MIT’s ecosystem helped him think about bringing the technology out of the lab.

“Being at MIT and in the Cambridge area probably made the leap to commercialization a bit easier than it would have been elsewhere,” Wang says. “A lot of the students are entrepreneurial, there’s that rich tradition, so that helped shape my mindset around commercialization.”

Weissman had developed a complementary, CRISPR-based technology that Wang and Sabatini knew would be useful for KSQ’s discovery platform. Around 2015, as the founders were starting the company, they also brought on co-founder William Hahn, a member of the Broad Institute of MIT and Harvard, a professor at Harvard Medical School, and the chief operating officer of the Dana-Farber Cancer Institute.

Since then, the company has advanced Wang’s method.

“They’re able to scale this to a degree that is not possible in any academic lab, even David’s,” Wang says. “The cell lines I used for my experiments were just what was easy to grow and what was in the lab, whereas KSQ is thinking about what therapies aren’t available in certain cancers and deciding what diseases to go after.”

KSQ’s gene evaluations include tens of millions of cells. The company says the data it collects has been predictive of past successes and failures in cancer drug development. Weissman equates the data to “a roadmap for finding cancer vulnerabilities.”

“Cancers have all these different escape routes,” Weissman says. “This is a way of mapping out those escape routes. If there are too many, it’s not a good target to go after, but if there is a small number, you can now start to develop therapies to block off the escape routes.”

From discovery to impact

KSQ’s lead drug candidate is in preclinical development. It targets a DNA-repair pathway identified using an updated version of Wang’s technique. The drug could treat multiple ovarian cancers as well as a disease called triple-negative breast cancer. KSQ is also currently developing a cell therapy to boost the immune system’s ability to fight tumors.

“I’ve always thought the best biotech companies start with information that other people don’t have,” Sabatini says. “I think biotech companies have to have some discovery to them. That’s enabled KSQ to go in different directions.”

The founders feel KSQ has already validated their approach and stimulated further interest in using CRISPR as a research tool.

“There’s a lot of interest in CRISPR as a therapeutic, and that’s an important aspect,” Weissman says. “But I’d argue equally important both in discovery and in therapeutics will be [using CRISPR] to identify the targets you want to go after to affect disease process. Your ability to engineer genomes or make drugs depends on knowing what genes you want to change.”

An on-off switch for gene editing
Eva Frederick | Whitehead Institute
April 9, 2021

Now, in a paper published online in Cell on April 9, researchers describe a new gene editing technology called CRISPRoff that allows researchers to control gene expression with high specificity while leaving the sequence of the DNA unchanged. Designed by Whitehead Institute Member Jonathan Weissman, University of California San Francisco assistant professor Luke Gilbert, Weissman lab postdoc James Nuñez and collaborators, the method is stable enough to be inherited through hundreds of cell divisions, and is also fully reversible.

“The big story here is we now have a simple tool that can silence the vast majority of genes,” says Weissman, who is also a professor of biology at MIT and an investigator with the Howard Hughes Medical Institute. “We can do this for multiple genes at the same time without any DNA damage, with great deal of homogeneity, and in a way that can be reversed. It’s a great tool for controlling gene expression.”

The project was partially funded by a 2017 grant from the Defense Advanced Research Projects Agency to create a reversible gene editor. “Fast forward four years [from the initial grant], and CRISPRoff finally works as envisioned in a science fiction way,” says co-senior author Gilbert. “It’s exciting to see it work so well in practice.”

Genetic engineering 2.0

The classic CRISPR-Cas9 system uses a DNA-cutting protein called Cas9 found in bacterial immune systems. The system can be targeted to specific genes in human cells using a single guide RNA, where the Cas9 proteins create tiny breaks in the DNA strand. Then the cell’s existing repair machinery patches up the holes.

Because these methods alter the underlying DNA sequence, they are permanent. Plus, their reliance on “in-house” cellular repair mechanisms means it is hard to limit the outcome to a single desired change. “As beautiful as CRISPR-Cas9 is, it hands off the repair to natural cellular processes, which are complex and multifaceted,” Weissman says. “It’s very hard to control the outcomes.”

