Phosphatidylinositol 3-Phosphate in the Regulation of Autophagic Membrane Remodeling

项目总结/文摘

PROJECT SUMMARY/ABSTRACT Autophagy is a fundamental cellular process mediating lysosome-dependent degradation of proteins, organelles, and intracellular pathogens. Dysfunction of autophagy is associated with many diseases, including cancer, neurodegeneration, diabetes, and chronic infections. Better elucidation of the molecular mechanisms for autophagy may inspire new therapeutic approaches to these diseases. Autophagy initiation occurs at endoplasmic reticulum (ER) subdomains enriched with phosphatidylinositol 3-phosphate (PI3P). Autophagy is blocked when depleting or inactivating VPS34, the phosphatidylinositol (PI) 3-kinase responsible for PI3P synthesis during autophagy. Nevertheless, how PI3P contributes to membrane remodeling events crucial for autophagosome biogenesis remains elusive. Much is also unknown about how PI3P synthesis and turnover are orchestrated at autophagy initiation sites. Dr. Hsieh recently discovered that the Legionella PI 3-kinase MavQ generates PI3P on the ER and drives membrane remodeling. He also found that MavQ is coupled with the Legionella PI 3-phosphatase SidP to spatiotemporally modulate PI3P levels at ER subdomains, inducing vesicle/tubule budding. The striking similarity between this pathological process and autophagy initiation prompts Dr. Hsieh to dissect the molecular and physical factors that PI3P brings for autophagic membrane remodeling. Moreover, the current technical hurdles in tackling this question can mostly be cleared by using MavQ and SidP as tools. In Dr. Hsieh’s proposed research, Aim 1 will develop optogenetic methods to control PI3P generation in the cell and determine how PI3P regulates autophagic membrane remodeling using live-cell imaging and systematic genetic perturbations. Aim 2 will combine in vitro biophysical assays, optogenetic control, and quantitative imaging to elucidate how PI3P domain formation leads to membrane remodeling. Aim 3 will reconstitute a reaction-diffusion system in vitro and use this system to determine how PI 3-kinases and phosphatases drive the formation of PI3P-enriched membrane subdomains. Dr. Hsieh’s career goal is to become a leader in cell biophysics and membrane biology, focusing on physiological and pathological processes involving membrane remodeling. Training during the award period will prepare him to lead an independent research group using cell biological, biochemical, and biophysical approaches to understand the mechanistic basis of autophagic membrane remodeling. UT Southwestern Medical Center provides an excellent environment to aid Dr. Hsieh’s proposed research and career development. Dr. Hsieh has also set up an advisory committee consisting of leading scientists with complementary research expertise. Under their guidance, Dr. Hsieh will receive the necessary research training, such as preparing and using various model membrane systems, and further develop professional skills during the award period. These will significantly facilitate Dr. Hsieh's transition into an independent investigator and ensure his future success.