Mechanism of action of steroidogenic acute regulatory protein (StAR)

描述（由申请人提供）：类固醇生成的限速步骤是胆固醇从线粒体外膜（OMM）移动到线粒体内膜（IMM）。甾体生成急性调节蛋白（StAR）通过一种未知的机制促进胆固醇从OMM到IMM的运动，从而调节底物流向P450scc进行甾体生成。StAR属于一个名为START （StAR相关脂质转运结构域）的蛋白家族，其中StAR是主要成员。StAR突变会导致一种潜在的致命疾病，称为先天性肾上腺增生症，在这种疾病中，所有类固醇激素的合成都会受损，胎儿在出生后不久就会死亡。StAR的活动与其在OMM的驻留时间成正比。我们的初步结果表明，i) StAR的进口机制是独特的，ii) StAR暂停序列在StAR折叠和活性中具有独特的作用，iii) StAR在一小时内运输100多个胆固醇分子，iv) StAR通过几种蛋白质的复合物进口，v) StAR的进口分两步进行。为了了解StAR加工和进入线粒体的机制，我们提出了三个具体目标。目的一是确定新合成的StAR的活性机制。这对于理解StAR如何以如此短的半衰期发挥作用，以及它如何在没有n端序列的情况下到达OMM至关重要。在目标2中，我们提出确定StAR作用的线粒体位点，以了解OMM驻留蛋白VDAC 1和2（电压依赖阴离子通道）对StAR的易位机制。在目标3中，我们提出通过OMM相关脂质膜确定StAR的展开机制，并揭示为什么特定的START-domain比其他START-domain更活跃。实现这些目标将：1)帮助我们扩展我们对StAR与OMM相互作用机制的认识；2)扩展我们对OMM在StAR展开和胆固醇运输动员中所起作用的理解。完成这些目标将有助于我们了解细胞脂质代谢的广泛方面，以及广泛的出生缺陷，胚胎发育障碍和遗传疾病。公共卫生相关性：这项拨款申请的目的是通过研究类固醇急性调节蛋白（StAR）的进口机制来确定胆固醇转运到线粒体的机制。StAR需要在线粒体外膜展开，因此拟议的工作将在其输入之前确定StAR与线粒体外居住蛋白电压依赖阴离子通道（VDAC）和脂质的相互作用。揭示StAR的输入机制将有助于我们了解细胞脂质代谢的广泛方面，因此，该项目的重要性远远超出了StAR和类固醇生成的调节。

DESCRIPTION (provided by applicant): The rate-limiting step in steroidogenesis is the movement of cholesterol from the outer mitochondrial membrane (OMM) to inner mitochondrial membrane (IMM). The steroidogenic acute regulatory protein (StAR) by an unknown mechanism facilitates the movement of cholesterol from the OMM to IMM, thus regulating the flow of substrate to P450scc for steroidogenesis. StAR belongs to a family of proteins called START (StAR related lipid transport domain), where StAR is the primary member. StAR mutations cause a potentially lethal disease called congenital adrenal hyperplasia, in which the synthesis of all steroid hormones is impaired, and the fetuses die shortly after birth. StAR activity is directly proportional to its residency time at the OMM. Our preliminary results show that i) the import mechanism of StAR is unique, ii) the StAR pause sequence has a distinct role in StAR folding and activity, iii) StAR transports more than 100 molecules of cholesterol in one hour, iv) StAR is imported through a complex of several proteins, and v) StAR is imported in a two step process. To understand the mechanism of StAR processing and import into mitochondria, we propose three specific aims. Aim 1 is to determine the mechanism of activity of newly synthesized StAR. This is critical in understanding how StAR functions with such a short half- life, and how it can reach the OMM with out the N-terminal sequence. In aim 2, we proposed to determine the mitochondrial site of StAR action, in order to understand the translocation mechanism of StAR by OMM resident proteins VDAC 1 and 2 (Voltage Dependent Anion Channel). In aim 3, we proposed to determine the unfolding mechanism of StAR by the OMM associated lipid membrane, and to uncover why a specific START-domain is more active than the other. Fulfilling these aims will, I) help us to expand our knowledge about the mechanism of interaction of StAR with the OMM and II) expand our understanding about the role of OMM that help in StAR unfolding and thus mobilization of cholesterol transport. Completing these aims will help us in the understanding of broad aspects of cellular lipid metabolism and also to a broad array of birth defects, disorders of embryogenesis and genetic diseases. PUBLIC HEALTH RELEVANCE: The narrative of this grant application is to determine the mechanism of cholesterol transport into mitochondria by studying the mechanism of import of steroidogenic acute regulatory protein (StAR). StAR requires unfolding at the outer mitochondrial membrane, so the proposed work will identify the interaction of StAR with outer mitochondrial resident protein voltage dependent anion channel (VDAC) and lipids prior to its import. Revealing the StAR import mechanism will help us in the understanding of broad aspects of cellular lipid metabolism and thus, the importance of this project extends far beyond StAR and regulation of steroidogenesis.