Molecular Identity of the Cardiac Mitochondrial Pore

项目摘要 线粒体功能障碍是心肌细胞死亡的根本原因，因此在心肌细胞死亡中起关键作用。 许多心脏病的发展。这种线粒体功能障碍通常是由开放的 线粒体渗透性转换（MPT）孔，这导致线粒体内部的快速增加， 膜透性这反过来又导致ATP耗竭，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 并了解调控这种复合物的分子机制。MPT孔最初是 提出由外膜中的电压依赖性阴离子通道（VDAC）、腺嘌呤 内膜中的核苷酸移位酶（ANT），加上内膜中的调节蛋白亲环素-D（CypD）， 矩阵然而，遗传学研究表明VDAC和ANT与MPT相关。因此CypD仍然 仍然是MPT孔复合物的唯一真正成员。重要的是，直接影响 调节CypD及其pro-MPT功能仍有待阐明。特别是CypD结合蛋白， 抑制CypD的功能还有待鉴定，CypD磷酸化在 调节MPT和细胞死亡的方法尚未全面解决。我们已经确定了线粒体 基质蛋白C1 qbp是一种新的CypD结合蛋白，我们的初步数据表明，它可以 抑制MPT和细胞死亡。相反，我们发现促死亡激酶GSK 3可以使细胞敏感， MPT和死亡，这些作用与CypD磷酸化有关。因此，我们的中央 假设C1 qbp抑制MPT和细胞死亡，而GSK 3促进MPT和细胞死亡是通过直接的 CypD的调节本申请的目的是利用遗传、生物化学、生理学 和药理学技术来系统地评估CypD抑制剂（C1 qbp）和CypD 活化剂（GSK 3）在MPT、心肌细胞死亡和心肌疾病进展中的作用。在目标1中， 确定C1 qbp在MPT反应和心肌细胞死亡中的功能参与。在目标2中， 将评估GSK 3依赖性CypD磷酸化对MPT和心脏细胞死亡的影响。的 这项研究的基本原理是，曾经调节MPT的关键线粒体蛋白质， 线粒体功能的鉴定，它们可以作为治疗一系列人类疾病的手段。 心脏病

PROJECT SUMMARY 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.