Developing Human gD-crystallins as an Experimental System for Studying Beta Sheet Folding and Cataract FormationM
Melissa S. Kosinski-Collins, Ph.D.
Professor of Biology
MS 008, SSC 016A
Waltham, MA 02454
The first time I met Jonathan, I was 21. I was an undergrad who had been lucky enough to be asked by my lab to attend the Biophysical Society annual meeting. It was in New Orleans, and I was a farm kid who had never been on a train. I was applying to grad school at places I had never dreamed of attending and about to meet some powerful, impressive Biophysicists. Maybe they would be my PI for grad school? Maybe they would be my future collaborators? My eyes were wide and my ignorance palpable.
At the time, my PI, Lila Gierasch was my hero. She was a successful, female Biophysicist who pushed me to be the best me, and a real powerhouse at UMass. She had been recruited to UMass from UT Southwestern, exploded professionally, and become a leader of research for BOTH the chemistry and Biochemistry departments. I was a senior doing honors research, a protein folder and a pretty successful student academically. I had started my research late in the game because I honestly didn’t even know bench research was “a thing” until junior year. I was extremely naive about the world of science. I worked hard, loved what I did, but had no real knowledge of how to make a career out of it. I saw Lila’s future as my future. PI, all the way. Writing grants, doing amazing science, and taking names the whole time. That would be my name projected up there some day welcoming attendees to the Biophysical Meeting. I should have had arrogant and idealistic written on my shirt.
As a former president of the society, Lila took me to some exclusive events at that conference. Lila introduced me to the incoming president, Chris Miller, and the outgoing president, Jonathan King. I was applying to both of their graduate programs and worshipped their work. It was like meeting the person who invented the Apple Watch while buying an Apple watch. Lila told me that she and Jon were doing an impromptu talk the next day about science education and she wanted me to show the audience some of the stuff I had been dabbling in while working for CHEW (the Chemistry Higher Education Workgroup) run by Bill Vining. As my work study job, I made money helping design and test interactive software for chemistry courses. Sure, no biggie. I could talk about that stuff in my sleep.
Lila and I spoke first. We talked about helping students understand, concept-based teaching, active learning, and added little demo to impress. We nailed it. We were followed by Jon. With his deep voice, Jon took the podium and, in moment I will never forget, changed the way I viewed science. Jon’s speech was short and sweet. He told us all we needed to stand up and fight for the community we serve. We all needed to educate the world about the importance of our research and show commitment to a broader world than just other scientists. We needed to be better. We needed to be not just scientists but advocates for change and educators of the world. How could we get funding or enact real change in scientific literacy if we just sat in our own Ivory Tower worried about only our own experiments? US scientists, he commented, are funded by the citizens of the US, and don’t have the luxury to be selfish. They are indebted and should be accountable to the community they serve. Who was this guy and how did he have the guts to say all this?
I took a crazy number of notes detailing the importance of explaining to the community who we are and what we do. I even presented it at a post-meeting journal club to such a detailed extent that Lila asked if I had gone to any other talks. I had, of course, I just thought Jon had the most important things to say. Little did I know that I was going follow in his footsteps and spend my life stressing the importance of science communication and outreach.
The very next year when I entered MIT as a graduate student, I ended up joining Jonathan’s lab. At the time I came, Jon wasn’t looking for graduate students. He had a post doc-centric lab and was clear that it would take a “special person” to break the 7-year lull he had supervising graduate students. As luck would have it, I am like a fungus. I am slightly strange and stay around despite efforts to get rid of me. Maybe it was the concussion playing on the MIT softball team during my rotation with Patricia Clark that made him feel bad for me, or maybe it was that I really was THAT persistent. Either way, I got to call the King lab my home for four years. During that time, Jon was my adviser, my mentor and truly my inspiration.
Very early on, Jon asked me to pick a project. He was adamant that he wasn’t in the business of assigning projects to students. He liked us to follow our interests. Given my main interest was in slapping DG’s and rate constants on every possible b-sheet protein under the sun, he recommended that I consider studying a relatively new system in the lab, Human gD crystallin (HgDC). In the months before I arrived, Jon had started a collaboration with George Benedek in the MIT physics department to study this protein. Jon helped the Benedek lab set up a recombinant model and purification system, and the Benedek lab was analyzing phase separation in juvenile-onset mutants. I wasn’t that excited about phase separation, but I was interested in looking at the folding and refolding patterns of the protein. I figured that if I could apply what I had learned in the Gierasch lab about fluorescence patterns, folding states, and single Trp mutants to crystallin, I could figure out the refolding kinetics of the molecule. As an added bonus, crystallin was involved in cataract formation and studying the kinetics and morphology of aggregates of the molecule seemed pretty interesting. The experiments were long and the work somewhat tedious, but that was okay. Staying late into the night absorbing all the thermodynamic knowledge I could from Steve Raso while he and I blasted Red Sox games throughout our bays, became my past time.