That’s where the researchers saw an opportunity for a different kind of gene editor — one that didn’t alter the DNA sequences themselves, but changed the way they were read in the cell.

This sort of modification is what scientists call “epigenetic” — genes may be silenced or activated based on chemical changes to the DNA strand. Problems with a cell’s epigenetics are responsible for many human diseases such as Fragile X syndrome and various cancers, and can be passed down through generations.

Epigenetic gene silencing often works through methylation — the addition of chemical tags to to certain places in the DNA strand — which causes the DNA to become inaccessible to RNA polymerase, the enzyme which reads the genetic information in the DNA sequence into messenger RNA transcripts, which can ultimately be the blueprints for proteins.

Weissman and collaborators had previously created two other epigenetic editors called CRISPRi and CRISPRa — but both of these came with a caveat. In order for them to work in cells, the cells had to be continually expressing artificial proteins to maintain the changes.

“With this new CRISPRoff technology, you can [express a protein briefly] to write a program that’s remembered and carried out indefinitely by the cell,” says Gilbert. “It changes the game so now you’re basically writing a change that is passed down through cell divisions — in some ways we can learn to create a version 2.0 of CRISPR-Cas9 that is safer and just as effective, and can do all these other things as well.”

Building the switch

To build an epigenetic editor that could mimic natural DNA methylation, the researchers created a tiny protein machine that, guided by small RNAs, can tack methyl groups onto specific spots on the strand. These methylated genes are then “silenced,” or turned off, hence the name CRISPRoff.

Because the method does not alter the sequence of the DNA strand, the researchers can reverse the silencing effect using enzymes that remove methyl groups, a method they called CRISPRon.

As they tested CRISPRoff in different conditions, the researchers discovered a few interesting features of the new system. For one thing, they could target the method to the vast majority of genes in the human genome — and it worked not just for the genes themselves, but also for other regions of DNA that control gene expression but do not code for proteins. “That was a huge shock even for us, because we thought it was only going to be applicable for a subset of genes,” says first author Nuñez.

Also, surprisingly to the researchers, CRISPRoff was even able to silence genes that did not have large methylated regions called CpG islands, which had previously been thought necessary to any DNA methylation mechanism.

“What was thought before this work was that the 30 percent of genes that do not have a CpG island were not controlled by DNA methylation,” Gilbert says. “But our work clearly shows that you don’t require a CpG island to turn genes off by methylation. That, to me, was a major surprise.”

CRISPRoff in research and therapy

To investigate the potential of CRISPRoff for practical applications, the scientists tested the method in induced pluripotent stem cells. These are cells that can turn into countless cell types in the body depending on the cocktail of molecules they are exposed to, and thus are powerful models for studying the development and function of particular cell types.

The researchers chose a gene to silence in the stem cells, and then induced them to turn into nerve cells called neurons. When they looked for the same gene in the neurons, they discovered that it had remained silenced in 90 percent of the cells, revealing that cells retain a memory of epigenetic modifications made by the CRISPRoff system even as they change cell type.

They also selected one gene to use as an example of how CRISPRoff might be applied to therapeutics: the gene that codes for Tau protein, which is implicated in Alzheimer’s disease. After testing the method in neurons, they were able to show that using CRISPRoff could be used to turn Tau expression down, although not entirely off.  “What we showed is that this is a viable strategy for silencing Tau and preventing that protein from being expressed,” Weissman says. “The question is, then, how do you deliver this to an adult? And would it really be enough to impact Alzheimer’s? Those are big open questions, especially the latter.”

Even if CRISPRoff does not lead to Alzheimer’s therapies, there are many other conditions it could potentially be applied to. And while delivery to specific tissues remains a challenge for gene editing technologies such as CRISPRoff, “we showed that you can deliver it transiently as a DNA or as an RNA, the same technology that’s the basis of the Moderna and BioNTech coronavirus vaccine,” Weissman says.

Weissman, Gilbert, and collaborators are enthusiastic about the potential of CRISPRoff for research as well.  “Since we now can sort of silence any part of the genome that we want, it’s a great tool for exploring the function of the genome,” Weissman says.

