Control of fat cell mass and fat cell size may result from signals generated in the brain or from other tissues but ultimately cellular events transpire that are translated into partitioning of fuels in a regulated manner, presumably due to the particular enzyme and metabolite concentrations that are present. Evidence in support of this concept is derived from the observation that, despite ad libitum access to food, most mammals maintain a constant weight and constant fat mass. The focus of the proposed studies is to understand regulation of partitioning of free fatty acids (FFA) between utilization (to produce energy or heat) and deposition (as triglyceride) in adipocytes and the impact of alterations in the putative key enzymes that regulate partitioning on fat accumulation. Partitioning between storage and disposal is subject to regulation at the enzymatic level on a minute-to-minute time scale, while enzyme levels are subject to transcriptional regulation, on a longer time scale. In particular, acyl CoA synthase is essential for the generation of long chain acyl CoA (LC-CoA), the precursor to both oxidation and complex lipid formation, while carnitine palmitoyl transferase-1 (CPT-1), acetyl CoA carboxylase and uncoupling proteins (UCPs) control the fate of the metabolically active form of FFA, LC-CoA. It is hypothesized that cells with low CPT-1 or UCP activity have lower rates of beta-oxidation, higher levels of malonyl CoA and LC-CoA, and a greater capacity to accumulate lipid than cells with high CPT-1 or UCP activity. Further, that altering enzyme expression, will change fuel partitioning. Preliminary data document differences in levels of LC-CoA, CPT-1 sensitivity and isoform distribution, and capacities for lipid accumulation and FFA uptake (even at the single cell level) in rat and human preadipocytes from different regions that have been differentiated in culture.
The Specific Aims will address the following questions using 3T3-L1 preadipocytes and rat adipocytes. 1. How does beta-oxidation vary with nutritional and transcriptional manipulations in fat cells? 2. Which enzymes regulate intracellular partitioning of FFA? 3. Do LC- and malonyl-CoA exert metabolic control over FFA partitioning? 4. Do FFA or LC-CoA modulate mitochondrial energy efficiency or the activity of UCP?
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