Experiment-wise, the King lab was amazing for me. I made a new expression system, mutants were easy to create, the protein was super stable, and the experiments just seemed to take off. I was able to determine certain refolding conditions that caused crystallin to adopt ordered aggregation patterns and began to define an ordered domain-based productive refolding pattern. This was a gold mine of new and uncharted experiments. I learned how to observe structures via AFM and EM, and I became the lab “go-to” for folding fluorescence. Within a couple years of beginning the project, Shannon Thol and Ishara Mills-Henry joined the lab, and, as the three amigos, we hunted the literature for new congenital mutants that we could unfold, refold and aggregate. Jon pushed me to try electrospray mass spec in Carol Robinson’s lab in Cambridge, and by the end of my four years, I even dabbled in a-crystallin chaperone interactions with HgDC. Jonathan was nothing but supportive of every crazy idea I had. Whenever I data-dumped on him in his office, he just smiled, tried to politely reel me in, and subtly direct me to reasonable pursuits in addition to my often-unreasonable experimental plans.
During my full immersion in the “wonderful world of crystallin”, I also realized I had a passion for teaching and science education. What’s amazing is that Jonathan seemed to recognize that in me before I even saw that in myself. Jon was like that, though. He seemed to be able to see something in people that may have not been visible to the rest of the world. He often hired people not just for their passion and ability in science, but for some other passion or ability they possessed that could bring a different chemistry and strength into our King-lab family. Now looking back on it, I think his mentoring goal was to train good, unique humans who could go out into the world and inspire a love for science in the community around them. Shannon and I spent quite a bit of time working with the MIT high school teacher program and the HHMI education group. Whereas some PIs would probably frown on time away from the bench, Jon encouraged it. Perhaps he saw how much I liked teaching and the classroom, or perhaps he just saw that those experiences kept me motivated in lab, but whatever it was, he constantly pushed me to hone my skills in the classroom.
Nearing the end of my third year of graduate school, Jon mistakenly came into the lab one afternoon to talk “postdocs” with me. It was time, he said, “To think about my future” and that meant lining up potential research postdocs as my next step. Perhaps it was the emotion of the bench or my sleepless night taking care of a sick toddler, but whatever it was, I think he was quite surprised when burst out into tears. “But I don’t want to do a research post doc!” I wailed. In his relaxed Jon fashion, he smiled, took a minute to process the news, and within days started helping me figure out what I did want to do. We explored teaching, science writing, policy, and even educational advocacy together. Jonathan never wanted me to be something I wasn’t or to follow a path that wasn’t my own. Not only did he support me as I tried to find myself, but he helped me test out different possibilities.
With Jon’s help, I took a science education postdoc right at MIT in the HHMI Education Group with Graham Walker. Jon said, if I missed the bench, I could always come back to lab and pick up refolding crystallin right where I left off. I could always “come home”, and there would always be a place for me if my heart wasn’t in teaching. I suspect he knew right from the start that I wasn’t coming back and that my career would be in the classroom. With a piece of chalk in my hand and a lecture hall of eager young minds in front of me, I never looked back. I spent two years in the education group and then took a non-tenure track faculty position at Brandeis teaching the introductory Biology lab sequence. I redesigned the entire fall course to be an inquiry-based lab focusing on crystallin mutagenesis, purification, and stability analysis. Using plasmids and constructs Jon graciously gave me, I put together the teaching experimental system he and I had always talked about together. Crystallin became the mechanism through which I was reaching my students. I tried every day to make them understand and love science the way that I do, focusing on both experiments and the skills of science communication. Further motivated by the King lab, now my students look at aggregation-based diseases like Huntington’s and cataracts through the lens of molecular biology and biochemistry so they can try to appreciate the value of basic research and take that understanding with them into the world. Every Brandeis life science major now spends a year developing a working knowledge of how protein structure and aggregation influence human disease.
I think it is also important to note, that Jonathan inspired something else in me during my time in graduate school, and that is the importance of being an advocate for others. I am not an intrinsically self-less person, in fact I would argue that in my early years in academia, I was quite selfish. Jonathan constantly reminded me of who I worked for and the reason I was supposed to be doing what I was doing. He consistently told me that my research was supposed to be for the common good and my data belonged to the world, just like he told the community in that very first conference where I met him. Jon was so passionate about equitable education and was so outspoken about equity and peace for all humans of the world, he planted a seed in my head that was too strong to ignore. I am not lying when I say, to this day, when someone asks a favor of me, I am faced with an ethical decision or I need to take a stand on a science education policy, I ask myself “how would Jon react?” He is the little voice inside my head that keeps me grounded and gives me a moral compass.
I have tried to take on roles in my professional career that ultimately will lead to a better scientific world. At Brandeis, I am the academic director of the STEM Posse, I am part of several campus initiatives for retention, equity, and inclusion for underserved students in the sciences, and I find many of my most rewarding moments mentoring students underserved or at risk in STEM. I know Brandeis is just a small, niche community for now, but I can see myself being involved in these types of initiatives at a much larger scale someday. The fate of world lies in the training of a new, diverse, and empowered group of STEM undergraduates. I want every student to know they can succeed in STEM regardless of their background or demographics. I want to be the same type of supportive, inspiring mentor that Jonathan was for me, and they need to be knowledgeable, confident, and capable to make change in the world.
I don’t know if Jonathan ever realized what an impact he had on me personally, professionally, and academically. I was truly changed during my years of graduate school. I only hope that someday when I am giving a talk about equity in the sciences, a young bright student hears Jonathan’s voice echoing through me and scribbles down pages of notes reflecting on who and what they want to be.