Plus, having a reliable system to alter a cell’s epigenetics could help researchers learn the mechanisms by which epigenetic modifications are passed down through cell divisions. “I think our tool really allows us to begin to study the mechanism of heritability, especially epigenetic heritability, which is a huge question in the biomedical sciences,” Nuñez says.

Eyeless roundworms sense color

C. elegans compares the ratio of wavelengths in its environment to avoid dangerous bacteria that secrete colorful toxins.

Raleigh McElvery | Department of Biology
March 4, 2021

Roundworms don’t have eyes or the light-absorbing molecules required to see. Yet, new research shows they can somehow sense color. The study, published on March 5 in the journal Science, suggests worms use this ability to assess the risk of feasting on potentially dangerous bacteria that secrete blue toxins. The researchers pinpointed two genes that contribute to this spectral sensitivity and are conserved across many organisms, including humans.

“It’s amazing to me that a tiny worm — with neither eyes nor the molecular machinery used by eyes to detect colors — can identify and avoid a toxic bacterium based, in part, on its blue color,” says H. Robert Horvitz, the David H. Koch Professor of Biology at MIT, a member of the McGovern Institute for Brain Research and the Koch Institute for Integrative Cancer Research, Howard Hughes Medical Institute Investigator, and the co-senior author of the study. “One of the joys of being a biologist is the opportunity to discover things about nature that no one has ever imagined before.”

The roundworm in question, Caenorhabditis elegans, is only about a millimeter long. Despite their minute stature and simple nervous system, these nematodes display a complex repertoire of behaviors. They can smell, taste, sense touch, react to temperature, and even escape or change their feeding patterns in response to bright, blue light. Although researchers once thought that these worms bury themselves deep in soil, it’s becoming increasingly clear that C. elegans prefers compost heaps above ground that offer some sun exposure. As a result, roundworms may have a need for light- and color-sensing capabilities after all.

The decomposing organic matter where C. elegans resides offers an array of scrumptious microbes, including bacteria like Pseudomonas aeruginosa, which secretes a distinctive blue toxin. Previous studies showed that worms in the lab feed on a lawn of P. aeruginosa for a few hours and then begin avoiding their food — perhaps because the bacteria continue to divide and excrete more of the colorful poison. Dipon Ghosh, Horvitz lab postdoc and the study’s first author, wondered whether the worms were using the distinctive color to determine if their meal was too toxic to consume.

Over the course of his experiments, Ghosh noticed that his worms were more likely to flee the colorful bacterial lawn if it was bathed in white light from a nearby LED bulb. This finding was curious on its own, but Ghosh wanted know if the blue toxin played a role as well.

To test this theory, he first exchanged the blue toxin for a harmless dye of the same color, and then for a clear, colorless toxin. On its own, neither substitute was sufficient to spur avoidance. Only together did they prompt a response — suggesting the worms were assessing both the toxic nature and the color of the P. aeruginosa secretions simultaneously. Once again, this behavioral pattern only emerged in the presence of the LED’s white light.

Intrigued, Ghosh wanted to examine what it was about the blue color that triggered avoidance. This time, he used two colored LED lights, one blue and one amber, to tint the ambient light. In doing so, he could control the ratio of wavelengths without changing the total energy delivered to the worms. The beam had previously contained the entire visible spectrum, but mixing the amber and blue bulbs allowed Ghosh to tweak the relative amounts of short-wavelength blue light and long-wavelength amber light. Surprisingly, the worms only fled the bacterial lawn when their environment was bathed in light with specific blue:amber ratios.

“We were able to definitively show that worms aren’t sensing the world in grayscale and simply evaluating the levels of brightness and darkness,” Ghosh says. “They’re actually comparing ratios of wavelengths and using that information to make decisions — which was thoroughly unexpected.”

It wasn’t until Ghosh ran his experiments again, this time using various types of wild C. elegans, that he realized the popular laboratory strain he’d been using was actually less color-sensitive compared to its close relatives. After analyzing the genomes of these worms, he was able to identify two genes in particular (called jkk-1 and lec-3) that contributed to these variations in color-dependent foraging.

Although the two genes play many important functions in a variety of organisms, including humans, they are both involved in molecular pathways that help cells respond to stress caused by damaging ultraviolet light.

