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A “model” parasite

Whitehead Institute researchers unravel the unique biology of apicomplexans — the parasites responsible for malaria, toxoplasmosis, and other diseases impacting global health.

Whitehead Institute
February 19, 2019

Apicomplexa: A brood of parasites

Malaria, cryptosporidiosis, and toxoplasmosis affect millions of people each year, killing an estimated 600,000 annually, mostly children under five in developing countries. Billions of dollars are spent each year to control and eliminate these diseases, according to the World Health Organization (WHO). Each of these diseases is caused by a different apicomplexan, a group of parasites that infect almost all animal species.

Toxoplasma gondii (T. gondii), which causes the disease toxoplasmosis, has a unique physiology that has allowed it to parasitize its hosts, yet it retains many features in common with other apicomplexans. Using T. gondii as a “model parasite”, Whitehead Member Sebastian Lourido is deciphering apicomplexans’ unique biology and uncovering aspects that could be harnessed to disrupt the parasites’ ability to proliferate and infect their hosts.

Apicomplexans’ toll on humans is staggering:

· Malaria, caused by several Plasmodium species of Apicomplexa, was responsible for over 200 million infections and more than 400,000 deaths, primarily in young children, in 2017 (WHO).

· Severe diarrhea kills an estimated 525,000 children under five each year (WHO). Over 200,000 of those deaths can be attributed to cryptosporidiosis, which is caused by the species of the apicomplexan Cryptosporidium (Sow et al., 2016, PLoS Negl Trop Dis.).

· 25% of the global population is infected with T. gondii with rates reaching over 60% in some areas (Pappas et al. 2009, Int. J. Parasitol.). Toxoplasmosis can cause an array of serious neurological disorders in those with weakened immune systems and can be lethal or lead to birth defects in a developing fetus. In an estimated 2% of infected individuals, toxoplasmosis causes retinal lesions (Holland, 2003, Am J Ophthalmol.).

A parasitic relationship, separated by a billion years of evolution

The diagram above depicts the evolutionary relationships between organisms — species separated by many branches are more distantly related than those divided by fewer branches. Apicomplexans and humans are separated by multiple branches and more than a billion years of evolution. In fact, apicomplexans are actually more closely related to plants than animals, having evolved from a non-parasitic ancestor about 700 to 900 million years ago that, like green plants, used photosynthesis to generate energy from sunlight. So far, scientists have studied only about half of the known apicomplexan genes, leaving the rest of their 8,000 predicted protein-coding genes uncharacterized.

Although key genes important for fundamental processes have remained fairly stable over the billion years since apicomplexans and their hosts diverged, other parts of the apicomplexan genomes evolved as they adapted to a parasitic lifestyle. The genes that emerged as unique to apicomplexans, such as those encoding factors involved in entering or exiting host cells, potentially represent therapeutic targets because curtailing their expression could hamstring — or even eliminate — the parasites without harming the host.

Analyzing a unique biology

Understanding apicomplexans and their distinct biology has been challenging at least in part because the tools — genomic analysis, genetic engineering, and culture systems — that scientists use to study and understand more traditional model organisms in the lab, such as mice, are difficult to apply in apicomplexans. Moreover, apicomplexans may spend different stages of their lives in different hosts, so studying a parasite’s complete life cycle may require studying and culturing multiple organisms or their tissues. For example, Plasmodium falciparum, which causes malaria, spends part of its life cycle in mosquitoes and another in humans.

Unlike the Plasmodium parasites that cause malaria, T. gondii is relatively easy to culture in the lab. Lourido, who is also an assistant professor of biology at Massachusetts Institute of Technology (MIT), and his lab are using this organism to unravel many elemental questions about apicomplexans: How do they infect their host cells? What do they require to reproduce? How do they break out of their host cell to infect more cells?

Adapted CRISPR/Cas9 gene editing system reveals first genome-wide glimpse of apicomplexan genomic profile

Researchers in Lourido’s lab are working to decipher the 50% of the T. gondii’s genome that remains to be characterized. To do so, they adapted the CRISPR/Cas9 gene editing system to work in T. gondii. Using CRISPR/Cas9, researchers can cut T. gondii’s DNA at specific sites to disable particular genes. With this approach, they were able to efficiently conduct genome-wide screens to identify genes that are functionally important to the parasite.

For this screen the Lourido lab used their adapted CRISPR/Cas9 gene editing system to remove the function — one at a time — of each of T. gondii’s ~8,000 protein-coding genes. The resulting altered parasites were then cultured with human host cells. After a period of time, the scientists tallied the number of parasites present with each modification to assess how disabling a particular gene’s function affects the parasites’ reproduction and survival. Altered parasites that successfully proliferated despite missing a gene’s function were deemed to have alterations in a gene that is dispensable, whereas modified parasites that did not thrive were deemed to have alterations in genes that are important for fitness.

Screen identifies apixomplexan-specific proteins

The initial screen of the T. gondii genome, led by Lourido lab research assistant Saima Sidik and postdoctoral researcher Diego Huet, identified a number of genes that encode indispensable conserved apicomplexan proteins, called ICAPs for short (Sidik et al. 2016, Cell). One ICAP identified by Sidik and Huet is an invasion factor called the claudin-like apicomplexan microneme protein (CLAMP).

In the same Cell paper, researchers described how they determined that CLAMP is critical to the initiation of host cell invasion by T. gondii. Working with Jacquin Niles and members of his lab in the MIT Department of Biological Engineering, the Lourido team also showed that CLAMP is required for the parasites that cause malaria to survive when grown in red blood cells.

