Research Area: Stem Cell and Developmental Biology

Yukiko Yamashita, unraveler of stem cells’ secrets

The MIT biologist’s research has shed light on the immortality of germline cells and the function of “junk DNA.”

Anne Trafton | MIT News Office
March 22, 2022

When cells divide, they usually generate two identical daughter cells. However, there are some important exceptions to this rule: When stem cells divide, they often produce one differentiated cell along with another stem cell, to maintain the pool of stem cells.

Yukiko Yamashita has spent much of her career exploring how these “asymmetrical” cell divisions occur. These processes are critically important not only for cells to develop into different types of tissue, but also for germline cells such as eggs and sperm to maintain their viability from generation to generation.

“We came from our parents’ germ cells, who used to be also single cells who came from the germ cells of their parents, who used to be single cells that came from their parents, and so on. That means our existence can be tracked through the history of multicellular life,” Yamashita says. “How germ cells manage to not go extinct, while our somatic cells cannot last that long, is a fascinating question.”

Yamashita, who began her faculty career at the University of Michigan, joined MIT and the Whitehead Institute in 2020, as the inaugural holder of the Susan Lindquist Chair for Women in Science and a professor in the Department of Biology. She was drawn to MIT, she says, by the eagerness to explore new ideas that she found among other scientists.

“When I visited MIT, I really enjoyed talking to people here,” she says. “They are very curious, and they are very open to unconventional ideas. I realized I would have a lot of fun if I came here.”

Exploring paradoxes

Before she even knew what a scientist was, Yamashita knew that she wanted to be one.

“My father was an admirer of Albert Einstein, so because of that, I grew up thinking that the pursuit of the truth is the best thing you could do with your life,” she recalls. “At the age of 2 or 3, I didn’t know there was such a thing as a professor, or such a thing as a scientist, but I thought doing science was probably the coolest thing I could do.”

Yamashita majored in biology at Kyoto University and then stayed to pursue her PhD, studying how cells make exact copies of themselves when they divide. As a postdoc at Stanford University, she became interested in the exceptions to that carefully orchestrated process, and began to study how cells undergo divisions that produce daughter cells that are not identical. This kind of asymmetric division is critical for multicellular organisms, which begin life as a single cell that eventually differentiates into many types of tissue.

Those studies led to a discovery that helped to overturn previous theories about the role of so-called junk DNA. These sequences, which make up most of the genome, were thought to be essentially useless because they don’t code for any proteins. To Yamashita, it seemed paradoxical that cells would carry so much DNA that wasn’t serving any purpose.

“I couldn’t really believe that huge amount of our DNA is junk, because every time a cell divides, it still has the burden of replicating that junk,” she says. “So, my lab started studying the function of that junk, and then we realized it is a really important part of the chromosome.”

In human cells, the genome is stored on 23 pairs of chromosomes. Keeping all of those chromosomes together is critical to cells’ ability to copy genes when they are needed. Over several years, Yamashita and her colleagues at the University of Michigan, and then at MIT, discovered that stretches of junk DNA act like bar codes, labeling each chromosome and helping them bind to proteins that bundle chromosomes together within the cell nucleus.

Without those barcodes, chromosomes scatter and start to leak out of the cell’s nucleus. Another intriguing observation regarding these stretches of junk DNA was that they have much greater variability between different species than protein-coding regions of DNA. By crossing two different species of fruit flies, Yamashita showed that in cells of the hybrid offspring flies, chromosomes leak out just as they would if they lost their barcodes, suggesting that the codes are specific to each species.

“We think that might be one of the big reasons why different species become incompatible, because they don’t have the right information to bundle all of their chromosomes together into one place,” Yamashita says.

Stem cell longevity

Yamashita’s interest in stem cells also led her to study how germline cells (the cells that give rise to eggs and sperm cells) maintain their viability so much longer than regular body cells across generations. In typical animal cells, one factor that contributes to age-related decline is loss of genetic sequences that encode genes that cells use continuously, such as genes for ribosomal RNAs.

A typical human cell may have hundreds of copies of these critical genes, but as cells age, they lose some of them. For germline cells, this can be detrimental because if the numbers get too low, the cells can no longer form viable daughter cells.

