Lipid Binding Proteins in Obesity/Diabetes Syndromes

描述（由申请人提供）：在美国，每三个人中就有一个被认为非常超重或肥胖。肥胖不仅仅是一个美容问题，它还与心血管疾病、高血压、动脉粥样硬化和糖尿病呈正相关。瘦素作为一种脂肪细胞源性分泌细胞因子的发现，使我们对脂肪细胞的认识超越了作为一个热量过剩的储存库，而成为一种动态的细胞类型，在评估和整合整体能量稳态中发挥作用。实验室的重点是肥胖、脂肪酸代谢和脂解之间的联系，重点是脂肪细胞中的脂肪酸运输和NIDDM。 该实验室研究脂肪细胞的脂肪酸载体，特别是FABP 4和FABP 5基因的产物。在脂肪细胞中，这些蛋白质与脂肪酸和其他疏水性单酰基脂质形成1：1非共价复合物。aP 2（FABP 4）和KLBP（FABP 5）各自与脂肪酶敏感性脂肪酶（HSL）形成物理复合物，HSL是将三酰甘油水解为脂肪酸的脂解的关键限速酶。与aP 2与HSL的关联一致，FABP 4缺失小鼠表现出降低的基础和脂肪酶刺激的脂解，表明脂肪酸结合蛋白通过从脂肪酶反应中去除终产物脂肪酸并促进其从细胞流出来影响该过程。 FABP 4基因敲除小鼠表现出对肥胖相关的胰岛素抵抗的保护，而不降低脂肪量或瘦素水平。双FABP 4-FABP 5缺失小鼠具有较低的脂肪量和瘦素水平。相反，在aP 2启动子下在脂肪细胞中过表达KLBP的转基因小鼠具有升高的脂肪细胞质量，并且比野生型动物更具胰岛素抗性。我们假设，FABPs刺激脂解通过与HSL的相互作用和相互作用的破坏结果在定量差异的分子种类和丰富的脂肪酸从脂肪细胞脂解。脂解衍生的脂肪酸的这种变化影响与胰岛素抵抗的发展相关的脂肪酸的可用性。具体目的A：从携带破坏的FABP基因或aP 2-KLBP转基因的低脂肪和高脂肪饲养的小鼠体内和原位评价脂解。在这些动物模型中，将脂解的速率和程度与胰岛素抵抗相关联。具体目的B：在体外检查FABP与HSL的物理相互作用。评估FABP在调节HSL活性中的作用，并绘制两种蛋白质的相互作用结构域。具体目的C：使用荧光共振能量转移评估293细胞和3 T3-L1脂肪细胞中FABP与HSL的关联。检查HSL的磷酸化/易位及其与FABP缔合的关系。

DESCRIPTION (provided by applicant): In the United States, one in every three individuals is considered very overweight or obese. More than a cosmetic issue, obesity is a positive correlate to cardiovascular disease, hypertension, atherosclerosis and diabetes. The discovery of leptin as an adipocyte-derived secreted cytokine has expanded our view of the fat cell beyond being a repository for caloric excess to a dynamic cell type functioning in the assessment and integration of overall energy homeostasis. The focus of the laboratory is on the linkage between obesity, fatty acid metabolism and lipolysis with an emphasis on fatty acid trafficking in the adipocyte and NIDDM. The laboratory studies fatty acid carriers of the adipocyte, specifically the products of the FABP4 and FABP5 genes. In fat cells, these proteins form 1:1 non-covalent complexes with fatty acids and other hydrophobic monoacyl lipids. aP2 (FABP4) and KLBP (FABP5) each form a physical complex with the hormone-sensitive lipase (HSL), the key rate-limiting enzyme of lipolysis that hydrolyzes triacylglycerol to fatty acids. Consistent with the association of aP2 with the HSL, FABP4 null mice exhibit reduced basal and hormone-stimulated lipolysis suggesting fatty acid binding proteins affect the process by removing end-product fatty acids from the lipase reaction and facilitating their efflux from the cell. FABP4 null mice exhibit protection from obesity-linked insulin resistance without reduction in fat mass or leptin levels. Double FABP4-FABP5 null mice have lower fat mass and leptin levels. Conversely, transgenic mice that overexpress KLBP in fat cells under the aP2 promoter, have elevated fat cell mass and are more insulin resistant than are wild type animals. We hypothesize that FABPs stimulate lipolysis via interaction with HSL and that the disruption of the interaction results in a quantitative difference in both the molecular species and abundance of fatty acids that are lipolyzed from the fat cell. Such changes in lipolysis-derived fatty acids affect the availability of fatty acids linked to the development of insulin resistance. To test this hypothesis we propose to: Specific Aim A: Evaluate lipolysis in vivo and in situ from low and high fat fed mice harboring either disrupted FABP genes or the aP2-KLBP transgene. Correlate the rate and extent of lipolysis to insulin resistance in such animal models. Specific Aim B: Examine the physical interaction of FABPs with HSL in vitro. Assess the role of FABPs in regulating the activity of HSL and map the interaction domains of the two proteins. Specific Aim C: Assess the association of FABPs with HSL in 293 cells and 3T3-L1 adipocytes using fluorescence resonance energy transfer. Examine the phosphorylation/translocation of HSL and its relationship to FABP association.