Author Archives: Zach Pracher

Coming into this program, I wondered if I would be able to contribute anything to the McClay lab’s work with my extremely limited knowledge, and wondered precisely how many things could (and would) go wrong along the way. I wondered if research was something I even wanted to work in, or if it was simply a lofty ideal stuck in my head, filled with distant figures in white coats. Throughout the summer, though, I got the opportunity to meet people who have gone through this exact struggle, and were compassionate and understanding in helping me answer these questions on my own, even if they didn’t know it at the time.

On one level, I’ve seen that research is not something anyone does alone. Some of the most valuable moments in the lab this summer have started when one person has an idea, thinks about it for a while, and then walks over to someone else to get their perspective on it. While the resulting conversations are certainly products of extreme intelligence and experience, they are also filled to the brim with creativity, which I’ve learned is essential to progressing the frontier of knowledge. This creativity, though, goes together with failure, and that’s ok. Many times, someone will say “Well, that might not work, because…” but then they work together and use their creativity to come up with yet another way to test their idea! Sometimes the failure is only realized at the bench, and then it’s simply time for another great conversation, and probably another few weeks of experiments. Best of all, these conversations have their fair share of funny comments and playful jabs along the way. Then, once the conversation’s finished, people ease back into the privacy of their thoughts to continue designing experiments to satisfy their wonder about a biological system, even if only for a moment.

These conversations, combined with the awesome faculty members that have come to talk to us through the summer, have also shown me an interesting juxtaposition in science: modern science is intrinsically collaborative, but it is also self-driven and critical. Generally, us students are used to other people pushing us forward, like teachers, parents, or coaches. But over this summer, I’ve discovered that no one has to push you in research. Not once did Dr. McClay look over my shoulder to make sure I was reading articles. Not once was I told to sit down and question everything I knew and had read so I could realize how little I didn’t know. Not once were any of the PIs that came to present to us told to be energetic and committed. Yes, research can be a glorious, collegial atmosphere of amazing scientific advances, but I realized that it is also largely what you make of it – a prospect at once daunting and invigorating, and one that I know I will continue to encounter and hopefully improve  on in my career, no matter the direction I take. Essentially, this summer taught me that scientific research sits at the intersection of drive, creativity, failure, and most of all, wonder. Given all I’ve learned and still have yet to learn, I can’t wait to come back in the fall and get back into the awesome environment that is scientific research, and maybe even go to graduate school and become a professional researcher. I know the path is hard, because I’ve talked to people that are traversing it right now, and there will certainly be moments of creativity and perhaps years of failures or faltering drive. Through it all, though, the experiences I’ve had and the people I’ve met this summer have taught me to make sure I keep doing one thing:

Wondering.

Zach Pracher

Mentor: Dave McClay, Ph.D.

Department of Biology

The developmental gene regulatory network (GRN) of the green sea urchin (Lytechinus variegatus) has been extensively investigated to illuminate the genetic interactions underlying embryonic development and has led to many insights in embryonic development and regeneration. Although the putative metalloprotease Astacin-4 is widely expressed in urchin embryonic blastocoelar cells, its developmental function and position in the established developmental GRN remains unknown. In our study, inhibiting Nodal with an antagonistic drug throughout embryonic development revealed that Nodal signaling between 4 and 6 hours post fertilization is critical to downstream Astacin-4 expression, blastocoelar cell formation, and overall embryo development. Separately, we repressed Astacin-4 expression using a morpholino to determine the potential role of Astacin-4 in embryonic and blastocoelar development and function. Embryo morphology and gene expression patterns were subsequently assessed using in situ hybridization and microscopy techniques. We hypothesize that repressing Astacin-4 inhibits normal blastocoelar cell formation, and we generally expect to elucidate the role of Astacin-4 in the epithelial-mesenchymal transition of blastocoelar cell precursors. Understanding the regulation and function of Astacin-4 in L. variegatus can enhance our knowledge of the genetic relationships underlying immune system development as well as evolutionary conservation of immune system development across the animal kingdom.

