Comparative functional analysis of endocytic membrane curvature-sensing BAR domain proteins in cancer and stem cell lines

Summer 2016

Jordan Said : Molecular and Cell Biology, Chemistry (minor), Music (minor)

Mentor: David Drubin

Clathrin-mediated endocytosis (CME) is a highly conserved cellular process responsible for nutrient uptake and membrane recycling in eukaryotic cells. After initiation, clathrin and adaptor proteins are recruited to form a clathrin-coated pit, which ultimately matures into a vesicle that is cleaved from the membrane by the concerted efforts of multiple proteins. Several of these endocytic proteins possess a conserved Bin-Amphiphysin-Rvs (BAR) domain, a protein element that can both sense and generate membrane curvature.

One of the key proteins in mammalian clathrin-mediated endocytosis is sorting nexin-9 (SNX9), a protein involved in cellular migration and membrane association but has received less study in the context of endocytosis. In clathrin-mediated endocytosis, SNX9 regulates oligomerization-dependent GTP hydrolysis by the cleavage protein dynamin at the end of the endocytic event, a necessary event for the scission of the invaginated membrane into an intracellular vesicle. It is unknown precisely how the BAR domain of SNX9 affects endocytosis and related protein-protein interactions with dynamin and other regularly endocytic proteins, such as the neural Wiskott–Aldrich Syndrome protein (N-WASp). This project will investigate the consequences of a BAR-mutated SNX9 on endocytosis and activity of SNX9 in induced pluripotent stem cells (iPSCs), in comparison to more commonly used mammalian cancer cell lines. iPSCs possess normal chromosome number and physiology, making them more ideal for cell research. This research will afford a greater understanding of the role of the BAR domain in endocytosis, allowing for some of still not well understood mechanisms underlying the complex process of CME to be characterized at the level of protein:protein interactions and cell dynamics.

I would like to extend deep gratitude to the Thye Foundation for providing this invaluable opportunity to devote myself full-time to research in a field I have become so passionate for. The funding, resources, and other forms of support provided by the Thye Foundation allow me to grow as a student researcher and further my journey toward a career in the applied health sciences. Joining and working in the Drubin Lab has been the paramount experience in my development as a scholar and scientist, providing me with bountiful experience in designing experiments, analyzing data, and forming logical scientific conclusions in accordance with relevant literature. My development in the Drubin Lab thus far and research this summer would not be possible without the generous support of the Thye Foundation, as well as my incredible Principle Investigator, David Drubin, and mentor, Jessica Marks.