The Dual Role of Pro-apoptotic BAD in Insulin Secretion and Beta Cell Survival

描述（由申请人提供）：越来越多的证据表明，线粒体生理学在葡萄糖刺激的胰岛素释放和β细胞的存活中都很重要。这些不断发展的文献提出了一个重要的问题：线粒体如何作为胰岛素分泌和生存的偶联剂发挥作用？在初步研究中，我们发现促凋亡蛋白BAD是细胞死亡调节因子BCL-2家族的一员，它存在于线粒体连接的含葡萄糖激酶复合物中，该复合物调节葡萄糖驱动的线粒体呼吸。此外，我们还发现Bad-null小鼠在葡萄糖刺激的胰岛素分泌和葡萄糖氧化代谢方面存在缺陷。本申请中提出的研究目的是检验BAD在β细胞中起双重作用的假设；它通过对有氧代谢的影响来调节胰岛素分泌，并通过参与其他BCL-2家族成员来调节细胞凋亡。这一假设产生了可测试的预测，我们建议使用遗传学和化学来解决。有两个特定的目的：Aim 1测试BAD在胰岛素分泌和细胞凋亡中的两种不同功能是通过选择性结构基序和/或翻译后修饰来指定的。这一预测将使用突变分析结合基因重构分析进行检验。这些研究将与线粒体功能的检查相结合，以确定BAD在胰岛素释放中的作用的代谢决定因素。作为这种结构-功能分析的补充方法，我们将采用新的细胞渗透性BAD肽模拟化合物（BAD SAHBs）作为化学工具，进一步剖析BAD在β细胞中葡萄糖代谢和胰岛素分泌中的作用。目的2验证BAD的代谢和凋亡功能与糖尿病细胞功能障碍的生理相关性。我们将使用几种现有的Bad遗传模型，包括敲除小鼠和敲入动物，在两种明确的2型糖尿病模型中，即高脂肪饮食诱导的糖尿病模型和瘦素不敏感的db/db遗传模型中，基因测试Bad对细胞功能和存活的贡献。在这个提议中描述的研究可能导致确定一个新的治疗靶点糖尿病。

DESCRIPTION (provided by applicant): A growing body of evidence indicates that mitochondrial physiology is important in both glucose-stimulated insulin release and survival of beta-cells. This evolving body of literature raises an important question: How do mitochondria function as coupling agents for insulin secretion and survival? In preliminary studies, we have shown that the pro-apoptotic protein BAD, a member of the BCL-2 family of cell death regulators, resides in a mitochondria-tethered glucokinase-containing complex that regulates glucose-driven mitochondrial respiration. We have shown moreover that Bad-null mice display defect in glucose-stimulated insulin secretion and oxidative metabolism of glucose. The objective of the studies proposed in this application is to test the hypothesis that BAD plays a dual role in beta-cells; it regulates insulin secretion through its effects on aerobic metabolism and it modulates apoptosis through engaging other BCL-2 family members. This hypothesis gives rise to testable predictions that we propose to address using both genetics and chemistry. There are two specific aims: Aim 1 tests the prediction that, the two distinctive functions of BAD in insulin secretion and apoptosis are specified by selective structural motifs and/or post translational modifications. This prediction will be tested using mutational analysis combined with genetic reconstitution assays. These studies will be integrated with examination of mitochondrial function to identify the metabolic determinants of BAD's role in insulin release. As a complementary approach to this structure-function analysis, we will employ novel cell permeable BAD peptido-mimetic compounds (BAD SAHBs) as chemical tools to further dissect the role of BAD in glucose metabolism and insulin secretion in beta-cells. Aim 2 tests the prediction that the metabolic and apoptotic functions of BAD are physiologically relevant to ¿-cell dysfunction in diabetes. We will use several available Bad genetic models, including knockout mice and knockin animals, to genetically test the contribution of BAD to ¿-cell function and survival in two well defined models of type 2 diabetes, namely the high fat diet-induced model of diabetes and the leptin insensitive db/db genetic model. The studies described in this proposal may lead to identification of a new therapeutic target in diabetes.