Analysis of Lipolytic Trafficking in Adipocytes.

Project Summary/Abstract Disruption of cellular neutral lipid metabolism promotes the progression of obesity, diabetes, fatty liver disease, and cancer. The long-term scientific goal of the project is to understand the molecular mechanisms that control lipid storage and mobilization in order to identify novel points for therapeutic intervention. ABHD5 regulates cellular lipid metabolism, including PNPLA2/ATGL, the rate-limiting triglyceride lipase in key metabolic tissues. Nonetheless, the mechanisms by which ABHD5, a protein lacking enzymatic activity, activates PNPLA2 (and other PNPLAs) remains an important mystery. We hypothesize that the remodeling of biological membranes is a general mechanism by which ABHD5 regulates enzyme access to membrane-delimited neutral lipid substrates. Mechanistically, we hypothesize that ligand binding stabilizes ABHD5 molecular and macromolecular conformations that target and alter membrane biophysical properties (tension and curvature) to allow the lipase access to specific substrates sequestered within lipid droplets (LDs). We will test this hypothesis using novel chemical probes of ABHD5 and informative genetic mutants in live adipocytes (Aim 1). We will directly assess the impact ABHD5 on the biophysical properties of membranes in model LD systems using an array of high resolution and high throughput approaches (Aim 2). These Aims are designed to be highly complementary and to provide strong cross- validation between experimental platforms. In addition, we present data demonstrating that ABHD5 is targeted to specific subcellular sites where LDs form upon fatty acid supplementation. Furthermore, the interaction of ABHD5 with PLIN5, driven by the ABHD5 ligand oleoyl-CoA, facilitates LD formation. Aim 3 will dissect the biochemical pathways promoted by ABHD5/PLIN5 complexes and, in concert with Aim 2, evaluate the impact of this interaction on the biophysical properties of model membranes. ABHD5 is emerging as a compelling therapeutic target for metabolic diseases and cancer. The results of this project will provide new insights into specific mechanisms by which ABHD5 regulates fatty acid flux at the cytosol/LD interface.