Molecular Identity of the Cardiac Mitochondrial Pore

描述（由申请人提供）：线粒体功能障碍是心肌细胞死亡的潜在原因，因此在许多心脏病理的发展中起着关键作用。这种线粒体功能障碍通常是由线粒体通透性转换（MPT）孔的打开介导的，这导致线粒体内膜通透性迅速增加。这反过来导致ATP消耗，活性氧产生，线粒体肿胀和破裂。因此，我们的长期目标是确定构成MPT孔的蛋白质，并了解这种复合物被调节的分子机制。MPT孔最初被提出由外膜上的电压依赖性阴离子通道（VDAC）、内膜上的腺嘌呤核苷酸转位酶（ANT）和基质上的调节蛋白亲环蛋白d （CypD）组成。然而，遗传学研究表明VDAC和ANT对于MPT是不可缺少的。因此CypD仍然是MPT孔复合物的唯一真正成员。重要的是，直接调节CypD及其促mpt功能的关键因素仍有待阐明。特别是，抑制CypD功能的CypD结合蛋白尚未被确定

DESCRIPTION (provided by applicant): Mitochondrial dysfunction is an underlying cause of cardiomyocyte death and therefore plays a critical role in the development of many cardiac pathologies. This mitochondrial dysfunction is often mediated by the opening of the mitochondrial permeability transition (MPT) pore, which causes a rapid increase in inner mitochondrial membrane permeability. This in turn leads to ATP depletion, reactive oxygen species production, mitochondrial swelling and rupture. Consequently, our long-range goal is to identify the proteins that make up the MPT pore and understand the molecular mechanisms by which this complex is regulated. The MPT pore was originally proposed to consist of the voltage-dependent anion channel (VDAC) in the outer membrane, the adenine nucleotide translocase (ANT) in the inner membrane, plus a regulatory protein cyclophilin-D (CypD) in the matrix. However, genetic studies have revealed that VDAC and ANT are dispensable for MPT. Thus CypD still remains as the only bona fide member of the MPT pore complex. Importantly, the key factors that directly regulate CypD and its pro-MPT function remain to be elucidated. In particular, CypD-binding proteins that act to inhibit CypD's function have yet to be identified and the additional role that CypD phosphorylation plays in regulating MPT and cell death has not been comprehensively addressed. We have identified the mitochondrial matrix protein C1qbp as a novel CypD-binding protein and our strong preliminary data indicate that it can inhibit MPT and cell death. Conversely, we have found that the pro-death kinase GSK3 can sensitize cells to MPT and death, and that these effects are associated with CypD phosphorylation. Consequently, our central hypothesis is that C1qbp inhibits, whereas GSK3 promotes, MPT and cell death through the direct regulation of CypD. The objective of the present application is to utilize genetic, biochemical, physiological, and pharmacological techniques to systematically evaluate the roles of a CypD inhibitor (C1qbp) and a CypD activator (GSK3) in MPT, cardiac cell death, and the progression of myocardial disease. In Aim 1 we will determine the functional involvement of C1qbp in the MPT response and cardiac myocyte death. In Aim 2 we will evaluate the effects of GSK3-dependent CypD phosphorylation on MPT and cardiac cell death. The rationale for the proposed research is that once key mitochondrial proteins that regulate MPT and mitochondrial function are identified, they can be targeted as a means of treating a whole array of human cardiac diseases.