Yamashita and her colleagues found that germline cells overcome this by tearing sections of DNA out of one daughter cell during cell division and transferring them to the other daughter cell. That way, one daughter cell has the full complement of those genes restored, while the other cell is sacrificed.

That wasteful strategy would likely be too extravagant to work for all cells in the body, but for the small population of germline cells, the tradeoff is worthwhile, Yamashita says.

“If skin cells did that kind of thing, where every time you make one cell, you are essentially trashing the other one, you couldn’t afford it. You would be wasting too many resources,” she says. “Germ cells are not critical for viability of an organism. You have the luxury to put many resources into them but then let only half of the cells recover.”

Uncovering the mysteries of methylation in plants
Eva Frederick | Whitehead Institute
January 11, 2022

Growing up is a complex process for multi-celled organisms — plants included. In the days or weeks it takes to go from a seed to a sprout to a full plant, plants express hundreds of genes in different places at different times.

In order to conduct this symphony of genes, plants rely in part on an elegant regulatory method called DNA methylation. By adding or removing small molecules called methyl groups to the DNA strand, the plant can silence or activate different regions of its genetic code without changing the underlying sequence.

In a new paper from the lab of Whitehead Institute Member Mary Gehring, researchers led by former Gehring lab postdoc Ben Williams (now an assistant professor at the University of California, Berkeley) tease apart the role of proteins governing this system of genetic control, and reveal how enzymes that regulate methylation can affect essential decisions for plants such as when to produce flowers. “We’re starting to see that there is actually a broader role for  demethylation [in plant development] than we thought,” Gehring said.

In the model plant Arabidopsis thaliana, methylation is regulated in part by enzymes encoded by  a family of four genes called the DEMETER genes. The protein products of these genes are in charge of demethylating, or removing those methyl groups from the DNA, allowing different parts of the strand to be expressed. “You have these enzymes that can come in and completely change the way the DNA is read in different cells, which I find super interesting,” Williams said.

But teasing apart the role of each DEMETER gene has proved difficult in the past, because one member of this gene family in particular, called DME, is essential for seed development. Knock out DME, and the seed is aborted. “We had to design a synthetic gene to get around that,” Williams said. “We had to create plants that would rescue the reproductive failure, but then still be mutated throughout the rest of the life cycle.”

The researchers accomplished this by putting the DME gene under the control of a genetic element called a promoter that allowed it to be expressed in a cell that only existed in the plant during seed development. Once the plant was past the critical point where DME was needed for development, the gene would no longer be expressed, allowing the plant to grow up as a dme knockout. “It was an exciting thing, finally being able to create this knockout,” Gehring said.

Now, for the first time, the researchers could create plants with any combination of the DEMETER family genes knocked out, and then compare them to try and understand what the enzymes produced by each of the four genes was doing.

As expected, plants missing any of the DEMETER demethylases ended up with areas of their genomes with too many methyl groups (this is called hypermethylation). These areas were often overlapping, suggesting that the four DEMETER genes shared responsibility for demethylating certain areas of the genome.

“When one of these enzymes is gone, the others are surprisingly good at knowing that they need to step forward and do the job instead,” Williams said. “So the system has flexibility built in, which makes sense if it’s going to be involved in making important decisions like when to make flowers. You’d want there to be multiple layers of responsibility, right? It’s like in an organization, you don’t want to load all responsibility on one person — you’d want a few people who can take on that responsibility.”

Williams hypothesizes that while the DEMETER enzymes could step in for each other when needed, each specialized in demethylating DNA in particular types of plant tissue. “If you look at the protein sequences,they are actually really similar,” he said. “What’s different about them is they’re expressed in different cell types.”

A crucial finding of the study came about when the researchers knocked out all four genes in the DEMETER family at the same time. “All flowering plants have this really important decision of when to make flowers,” Williams said. “For plants out in the wild, that decision is usually dependent on temperature and pollinators. What we found really strange is that these mutants just flowered straight away. It’s almost like they weren’t even putting any effort into the decision. They made a few leaves, then boom, flower.”

