Breaking Through Winter 2022

Finding New Ways to Light Up Biology |

WINTER 2022

4 DISABLING COVID-19 SPIKE PROTEINS

A new path to treatment for COVID-19 8 FINDING NEW WAYS TO LIGHT UP BIOLOGY

How fluorophore technology advances live imaging 12 LAB NOTES:

Collaborating to Identify the Cause of Chronic Kidney Disease of Unknown Etiology

16 THIS IS WHY:

Hannah Lust:

Coming Full Circle

Introducing the NEW mdibl.org We’re excited to announce the launch of our new website. Log on to learn about our impact, our world-class research, upcoming courses, conferences and events, and meet some of the generous supporters who make our work possible. Simply visit mdibl.org

FRONT COVER: The germ cells of a male C. elegans, stained with anti-a-tubulin (green) and anti-pH3 (red) fluoresce under a microscope. Learn more about the technologies that are helping light up biology on page 9.

FROM THE PRESIDENT

Celebrating Our Accomplishments As we begin a new year and continue to navigate these challenging times, I want to thank you for your ongoing support and engagement. Despite lingering uncertainties, we have made tremendous progress and I am grateful for the opportunity to work with you to ensure the Laboratory’s future success.

Strengthening Our Research Programs.

Over the last year we have recruited excellent scientists, developed new animal models, established new international collaborations and programs and made fascinating discoveries. I am pleased to share the details of some of our recent developments in this issue of Breaking Through.

Expanding our visiting scientist program is another important component of strengthening our research capacity.

For more than 100 years, MDI Biological Laboratory has served as an international gathering place for renowned researchers from across the U.S., Canada and Europe. This past year we had the opportunity to welcome back Nishad Jayasundara, Ph.D., Assistant Professor at Duke University to the MDIBL campus as a visiting scientist collaborating with Iain Drummond, Ph.D. Nishad is no stranger to MDIBL, having spent several summers working with us as one of our early INBRE fellows while a student at College of the Atlantic. It was a pleasure to see Nishad return to our campus with his own student.

Renovating Our Campus.

Expansion of our research and educational programs requires that we make a major investment in our housing and scientific infrastructure. Over the last three years we renovated seven of our campus cottages, providing adequate housing for our graduate students and visiting scientists. We are now embarking on an effort to restore all of our historic cottages. Presently, we are in the midst of a complete restoration of Bowen Hall, the oldest cottage on our campus, to serve as an Education and Outreach Center (see back cover).

Opportunities and Challenges.

We continue to work to address the ongoing challenges of the COVID-19 virus. While variants such as Omicron and Delta continue to cause concern, we remain confident in the ability of vaccines to prevent serious illness.

Since the onset of the pandemic, I have found myself in the unexpected position of developing a potential treatment that interferes with the ability of the spike protein to bind to the host cell surface, preventing the systematic complications that occur when the virus enters the bloodstream. Details of this ongoing research project are outlined on page four.

The scientific atmosphere at MDIBL is vibrant and our research and education programs are thriving. None of this would be possible without your ongoing support. I look forward to working with you to ensure we have the resources to meet future challenges head on and secure our continued success.

With appreciation and gratitude,

Hermann Haller, M.D. President

BREAKING THROUGH WINTER 2022 › 3

Disabling COVID-19

SPIKE PROTEINS A New Path to Treatment for COVID-19

By now, almost everyone is familiar with the image of the crown-like, reddish spike proteins protruding from the outer envelope of the coronavirus that lend its name (corona is Latin for “crown”) and are responsible for its virulence. These spike proteins, which are characteristic of coronaviruses, allow the COVID-19 virus to latch onto the surface of the host’s cell, penetrate the cell’s external membrane and hijack its molecular machinery to make new virus particles. They may also be its undoing.

4 ‹ BREAKING THROUGH WINTER 2022

Having studied the role of the glycocalyx in diabetic kidney disease, Haller has turned his attention to its role in COVID-19.