In a recent paper published in the journal eLife, Lourido and first author Huet, identified a protein in apicomplexans that is crucial for creating adenosine triphosphate (ATP), the universal energy storage unit of cells. ATP is essential for cells’ survival and without it, cellular processes would stall. Most organisms have an enzyme, called ATP synthase, that creates ATP by converting the energy of a proton gradient across a membrane into mechanical energy. As the protons move through the ATP synthase, it spins like a turbine. This movement powers the formation of ATP. For the enzyme to work properly, a portion of the ATP synthase acts as a scaffold, or stator, by counteracting the rotation of the turbine-like part of the enzyme.

Although most components of the ATP synthase are conserved between apicomplexans and humans, scientists had been unable to pinpoint the gene encoding the essential stator portion – no DNA sequence in the apicomplexan genome resembles the sequences of known stator genes. Using a genomic approach, Huet and Lourido analyzed the predicted function and structure of the ICAPs  present in the mitochondrion, the ATP synthase’s home in all organisms. To their surprise, the predicted shape of one of the ICAPs resembles a stator subunit found in yeast and mammals.

When Huet and Lourido mutated or removed the stator subunit, the parasite’s ATP synthase failed to function properly, damaging the structure and performance of its mitochondria and halting the the parasite’s growth.

The beginning of a parasitic relationship

For Lourido and his lab, T. gondii’s unique stator protein is just one example of how these extraordinary apicomplexan organisms have evolved and adapted. By tailoring current tools and inventing new ones, Lourido’s investigations into T. gondii’s biology have the potential to reveal important insights into this family of parasites that impacts millions of people each year.

Credits

Written and produced by Nicole Giese Rura and Whitehead Institute

Illustrations and animations by Andrew Tubelli

Cover image courtesy of Clare Harding/Whitehead Institute

Special thanks to Sebastian Lourido and his lab, especially Clare Harding, Diego Huet, and Saima Sidik

References

WHO: Global Health Observatory (GHO data) for number of malaria cases

UNICEF: Diarrhoea as a cause of death in children under 5

Holland GN. (2003) Ocular toxoplasmosis: a global reassessment. Part I: epidemiology and course of disease. Am J Ophthalmol. Dec;136(6): 973-988. https://doi.org/10.1016/j.ajo.2003.09.040

Huet D, Rajendran E, van Dooren GG, Lourido S. (2018) Identification of Cryptic Subunits from an Apicomplexan ATP Synthase. eLife. 2018;7:e38097. https://doi:10.7554/eLife.38097

Pappas G, Roussos N, Falagas ME. (2009) Toxoplasmosis Snapshots: Global Status of Toxoplasma gondii Seroprevalence and Implications for Pregnancy and Congenital Toxoplasmosis. Int J Parasitol. Oct;39(12):1385-94. https://doi: 10.1016/j.ijpara.2009.04.003

Sidik SM, Huet D, Ganesan SM, Huynh MH, Wang T, Nasamu AS, Thiru P, Saeij JPJ, Carruthers VB, Niles JC, Lourido S. (2016) A Genome-wide CRISPR Screen in Toxoplasma Identifies Essential Apicomplexan Genes. Cell. Sep 8;166(6):1423-1435.e12. https://doi:10.1016/j.cell.2016.08.019

Sow SO, Muhsen K, Nasrin D, Blackwelder WC, Wu Y, Farag TH, et al. (2016) The Burden of Cryptosporidium Diarrheal Disease among Children < 24 Months of Age in Moderate/High Mortality Regions of Sub-Saharan Africa and South Asia, Utilizing Data from the Global Enteric Multicenter Study (GEMS). PLoS Negl Trop Dis. 10(5): e0004729. https://doi.org/10.1371/journal.pntd.0004729

Yahata K, Treeck M, Culleton R, Gilberger T-W, Kaneko O (2012) Time-Lapse Imaging of Red Blood Cell Invasion by the Rodent Malaria Parasite Plasmodium yoelii. PLoS ONE. 7(12): e50780. https://doi.org/10.1371/journal.pone.0050780

Start signal for sex cell creation
Greta Friar | Whitehead Institute
February 27, 2019

Cambridge, MA — Cells can divide and multiply in two ways: mitosis, in which the cell replicates itself, creating two copies identical to the original; or meiosis, in which the cell shuffles its DNA and divides twice, creating four genetically unique cells, each with half of the original cell’s number of chromosomes. In mammals, these latter cells become eggs and sperm.

How do germ line cells, the repository of cells that create eggs and sperm, know when to stop replicating themselves and undergo meiosis? Researchers had been aware that a protein called STRA8, which is only active in germ line cells, was involved in initiating meiosis, but they did not know how. New research from Whitehead Member and Institute Director David Page, also a professor of biology at Massachusetts Institute of Technology and an investigator with Howard Hughes Medical Institute; Mina Kojima, formerly a Massachusetts Institute of Technology graduate student and now a postdoctoral researcher at Yale; and visiting scientist Dirk de Rooij has revealed that in mice, STRA8 initiates meiosis by activating and amplifying a network of thousands of genes. This network includes genes involved in the early stages of meiosis, DNA replication, and other cell division processes. The research was published in eLife on February 27, 2019.

In the past, researchers have had difficulty collecting enough cells on the cusp of meiosis to investigate STRA8’s role. In mammals, germ line cells are inside the body, difficult to access, and they begin meiosis in staggered fashion so few cells are at the same stage during an extraction. Researchers in Page’s lab had previously come up with an approach to solve this problem using developmental synchronization, manipulating the cells’ exposure to the chemical that triggers their development in order to prompt all of the cells to begin meiosis simultaneously. Once the cells were synced up, first author Kojima could get a large enough sample to observe patterns in gene expression leading up to and during meiosis, and to figure out where STRA8 is binding.

She found that STRA8 binds to the regulatory portions of DNA called promoter regions, which initiate or increase transcription of adjacent genes, of most critical meiosis genes. With some exceptions, STRA8 does not switch genes from off to on. Rather, genes in the STRA8-regulated network are already expressed at low levels and STRA8 binding massively ramps up their production. The researchers posit that meiosis is then initiated once the genes reach a threshold of expression. This finding sheds light on instances in previous studies in which researchers found meiosis-related genes active in cells not yet undergoing meiosis.