When the researchers dove deeper, they saw that one area of the genome in particular that controls flowering time is under very careful and continuous regulation by methylating and demethylating enzymes. “We don’t really know why they’re doing that,” he said. “But when you knock out the demethylases, that gene just becomes methylated, and it’s then switched off. And that just sends plants into an automatic flowering state.”

In the future, the researchers plan to investigate other outcomes associated with their quadruple knockout of the DEMETER genes. “When we knocked out all four of the enzymes, it led to a lot of interesting phenotypes and tons of stuff to study,” Williams said. “We’ve learned through doing this that with DEMETER, like many gene families, we had to knock out all the players to find out the importance of what they are doing.”

Gehring will continue the research at Whitehead Institute. Williams recently started his own lab at the University of California, Berkeley. “I feel very lucky because this project has given me two or three different avenues that I can pursue in my new lab,” Williams said. “It has opened a lot of doors, which is very rewarding.”

3 Questions: Kristin Knouse on the liver’s regenerative capabilities

The clinically-trained cell biologist exploits the liver’s unique capacities in search of new medical applications.

Grace van Deelen | Department of Biology
December 15, 2021

Why is the liver the only human organ that can regenerate? How does it know when it’s been injured? What can our understanding of the liver contribute to regenerative medicine? These are just some of the questions that new assistant professor of biology Kristin Knouse and her lab members are asking in their research at the Koch Institute for Integrative Cancer Research. Knouse sat down to discuss why the liver is so unique, what lessons we might learn from the organ, and what its regeneration might teach us about cancer.

Q: Your lab is interested in questions about how body tissues sense and respond to damage. What is it about the liver that makes it a good tool to model those questions?

A: I’ve always felt that we, as scientists, have so much to gain from treasuring nature’s exceptions, because those exceptions can shine light onto a completely unknown area of biology and provide building blocks to confer such novelty to other systems. When it comes to organ regeneration in mammals, the liver is that exception. It is the only solid organ that can completely regenerate itself. You can damage or remove over 75 percent of the liver and the organ will completely regenerate in a matter of weeks. The liver therefore contains the instructions for how to regenerate a solid organ; however, we have yet to access and interpret those instructions. If we could fully understand how the liver is able to regenerate itself, perhaps one day we could coax other solid organs to do the same.

There are some things we already know about liver regeneration, such as when it begins, what genes are expressed, and how long it takes. However, we still don’t understand why the liver can regenerate but other organs cannot. Why is it that these fully differentiated liver cells — cells that have already assumed specialized roles in the liver — can re-enter the cell cycle and regenerate the organ? We don’t have a molecular explanation for this. Our lab is working to answer this fundamental question of cell and organ biology and apply our discoveries to unlock new approaches for regenerative medicine. In this regard, I don’t necessarily consider myself exclusively a liver biologist, but rather someone who is leveraging the liver to address this much broader biological problem.

Q: As an MD/PhD student, you conducted your graduate research in the lab of the late Professor Angelika Amon here at MIT. How did your work in her lab lead to an interest in studying the liver’s regenerative capacities?

A: What was incredible about being in Angelika’s lab was that she had an interest in almost everything and gave me tremendous independence in what I pursued. I began my graduate research in her lab with an interest in cell division, and I was doing experiments to observe how cells from different mammalian tissues divide. I was isolating cells from different mouse tissues and then studying them in culture. In doing that, I found that when the cells were isolated and grown in a dish they could not segregate their chromosomes properly, suggesting that the tissue environment was essential for accurate cell division. In order to further study and compare these two different contexts — cells in a tissue versus cells in culture — I was keen to study a tissue in which I could observe a lot of cells undergoing cell division at the same time.

So I thought back to my time in medical school, and I remembered that the liver has the ability to completely regenerate itself. With a single surgery to remove part of the liver, I could stimulate millions of cells to divide. I therefore began exploiting liver regeneration as a means of studying chromosome segregation in tissue. But as I continued to perform surgeries on mice and watch the liver rapidly regenerate itself, I couldn’t help but become absolutely fascinated by this exceptional biological process. It was that fascination with this incredibly unique but poorly understood phenomenon — alongside the realization that there was a huge, unmet medical need in the area of regeneration — that convinced me to dedicate my career to studying this.