This week everyone in BSURF gave quick presentations to the rest of the program, essentially summarizing our work so far this summer in our respective labs. Specifically, I wanted to highlight Ben Johns’ chalk talk about his work on the cell-cell interactions underlying collective cell migration in the Hoffman lab. Ben’s research focuses on a protein called “vinculin,” which basically forms part of a larger scaffold connecting the actin cytoskeletons of neighboring cells, allowing them to coordinate their individual movements into a larger-scale phenomenon called “collective cell migration,” or CCM for short. With a palpable and contagious excitement, Ben explained to us how CCM is an integral part of essential processes like wound healing (regeneration) and morphogenesis, plus how his work in elucidating the localization and function of vinculin can enhance our understanding of CCM in general. This research particularly stuck out to me since my research this summer also focuses on development (albeit in sea urchins) and I find the whole field of regeneration fascinating, but hadn’t explored much how mechanobiology might play a role in either of these systems. He drew me in further, though, when he offhandedly mentioned the presence of alpha-catenin in CCM, because earlier this week my mentor had mentioned the role of beta-catenin in initiating movement during a developmental process our lab studies, and it was listed as a central element in the GRNs our lab has helped construct over the years. During the break after his presentation, we got to talking and found some pretty great connections between our work that might’ve gone undiscovered had Ben never mentioned it.

While these connections between Ben’s work and my own struck me as incredibly unexpected at the time, I’ve since come to realize they’re completely natural and, in fact, fully expected. It can be easy to fall into a rigid mindset sometimes, where you feel like you’re studying this specific field and nothing else, but Ben’s presentation reminded me of the intrinsic harmony across biological research. His project focuses on protein-protein interactions and a much more mechanical/physical model than the transcriptional GRNs of my work, and yet we’ve found this commonality where mechanobiology plays a role in development, and developmental GRNs play a role in operating and creating mechanical systems. Further, this symbiosis between our research interests has helped us both begin to understand each other’s fields of research better, which could alter the way each of us conducts research and think about problems that are archetypal to each of our fields which may require interdisciplinary solutions. Overall, Ben’s presentation helped energize me to continue looking into research that may just seem cool, if not completely unrelated to anything I’m working on, because sometimes that’s where the best and most valuable connections lie. Ben’s contagious enthusiasm for his work in mechanobiology, combined with these unexpected but awesome insights into our shared interests, meant that his presentation was a particular highlight of my week, and not one I’ll soon forget.

During our interview, Dr. McClay said keeping up with the perpetually-advancing frontier of science was easy because “The game is fun.” Now, after three weeks, it’s time for me to recount my own opening moves in the game of scientific research. Any competitive gamer or athlete could tell you that before you can make a move, you must study it extremely thoroughly, or else risk the entire game. In research, this translates to comprehensively and continuously reading the relevant (or even seemingly irrelevant) published work. Each article, then, becomes a kind of minigame to showcase a new move, evaluate its merits and shortcomings, and determine how it’s helped the game progress.

My reading has generally focused on understanding the specific interactions in the gene regulatory networks (GRNs) underlying sea urchin embryonic development, but I’m beginning to read some more about developmental biology in general, which has lately involved some interesting computational and synthetic approaches! Since week 2, I’ve also begun delving into other scientific areas where my gene of interest (Astacin-4) has shown up, to see if there are potentially homologous functions I might be able to look for in my own experiments. Typically I’ll play this article minigame only a couple times a day, because the real work comes in transitioning from understanding the minigame to understanding it in the context of the other information collected (the other possible moves). This synthesis likely takes up the rest of my “study” time, and, together with the reading itself, accounts for the solid majority of an average day in the McClay Lab.

Of course, once you’ve studied your options, the next logical step is to actually make a move. To start an experiment, I collect urchin eggs and sperm, fertilize, and then transfer the embryos to seawater containing a certain gene inhibitor (the specific timing has varied by experiment). When the treatment is finished (usually a day after), I put the embryos through a 3-day washing procedure, marking them with certain genetic probes along the way, and then use a microscope to analyze the stained embryos for changes in gene expression patterns that might help me illuminate relationships between known sea urchin embryonic GRN elements and Astacin-4. Once I’ve collected that data, or even simply just finished a particularly interesting article, I’ll spend a little while talking to Dr. McClay or Esther (our lab technician) about possible interpretations, lessons for next time (of which there are always plenty), and even just random things about science in general, like the difficulty of publishing contrary results.

