Obesity has reached epidemic proportions and is directly linked to several systemic diseases including type 2 diabetes and coronary artery disease. Energy homeostasis is regulated by a complex neuroendocrine system involving peripheral signals such as the adipocyte-derived hormone, leptin, as well as several appetite-regulating neuropeptides. Obesity can result from dysregulation of either the peripheral or central signals. Our current understanding of the regulation of leptin is very limited. The significance of understanding the cell biology of leptin is based on its ability to relay information regarding the status of fat depots from the adipocyte to the brain. The overall goal of this proposal is to extend our understanding of the mechanisms of leptin synthesis and secretion. The specific objectives are: 1) To study transcriptional regulation of the leptin gene promoter. Recently, we showed that the appetite-stimulating neuropeptide, melanin-concentrating hormone (MCH) stimulates an acute increase in leptin mRNA in rat adipocytes and also stimulates an increase in leptin promoter-driven luciferase activity in transiently transfected 3T3-L1 adipocytes. These data strongly suggest the presence of a MCH-responsive element in the leptin promoter. To further define transcriptional regulation of the leptin promoter, we will identify the putative MCH responsive element through transient transfections of 3T3- L1 adipocytes with various point mutants of the leptin promoter construct. 2) To determine post-transcriptional control mechanisms that regulate leptin production. Preliminary data indicate that targeted degradation of leptin mRNA occurs via an instability element present in its 3'-untranslated region. Insulin and beta-3-adrenergic receptor agonists are known to stimulate and inhibit leptin production respectively via posttranscriptional and transcriptional control mechanisms. The ability of these agents to modulate leptin production in primary rat adipocytes via effects at the level of this instability element will be determined. 3) To study in vivo regulation of leptin production. We have shown that MCH stimulates leptin production in vitro. We will address the biological significance of this observation by determining the effects of both central and peripheral administration of MCH on leptin gene expression and circulating leptin levels in wild-type and MCH-knockout mice respectively.
