Analysis of Fis1 in Mitophagy in Mammals

 描述（由申请人提供）：由于线粒体在细胞代谢和其他功能中具有重要作用，因此维持细胞内线粒体群体的质量至关重要。线粒体自噬（Mitophagy）是通过自噬降解线粒体，被认为是剔除功能障碍或受损线粒体的主要途径。研究得最多的线粒体自噬途径涉及Pink 1和Parkin，但关于这一途径的生理重要性仍存在疑问。Pink 1和Parkin基因敲除小鼠仅显示基础线粒体功能的细微缺陷，与线粒体自噬的广泛缺陷不一致。这一观察结果表明，有替代或多余的途径线粒体自噬。我们的初步数据表明，Fis 1，一种以前被认为参与线粒体分裂的蛋白质，是线粒体自噬的主要参与者，包括Pink 1/Parkin独立的线粒体自噬。此外，缺乏Fis 1的小鼠具有严重的生理缺陷。因此，对Fis 1突变小鼠的研究可能揭示线粒体自噬的生理功能。本研究项目的长期目标是了解Fis 1依赖的线粒体自噬的分子机制以及线粒体自噬在小鼠发育中的作用。在目的1中，我们开发了一种功能分析来剖析哺乳动物线粒体单亲遗传的分子机制。使用这个系统，我们将确定参与消除早期小鼠胚胎中的父系线粒体的分子。在目标2中，我们开发了一种细胞系统，其中增加氧化磷酸化刺激线粒体自噬。我们将使用这个细胞系统来了解线粒体自噬的分子机制，包括Fis 1的作用，泛素化的作用，以及自噬适配器p62的招募。在目标3中，我们利用Fis 1基因敲除小鼠模型来了解线粒体自噬的生理功能。我们将研究Fis 1在男性生殖细胞成熟中的作用，Fis 1的缺失导致线粒体和过氧化物酶体密度增加。两者合计，这些目标将导致深入了解的Fis 1在体内的作用和线粒体自噬在哺乳动物生理学中的作用。

 DESCRIPTION (provided by applicant): Because mitochondria have vital roles in cell metabolism and other functions, it is critical to maintain the quality of the mitochondrial population within cells. Mitophagy, the degradation of mitochondria via autophagy, is thought to be the major pathway for culling dysfunctional or damaged mitochondria. The best- studied mitophagy pathway involves Pink1 and Parkin, but questions remain about the physiological importance of this pathway. Mouse knockouts of Pink1 and Parkin show only subtle defects in basal mitochondrial function, inconsistent with a widespread defect in mitophagy. This observation suggests that there are alternative or redundant pathways for mitophagy. Our preliminary data indicate that Fis1, a protein previously thought to be involved in mitochondrial fission, is a major player in mitophagy, including mitophagy that is Pink1/Parkin-independent. Moreover, mice lacking Fis1 have severe physiological defects. Therefore, the study of Fis1 mutant mice may reveal the physiological functions of mitophagy. The long-term goals of this research project are to understand the molecular mechanism of Fis1-dependent mitophagy and the role of mitophagy in mouse development. In Aim 1, we develop a functional assay to dissect the molecular mechanism underlying uniparental inheritance of mitochondria in mammals. Using this system, we will identify molecules involved in elimination of paternal mitochondria in the early mouse embryo. In Aim 2, we develop a cellular system in which increased oxidative phosphorylation stimulates mitophagy. We will use this cellular system to understand the molecular mechanism of mitophagy, including the role of Fis1, the role of ubiquitination, and the recruitment of the autophagy adaptor p62. In Aim 3, we utilize a Fis1 knockout mouse model to understand the physiological functions of mitophagy. We will study the role of Fis1 in male germ cell maturation, where loss of Fis1 results in increased mitochondrial and peroxisomal density. Taken together, these Aims will lead to a deep understanding of the in vivo roles of Fis1 and the role of mitophagy in mammalian physiology.