Looking back, the composition of this post approximates pretty well most days of playing the game in the McClay Lab. It’s about 70% reading and thinking, 5% talking about it, and 25% actually doing anything I read, thought, or talked about. Overall, my daily schedule (though hardly routine) emphasizes 2 essential concepts of research: the “game” of research is almost entirely mental, while the physical aspect comprises a small yet essential fraction of the work, and, each day, you only progress as much as you allow yourself to.

A virtual shoo-in at Penn State thanks to his dad’s faculty status, then-young Dave McClay had little certainty about what to study in college. He wandered through five majors and their associated coursework during undergrad, unable to decide what most interested him, until he took a genetics course with an awesome professor which started him down the path to graduating with a degree in biology. The next logical step, then, was graduate school at the University of Vermont. Why? To ski, of course! And, you know, maybe do some research or take courses or something, too. Yet, while he may not have intended it originally, the clear, cold mountain air made helped McClay start asking himself what he actually wanted to do with his life. He had always admired one of his father’s colleagues, who was more of a researcher than his father, an administrator, and McClay could seriously envision himself working in research for the rest of his life. But the slopes of snowy Vermont brought him clarity of a more disturbing, if not motivating, nature: he was never going to succeed at this unless he started putting the work in.

And so it came to be that Dave McClay found himself in his advisor’s office, discussing his transition to the up-and-coming field of developmental biology, when the phone rang, asking for someone who might be able to teach anatomy to nursing students. Dave accepted, and thus began his now long and venerated teaching career. Eventually, he decided that research was the life for him and moved to the University of Chicago to do PhD work on cell adhesion and discovery of molecules that affected cell-cell adhesion under a less-than-ideal (read: tyrannical) mentor. Over time, now-Dr. McClay gained more independence and transitioned his research from discovering particular molecules governing adhesion to the network control of cell adhesion, leading to the McClay lab’s current focus on embryonic sea urchin gene regulatory networks. And along the way, he got his first faculty position job at some university in Durham, North Carolina, where he’s been happily teaching and researching ever since.

Earlier, I said “over time” in describing the evolution of Dr. McClay’s research, but what that really meant was, “over forty years in which the field of biology was radically changed forever.” During that time, RNA suddenly acquired biological meaning, computers started becoming widespread tools for research, and humans figured out how to read and eventually edit our own genetic code –  and Dr. Dave McClay was there for every second of it. When asked how he kept up with these revolutionary changes, Dr. McClay simply said, “The game is fun.” For him, keeping up with the advancing frontier is nowhere near a chore because science easily continues to fascinate him enormously even after all these years. More than that, he gets to learn about these awesome, new wonders of science and then turn around to teach it to the next generation, inspiring them to learn more about it in turn. As great as it is to marvel at the incredible features of life that we now understand, though, Dr. McClay truly loves everything we haven’t learned yet; all the knowledge that remains to be known. To Dr. McClay, the best part of science isn’t the high of figuring out something new (although that can be pretty great), but rather being able to come up with questions that you didn’t even know to ask before, and then getting to set up an entirely new investigation to just begin answering those questions, gladly entering a seemingly perpetual cycle of wonder, inquiry, and discovery. Simply put, Dr. McClay’s favorite part of being a researcher, after dedicating most of his life to developmental biology, is “tomorrow’s experiment.”

I’m sure the last thing anyone wants to read about right now is more immunology. A lot of people (myself included) have thought something like this at least once in the past few months: “If I hear the word ‘antibody’ one more time, I will walk out of this room right now.” Well, the bad news is I’m still going to be talking amount immunology, but the good news is my research has to do with a different kind of immune system! While antibodies are essential to fine-tuning humans’ adaptive immune response to very specific pathogens, there is another, broader kind of immune system called the innate immune response. The innate system recognizes general biological traits associated with pathogens, like glycan (a component of many bacterial cell walls), unlike the very specific adaptive system. Given the recent explosion of coverage concerning antibodies, it would be easy to think the innate response is just irrelevant, but that wouldn’t be doing justice to the evolution and ubiquity of innate immunity. Immunologists and developmental biologists discovered that innate immunity is way older than adaptive, and is present in a vast array of organisms compared to adaptive immunity, which is really only present in vertebrates (that’s us!). Because adaptive immunity likely evolved from the innate response, it also means that the two are far more interconnected than anyone previously thought.