ACTING LIKE LOCK PICKS, the spike proteins gain entry to the host’s cells by modifying their shape in order to interact with receptors on the host cell’s surface. Once the spikes are attached, the virus clings tightly — 10 to 20 times more tightly than its close relative, the SARS (Severe Acute Respiratory Syndrome) virus — which explains why it spreads more easily than coronaviruses like SARS or the common cold. The appearance of the Delta and Omicron variants, both exponentially more transmissible than the original virus, have created additional complexity and urgency for scientists working to prevent the spread of this deadly disease.

“This research is so exciting because we have the opportunity to contribute to a treatment for a virus that has taken the lives of nearly five and a half million people around the world,” Haller says.

“It also demonstrates why basic research is so important: when scientists have the opportunity to look deeply into a problem over the course of time, they often find that their research can lead to solutions to novel problems.”

Since the onset of the pandemic, the mechanism that allows spike proteins to attach to the host’s cells has come under intense scientific scrutiny.

While COVID-19 was initially thought to be a respiratory disease, scientists now know it can directly affect the microvascular endothelium, or membrane, in the kidney

In addition to contributing to the development of vaccines, an increased understanding of the way in which these spike proteins function holds the potential to prevent the microvascular, or small blood vessel, damage that is a major contributor to mortality from COVID-19, as well as to its long-term health consequences, including damage to the heart, kidney, liver, lung and gastrointestinal, hematological and nervous systems.

If research on spike proteins is new ground for the scientists who have turned to the study of the coronavirus since the onset of the pandemic, it is familiar territory for Hermann Haller, M.D., president of the MDI Biological Laboratory and a nephrologist, or kidney specialist, who studies how maintaining healthy cell surfaces can offset, and even prevent, diabetic nephropathy, a common microvascular complication of diabetes.

THE GLYCOCALYX AND COVID-19

Haller’s research focuses on the glycocalyx, a mixture of sugar chains and proteins that makes up the protective surface of the endothelial cells that line the small blood vessels of the kidney and other organs.

6 ‹ BREAKING THROUGH WINTER 2022

40%

of patients hospitalized with severe infections develop acute kidney injury

and other organs. Though patients with mild symptoms are spared kidney damage, 20 percent of infected patients experience life-threatening kidney damage, with close to 40 percent of patients who are hospitalized with severe infections developing acute kidney injury.

Though the mechanisms behind COVID-19-associated kidney damage are believed to be multifactorial, a big factor is damage to the endothelium, which as the interface between blood and tissues regulates the defensive functions of the cell, including vascular inflammation. When the endothelium is damaged, these defensive functions are impaired — or worse, turn against the host, causing inflammation, thrombosis or a severe reaction called a “cytokine storm.”

20%

of patients hospitalized with severe infections experience life-threatening kidney damage

In research over three years at the MDI Biological Laboratory and at Hannover Medical School in Hannover, Germany, where he is a professor and chair of the division of nephrology, Haller has been studying how a drug called calcium dobesilate (trade name Doxium,®) produced by the Swiss company Vifor Pharma, interferes with a binding particle called heparan sulfate (HS) on the glycocalyx of endothelial cells in patients with diabetic kidney disease.

The onset of the pandemic provided an opportunity to apply this knowledge to the treatment of COVID-19. While some scientists are studying other binding particles on the surfaces of the host’s cells, none have studied HS, which Haller describes as “overlooked.” His goal is to bring calcium dobesilate, which has traditionally been used as a treatment for diabetic retinopathy, a diabetes-related eye disease, to market as a treatment for COVID-19.

“At the moment, we are the only ones with data on a drug that protects endothelial integrity by interfering with the ability of the spike proteins to bind with heparan sulfate particles on the surfaces of the host’s cells,” Haller said. “In the lock analogy, calcium dobesilate functions like a high security feature within the locking mechanisms on the surfaces of the host’s cells that makes them resistant to forced entry by the coronavirus’ spike proteins.”

‘WHERE THE MEDICAL NEED IS’

Though calcium dobesilate may also have potential for preventing coronavirus infection, Haller’s research focuses on preventing the systemic complications that occur when the virus enters the bloodstream, where it can lead to organ-specific microvascular damage for which there is currently no treatment.

Clinical trials on the use of calcium dobesilate for treatment of COVID-19 complications are now underway in Switzerland, Haller said.