“We’ve discovered that the color of light in the worm’s environment can influence how the worm navigates the world,” Ghosh says. “But our work suggests that many genes, in addition to the two we’ve already identified, can affect color sensitivity, and we’re now exploring how.”

The notion that worms can sense color is “astounding” and showcases nature’s innovation, according to Leslie Vosshall, Robin Chemers Neustein Professor and Howard Hughes Medical Institute Investigator at The Rockefeller University, who was not involved in the study. “These worms are sliding around in a dim muck with colorful, toxic bacteria. It would be helpful to see and avoid them, so the worms somehow evolved a completely new way to see.”

Vosshall is curious about which cells in C. elegans help discriminate light, as well as the specific roles that the jkk-1 and lec-3 genes play in mediating light perception. “This paper, like all important papers, raises many additional questions,” she says.

Ghosh suspects the lab’s findings could generalize to other critters besides roundworms. If nothing else, it’s clear that light-sensitivity does not always require vision — or eyes. C. elegans are seeing the light, and now so are the biologists.

This research was funded by the Howard Hughes Medical Institute and National Institute of General Medical Sciences.

Cells are known by the company they keep
Eva Frederick
March 2, 2021

In the paper, published online March 1 in the journal Cell Metabolism, researchers at Whitehead Institute and the Morgridge Institute for Research performed CRISPR-based genetic screens of cells cultured in either traditional media or a new physiologic medium previously designed in the Sabatini Lab at Whitehead Institute designed to more closely reflect the nutrient composition of human blood. The screen revealed that different genes became essential for survival and reproduction in the various conditions.

“This work underscores the importance of using more human-like, physiologically relevant media for culturing human cancer cell lines,” said Whitehead Institute Member and co-senior author David Sabatini, who is also a professor of biology at the Massachusetts Institute of Technology and an investigator of the Howard Hughes Medical Institute. “The information we can learn from screens in human plasma-like media — or media designed to mimic other fluids throughout the body — may inform new therapeutic methods down the line.”

The widespread use of a human plasma-like medium could open the door for many researchers to conduct experiments in the lab that could have more relevance to human disease, but without complicated methods or prohibitive costs.

“Medium composition is both relatively accessible and quite flexible,” said co-senior author Jason Cantor, an Investigator at the Morgridge Institute for Research and an assistant professor of biochemistry at the University of Wisconsin-Madison, and a former postdoc in Sabatini’s lab. “Not all researchers have access to specialized tissue culture incubators, nor can everyone easily pursue some of the more complex 3D and co-culture methods, but most can get their hands on a bottle of media.”

The big screen

The idea that different environmental conditions may lead to different genes being essential is not a new one. “People have done this in microorganisms, and shown that if you throw [bacteria] into different growth conditions — the contributions of different genes to cell fitness can change,” Cantor said.

With this reasoning in mind — that medium composition could affect which genes become necessary for human cells to grow — the researchers set up screens to identify essential genes in a single leukemia cell line in different kinds of culture media. One batch was grown in a traditional medium, and another cultured in the lab’s new medium called Human Plasma-Like Medium, or HPLM, which has a metabolic composition more reflective of that in human blood.

The approach they used — called a CRISPR screen —  takes advantage of CRISPR-Cas9 gene editing technology to systematically snip and knock out genes across the genome, with the goal of creating a population of cells in which every possible gene knockout is represented. The expression of some genes is essential to survival, and cells grow substantially slower or die when those genes are deleted. Other cells may have trouble functioning, and some may grow even faster. Once the pooled cells have had a chance to grow and proliferate, researchers sequence the genetic material of the entire population to determine which genes were critical for survival within the given screen.

Once they completed the initial screens, the researchers identified hundreds of genes that were conditionally essential — that is, necessary for cell growth in one medium versus another. Interestingly, these medium-dependent essential genes collectively had roles in a number of different biological processes.

To determine how much these genes were dependent on the type of cells studied, the researchers then ran similar screens across a panel of human blood cancer cell lines, and then pursued follow-up work to understand why certain genes were identified as conditionally essential.

Ultimately, they uncovered the underlying gene-nutrient interactions, and specifically for these hit genes, traced the effects to availability of certain metabolites — the nutrients and small molecules necessary for metabolism — that are uniquely defined in HPLM versus the traditional media.