Because the innate system is so old, it’s had a lot of time to diversify the molecules and cells involved, making it hard to draw evolutionary connections between the immune systems of humans, which have both innate and adaptive immune systems, and those of sea urchins, which have only an innate system. So instead of looking only at the kinds of immune cells and molecules produced, essentially the “end results” of immunity, developmental biologists and immunologists have turned to gene regulatory networks (GRNs) that determine when and how these immune cell types develop. Through these GRNs, we can better understand both how immune systems evolved and what role each gene plays in immune cell development and function.

Recently, the McClay Lab discovered a gene that is very highly expressed in a certain cell type of the embryonic sea urchin innate immune system. There’s also a kind of “master signal” at the beginning of sea urchin development which has been really thoroughly investigated over the past couple of decades. My work in the McClay Lab this summer focuses on finding out if there is a significant connection between this original “master signal” and this specific “end product” gene in the immune cells. If the genes can affect each other, it means there is probably a GRN connecting them, but we have very little idea of how many gene components are in the circuit, what they are, or what they do – all out there to be discovered. If there isn’t an observable connection between the master and end genes, then the end gene could be the tail of a completely unexpected GRN, which poses an equally exciting opportunity for research and discovery! I’ll be using a battery of microscopy, molecular biology, and moving-colorless-liquids-back-and-forth techniques to get at this GRN, and probably producing some really cool pictures of colorful embryos along the way (stay tuned)! Although this project may seem daunting, characterizing this genetic circuitry could help us better understand the incredible harmony between diversity and unity in immune systems across all domains of life, and provide some really awesome insights into how to analyze rapidly evolving biological systems, like the immune system. The past week working on this project has been a complete joyride, and I can’t wait to keep it going through the summer!

Reading biological research papers and hanging at the fringes of the research community, you start to get accustomed to the model organisms used in research (I’m looking at you, D. melanogaster) – the sea urchin, however, was not one I’d ever encountered before. Since starting with the McClay Lab, though, I’ve begun to understand the elegant complexity of the sea urchin. Looking through the microscope on my first day down at newly-fertilized embryos, then watching the first embryonic cleavage, I was struck by how simple it all appeared. Where before there was one circle, now there were two conjoined ovals. Then I swiveled my chair around to look at one of the many copies of the embryonic gene regulatory network posted around the lab, and was struck by the extreme complexity underlying this ostensibly simple process. All of these complex interactions between transcription factors of constantly differentiating cells, all their neighbors, and even cells clear across the embryo regulate this dynamic, yet elegant, process of embryonic development.

This contrast of elegant complexity is the basis of my own work in the McClay Lab for this summer. I’ll be looking at where previously understood cellular regulators (like transcription factors) affect the development of the urchin innate immune system cell types. Over the summer, I want to develop my ability to constantly ask probative questions to fully understand and critically evaluate the logic and discoveries of cutting-edge research, rather than simply accepting previous work as truth. By developing my skepticism and understanding of the research tools currently available, I also hope to become a more independent lab member, even designing some experiments of my own in the future, thus beginning to carve my own path through the landscape of scientific research by asking (and even perhaps beginning to answer) some of my own questions about the elegant complexity of life.

But I know the world of research isn’t filled to the brim with glowing accomplishments, earth-shattering questions, and “Eureka!” moments. I expect to spend a not-insignificant portion of my time just trying to figure out precisely what went wrong, and then trying an experiment again (and again, and again if necessary) to see if something else goes wrong. Humility, particularly the ability to admit that you were wrong, no matter how horribly wrong you were, is integral to science, and I look forward to developing that skill among my immensely talented and compassionate peers in the McClay Lab. By the end of the summer, I want to begin humbly asking interesting and valuable questions, be independent and driven while enthusiastically collaborating with others, and always ask for help when I need it, all while contributing to our understanding of the elegant complexity that comprises life. In short, I want to start to start becoming a scientist.