LECTIN / DAPI

“Over the last 18 months we have seen a tremendous increase in our understanding of SARS-CoV2 virus. This has led to the development of the first mRNA vaccines, to one FDA approved drug and several emergency use authorizations for several others. But more treatment options are needed as we begin to transition into long-term management of this disease.”

“We have characterized a new molecule that prevents binding of the viral spike protein in endothelial cells and have successfully reproduced these findings in vivo. We now have convincing data on a brand new therapeutic approach for treating COVID-19 patients.”

Calcium dobesilate may also have applications for the treatment of other pathological conditions of the microvasculature as well as for other viruses, making it what Haller calls an “anti-virus strategy for the future, in general.”

In the now-iconic image of the coronavirus, the spikes cast long shadows, which the artists who created it included to convey the “gravity of the situation.”

In life, these spikes cast long shadows as well — shadows that can lead to death and disability. Though vaccines mitigate the immediate threat, research at the MDI Biological Laboratory seeks to vanquish the danger that COVID-19 and other coronaviruses pose over the long term.

LECTIN / SPIKE / DAPI ABOVE: Mouse kidney before and after perfusion with recombinant SARS-CoV-2 spike protein (green).

Spike protein binds and breaches the endothelial cell surface (red) in small blood vessels. [Cell nuclei (blue)].

“While COVID-19 was initially thought to be a respiratory disease, scientists now know it can directly affect the microvascular endothelium, or membrane, in the kidney and other organs.”

HERMANN HALLER, M.D. NEPHROLOGIST / PRESIDENT, MDI BIOLOGICAL LABORATORY

BREAKING THROUGH WINTER 2022 › 7

“It was as if scientists could now see where the workers were and what they were doing instead of having to infer it.”

DUSTIN L. UPDIKE, PH.D.

IN THE 1990s, two scientists from the University of California published a widely circulated analogy comparing the geneticist’s and a biochemist’s approach to biomedical research. In it, the cell is compared to an auto factory and the geneticist and biochemist to observers standing on a hill overlooking the factory, trying to figure out how it works by observing the employees who enter and the cars that exit.

In the analogy, the autoworkers are genes and the parts they make are proteins manufactured in the cell that provide it with shape and structure and transmit information about basic biological functions. The analogy highlights how genetic and biochemical approaches can provide insight into the complex processes that are hidden behind the walls of the biological factory of the cell.

In recent years, green fluorescent protein (GFP) technology has witnessed new advancements, most notably the development of the CRISPR/Cas9 gene-editing technology, which allows fluorescent proteins to easily be inserted into the genomes of model animals such as C. elegans, and split fluorophore technology, which allows biologists to track protein-protein interactions within defined cell types of living animals.

“It was as if scientists could now see where the workers were and what they were doing instead of having to infer it,” says Dustin L. Updike, Ph.D., a MDI Biological Laboratory scientist who uses fluorescent proteins to study C. elegans, a tiny, transparent roundworm that is a popular model in biomedical research.

BREAKING THROUGH WINTER 2022 › 9

Meet our experts Working together for five years, they’ve refined fluorophore technology and created about 40 tagged strains of C. elegans.

DUSTIN UPDIKE, PH.D. LEAD SCIENTIST

CATHERINE SHARP RESEARCH ASSISTANT

10 ‹ BREAKING THROUGH WINTER 2022

A new refinement to fluorophore technology Now, in a further refinement to fluorophore technology, Updike and his research assistant, Catherine S. Sharp, in collaboration with scientists from Calico Life Sciences, Google’s cutting-edge R&D anti-aging venture, have expanded the split fluorophore technology toolkit, making it faster, easier, more reliable and more cost-effective to label C. elegans proteins with genetically expressed fluorescent tags.

The split fluorophore technology works a little like a bulb that lights up when wires are connected. The mechanism is the structure of the fluorescent tag, which takes the form of a barrel with 11 slats, or strands, all of which are required for bioluminescence. When 10 strands of one protein and one strand of another are tagged, the proteins light up when they fuse, enabling tissue-specific expression and the study of protein-protein interactions.