The next steps

CRISPR screens can help scientists identify potential drug targets and map out important cellular interactions to inform cancer therapies. “There are so many ways that people use CRISPR screens right now,” said Cantor. “What this study is showing is that the availability metabolites can have a major impact on which genes are important for cell growth, and so I think there are a lot of implications here in terms of how these types of screens could be performed in the future in order to potentially increase the fidelity of what we see in the lab and what might happen in the body.”

The research also establishes more nuanced relationships between cells’ genes and their environment. “What this allows us to do down the line, theoretically, is to tune how important a gene is — how important the encoded protein is — by manipulating metabolite levels in the blood,” said Cantor. “That’s one of our bigger-picture ideas.”

In the future, these relationships could inform cancer treatments. For example, if scientists could “tune” the importance of a specific gene for cancer cell growth, then the protein encoded by that gene could become a more promising drug target — in effect, tricking cancer cells into revealing possible context-dependent vulnerabilities. “The idea of targeting metabolites to treat cancer isn’t itself new — in fact, it [underlies] a well-established anti-cancer therapeutic enzyme still in use today — but I think our work maybe enables us to look for ways to couple this type of approach with other targeted therapies.”

“At our core, we are a basic cell biology lab,” added Nicholas Rossiter, a technician in Cantor’s lab and the first author of the study. “But whenever you’re studying basic cell biology, there’s the potential to translate it into therapeutic strategy. Our plan is just to keep on chugging along in our lab and studying how exactly cell biology can be influenced by these environmental factors. We do the basics, and then there will hopefully be some auspicious findings that can be carried on into therapeutics.”

Snake sex chromosomes say less about sex and more about survival
Eva Frederick | Whitehead Institute
January 21, 2021

Sex-specific chromosomes are a dangerous place to be, if you’re a gene. Because these chromosomes — Y chromosomes, in humans — do not have a matching chromosome with which to exchange genetic information, they are prone to losing non-essential genes left and right in a process called genetic decay.

Now, a new study from research scientist Daniel Winston Bellott in the lab of Whitehead Institute Member David Page broadens our understanding of what makes a gene able to survive on a sex-specific chromosome by looking at one especially slithery branch of the evolutionary tree: snakes.

Comparing surviving genes on snake sex-specific chromosomes to those that are lost to the ravages of time can teach scientists about the evolutionary pressures that shaped sex chromosomes as we know them today. “You might think, ‘These are sex chromosomes, so the surviving genes should have something to do with sex, right?’” Bellott said. “But they don’t.”

Instead, many of these genes are essential to the survival of the animal, and take part in key developmental processes. “It turns out that these survivor genes on sex-specific chromosomes may play a very big role in governing how all of the genes across all the chromosomes are read, interpreted and expressed,” said Page, who is also a professor of biology at Massachusetts Institute of Technology (MIT) and an Investigator of the Howard Hughes Medical Institute. “Winston’s study is absolutely foundational to our understanding of what the sex chromosomes are, how the two sexes come to be, and how health and disease traits play out similarly or differently in males and females.”

What is a sex chromosome, anyway? 

Over the course of evolution, all sex chromosomes start out as regular, matching chromosomes called autosomes. Then, somewhere along the line, a mutation happens, and one of the chromosomes gains a “switch,” that, when present, causes an embryo to to develop as a specific sex. “It’s actually really easy to make a sex chromosome,” said Bellott. “In most cases, you only need to change one or two genes and you’ve started the sex chromosome system.”

This process has happened numerous times during the course of evolution. It makes sense; sexual reproduction is an efficient way to ensure genetic diversity. But the whole thing is a bit mysterious; are, for example, certain chromosomes predisposed to become sex chromosomes?

That’s where Bellott thought snakes could be especially helpful. “Snakes have a relatively old system of sex chromosomes, where you have a lot of time for the chromosomes to diverge,” Bellott said. “Time has swept away all the genes that aren’t important, and you can see what kind of genes are left.”

Their sex chromosome system also evolved from different autosomes, some 100 million years after humans’, and thus would provide a useful vantage point from which to consider our own genomes.

