ABSTRACT
First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping researchers promote themselves alongside their papers. Mitchell Leih is first author on ‘ Disordered hinge regions of the AP-3 adaptor complex promote vesicle budding from the late Golgi in yeast’, published in JCS. Mitchell is a PhD Student in the lab of Greg Odorizzi at the University of Colorado, Boulder, investigating adaptor protein complexes and their dynamics in membrane trafficking.
Mitchell Leih
How would you explain the main findings of your paper in lay terms?
Adaptor proteins recognize protein cargoes, package them into vesicles and ensure these cargos end up where they need to go. This mechanism can be broken down into the steps of vesicle budding, trafficking, tethering and fusion, as these vesicles form at a donor membrane and move to their target cellular compartment. In the case of the adaptor protein complex AP-3, tethering and fusion have been studied in-depth, and specific factors that guide these processes have been identified. In our study, we show that the mechanism for successful AP-3-mediated budding and trafficking is dependent on disordered hinge regions of the AP-3 complex, areas of the proteins that are unstructured and flexible. By truncating these hinges, we show that AP-3 accumulates onto Golgi membranes because AP-3 vesicles fail to bud. This failure to bud drastically reduces the number of motile AP-3 vesicles and leads to the missorting of AP-3 protein cargoes.
Were there any specific challenges associated with this project? If so, how did you overcome them?
Previously, tracking of AP-3 vesicles in live cells has been limited to timelapse imaging with multi-second resolution. When observed in real-time, we found that AP-3 vesicle trafficking is much faster than these time scales can capture. With newer imaging technologies, we were able to take images at 50 ms intervals, which is 100 times faster than previous studies that looked at AP-3. The results of this approach solidified the key finding in the manuscript that AP-3 hinge truncations halt AP-3 vesicle budding from the Golgi and drastically decrease the rate of vesicle displacement in the cell.
When doing the research, did you have a particular result or ‘eureka’ moment that has stuck with you?
I have spent a lot of time imaging on the microscope, and we were incredibly excited when we first saw AP-3 accumulating in donut-shaped structures in hinge truncation mutant cells. However, when we moved to super-resolution imaging, we noticed these donuts were incomplete. AP-3 seemed to form dotted lines in a circle around Golgi structures. Once we found that these dotted circles could be filled in by the Golgi-associated adaptin GGA, I was pretty excited. This finding led us to perform experiments on AP-3 in the context of Golgi maturation, which really helped to fill out the paper.
Why did you choose Journal of Cell Science for your paper?
We chose Journal of Cell Science because of its reputation for recruiting excellent reviewers and its commitment to scientific excellence.
Super resolution microscopy image showing the distribution of AP-3 in wild-type yeast (left) and in our hinge truncation mutant, apl6-734Δ (right). Apl5-GFP is shown in green, and the vacuole lumen is shown in blue. The cell outline is indicated with a white dashed line.
Have you had any significant mentors who have helped you beyond supervision in the lab? How was their guidance special?
My co-mentors Greg Odorizzi and Alex Merz have greatly influenced my view of science and experimental rigor. They taught me how to have active discussions, think broadly and remember the decades of work that have brought science to where it is today.
What motivated you to pursue a career in science, and what have been the most interesting moments on the path that led you to where you are now?
My desire to pursue a career in science began in an Advanced Placement biology class I took in high school. I don't remember exactly when it happened, but somewhere along the line, pieces began falling in place and opened my mind to the patterns of biology, structure and function. When I pursued my undergraduate degree, these pieces continued to come together to create a picture of complex but beautiful systems. However, when I started discussing my fascinations with folks not in science, I didn't get the same enthusiasm from them − that was, until I sat down with them and explained what was so fun and important about what I was studying. To this day, I strive to push for understanding with those who think science is beyond them and generate inclusive conversations to demystify what goes into scientific discovery. In this way, I love both pursuing basic science in my research and, in parallel, communicating science in an accessible and relevant way to different audiences, regardless of their educational background.
Who are your role models in science? Why?
My dad, Dr Mike Leih, is my most influential role model in science. Although he studies information technology rather than biology, he has been and continues to be a model of resilience, patience and advocacy for those who want to learn.
What's next for you?
I recently accepted a postdoctoral position with Dr Kelsie Eichel, where I will focus on membrane trafficking in neurons.
Tell us something interesting about yourself that wouldn't be on your CV
I try to learn a new practical skill every year. Since starting grad school, I have learned woodworking and bartending and am currently learning to sew.
Mitchell Leih’s contact details: 347 University of Colorado, Boulder, CO 80309, USA.
E-mail: [email protected]