The researchers were surprised to find that STRA8 also amplifies many genes involved in mitosis. However, they suggest that the meiosis-specific genes activated by STRA8 take precedence in determining which of the two cell-cycle processes the cell will undergo. STRA8 regulates certain critical genes, such as Meioc and Ythdc2, which help to establish a meiosis-specific cell-cycle program.

This research enriches our understanding of the process of sexual reproduction. Identifying the expansive STRA8-regulated network has elucidated the start of meiosis: the moment a cell commits to recombining and dividing, relinquishing its genetic identity for the chance to create something — or someone — new.

This work was supported by the National Science Foundation and the Howard Hughes Medical Institute.

 

Written by Greta Friar

***

David Page’s primary affiliation is with Whitehead Institute for Biomedical Research, where his laboratory is located and all his research is conducted. He is also a Howard Hughes Medical Institute Investigator and a Professor of Biology at the Massachusetts Institute of Technology.

***

Full citation:

“Amplification of a broad transcriptional program by a common factor triggers the meiotic cell cycle in mice”

eLife, February 27, 2019, https://doi.org/10.7554/eLife.43738

Mina L. Kojima (1,2), Dirk G. de Rooij (1), and David C. Page (1,2,3)

1. Whitehead Institute, 455 Main Street, Cambridge, MA 02142, USA

2. Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA

3. Howard Hughes Medical Institute, Whitehead Institute, Cambridge, MA 02142, USA

Pumping up red blood cell production
Greta Friar | Whitehead Institute
February 28, 2019

Cambridge, MA — Red blood cells are the most plentiful cell type in our blood and play a vital role transporting oxygen around our body and waste carbon dioxide to the lungs. Injuries that cause significant blood loss prod the body to secrete a one-two punch of signals – stress steroids and erythropoietin (EPO) – that stimulates red blood cell production in the bone marrow. These signals help immature cells along the path to becoming mature red blood cells. In a healthy individual, as much as half of their blood volume can be replenished within a week. Despite its importance, scientists are still working to unravel many aspects of red blood cell production. In a paper published online February 28 in the journal Developmental Cell, Whitehead Institute researchers describe work that refines our understanding of how stress steroids, in particular glucocorticoids, increase red blood cell production and how early red blood cell progenitors progress to the next stage of maturation toward mature red blood cells.

These findings are especially important for patients with certain types of anemia that do not respond to clinical use of EPO to stimulate the final stages of red cell formation, such as Diamond-Blackfan anemia (DBA). In this rare genetic disorder usually diagnosed in infants and toddlers, the bone marrow does not produce enough of early red blood cell progenitors, called burst forming unit-erythroids (BFU-Es), that respond to glucocorticoids. In both healthy people and DBA patients, these BFU-Es divide several times and mature before developing into colony forming unit-erythroids (CFU-Es) that that, stimulated by EPO, repeatedly divide and produce immature red blood cells that are released from the bone marrow into the blood. But the lack of BFU-Es in DBA patients means that the glucocorticoid signal has a limited target, and the cascade of cell divisions that should result in plentiful red blood cells is contracted and instead produces an insufficient amount.

One of the standard treatments for DBA is boosting red blood cell production with high doses of synthetic glucocorticoids, such as prednisone or prednisolone. But the mechanisms behind these drugs and their normal counterparts are not well understood. By deciphering the mechanisms by which glucocorticoids stimulate red cell formation, scientists may be able identify other ways to stoke CFU-E production – and ultimately red blood cell production – without synthetic glucocorticoids and the harsh side effects that their long-term use can cause, such as poor growth in children, brittle bones, muscle weakness, diabetes, and eye problems.

For more than two decades, Whitehead Institute Founding Member Harvey Lodish, has investigated glucocorticoids’ effects on red blood cell production. In his lab’s most recent paper, co-first authors and postdocs Hojun Li and Anirudh Natarajan, describe their research, which helps decipher how BFU-Es progress through their maturation process.

For more than 30 years, scientists have thought that glucocorticoids bestowed BFU-Es with a stem cell-like ability to divide until an unknown switch flipped and the cells matured to the CFU-E stage. By looking at gene expression in individual BFU-Es from normal mice, Li and Natarajan determined that the developmental progression from BFU-E to CFU-E is instead a smooth continuum. They also found that in mice glucocorticoids exert the greatest effect on the BFU-Es at the beginning of the developmental continuum by slowing their developmental progression without affecting their cell division rate. In other words glucocorticoids are able to effectively compensate for a decreased number of BFU-Es by allowing those that do exist, while still immature, to divide more times, producing in mice up to 14 times more CFU-Es than BFU-Es lacking exposure to glucocorticoids.

Li and Natarajan’s work reveals previously unknown aspects of the mechanism by which glucocorticoids stimulate red blood cell production. With this better understanding, scientists are one step closer toward pinpointing more targeted approaches to treat certain anemias such as DBA.

This work was supported by the National Institutes of Health (NIH grants DK06834813 and HL032262-25) and the American Society of Hematology and was performed with the assistance of Whitehead Institute’s Fluorescence Activated Cell Scanning (FACS) Facility and Genome Technology Core facility. Styliani Markoulaki, head of the Whitehead Genetically Engineered Models Center, and M. Inmaculada Barrasa of Bioinformatics and Research Computing (BaRC) are also co-authors of the paper.

 

Written by Nicole Giese Rura

***

Harvey Lodish’s primary affiliation is with Whitehead Institute for Biomedical Research, where his laboratory is located and all his research is conducted. He is also a professor of biology and a professor of biological engineering at Massachusetts Institute of Technology (MIT). Lodish serves as a paid consultant and owns equity in Rubius, a biotech company that seeks to exploit the use of modified red blood cells for therapeutic applications.