“The system gives us a way to cheaply and efficiently find these proximity interactions in real time in the living animal ­­— to see in what tissue, in what cell type and in what areas of the cell these interactions are happening, and to see how dynamic they are,” Updike says. “In the auto factory analogy, it would be as if the parts

light up when they are assembled, allowing us to understand the function of each and how they work together.”

The cost savings come into play because it is more difficult and costly to insert a fluorophore-sized DNA segment into a gene (the entire barrel) than to make small gene edits (the single strand). Using the new technology developed at the MDI Biological Laboratory and Calico, the small fragment, which can quickly and easily be fused to almost any protein of interest, will be visible wherever the large fragment is expressed.

The development of the new technology grew out of Updike’s research on a gene in germ cells, which are the precursors of sperm and eggs and the only cells with the potential to create an entirely new organism. The stem-cell-like ability of germ cells to transform into any kind of cell, which is called “totipotency,” could someday be exploited for the regeneration of diseased or injured tissues and organs.

Updike’s collaborators at Calico, lead author Jérôme Goudeau, Ph.D., of the laboratory of Cynthia Kenyon, Ph.D., and scientists working with Maria Ingaramo, a Calico research specialist, developed an additional color — a bright scarlet modeled after a protein from a bioluminescent coral. The resulting paper, whose authors

include Updike and Sharp (see sidebar), was recently published in the journal Genetics.

The role of INBRE Updike credits students from the University of Maine campuses at Fort Kent and Presque Isle with piloting the new split fluorophore technology during a 2020 Maine INBRE course on the CRISPR/Cas9 gene-editing technology.

SUPPORT OUR WORK › Your gift has a double impact — furthering scientific progress and providing quality mentorship to young scientists like Catherine. Give today at: mdibl.org/donate

Maine INBRE (IDeA Network of Biomedical Research Excellence) is a collaborative network of educational and research institutions led by the MDI Biological Laboratory whose goals include creating a technically skilled workforce through undergraduate biomedical research training. INBRE is sponsored by the National Institute of General Medical Sciences and the National Institutes of Health.

CATHERINE SHARP:

‘Like seeing a familiar face in the crowd’ In awarding the Nobel Prize to the scientists who discovered fluorescent tags, the Royal Swedish Academy of Sciences called such tags a “guiding star” that has allowed science to map the roles of the proteins that control what happens in the cell.

If fluorophore technology is a “guiding star,” the creation of that star depends on research assistants like Catherine S. Sharp who insert the tags into the genomes of laboratory animals.

Using CRISPR/Cas9 gene-editing technology, she has created approximately 40 fluorescent strains of C. elegans. The tagged roundworms’ progeny will include strains that display the selected-for traits, and these have been used

by MDI Biological Laboratory scientists as well as by scientists from around the world.

She compares developing research tools such as split fluorophore technology to building the foundation of a pyramid of biological knowledge “when you have no idea what the top of the pyramid” — or the discoveries enabled by those tools — will look like.

AT LEFT: Male germ cells were co-immunostained with anti-a-tubulin (green) and anti-pH3 (red). DNA was counter-stained with DAPI (blue).

The Genetics paper, which was authored

by Updike, Sharp and scientists at the world-famous aging laboratory at Calico headed by Cynthia Kenyon, Ph.D., provides an intriguing glimpse of what the peak of the pyramid may look like.

“When you do something every day it becomes commonplace,” Sharp says. “The paper highlights the fact that what I do can go out into the world and be used by someone else to do something completely different.”

“It’s like spotting a friend in the crowd at a baseball game on TV,” she continues. “It’s like, ‘Wait a minute, I know them!’”

BREAKING THROUGH WINTER 2022 › 11

PICTURED: Hannah Lust, recently awarded her Ph.D. by Dartmouth College, is thrilled to be back in her home state of Maine, and at MDI Biological Laboratory, where she spent time as an undergraduate student.

THIS IS WHY YOUR GIFT MATTERS

Hannah Lust: Coming Full Circle Hannah Lust, Ph.D., is not the first student to have “boomeranged” back to the MDI Biological Laboratory.

Lucky for us, she’s unlikely to be the last.

