Project Abstract/Summary The rising prevalence of obesity and type 2 diabetes threatens to limit human healthspan by increasing the risks for cancer and cardiometabolic disease and to impose overwhelming economic burdens. New therapeutic strategies are urgently needed. Since the discovery of functional brown and beige adipocytes in adult humans, much attention has focused on exploiting the ability of these thermogenic adipocytes to dissipate excess energy as heat through uncoupled mitochondrial respiration. Advanced imaging in humans has revealed a favorable correlation between brown fat mass and cardiometabolic risk factors. A major gap in the field is a safe and effective pharmacological strategy to activate brown and/or beige adipocytes to promote negative energy balance, reverse obesity, and mitigate obesity-related metabolic disorders, such type 2 diabetes, cardiovascular disease, and nonalcoholic fatty liver disease. The overall goal of this application is to provide proof-of-concept for one such strategy that has been suggested by the lab’s long- standing research program on Perilipin 5 (PLIN5), a member of the Perilipin family of lipid droplet proteins that is expressed in oxidative tissues, including brown adipose tissue (BAT). A growing body of literature from our lab and others has implicated PLIN5 not only in the regulation of lipolysis at the lipid droplet surface, but also in the regulation of gene expression via interactions in the nucleus with SIRT1 and PGC1a and in the tethering of lipid droplets to mitochondria. Our published work in mice has shown that PLIN5 is required for the metabolic, transcriptional, and mitochondrial adaptations of BAT to cold stress. We have also shown that PLIN5 gain-of-function in BAT of mice can prevent glucose intolerance and fatty liver from high-fat diet and promote healthy remodeling of white adipose tissue (WAT) with prevention of adipocyte hypertrophy. In this renewal application we propose to test the hypothesis that promoting PLIN5 expression in BAT in conjunction with b3 adrenergic receptor agonist treatment of diet-induced obese mice will reverse adipocyte hypertrophy in WAT, reverse obesity, and reverse glucose intolerance. Aim 1 will elucidate the metabolic and signaling pathways responsible for the effects of PLIN5 on mitochondrial form and function in BAT and on systemic lipid and glucose metabolism by means of genetic mouse models, in vivo structure- function studies, and mechanistic mitochondrial experiments. Aim 2 will interrogate the signaling pathways and physiological responses associated with treatment of diet-induced obesity with a 2-hit intervention built on augmentation of PLIN5 in BAT in synergistic combination with b3 adrenergic receptor agonist treatment. Successful completion of these Aims may establish the conceptual foundation for a new therapeutic paradigm for treatment of obesity that is efficacious but at lower, non-toxic doses of existing medications.