***

Citation:

“Rate of Progression through a Continuum of Transit-Amplifying Progenitor Cell States Regulates Blood Cell Production”

Developmental Cell, online February 28, 2019, https://doi.org/10.1016/j.devcel.2019.01.026

Hojun Li*, Anirudh Natarajan*, Jideofor Ezike, M. Inmaculada Barrasa, Yenthanh Le, Zoë A. Feder, Huan Yang, Clement Ma, Styliani Markoulaki, and Harvey F. Lodish.

*These authors contributed equally

Committed to service and science

When senior Julia Ginder isn’t investigating the mystery of her own allergies, she’s volunteering to help young people reach their goals.

Gina Vitale | MIT News correspondent
February 25, 2019

Julia Ginder has to avoid a lot of foods due to allergies. From a young age, she got used to bringing her own snacks to birthday parties and group outings. But she didn’t really know the science behind her allergies until high school, when she read a chapter for class on immunology.

“I read it, and then I read it again, and I went running downstairs to tell my mom, ‘This is what’s wrong with me!’” she recalls.

From them on, Ginder was driven to learn about what made her body react so severely to certain stimuli. Now a biology major, she does research in the lab of Christopher Love, in the Koch Institute for Integrative Cancer Research, where she studies peanut allergies — one of the few food allergies she actually doesn’t have.

“I really enjoy figuring out, what’s the perspective from the biology side? What is the contributing chemistry? And how do those fit together?” she says. “And then, when you take a step back, how do you use that knowledge and perhaps the technology that comes out of it, and actually apply that in the real world?”

Nuts about research

In the Love lab, researchers look at how individual immune cells from people with peanut allergies react when stimulated with peanut extracts. More recently, they’ve been analyzing how the stimulated cells change over the course of treatment, evolving from one state to the next.

“You can watch the activation signals change over time in individual cells from peanut-allergic patients compared to healthy ones,” Ginder explains. “You can then dig deeper and look at distinct populations of cells at a single time point. With all of this information, you can start to get a sense of what critical cell types and signals are making the allergic person maintain a reaction.”

The researchers aim to figure out which cell types are associated with the development of tolerance so that more effective treatments can be developed. For instance, allergic people are sometimes given peanuts in small doses as a sort of biological exposure therapy, but perhaps if more key cell states are identified, targeted drug treatments can be added on top of that to induce those cell states.

Further pursuing her interest in health, Ginder spent the Independent Activities Period of her sophomore year volunteering for Boston Medical Center. The program she worked for helped families learn how to be advocates for their children with autism. For instance, it provided guidance on how to negotiate an appropriate accommodations agreement with their child’s school for their individual needs.

“It [the BMC experience] made it clear to me that for a child to succeed, they need to have support from both the educational side and the health side,” she says. “And it might seem obvious, but, especially for a child who might be coming from a less privileged background, those are two really important angles for ensuring that they are given the opportunity to reach their full potential.”

“The most helpful thing you can do is simply be there.”

Ginder became a swim coach and tutor for Amphibious Achievement in the fall of her first year, almost immediately after arriving at MIT. It’s a program that aims to help high schoolers reach both their athletic and academic goals. The high schoolers, often known as Achievers, are assigned a mentor like Ginder who helps with the academic and the athletic activities.

Local students come to MIT early Sunday morning to practice swimming or rowing, head to the Maseeh dining hall for lunch, participate in an afternoon academics lesson, reflect on their goals, and then spend a half an hour one-on-one with their mentor. It’s a big commitment for both the Achievers and mentors to spend almost six hours every Sunday with the program, but Ginder, who completed her two-year term as one of the co-executive directors this fall, has seen the importance of showing up week after week.

“The most helpful thing you can do is simply be there. Listen if they want to tell you anything, but really just being consistent — every single Sunday, being there.”

Ginder played on the field hockey team during her first year. However, when a practice during her sophomore year left her with a concussion and unable to play, she used the newfound spare time to start volunteering for Camp Kesem (CK). Having really enjoyed her experience at Amphibious Achievement, she was eager to be a counselor for the camp, which serves children whose parents are affected by cancer.

“Being there for someone, whether they are having a tough time or a great day, is really important to me. I felt that CK really aligned with that value I hold, and I hoped to meet even more people at MIT who felt that way. And so I joined, and I’ve loved it,” she says.

Management and moving west

Eventually, Ginder would like to become a physician, possibly in the fields of pediatrics and allergies. However, with a minor in public policy, she’s interested in developing areas outside of science as well. So, for the next couple of years, she’ll be moving westward to work as an associate consultant for Bain and Company in San Francisco.

“The reason I’m most interested in consulting is that there is this strong culture of learning and feedback. I want to improve my ability to be a strong team member, leader, and persuader. I think these are areas where I can continue to grow a lot,” she says. “It may sound silly, but I think for me, as someone who is 5’2” and hoping to become a pediatrician, it’s important to cultivate those professional skills early. I want to also serve as a leader and advocate outside of the clinic.”

As Ginder admits, the move is quite the geographic leap. Right now, her entire family is between a 20-minute and two-hour drive away. Moving to the opposite side of the country will be difficult, but she isn’t one to shy away from a challenge.

“I think it’ll be a bit sad because I’m not going to be as close to my family, but I think that it’ll really push me to be as independent as possible. I’ll need to look for my own opportunities, meet new people, build my network, and be my own person,” she says. “I’m really excited about that.”

Why too much DNA repair can injure tissue

Overactive repair system promotes cell death following DNA damage by certain toxins, study shows.

Anne Trafton | MIT News Office
February 14, 2019

DNA-repair enzymes help cells survive damage to their genomes, which arises as a normal byproduct of cell activity and can also be caused by environmental toxins. However, in certain situations, DNA repair can become harmful to cells, provoking an inflammatory response that produces severe tissue damage.