Q: What kinds of clinical applications might a better understanding of organ regeneration lead to, and what role do you see your lab playing in that research?

A: The most proximal medical application for our work is to confer regenerative capacity to organs that are currently non-regenerative. As we begin to achieve a molecular understanding of how and why the liver can regenerate, we put ourselves in a powerful position to identify and surmount the barriers to regeneration in non-regenerative tissues, such as the heart and nervous system. By answering these complementary questions, we bring ourselves closer to the possibility that, one day, if someone has a heart attack or a spinal cord injury, we could deliver a therapy that stimulates the tissue to regenerate itself. I realize that may sound like a moonshot now, but I don’t think any problem is insurmountable so long as it can be broken down into a series of tractable questions.

Beyond regenerative medicine, I believe our work studying liver regeneration also has implications for cancer. At first glance this may seem counterintuitive, as rapid regrowth is the exact opposite of what we want cancer cells to do. However, the reality is that the majority of cancer-related deaths are attributable not to the rapidly proliferating cells that constitute primary tumors, but rather to the cells that disperse from the primary tumor and lie dormant for years before manifesting as metastatic disease and creating another tumor. These dormant cells evade most of the cancer therapies designed to target rapidly proliferating cells. If you think about it, these dormant cells are not unlike the liver: they are quiet for months, maybe years, and then suddenly awaken. I hope that as we start to understand more about the liver, we might learn how to target these dormant cancer cells, prevent metastatic disease, and thereby offer lasting cancer cures.

How sea stars get their symmetry
Greta Friar | Whitehead Institute
November 4, 2021

In a paper published Nov. 4 in the journal Current Biology, Zak Swartz, a postdoctoral researcher at Whitehead Institute, along with researchers in the lab of Whitehead Institute Member Iain Cheeseman and collaborators at the Massachusetts Institute of Technology (MIT), the University of Miami, and the Marine Biological Laboratory Embryology Course delve into the origins of the initial polarity in an animal’s first cell, which establishes an axis of symmetry for the developing organism and underlies the first steps of development. Their research reveals how a specific protein, called Dishevelled, localizes in a cell to help create this polarity.

All multicellular organisms begin as a single cell — the oocyte, precursor cell to the egg — which carries within it a “plan” for the fully developed, complex creature it will become. “How that multifunctional body plan is created is one of the deepest questions in developmental biology,” said Swartz.

“Sea stars, and a huge diversity of other animals, have an incredibly complex body plan, none of which is possible without the polarity of the initial cell,” said Cheeseman. “This work shows how the polarity originates as early as the meiotic divisions in the developing oocyte through an unexpected strategy to break its symmetry and achieve the asymmetric distribution of developmental factors.”

To study the intricate process of body patterning, Cheeseman Lab researchers used a type of sea star called the bat star, or Patiria miniata. These colorful animals are radially symmetric as adults — they usually have five arms, sometimes more — but as larvae they are bilaterally symmetric like humans.

The sea star larvae’s mirror-image symmetry is established when they are egg cells, called oocytes. A key step in the development of this organization involves a protein called Dishevelled, which localizes to the vegetal, or “bottom” end of the oocyte (which will define the posterior end of the embryo) as the cell gets ready to divide into two daughter cells.

Dishevelled — so named because a mutation in the homologous protein in fruit flies lends their tiny hairs a messy, tousled look  — is a component of a common signaling pathway called the Wnt pathway, which is found in many creatures throughout the animal kingdom. The pathway serves various purposes in the cells, from body patterning to cell proliferation. “The Wnt pathway is evolutionarily ancient,” Swartz said. “Jellyfish use it, sea stars use it, people use it, and I think that’s really quite profound.”

In the sea stars, the pathway provides a link between the initial asymmetry of the oocyte and the polarity of the resulting embryo. Dishevelled serves as a messenger on the inside of the sea star’s cells, relaying external signals that are then transmitted through a molecular pathway to the cells’ nuclei.