To learn more about the evolution of these chromosomes, Bellott and Page first gathered a list of “ancestral genes,” which were likely on the chromosome from which the snake sex chromosomes evolved. New sequencing data for several species of animals distantly related to snakes meant that they had a more complete list of these genes — 1,648 to be exact.

Bellott began painstakingly sifting through the genes that remained on the sex-specific chromosomes of three species of snake: the pygmy rattlesnake, mountain garter snake, and the five-pacer viper. He eventually identified 103 ancestral genes that had survived as long as 90 million years of evolution on the snakes’ sex chromosomes. With this list in hand, Bellott could then ask what these surviving genes had in common that set them apart from the hundreds of genes that were swept off the snakes’ sex chromosomes by genetic decay.

What makes a survivor? 

To Bellott’s surprise, the genes that remained on the snakes’ sex-specific chromosome had nothing to do with sex determination; neither were they expressed more often in sex-specific tissues, or more often in one sex than the other.

Instead, Bellott and Page’s research identified three key properties that led to a gene’s survival on snake sex-specific chromosomes. First, the gene must be dosage sensitive. In other words, the snake’s body depends on its cells to produce an exact amount of that gene’s protein product. Any more, or any less, and the snake experiences illness or death. Second, a surviving gene is likely broadly expressed in different tissues across the body, not localized to one specific organ or area. And third, surviving genes are subject to strong purifying, or negative, selection. Simply put, this means that if something goes wrong with one of these genes, the snake has a slim chance of survival or producing offspring.

When Bellott dove deeper into the genes’ function, he discovered that for many of them the equivalent gene in humans played a role in key developmental processes such as the formation of the face. When these genes were mutated in humans, their faces — and other essential parts of the body — would not develop properly. “What Winston is seeing here is that the genes that were preserved on the sex specific chromosomes in snakes are disproportionately involved in birth defects in people,” Page said. “We think that nature is selecting for the survival of [sex chromosome] genes whose dosage in certain parts of embryonic development is especially critical.”

In time, Bellott said, this may allow scientists to predict genes whose role in developmental disorders is yet to be discovered.  “In some sense, you get to the place where you’re starting to work the experiment backwards in your mind, and say, ‘Let’s take the set of genes that are on sex specific chromosomes in snakes and birds, but that have not yet been implicated in birth defects in humans,’” Page said. “They might be prime candidates to be responsible for heretofore unexplained birth defects.”

From snakes to humans

Next, the researchers sought to broaden their scope. They compared ancestral genes across the three species of snakes and 38 species of birds and mammals with a larger pool of genes that made it to the present day. Many of the surviving genes on bird and mammal chromosomes had different functions than those on snake chromosomes, but again, most had little to do with sex determination.

“Adding the snakes in with the birds and the mammals gave Winston enough data points to be able to see further and to see more precisely, and now for the first time, he was able to confirm something that we had been suspecting for a long time but really didn’t have sufficient data to pin down,” Page said. “And that is that the chromosomes that became sex chromosomes were not sort of inclined to function in sex differences. Before they got picked out of the crowd, they weren’t specialized towards differentiating between the sexes in much of any way.”

Then, as genes were lost over time, evolutionary pressures ensured that the same sort of genes survived. This idea that sex chromosomes — besides their key developmental switch — have little do do with sex determination challenges the common notion of what a sex chromosome actually is.

“I hope people will pick up on this idea that the chromosomes that became sex chromosomes weren’t in any way preordained,” Page said. “They were just ordinary chromosomes out for a walk in the park, and something happened.”

In the future, Bellott and Page plan to further broaden their scope to include other animals, toward the ultimate goal of understanding our own sex chromosomes. “We take these results, and we turn them into a lens through which we look at sex differences in health and disease in our own species,” Page said. “This research really refines our ideas about what it means to be a gene on the human X or Y chromosome, and how we should think about those genes that survive.”

Notes

Bellott, D.W. and Page, D.C. “Dosage-sensitive functions in embryonic development drove the survival of genes on sex-specific chromosomes in snakes, birds, and mammals.” Genome Research, Jan. 21, 2021. DOI: 10.1101/gr.268516.120.