MIT Professor Leona Samson has now determined that inflammation is a key component of the way this damage occurs in photoreceptor cells in the retinas of mice. About 10 years ago, she and her colleagues discovered that overactive initiation of DNA-repair systems can lead to retinal damage and blindness in mice. The key enzyme in this process, known as Aag glycosylase, can also cause harm in other tissues when it becomes hyperactive.

“It’s another case where despite the fact that inflammation is there to protect you, in some circumstances it can actually be harmful, when it’s overactive,” says Samson, a professor emerita of biology and biological engineering and the senior author of the study.

Aag glycosylase helps to repair DNA damage caused by a class of drugs known as alkylating agents, which are commonly used as chemotherapy drugs and are also found in pollutants such as tobacco smoke and fuel exhaust. Retinal damage from these drugs has not been seen in human patients, but alkylating agents may produce similar damage in other human tissues, Samson says. The new study, which reveals how Aag overactivity leads to cell death, suggest possible targets for drugs that could prevent such damage.

Mariacarmela Allocca, a former MIT postdoc, is the lead author of the study, which appears in the Feb. 12 issue of Science Signaling. MIT technical assistant Joshua Corrigan, former postdoc Aprotim Mazumder, and former technical assistant Kimberly Fake are also authors of the paper.

A vicious cycle

In a 2009 study, Samson and her colleagues found that a relatively low level of exposure to an alkylating agent led to very high rates of retinal damage in mice. Alkylating agents produce specific types of DNA damage, and Aag glycosylase normally initiates repair of such damage. However, in certain types of cells that have higher levels of Aag, such as mouse photoreceptors, the enzyme’s overactivity sets off a chain of events that eventually leads to cell death.

In the new study, the researchers wanted to find exactly out how this happens. They knew that Aag was overactive in the affected cells, but they didn’t know exactly how it was leading to cell death or what type of cell death was occurring. The researchers initially suspected it was apoptosis, a type of programmed cell death in which a dying cell is gradually broken down and absorbed by other cells.

However, they soon found evidence that another type of cell death called necrosis accounts for most of the damage. When Aag begins trying to repair the DNA damage caused by the alkylating agent, it cuts out so many damaged DNA bases that it hyperactivates an enzyme called PARP, which induces necrosis. During this type of cell death, cells break apart and spill out their contents, which alerts the immune system that something is wrong.

One of the proteins secreted by the dying cells, known as HMGB1, stimulates production of chemicals that attract immune cells called macrophages, which specifically penetrate the photoreceptor layer of the retina. These macrophages produce highly reactive oxygen species — molecules that create more damage and make the environment even more inflammatory. This in turn causes more DNA damage, which is  recognized by Aag.

“That makes the situation worse, because the Aag glycosylase will act on the lesions produced from the inflammation, so you get a vicious cycle, and the DNA repair drives more and more degeneration and necrosis in the photoreceptor layer,” Samson says.

None of this happens in mice that lack Aag or PARP, and it does not occur in other cells of the eye or in most other body tissues.

“It amazes me how segmented this is. The other cells in the retina are not affected at all, and they must experience the same amount of DNA damage. So, one possibility is maybe they don’t express Aag, while the  photoreceptor cells do,” Samson says.

“These molecular studies are exciting, as they have helped define the underlying pathophysiology associated with retinal damage,” says Ben Van Houten, a professor of pharmacology and chemical biology at the University of Pittsburgh, who was not involved in the study. “DNA repair is essential for the faithful inheritance of a cell’s genetic material. However, the very action of some DNA repair enzymes can result in the production of toxic intermediates that exacerbate exposures to genotoxic agents.”

Varying effects

The researchers also found that retinal inflammation and necrosis were more severe in male mice than in female mice. They suspect that estrogen, which can interfere with PARP activity, may help to suppress the pathway that leads to inflammation and cell death.

Samson’s lab has previously found that Aag activity can also exacerbate damage to the brain during a stroke, in mice. The same study revealed that Aag activity also worsens inflammation and tissue damage in the liver and kidney following oxygen deprivation. Aag-driven cell death has also been seen in the mouse cerebellum and some pancreatic and bone marrow cells.

The effects of Aag overactivity have been little studied in humans, but there is evidence that healthy individuals have widely varying levels of the enzyme, suggesting that it could have different effects in different people.

“Presumably there are some cell types in the human body that would respond the same way as the mouse photoreceptors,” Samson says. “They may just not be the same set of cells.”

The research was funded by the National Institutes of Health.

Nedivi named to new professorship
Picower Institute
February 8, 2019

Elly Nedivi, a professor in the Picower Institute and the Departments of Brain and Cognitive Sciences and Biology, has been named the inaugural William R. (1964) & Linda R. Young Professor of Neuroscience, the MIT School of Science announced.

Nedivi, an MIT faculty member since 1998, studies the cellular mechanisms that underlie activity-dependent plasticity in the developing and adult brain through studies of neuronal structural dynamics, identification of the participating genes, and characterization of the proteins they encode.

Her work to identify “candidate plasticity genes” has yielded many insights, including elucidating the neuronal and synaptic function of two previously unknown CPGs: CPG2 and CPG15. In a study published earlier this year, her lab showed that the protein CPG2 is significantly less abundant in the brains of people with bipolar disorder and showed how specific mutations in the SYNE1 gene that encodes CPG2 undermine the protein’s expression and its function in neurons, potentially contributing to disease.

Motivated by the large number of CPGs that affect neuronal structure, her lab has also been collaborating with that of Peter So’s in MIT’s Department of Mechanical Engineering to develop multi-color two photon microscopy for large volume, high resolution imaging of dendritic arbor and synaptic structural dynamics in vivo. Nedivi’s lab was the first to show unambiguous evidence of dendritic arbor remodeling in the adult brain, and identify inhibitory connections as the most plastic component of experience-dependent circuit rearrangements.