Lust came to MDIBL as an INBRE fellow from the University of Maine Farmington. It was here that her passion for science was ignited. “I was fully immersed; completely surrounded by peers and mentors who were just as enthusiastic and excited as I was.” Lust says. “It sounds cliché, but that’s when I fell in love with science.” Her 10 weeks in Salisbury Cove were so formative that she changed her plans of becoming a medical doctor to becoming a scientist. In 2021 she received her Ph.D. in Microbiology and Immunology from Dartmouth College.

When the opportunity to come back to MDIBL as an employee arose last fall, Lust jumped at the chance. Her new role will see her heavily involved in the federally funded All About Arsenic SEPA (Science Education Partnership Award) program.

The data-literacy program trains educators and students throughout Maine and

New Hampshire in real-world research skills. Participants collect and test private well water samples for the presence of arsenic, analyze data and present their findings, along with mitigation recommendations, to the community.

It’s a change for Lust, who spent her doctoral years focused on T cell immunology, but she’s eager for the challenge. “During graduate school I realized I really enjoy helping other students fall in love with science,” she explains. “I feel privileged to be introducing them to science and instilling a sense of curiosity and confidence.”

Lust is excited to make science more accessible for Maine students and help them see the impact it, and they, can have. She has traveled full circle back to MDIBL and Maine, and who knows — maybe she will be the spark that ignites another young scientist’s passion.

SUPPORT OUR WORK › Read more about the SEPA program and how it benefits Maine communities at: mdibl.org/SEPA

“MDI Biological Laboratory is where I fell in love with science, and now I get to serve as a catalyst to other students making their own discoveries.” – HANNAH LUST, PH.D. BREAKING THROUGH WINTER 2022 › 13

LAB NOTES

Community news about the developments and scientists that are shaping the future of the MDI Biological Laboratory.

For the latest news, visit mdibl.org/news

ENVIRONMENTAL SPOTLIGHT ›

Working Together to Explain a Mysterious Kidney Ailment Plaguing Farmers Around the World Environmental toxicologist Nishad Jayasundara, Ph.D. and kidney researcher Iain Drummond, Ph.D. are working together to solve a mystery.

During field work, Jayasundara (Duke University) found that Sri Lankan farmers were suffering from a chronic kidney disease of unknown etiology (CKDu). Suspecting it was linked to toxic chemicals in drinking water, but not knowing which, if any, was the culprit, he asked Drummond (then at Harvard Medical School/MGH) to screen five blind water samples. One sample produced kidney dysfunction in the zebrafish model — the one that came from a region where CKDu was endemic.

“It was like a detective story — we had the match,” said Drummond, who is now director of the MDI Biological Laboratory’s Kathryn W. Davis Center for Regenerative Biology and Aging. “Though there’s more to do, the results showed we were on track in developing a tool to identify the early signs of environmentally related kidney disease.”

With their research published in 2020, this past summer Jayasundara and Drummond teamed up again, demonstrating the value of ongoing scientific collaboration.

stress-induced dehydration and by socioeconomic factors such as inadequate nutrition and medical care. The same thing may be happening elsewhere, but at a lower, more chronic level that may go undetected.”

Jayasundara and Drummond consider CKDu to be a sentinel disease for the effects of climate change on health. Through its study, scientists working in the emerging field of climate medicine can investigate how climate change affects health; how organisms, including humans, adapt to climate change; and how to prevent or mitigate climate change-related health issues. “The main goal is to prevent progression,” Drummond said.

In some Sri Lankan communities, up to 20

“Once we have developed the tools to identify

percent of the population, including children, have

the contaminants, we can go back to affected

displayed symptoms. CKDu has also been

communities and say, ‘These are the ones you

identified in other areas of the world experiencing

need to worry about.’”

unprecedented warming, including Africa, Central and South America, India, the Middle East and even the United States.

“I don’t think we fully appreciate the role of

Jayasundara plans to continue his research with Drummond in 2022, working to refine the zebrafish as a screening tool for the early detection of kidney dysfunction, develop the potential

environmental contaminants in kidney disease,”

of using the model for evaluating the effects of

Jayasundara explains. “With equatorial CKDu,

other contaminants on kidney function, and

we may be seeing the most severe form —

gain a deeper understanding of the mechanisms

a form exacerbated by concentrated levels of

that underlie the negative health effects of

toxic compounds in drinking water, by heat

environmental contaminants.

