Role of caveolae in signaling in fat cells

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DESCRIPTION: (provided by applicant) Obesity is the major factor predisposing people to insulin resistance and type II diabetes, despite the fact that it is the failure of skeletal muscle to respond to insulin which prevents glucose uptake and results in hyperglycemia and diabetes. From a mechanistic viewpoint, it is the availability of lipids (fatty acids) from fat stores that produces muscle insulin resistance, in part if not entirely. Insulin resistant skeletal muscles contain more fat than normal muscle, and acute perfusion of fatty acids into muscle will rapidly produce this resistance. Indeed, Richard Bergman and colleagues have postulated that inhibition of lipid release from fat cells is rate limiting with respect to insulin's organismal actions (the so-called single gateway hypothesis to explain the rate limiting step of insulin action). Dennis MeGarry has also emphasized the role of free fatty acids in muscle insulin resistance and the failure, in diabetes, of insulin to suppress fatty acid release from adipocytes. Moreover, the recent discovery of the role of the adipocyte with regard to leptin secretion has added further to the importance of this cell in the regulation of metabolic homeostasis. Thus, while there remains incomplete agreement about cause and effect in type II diabetes, no one would argue that obesity and fat cell metabolism are not critically relevant. The mechanism(s) by which fatty acids are taken up (and released) by adipocytes is not clear. Published data as well as data in this application suggest that structures abundant in adipocytes, called caveolae, may be the Site of lipid (fatty acid) entry and egress in these cells and may play a role in regulating lipid flux. Caveolae (little caves) are sac like structures that protrude into the cell interior from the cell surface. They are an anatomical feature of most cells whose overall physiological role is still unclear and controversial. It has been shown that caveolae bind fatty acids, and caveolae have been postulated as the site of cholesterol release from cells. We have raised a novel monoclonal antibody with which we can irnmuno-isolate caveolae. We are using this new tool to characterize the composition and physiological function of caveolae. In support of a role for caveolae in lipid metabolism, we have identified a putative fatty acid Lransporter (FAT/CD36) as a major protein component. We propose three specific aims: 1. to further characterize the protein constituents of caveolae in primary and cultured adipocytes. 2. to determine the physiological function of these proteins. 3. to modulate the expression of caveolae and determine the effects of this on the function of specific proteins as weU as on overall fat cell metabolism. Such studies address fundamental questions concemin2 insulin resistance as well as the cell biologv of caveolae.