Nedivi thanked the Youngs for their support of neuroscience research at MIT.

“I recently met the Youngs, and share their view that study of the brain and mind is an area of science with tremendous potential to improve people’s lives,” she said. “I respect their wish to give back to MIT, and am deeply honored to be named the inaugural William R. (1964) & Linda R. Young Professor of Neuroscience.”

Biologists answer fundamental question about cell size

The need to produce just the right amount of protein is behind the striking uniformity of sizes.

Anne Trafton | MIT News Office
February 7, 2019

MIT biologists have discovered the answer to a fundamental biological question: Why are cells of a given type all the same size?

In humans, cell size can vary more than 100-fold, ranging from tiny red blood cells to large neurons. However, within each cell type, there is very little deviation from a standard size. In studies of yeast, MIT researchers grew cells to 10 times their normal size and found that their DNA could not keep up with the demands of producing enough protein to maintain normal cell functions.

Furthermore, the researchers found that this protein shortage leads the cells into a nondividing state known as senescence, suggesting a possible explanation for how cells become senescent as they age.

“There are so many hypotheses out there that try to explain why senescence happens, and I think this data provides a beautiful and simple explanation for senescence,” says Angelika Amon, the Kathleen and Curtis Marble Professor in Cancer Research in the Department of Biology and a member of the Koch Institute for Integrative Cancer Research.

Amon is the senior author of the study, which appears in the Feb. 7 online edition of Cell. Gabriel Neurohr, an MIT postdoc, is the lead author of the paper.

Excessive size

To explore why cell size is so tightly controlled, the researchers prevented yeast cells from dividing by modifying a gene critical for cell division, so that it could be turned off at a certain temperature. These cells continued to grow, but they could not divide and they did not replicate their DNA.

The researchers discovered that as the cells expanded, their DNA and their protein-building machinery could not keep pace with the needs of such a large cell. This failure to produce enough protein led to the dilution of the cytoplasm and disruption of cell division. The researchers believe that many other fundamental cell processes that rely on cellular molecules finding and interacting with each other may also be impaired when cells are too big.

“Theoretical models predict that diluting the cytoplasm will decrease reaction rates. Every chemical reaction would occur more slowly, and some threshold concentrations of certain proteins may not be reached, so certain reactions would never happen because the concentrations are lower,” Neurohr says.

The researchers showed that yeast cells with two sets of chromosomes were able to grow to twice the size of yeast cells with just one set of chromosomes before becoming senescent, suggesting that the amount of DNA in the cells is the limiting factor in the cells’ ability to grow.

Experiments with human cells yielded similar results: In a study of human fibroblast cells, the researchers found that forcing the cells to grow to excessive sizes (eight times their normal size) disrupted many functions, including cell division.

“It’s been clear for some time that cells do control their size, but it’s been unclear what the various physiological reasons are for why they do so,” says Jan Skotheim, an associate professor of biology at Stanford University, who was not involved in the research. “What’s nice about this work is it really shows how things go wrong when cells get too big.”

Age-related disease

Amon says excessive growth likely plays a major role in the development of senescence, which occurs in many types of mammalian cells and is thought to contribute to age-related organ dysfunction and chronic age-related diseases.

Senescent cells are often larger than younger cells, and this study, which showed that unchecked cell growth leads to senescence, offers a possible explanation for this observation. Human cells tend to grow slightly larger throughout their lifetimes, because every time a cell divides, it checks for DNA damage, and if any is found, division is halted while repairs are made. During each of these delays, the cell grows slightly larger.

“Over the lifetime of a cell, the more divisions you make, the higher your probability is of having that damage, and over time cells will get larger,” Amon says. “Eventually they get so large that they start diluting critical factors that are important for proliferation.”

A difficult question that remains unanswered is how different types of cells maintain the appropriate size for their cell type, which the researchers now hope to study further.

The research was funded, in part, by the National Institutes of Health, the Howard Hughes Medical Institute, the Paul F. Glenn Center for Biology of Aging Research at MIT, a National Science Foundation graduate research fellowship, the William Bowes Fellows program, and the Vilcek Foundation.

Origin story

Junior Leah McKinney practiced kitchen microbiology on her ranch in Nevada before exploring the intricacies of DNA replication initiation in bacteria at MIT Biology.

Raleigh McElvery
February 6, 2019

Leah McKinney grew up on a 50,000-acre cattle ranch in Nevada — vaccinating sheep, roping calves, digging for fossils, and occasionally hauling home old bovine femurs. She saddled horses, treated sick lambs, and helped ewes struggling to give birth. One Christmas, she even asked Santa for a fetal pig. “He delivered,” McKinney, now a junior in Course 7, recalls with a laugh.

When she was 12 years old, she saved up enough birthday money to purchase a microscope. Even though she permanently dyed the kitchen sink a distinct shade of blue while making slides, her parents (who both hold degrees in animal science) didn’t mind. They even let her grow bacteria in the heater closet and tally them on the kitchen counter — all in the name of the elementary school science fair.

“They were always encouraging my weird scientific endeavors,” she says. “I think my love for science, and microbiology specifically, came out of my agricultural upbringing.”

She grew to appreciate basic science because it allowed her to study the fundamental mechanisms behind key biological processes. She arrived at MIT in 2016 determined to major in Biology, and hasn’t wavered in her decision. Although she relishes the subject matter, she initially feared the classes would be tedious and memorization-heavy.

“I was quite happy to learn that’s not the case here,” she says. “MIT Biology values problem-solving over rote memorization, and encourages you to take the information you’ve learned in class and apply it to interesting problems. And that mindset extends from the classroom into the lab.”

