This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Obesity is a disorder that imposes increased risk of diabetes, cardiovascular disease, and cancer. Leptin, a protein produced by fat tissue, plays an important role in regulating food intake and appetite by acting on specific receptors in the brain. Differences in the fat's production rate of leptin, together with resistance to leptin at its site of action or a combination of these factors can influence eating behaviors and energy use to cause obesity. Leptin also functions to regulate sexual maturity and reproduction, immune responses and bone formation. Our research focuses in understanding the mechanisms that govern leptin production and secretion in adipose (fat) tissue. In order to study protein-protein interactions that regulate leptin synthesis, traffic and secretion we utilized the yeast two hybrid technique to identify proteins that bind to leptin in adipocytes. We constructed and screened an adipose cDNA library with the full length leptin cDNA as a bait. Positive clones were verified in liquid -galactosidase assay and subsequently DNA has been isolated from yeast for insert analysis and sequencing. We have identified several proteins that interact with leptin. Biochemical and in vitro functional studies will be used to confirm these interactions and to examine the role that these proteins play in the regulation of leptin and other adipokines. These protein interactions will be further characterized using NMR structural stu
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