Adipose tissue triacylglycerol (TAG) is the major source of energy in higher eukaryotes. The ability of adipocytes to mobilize free fatty acids (FFAs) from TAG stores during times of demand is essential for survival. The altered rates of FFA release are strongly associated with the development of obesity and type 2 diabetes, both in humans and in rodents. Lipolysis, the process in which TAG is hydrolyzed to FFAs and glycerol, has been a subject of intense investigation for decades, with much focus on hormone-sensitive lipase (HSL). With the discovery of adipose triglyceride lipase (ATGL) as the rate-limiting enzyme in lipolysis, a clearer understanding of the molecular processes governing TAG mobilization emerged. Although recent studies identified CGI-58 as a co-activator of ATGL, the mechanisms that regulate the ATGL enzyme action remain insufficiently understood. In this regard, our preliminary studies have uncovered a novel mechanism for the control of ATGL-mediated lipolysis. We have obtained compelling evidence that a protein encoded by G0/G1 switch gene 2 (G0S2) is a selective regulator of ATGL. G0S2 is highly expressed in adipose tissue and differentiated adipocytes. It specifically interacts with ATGL, leading to the inhibition of its TAG lipase activity. Knockdown and overexpression experiments demonstrate that G0S2 functions to attenuate ATGL-mediated lipolysis in both adipocytes and nonadipocyte cells. The overall goal of this proposal is to further explore the molecular details of ATGL inhibition by G0S2, and more importantly, the physiologic roles of G0S2 in TAG lipolysis in the context of various functional settings. The goal will be accomplished by (1) determining the biochemical mechanisms by which ATGL activity is regulated by G0S2; (2) testing the hypothesis that G0S2 and CGI-58 play opposing roles in ATGL-mediated lipolysis; and (3) evaluating the effects of adipose overexpression of G0S2 on energy metabolism and insulin sensitivity. In aim 1, we will perform both enzyme kinetic studies and substrate interaction assays. We will also map the precise sequence motif(s) in ATGL for G0S2 binding. For aim 2, we will manipulate relative levels of G0S2 and CGI-58 in various cellular setting and examine the effects on ATGL-mediate lipolysis and the related functional consequences. Aim 3 will be accomplished through diet treatment studies using a transgenic animal model overexpressing G0S2 fat specifically. These studies will contribute to our understanding of TAG metabolism, and to the development of new therapeutic strategies against obesity and type 2 diabetes. PUBLIC HEALTH RELEVANCE: Obesity is a prevalent disorder of energy balance in which excess energy, in the form of triacylglycerol, accumulates in adipose tissue. Despite a need for therapeutic approaches to treat obesity, little is known about the mechanisms that control fat storage and release from adipose tissue. The proposed study will focus on investigating the important role of a new protein that regulates ATGL, the key enzyme for fat mobilization in adipose tissue.