One of the things that drew McKinney to MIT was the institute’s Undergraduate Research Opportunities Program (UROP), which allows students to join labs and collaborate with faculty as early as their first year. She recalls that, while other universities touted similar opportunities, MIT placed theirs front and center.

“I’d heard that all you had to do was email a professor and ask to join the lab, but I didn’t believe it — that just seemed way too easy,” McKinney says. “But when I was looking for a UROP, I just emailed my current principal investigator to set up a time to talk, and now I’ve been in his lab for over a year.”

McKinney is part of Department Head Alan Grossman’s lab, which investigates the molecular mechanisms and regulation underlying basic cellular processes in bacteria. The entire group works with the rod-shaped Bacillus subtilis, but some members study horizontal gene transfer while others focus on DNA replication and gene expression. McKinney and her graduate student mentor Mary Anderson are in this second category, examining a protein called DnaA that is required to initiate DNA replication and also modulates the expression of several genes.

In order to successfully grow and reproduce, a bacterium must first replicate its single chromosome before dividing into two identical daughter cells. DnaA is responsible for beginning DNA replication in all bacteria. It binds to the origin of replication on the chromosome, unwinds some of the nearby DNA, and recruits the other proteins needed to copy the chromosome.

This operation is highly regulated to ensure that each daughter cell receives only a single chromosome. B. subtilis controls replication via several proteins, including YabA. When YabA binds to DnaA, it prevents replication from ever getting started.

Since DnaA also serves as a transcription factor — binding to other DNA sequences called promoters to increase or decrease expression of certain genes, including its own gene dnaA — YabA may also impact DnaA’s gene targets. McKinney hopes to eventually determine exactly how.

While McKinney discovers something new about her bacteria each time she conducts a successful experiment, she learns almost as much when her tests go awry. “I’ve had to practice a lot of troubleshooting,” she says, “and that’s not something you can learn in class. But everyone in the lab is incredibly friendly and always willing to answer questions or give advice.”

As a teaching assistant for the lab class 7.02 (Introduction to Experimental Biology and Communication), McKinney had the chance to help other students conduct experiments, answering their questions and grading their lab notebooks. She took 7.02 last spring, but says it’s been enlightening to experience the class through a different lens. She adds: “I definitely understand the material more deeply than I did before.”

In addition to TAing, McKinney teaches an SAT preparatory program run by MIT students. “At first, standing up and talking in front of a 20-person section was rather terrifying, but it’s become so much easier,” she says. “The experience has been really good for me.”

After she graduates, McKinney knows she wants to go to graduate school — likely for microbiology — but beyond that, nothing is concrete. She is sure of one thing, though: joining the Grossman lab was one of the best decisions she’s made at MIT.

She advises all current and prospective students to do a UROP. “Find something you’re really interested in,” she says. “It’s okay not to know a lot coming in; you’re going to learn so much, including topics and techniques you won’t learn in class. And don’t be too disappointed when things don’t work; that’s just part of the process. And when you finally get something to work that you’ve been troubleshooting for a while, the feeling is absolutely amazing.”

Posted 2.5.19
Biologist Adam Martin studies the mechanics of tissue folding

The dynamic process is critical to embryonic development and other cellular phenomena.

Anne Trafton | MIT News Office
February 1, 2019

Embryonic development is tightly regulated by genes that control how body parts form. One of the key responsibilities of these genes is to make sure that tissues fold into the correct shapes, forming structures that will become the spine, brain, and other body parts.

During the 1970s and ’80s, the field of embryonic development focused mainly on identifying the genes that control this process. More recently, many biologists have shifted toward investigating the physics behind the tissue movements that occur during development, and how those movements affect the shape of tissues, says Adam Martin, an MIT associate professor of biology.

Martin, who recently earned tenure, has made key discoveries in how tissue folding is controlled by the movement of cells’ internal scaffolding, known as the cytoskeleton. Such discoveries can not only shed light on how tissues form, including how birth defects such as spina bifida occur, but may also help guide scientists who are working on engineering artificial human tissues.

“We’d like to understand the molecular mechanisms that tune how forces are generated by cells in a tissue, such that the tissue then gets into a proper shape,” Martin says. “It’s important that we understand fundamental mechanisms that are in play when tissues are getting sculpted in development, so that we can then harness that knowledge to engineer tissues outside of the body.”

Cellular forces

Martin grew up in Rochester, New York, where both of his parents were teachers. As a biology major at nearby Cornell University, he became interested in genetics and development. He went on to graduate school at the University of California at Berkeley, thinking he would study the genes that control embryonic development.

However, while in his PhD program, Martin became interested in a different phenomenon — the role of the cytoskeleton in a process called endocytosis. Cells use endocytosis to absorb many different kinds of molecules, such as hormones or growth factors.

“I was interested in what generates the force to promote this internalization,” Martin says.

He discovered that the force is generated by the assembly of arrays of actin filaments. These filaments tug on a section of the cell membrane, pulling it inward so that the membrane encloses the molecule being absorbed. He also found that myosin, a protein that can act as a motor and controls muscle contractions, helps to control the assembly of actin filaments.

After finishing his PhD, Martin hoped to find a way to combine his study of cytoskeleton mechanics with his interest in developmental biology. As a postdoc at Princeton University, he started to study the phenomenon of tissue folding in fruit fly embryonic development, which is now one of the main research areas of his lab at MIT. Tissue folding is a ubiquitous shape change in tissues to convert a planar sheet of cells into a three-dimensional structure, such as a tube.

In developing fruit fly embryos, tissue folding invaginates cells that will form internal structures in the fly. This folding process is similar to tissue folding events in vertebrates, such as neural tube formation. The neural tube, which is the precursor to the vertebrate spinal cord and brain, begins as a sheet of cells that must fold over and “zip” itself up along a seam to form a tube. Problems with this process can lead to spina bifida, a birth defect that results from an incomplete closing of the backbone.