14 ‹ BREAKING THROUGH WINTER 2022

ABOVE Nishad Jayasundara and Iain Drummond will continue their collaboration in 2022.

Jayasundara’s journey with MDI Biological Laboratory started as a College of the Atlantic student, continued as a trainee, and now he has come full circle as he returns to MDIBL as a visiting scientist with students of his own.

LEARN MORE › Partnerships between MDI Biological Laboratory faculty and visiting scientists are advancing scientific progress. Read their stories at: mdibl.org/ collaboration

ABOUT US

We aim to improve human health by discovering novel mechanisms of tissue repair, aging and regeneration, translating our discoveries for the benefit of medicine and society and developing the next generation of scientific leaders.

BOARD OF TRUSTEES Alan W. Kornberg, Esq., Chairman Paul, Weiss, Rifkind, Wharton & Garrison LLP

Edward J. Benz Jr., M.D., Vice Chairman Dana Farber Cancer Institute

Thomas A. Boyd, Ph.D., Treasurer Anthology BioDevelopment LLC

Janis L. Coates, Ph.D., Secretary Island Readers and Writers

Hermann Haller, M.D., ex officio MDI Biological Laboratory, President

Peter J. Allen, M.D. Duke University School of Medicine

James L. Boyer, M.D. Yale University

Phoebe C. Boyer The Children’s Aid Society

Terence C. Boylan The River Press

Ruth Cserr Sigmar H. Gabriel John A. Hays Christie’s

Anne H. Lehmann Alan B. Miller, Esq. I. Wistar Morris III Margaret Myers, M.D. Dennis L. Shubert, M.D., Ph.D. Christopher P. Sighinolfi Bruce A. Stanton, Ph.D. Geisel School of Medicine at Dartmouth

Clare Stone Daphne W. Trotter, Esq. McDermott Will & Emery LLP

Robert Taft Whitman Published by the Office of Development and Public Affairs Editors + Writers Jeri Bowers, Pippa Hansen and Stefanie Matteson Design Cushman Creative

DONOR SPOTLIGHT

Selfless Gift Buoys the Future of Scientific Discovery It was touching to hear this extraordinary news from long time supporters, Mike and Mary Hays — they made a planned gift to the MDI Biological Laboratory.

Members of our Star Point Society, the Hays believe in the importance of biomedical research and the need for public engagement with science. Now, as members of the John S. Kingsley Society, this conviction will become part of their legacy.

Mike and Mary identified a small number of nonprofits they feel do exemplary work and included these organizations in their estate plan. By doing so, the Hays have peace of mind knowing their assets will help the causes most meaningful to them.

Photography Kevin Bennett, Jerome Goudeau, Hermann Haller, Anna Farrell, James Maloney-Hawkins, Hyemin Min, Marko Pende, Rogier van Bakel, Michael York

MDI Biological Laboratory takes its role of stewarding the generosity of donors like Mike and Mary Hays seriously, and it is an honor to be entrusted with their legacy. We are grateful for their help in ensuring MDIBL’s research, education and outreach programs continue for many years to come.

MDI Biological Laboratory PO Box 35, Salisbury Cove, ME 04672 Website: mdibl.org Email: breakingthrough@mdibl.org

To learn more about how to leave your legacy at MDIBL, please contact Jeri Bowers, Director of Development, at jeri@mdibl.org or 207-288-3147.

MDI Biological Laboratory PO Box 35 Salisbury Cove, ME 04672 mdibl.org

Looking Back to Move Forward Breathing new life into Bowen Hall Bowen Hall is a central and visible part of the MDI Biological Laboratory’s history.

2021 marked the 100th anniversary of the institution’s relocation to Salisbury Cove.

Thus it seems fitting to breathe new life into Bowen Hall, the oldest building on our campus. With your help, we will bring it back to its rightful place in the center of community life as a modern, multi-use space.

Support Bowen’s renovation with a donation today at mdibl.org/donate

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