The researchers used time-lapse imaging to visualize how Dishevelled moved around the oocyte as the cell went through different phases of its development. When the sea star oocyte was in a non-dividing phase, Dishevelled could be found distributed uniformly in small aggregations throughout the cytoplasm.

As the oocyte got ready to divide, however, Dishevelled aggregations dissolved and then reformed at the bottom of the cell at the furthest point from the nucleus.  This provided a clear difference between the two ends of the oocyte.

Swartz was curious about how exactly the protein was localizing to the bottom of the oocyte. There were a number of protein transport options to investigate, so he began systematically ruling them out; the protein was not transported by the cell’s cytoskeleton (“You can think of these like little railroad tracks,” Swartz said), nor was it buoyed along on cytoplasmic currents, nor repelled by some factor at the “top” of the oocyte.

At this interval, Swartz reached out to two collaborators in MIT’s physics department, who helped design experiments to further probe the behavior of Dishevelled in the oocytes. “That’s when we started to consider the idea of dissolution and reassembly, which is kind of the punchline of the paper,” Swartz said. “You can think of it like salt crystals dissolving in water — rather than taking a pre-assembled thing and physically transporting it down [to the bottom of the oocyte], the idea is that these Dishevelled assemblies start out everywhere, get dissolved into their individual components, and then selectively reform in the vegetal region.”

The exact mechanism of this dissolution and reformation is not yet clear. Swartz was able to show that the reformation could not take place in the absence of another Wnt pathway protein called Frizzled, but because Frizzled is not exclusive to the bottom of the oocyte, it is not the only thing driving the reassembly.

In the future, Swartz plans to investigate whether the Dishevelled aggregates are formed in precise structures, or whether they group together as phase-separated droplets such as the RNA molecules studied in Whitehead Institute Member Ankur Jain’s lab or the protein molecules involved in transcription from Whitehead Institute Member Richard Young’s lab. “I’m interested in the broader composition of these structures,” he added. “Do they only contain Dishevelled, or are there other ingredients?”

Regardless of how the assemblies form, the new information on how Dishevelled localizes shines a light on a previously mysterious step in how the Wnt pathway plays a role in early body patterning in sea stars.

“It’s quite striking that Dishevelled localization seems to be an important feature in the Wnt pathway in sea stars, but also in distantly related vertebrates,” Swartz said. “My feeling is that the ability to activate this pathway in selective parts of the early embryo by interpreting polarity built into the oocyte may be a really critical feature of the evolution of the animal body plan.”

A “tail” of two RNA regulatory systems
Greta Friar | Whitehead Institute
July 12, 2021

In new research, published in eLife on July 2, Bartel, who is also a professor of biology at Massachusetts Institute of Technology and an investigator with the Howard Hughes Medical Institute, and Bartel lab member Kehui Xiang, a CRI Irvington Postoctoral Fellow, have now discovered how cells establish this early gene regulatory regime and what conditions prompt a switch as the embryos mature. The researchers have observed the same regulatory switch in fish, frogs, and flies, and because the switch occurs across the animal kingdom, they would expect to see that the mechanism applies in other species including mammals.

“When I joined the lab, they had discovered that egg cells and early embryos had this different regulatory regime, and I wanted to know why,” Xiang says. “There must be fundamental changes to the cell, or to the molecules in the cell, that define this.”

The difference in how mRNAs are regulated during and after early development has to do with the length of their tails. mRNAs have tails made up of strings of adenines, one of the building blocks of RNA. Tail length varies between mRNAs from different genes and even between mRNAs from the same gene. Usually, the length of this “poly(A)-tail” corresponds to how long an mRNA lasts before getting degraded. An mRNA with a long tail is more stable, and will generally last longer. However, researchers had also observed that in some cases mRNA tail length corresponds to how readily an mRNA is used to make protein. Bartel’s earlier research had helped define when each of these connections occurs: mRNA tail length affects translational efficiency only in immature eggs and early embryos, and in other stages, it affects mRNA stability or lifespan.

In their new research, Xiang and Bartel uncovered three conditions that are required for the mRNA regulatory regime that exists in early development.