When Martin began working in this area, scientists had already discovered many of the transcription factors (proteins that turn on networks of specific genes) that control the folding of the neural tube. However, little was known about the mechanics of this folding.

“We didn’t know what types of forces those transcription factors generate, or what the mechanisms were that generated the force,” he says.

He discovered that the accumulation of myosin helps cells lined up in a row to become bottle-shaped, causing the top layer of the tissue to pucker inward and create a fold in the tissue. More recently, he found that myosin is turned on and off in these cells in a dynamic way, by a protein called RhoA.

“What we found is there’s essentially an oscillator running in the cells, and you get a cycle of this signaling protein, RhoA, that’s being switched on and off in a cyclical manner,” Martin says. “When you don’t have the dynamics, the tissue still tries to contract, but it falls apart.”

He also found that the dynamics of this myosin activity can be disrupted by depleting genes that have been linked to spina bifida.

Breaking free

Another important cellular process that relies on tissue folding is the epithelial-mesenchymal transition (EMT). This occurs during embryonic development when cells gain the ability to break free and move to a new location. It is also believed to occur when cancer cells metastasize from tumors to seed new tumors in other parts of the body.

During embryonic development, cells lined up in a row need to orient themselves so that when they divide, both daughter cells remain in the row. Martin has shown that when the mechanism that enables the cells to align correctly is disrupted, one of the daughter cells will be squeezed out of the tissue.

“This has been proposed as one way you can get an epithelial-to-mesenchymal transition, where you have cells dissociate from native tissue,” Martin says.  He now plans to further study what happens to the cells that get squeezed out during the EMT.

In addition to these projects, he is also collaborating with Jörn Dunkel, an MIT associate professor of mathematics, to map the network connections between the myosin proteins that control tissue folding during development. “That project really highlights the benefits of getting people from diverse backgrounds to analyze a problem,” Martin says.

Bacteria promote lung tumor development, study suggests

Antibiotics or anti-inflammatory drugs may help combat lung cancer.

Anne Trafton | MIT News Office
January 31, 2019

MIT cancer biologists have discovered a new mechanism that lung tumors exploit to promote their own survival: These tumors alter bacterial populations within the lung, provoking the immune system to create an inflammatory environment that in turn helps the tumor cells to thrive.

In mice that were genetically programmed to develop lung cancer, those raised in a bacteria-free environment developed much smaller tumors than mice raised under normal conditions, the researchers found. Furthermore, the researchers were able to greatly reduce the number and size of the lung tumors by treating the mice with antibiotics or blocking the immune cells stimulated by the bacteria.

The findings suggest several possible strategies for developing new lung cancer treatments, the researchers say.

“This research directly links bacterial burden in the lung to lung cancer development and opens up multiple potential avenues toward lung cancer interception and treatment,” says Tyler Jacks, director of MIT’s Koch Institute for Integrative Cancer Research and the senior author of the paper.

Chengcheng Jin, a Koch Institute postdoc, is the lead author of the study, which appears in the Jan. 31 online edition of Cell.

Linking bacteria and cancer

Lung cancer, the leading cause of cancer-related deaths, kills more than 1 million people worldwide per year. Up to 70 percent of lung cancer patients also suffer complications from bacterial infections of the lung. In this study, the MIT team wanted to see whether there was any link between the bacterial populations found in the lungs and the development of lung tumors.

To explore this potential link, the researchers studied genetically engineered mice that express the oncogene Kras and lack the tumor suppressor gene p53. These mice usually develop a type of lung cancer called adenocarcinoma within several weeks.

Mice (and humans) typically have many harmless bacteria growing in their lungs. However, the MIT team found that in the mice engineered to develop lung tumors, the bacterial populations in their lungs changed dramatically. The overall population grew significantly, but the number of different bacterial species went down. The researchers are not sure exactly how the lung cancers bring about these changes, but they suspect one possibility is that tumors may obstruct the airway and prevent bacteria from being cleared from the lungs.

This bacterial population expansion induced immune cells called gamma delta T cells to proliferate and begin secreting inflammatory molecules called cytokines. These molecules, especially IL-17 and IL-22, create a progrowth, prosurvival environment for the tumor cells. They also stimulate activation of neutrophils, another kind of immune cell that releases proinflammatory chemicals, further enhancing the favorable environment for the tumors.

“You can think of it as a feed-forward loop that forms a vicious cycle to further promote tumor growth,” Jin says. “The developing tumors hijack existing immune cells in the lungs, using them to their own advantage through a mechanism that’s dependent on local bacteria.”

However, in mice that were born and raised in a germ-free environment, this immune reaction did not occur and the tumors the mice developed were much smaller.

Blocking tumor growth

The researchers found that when they treated the mice with antibiotics either two or seven weeks after the tumors began to grow, the tumors shrank by about 50 percent. The tumors also shrank if the researchers gave the mice drugs that block gamma delta T cells or that block IL-17.

The researchers believe that such drugs may be worth testing in humans, because when they analyzed human lung tumors, they found altered bacterial signals similar to those seen in the mice that developed cancer. The human lung tumor samples also had unusually high numbers of gamma delta T cells.

“If we can come up with ways to selectively block the bacteria that are causing all of these effects, or if we can block the cytokines that activate the gamma delta T cells or neutralize their downstream pathogenic factors, these could all be potential new ways to treat lung cancer,” Jin says.

Many such drugs already exist, and the researchers are testing some of them in their mouse model in hopes of eventually testing them in humans. The researchers are also working on determining which strains of bacteria are elevated in lung tumors, so they can try to find antibiotics that would selectively kill those bacteria.

The research was funded, in part, by a Lung Cancer Concept Award from the Department of Defense, a Cancer Center Support (core) grant from the National Cancer Institute, the Howard Hughes Medical Institute, and a Margaret A. Cunningham Immune Mechanisms in Cancer Research Fellowship Award.