Role of Mitophagy in Mitochondrial Protein Quality Control

描述(申请人提供)：线粒体功能障碍经常发生在老年人身上，是帕金森氏症、阿尔茨海默氏症、癌症和糖尿病等疾病的标志。生物体具有应对线粒体功能障碍的机制，包括PINK1-Parkin途径，它通过自噬(有丝分裂)促进溶酶体对线粒体蛋白的周转，以响应线粒体功能障碍。然而，在理解这一途径如何发挥作用以促进线粒体完整性和预防疾病形成方面存在着根本的差距。这项应用的目的是利用酿酒酵母确定损伤诱导的有丝分裂吞噬促进线粒体质量控制的机制。以前的研究发现酵母中存在一个吞丝分裂途径，但该途径与PINK1-Parkin途径不是同源的，并且对线粒体功能障碍没有反应，这限制了它作为模型系统的实用性。申请人最近在酵母中发现了第二条与PINK1-Parkin途径功能相同的有丝分裂途径，并促进了有丝分裂途径，以应对因溶酶体样空泡酸性丧失而导致的AGE诱导的线粒体功能障碍。申请者将使用这一新的途径来测试这一申请的中心假设，即有丝分裂选择性地改变线粒体蛋白质组，以维持线粒体的完整性，以应对线粒体功能障碍。了解酵母中有丝分裂如何预防线粒体功能障碍将迅速推进我们对人类这一过程的理解，并促进与线粒体功能障碍相关疾病的治疗发展。这项应用的目标将通过追求以下三个具体目标来实现：目标1：确定空泡酸度降低如何导致线粒体功能障碍。抑制子筛选和营养调节将被用来描述在没有空泡酸性的情况下导致线粒体功能障碍的途径。目的2：确定线粒体蛋白是如何被有丝分裂降解的。这一目标将使用基于显微镜的有丝分裂分析和质谱学来进一步表征损伤诱导的有丝分裂吞噬途径，并识别其功能所需的新基因。目的3：确定线粒体的哪些成分被有丝分裂降解。这一目标将使用基于显微镜的技术来识别因有丝分裂而降解的线粒体蛋白，并确定有丝分裂是否针对先前存在的或新合成的蛋白质。在这项申请中提出的研究是创新的，因为它利用发芽酵母中先前未知的PINK1-Parkin通路的功能等价物剖析了有丝分裂在线粒体质量控制中的作用，从而将酵母遗传学的力量带入了损伤诱导的有丝分裂领域。这一点意义重大，因为它将提供对线粒体蛋白质质量控制途径的深入分子理解，该途径可以保护细胞免受年龄诱导的线粒体功能障碍的影响，并防止帕金森氏症的发展。

DESCRIPTION (provided by applicant): Mitochondrial dysfunction frequently occurs in aged individuals and is a hallmark of diseases such as Parkinson's, Alzheimer's, cancer, and diabetes. Organisms are equipped with mechanisms to cope with mitochondrial dysfunction, including the PINK1-Parkin pathway that promotes lysosomal turnover of mitochondrial proteins by autophagy (mitophagy) in response to mitochondrial dysfunction. However, there is a fundamental gap in understanding how this pathway functions to promote mitochondrial integrity and prevent disease formation. The objective of this application is to determine the mechanism through which damage- induced mitophagy promotes mitochondrial quality control using S. cerevisiae. Previous studies identified a mitophagy pathway in yeast, but this pathway is not homologous to the PINK1-Parkin pathway and does not respond to mitochondrial dysfunction, which limits its usefulness as a model system. The applicant recently discovered a second mitophagy pathway in yeast that is functionally equivalent to the PINK1-Parkin pathway and promotes mitophagy in response to age-induced mitochondrial dysfunction caused by loss of lysosome- like vacuolar acidity. The applicant will use this novel pathway to test the central hypothesis of this application that mitophagy selectively alters the mitochondrial proteome to maintain mitochondrial integrity in response to mitochondrial dysfunction. Understanding how mitophagy prevents mitochondrial dysfunction in yeast will rapidly advance our understanding of this process in humans and facilitate the development of treatments for diseases associated with mitochondrial dysfunction. The objective of this application will be accomplished by pursuing the following three specific aims: Aim 1: Determine how reduced vacuolar acidity causes mitochondrial dysfunction. Suppressor screens and nutrient modulation will be used to characterize pathways that contribute to mitochondrial dysfunction in the absence of vacuolar acidity. Aim 2: Determine how mitochondrial proteins are degraded by mitophagy. This aim will use a microscopy-based mitophagy assay and mass spectrometry to further characterize the damage-induced mitophagy pathway and identify novel genes required for its function. Aim 3: Identify what components of the mitochondria are degraded by mitophagy. This aim will use microscopy-based techniques to identify mitochondrial proteins degraded by mitophagy and determine if mitophagy targets preexisting or newly synthesized proteins. The research proposed in this application is innovative because it dissects the role of mitophagy in mitochondrial quality control using a previously unknown functional equivalent of the PINK1-Parkin pathway in budding yeast and thus brings the power of yeast genetics to the damage-induced mitophagy field. This is significant because it will provide a deep molecular understanding of a mitochondrial protein quality control pathway that protects cells against age-induced mitochondrial dysfunction and prevents the development of Parkinson's disease.