Nix Is Critical to Two Distinct Phases of Mitophagy, Reactive Oxygen Species-mediated Autophagy Induction and Parkin-Ubiquitin-p62-mediated Mitochondrial Priming
Nix Is Critical to Two Distinct Phases of Mitophagy, Reactive Oxygen Species-mediated Autophagy Induction and Parkin-Ubiquitin-p62-mediated Mitochondrial Priming
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DOI：
10.1074/jbc.m110.119537
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发表时间：
2010-09-03
影响因子：
4.8
通讯作者：
Yin, Xiao-Ming
中科院分区：
文献类型：
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作者：
Ding, Wen-Xing;Ni, Hong-Min;Yin, Xiao-Ming
受损的线粒体可以通过自噬（即线粒体自噬）消除，这对细胞稳态和细胞存活很重要。尽管事实上已经发现许多因素对哺乳动物细胞中的线粒体自噬很重要，但它们在该过程中的单独作用尚未明确定义。帕金蛋白是一种泛素-蛋白质异肽连接酶，能够转移到要去除的线粒体。我们在这里表明，在一个化学缺氧模型的线粒体自噬诱导的解偶联剂，羰基氰间氯苯腙（CCCP），帕金易位导致线粒体泛素化和p62招聘的线粒体。小抑制性RNA介导的p62敲除显着减少了自噬机制对线粒体的识别和随后的消除。因此，帕金蛋白、泛素和p62在线粒体自噬的准备中起作用，这里称为线粒体启动。然而，这些分子并不是诱导自噬机制所必需的。Parkin和p62似乎都不影响CCCP诱导的自噬。相反，我们发现Nix是自噬诱导所必需的。Nix促进CCCP诱导的线粒体去极化和活性氧的产生，从而抑制mTOR信号传导并激活自噬。Nix还通过控制Parkin的线粒体易位来促进线粒体启动，尽管活性氧的产生不参与这一步骤。C-末端膜靶向序列的缺失而不是BH 3结构域中的突变使Nix丧失了这些功能。因此，我们的工作区分了负责线粒体自噬不同阶段的分子事件，并将Nix置于事件的上游。
Damaged mitochondria can be eliminated by autophagy, i.e. mitophagy, which is important for cellular homeostasis and cell survival. Despite the fact that a number of factors have been found to be important for mitophagy in mammalian cells, their individual roles in the process had not been clearly defined. Parkin is a ubiquitin-protein isopeptide ligase able to translocate to the mitochondria that are to be removed. We showed here in a chemical hypoxia model of mitophagy induced by an uncoupler, carbonyl cyanide m-chlorophenylhydrazone (CCCP) that Parkin translocation resulted in mitochondrial ubiquitination and p62 recruitment to the mitochondria. Small inhibitory RNA-mediated knockdown of p62 significantly diminished mitochondrial recognition by the autophagy machinery and the subsequent elimination. Thus Parkin, ubiquitin, and p62 function in preparing mitochondria for mitophagy, here referred to as mitochondrial priming. However, these molecules were not required for the induction of autophagy machinery. Neither Parkin nor p62 seemed to affect autophagy induction by CCCP. Instead, we found that Nix was required for the autophagy induction. Nix promoted CCCP-induced mitochondrial depolarization and reactive oxygen species generation, which inhibited mTOR signaling and activated autophagy. Nix also contributed to mitochondrial priming by controlling the mitochondrial translocation of Parkin, although reactive oxygen species generation was not involved in this step. Deletion of the C-terminal membrane targeting sequence but not mutations in the BH3 domain disabled Nix for these functions. Our work thus distinguished the molecular events responsible for the different phases of mitophagy and placed Nix upstream of the events.