A competitive environment

The first condition is that there has to be a limited availability of a protein that binds to mRNA tails called cytoplasmic poly(A)-binding protein (PABPC). PABPC is known to help activate the translation of mRNA into protein. It binds to the mRNA tail and—in embryos—helps to increase translational efficiency; the researchers propose that it may do this by promoting a more favorable structure for translation. When PABPC is in limited supply, as it is in early embryos, then short-tailed mRNAs are less likely to bind any of the protein, as they will be outcompeted by long-tailed mRNAs, explaining the correlation between tail length and translational efficiency. Later in development, PABPC is in such ready supply that all of the mRNAs are able to bind at least one, decreasing the competitive edge of long-tailed mRNAs.

Early durability

However, the researchers observed that reducing the amount of PABPC in adult cells so that it becomes limiting in these cells did not cause mRNAs with longer tails to be translated more efficiently, which showed that other conditions must also contribute to early embryos’ unique regulation. The second condition that Xiang identified is that mRNAs must be relatively stable in spite of their inability to compete for PABPC. In adult cells, RNAs without PABPC bound to their tails are very unstable, and so are likely to degrade. If the same were true in early embryos, then the short-tailed mRNAs would degrade quickly because they are outcompeted for binding PABPC, and so one would again see a link between tail length and stability, rather than between tail length and translational efficiency—short-tailed mRNAs would be eliminated rather than poorly translated. However, the processes that would normally degrade mRNAs without PABPC have not yet started occurring in early embryos, allowing the short-tailed mRNAs to survive.

Big fish in a small pond

Finally, Xiang discovered that in order for tail length and translational efficiency to be linked, PABPC has to be able to affect translational efficiency. He found that in adult cells PABPC does not appear to boost translational efficiency the way it does in embryos. The researchers hypothesize that this is because the process of translating mRNAs in adult cells is already so efficient that the small boost from binding PABPC does not make a significant difference. However, in early embryos PABPC is more of a big fish in a small pond. The cells do not have all of the machinery to maximize translational efficiency, so every bit of improvement, such as the benefit of binding PABPC, makes a noticeable difference.

Together, these three conditions enable early eggs and embryos to regulate their mRNA in a unique fashion that can control how much protein is made from each gene without destroying the limited pool of mRNA available. In the future, the researchers hope to recreate the three conditions in non-embryonic cells to confirm that the conditions Xiang identified are not only necessary but also sufficient to cause the switch in regulatory regimes.

“Knowing which function the poly(A)-tail is performing in a specific cell or scenario—providing mRNA stability or translational efficiency—is really critical for understanding how genes are regulated in the different cells,” Bartel says. “And understanding that is important for answering all kinds of questions about cells, from their functions to what can go wrong with them in diseases.”

Shaping life

Patterns of myosin and F-actin proteins across developing embryos promote tissue folding and shape new life.

Raleigh McElvery
June 14, 2021

Virtually all multicellular organisms, including humans, begin as a single cell that rapidly divides and begets trillions of others. These cells work together, stretching, squishing, and migrating to sculpt organs and tissues. In the case of the fruit fly embryo, it only takes a few hours for life to take shape. First, the multiplying cells form an oblong sphere akin to a football. Then, mechanical forces cause a band of cells along the “belly” side of the developing fly to furrow inwards. These “mesoderm” cells form a new layer that will later give rise to muscles. Although the folding process transpires in less than 20 minutes, it’s crucial for determining where the cells will go and what roles they will assume.

Scientists previously identified two important proteins that generate the force needed to fold the tissue. The first, myosin, has a characteristic rod shape with feet hanging off both ends. It can “walk” along the cell’s inner scaffolding, composed of a second, rope-like protein called filamentous actin (F-actin). As it walks, myosin tugs on the F-actin and constricts the tissue. Researchers are probing the distribution of myosin and F-actin across the developing embryo, an important step towards understanding how these proteins drive constriction in the proper places to fold the tissue.

