The crosstalk of PPT1, APT1, and APT2 depalmitoylases in melanoma cells

Jennifer working inside vent hood

Students

2021
College

Faculty

Associate Professor of Medicine

Project Summary

For the summer of 2018, I worked in the Amaravadi Lab, an oncology lab that primarily focuses on autophagy in melanoma cells, with the support of my principal investigator Dr. Ravi Amaravadi and mentor Dr. Rani Ojha. While I had been working in this lab since the previous summer, during my three months this time, I was able to take on an extensive project with Dr. Ojha. Our project focused on three certain proteins that may alter autophagic function, the process which cells use to recycle old organelles and machinery; moreover, cancer cells often use autophagy to gain nutrients and continue their proliferation. The three proteins we examined were protein-palmitoyl thioesterase 1 (PPT1), acyl-protein thioesterase 1 (APT1), and acyl-protein thioesterase 2 (APT2). We ran numerous experiments to understand both how these proteins relate and interact and how they may alter levels of autophagy.

While I had already mastered cell culturing, western blots, and other common biomedical research protocols, this summer, I was able to learn many more. In preparation for in vivo experiments, I underwent mouse training, which was a frightening but rewarding experience. Outside of the mouse facility, I primarily worked at the bench and cell culture hood. The MTT assay and cell colony formation assay were important for me to learn, in that I am now able to analyze the functional importance of certain proteins in organisms. Mastering the immunoprecipitation and PLA protocols allows me to analyze protein interactions, and I also continued to build my experience with the acyl-biotin exchange assay, which is specific to measuring the lipidated characteristic of the three proteins under experiment. This protocol was especially satisfying, as I was able to combine the skills and knowledge I gained from other protocols in a cohesive manner.

However, no matter the numerous protocols and technical skills I acquired, procuring and analyzing the results was the most exciting. Experiments rarely provide clear cut data, so attempting to apply prior knowledge and develop a story was a challenge that I enjoyed. I would spend long afternoons sitting with Dr. Ojha trying to make sense of our data, and our heads would often hurt by the end of the day. Nevertheless, through this research experience I learned what it means to critically think and be proactive in the research field. I am excited to continue working on this project and better understand what it means to research independently.