Myosin appears in a gradient across the belly and back of the developing fly. Since myosin and F-actin work together, many scientists assumed they would display the same pattern. However, new work from MIT’s Department of Biology and Department of Mathematics suggests otherwise. The study, published in the journal Development, shows how gene expression patterns dictate a unique distribution of F-actin across the mesoderm, which exhibits peaks and valleys. In combination with the myosin present, this F-actin pattern causes the cells to stretch, squish, or maintain their shape in just the right places to bend the tissue.

“We’ve known for decades that mechanical proteins like myosin and F-actin regulate tissue curvature during development,” says Adam Martin, an associate professor of biology and the study’s senior author. “But what hasn’t been appreciated is the extent to which these two proteins are intricately patterned during the tissue folding process. Our finding that F-actin has a different pattern than myosin was quite surprising.”

The researchers, led by graduate student Marlis Denk-Lobnig, began by focusing on two well-known transcription factor proteins, Twist and Snail, which bind to DNA to control gene expression. These transcription factors are known to dictate cell shape and fate during tissue folding, and Denk-Lobnig wondered how they affected F-actin levels.

By imaging live and fixed cells, the researchers observed that Snail and Twist drove a different pattern of F-actin density across the mesoderm compared to previously described myosin gradients. Two to three hours after the fruit fly eggs are laid, Snail depletes F-actin levels across the mesoderm. But, as Twist activates its transcriptional targets, F-actin and myosin levels rise in a subset of the mesoderm cells along the belly of the developing fly — constricting them and folding that swath of the tissue. The more F-actin and myosin a cell contains, the more compressed and wedge-shaped it becomes.

Denk-Lobnig also targeted another protein, RhoA, that tunes F-actin and myosin levels. RhoA activation is ultimately controlled by the ratio of two other molecules, C-GAP and RhoGEF2. The researchers adjusted the levels of C-GAP and RhoGEF2 in live cells, and watched the subsequent changes in myosin and F-actin distribution in real time.

To continue disentangling the effects of each protein on tissue curvature, they leveraged a computer simulation of a developing embryo designed by Associate Professor of Mathematics Jörn Dunkel, former grad student Pearson Miller PhD ’20, and postdoctoral researcher Jan Totz. The model allowed the team to adjust patterns of force and protein activity, in order to determine how the changes that they’d witnessed in real embryos affected tissue shape.

“The main takeaway is that you need this elegant coordination between cells during development,” Denk-Lobnig says. “We’ve shown how force generation patterns change the shape of individual cells — and how this leads to shape changes across entire tissues.”

Top image: Cross sections of three fruit fly embryos undergoing tissue folding. Nuclei are in blue, the transcription factor Snail is in red, and the junctional protein Armadillo is in green. Credit: Marlis Denk-Lobnig.
Video: An early stage fruit fly embryo has a band of cells on its surface that furrows inward to form a fold. Credit: Marlis Denk-Lobnig.

Citation:
“Combinatorial patterns of graded RhoA activation and uniform F-actin depletion promote tissue curvature”
Development, online June 14, DOI: 10.1242/dev.199232
Marlis Denk-Lobnig, Jan F. Totz, Natalie C. Heer, Jörn Dunkel, and Adam C. Martin

Posted: 6.14.21
Yadira Soto-Feliciano

Education

  • PhD, 2016, MIT
  • BS, 2008, Chemistry, University of Puerto Rico-Mayagüez 

Research Summary

We study chromatin — the complex of DNA and proteins that make up our chromosomes. We aim to understand how post-translational modifications to these building-blocks, as well as the factors that regulate these events, play essential roles in maintaining the integrity of cells, tissues, and ultimately entire organisms. We implement a combination of functional genomics, biochemical, genetic, and epigenomic approaches to study how chromatin and epigenetic factors decode the chemical language of chromatin, and how these are dysregulated in diseases such as cancer.

Awards

  • AACR Gertrude B. Elion Cancer Research Award, 2023
  • V Foundation Award, 2022
  • NIH MOSAIC K99/R00 Postdoctoral Career Transition Award, 2021
  • Eddie Méndez Scholar Award, Fred Hutchinson Cancer Research Center, 2020
  • Damon Runyon-Sohn Pediatric Cancer Fellowship, Damon Runyon Cancer Research Foundation, 2017
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