Removal of damaged mitochondria by alternative autophagy

通过替代自噬去除受损的线粒体

• 批准号: 9978602
• 负责人: Junichi Sadoshima
• 金额: $ 60.1万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-15 至 2021-07-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9978602
• 关键词: Address; Affect; Autophagocytosis; Autophagosome; Biogenesis; Cardiac; Cardiac Myocytes; Cell Death; Detection; Development; Electron Microscopy; Excision; Fasting; Goals; Golgi Apparatus; Heart; Heart failure; Homeostasis; Injury; Intervention; Ischemia; Knock-in Mouse; Knockout Mice; Knowledge; Lead; Maintenance; Mediating; Membrane; Metabolism; Mitochondria; Molecular; Myocardial Ischemia; Myocardial dysfunction; Organelles; Pathologic; Patients; Phosphorylation; Play; Production; Quality Control; RIPK1 gene; Reactive Oxygen Species; Reporter; Reporting; Role; Starvation; Stimulus; Stress; Testing; Vesicle; cardioprotection; cell type; heart cell; heart function; ischemic injury; loss of function; mitochondrial autophagy; mitochondrial dysfunction; mouse model; mutant; novel; novel strategies; parkin gene/protein; recruit; release factor; response; scaffold

Autophagy is a major mechanism of degradation of damaged mitochondria. Without elimination of damaged mitochondria, depolarized mitochondria and reactive oxygen species (ROS) rapidly affect healthy mitochondria, leading to wide-spread mitochondrial dysfunction and cell death. Understanding how damaged mitochondria are removed will provide a key to achieving healthier mitochondria in cardiomyocytes (CMs) and developing novel treatments for heart failure. Although it is believed that damaged mitochondria are degraded primarily by Pink1-Parkin-mediated mitophagy, we have discovered that CMs degrade mitochondria through an Atg7-independent and Ulk1-dependent form of autophagy that is homologous to the “alternative” autophagy previously reported by Nishida, and that this form of autophagy plays a significant role in the elimination of damaged mitochondria in response to starvation. However, neither the molecular mechanism nor the functional significance of mitophagy mediated through alternative autophagy has been clearly established in CMs yet. Thus, the goal of this project is to demonstrate the functional significance of alternative autophagy in eliminating damaged mitochondria in the heart in response to relevant stresses and to elucidate the underlying molecular mechanisms. Hypothesis 1: In response to myocardial ischemia, the heart activates Atg7- independent/Ulk1-dependent alternative autophagy, which plays an essential role in mediating the clearance of damaged mitochondria and protects the heart from myocardial ischemia. Hypothesis 2: Ulk1 phosphorylated at Ser555 acts as a scaffold to induce Rab9 interaction for autophagosome formation and phosphorylation of Drp1 at Ser616 for mitochondrial fission, both of which are important in mediating mitochondrial autophagy in response to myocardial ischemia. We will: Aim 1: Demonstrate that the atg7-independent and ulk1-dependent alternative autophagy is activated by myocardial ischemia. Evaluate whether alternative autophagy protects the heart during myocardial ischemia. To this end, we will use cardiac-specific atg7- and ulk1-knockout mice, electron microscopy, specific reporters of alternative autophagy and mitochondrial autophagy, and functional analyses of mitochondria. We will show that damaged mitochondria are degraded primarily through alternative autophagy during myocardial ischemia. Aim 2: Evaluate whether phosphorylation of Ulk1 at Ser555 plays an essential role in mediating alternative autophagy and cardioprotection during myocardial ischemia by stimulating interaction with Rab9 and Ser616-phosphorylated Drp1. To this end, we will use loss-of-function and knock-in mouse models and unique and reliable reporters for alternative autophagy and lysosomal degradation of mitochondria. The knowledge obtained from this aim should lead to development of specific interventions to modulate mitophagy during myocardial ischemia. In summary, our study will demonstrate that alternative autophagy is a novel and predominant mechanism of mitochondrial degradation, which is essential for the maintenance of mitochondrial quality in the heart during ischemia.

自噬是线粒体受损的降解的主要机制。没有消除损坏 线粒体，线粒体加深和活性氧（ROS）迅速影响健康 线粒体，导致广泛的线粒体功能障碍和细胞死亡。了解损坏程度 切除线粒体将为实现心肌细胞（CMS）中更健康的线粒体提供关键 为心力衰竭开发新的治疗方法。尽管据信损坏的线粒体会降解 主要是通过Pink1-Parkin介导的线粒体，我们发现CMS通过一种降解线粒体 与“替代”自噬同源的自噬的无关和ULK1依赖性形式 Nishida先前报道的，这种形式的自噬在消除中起着重要作用 响应饥饿的线粒体受损。但是，分子机制和功能均不 通过替代自噬介导的线粒体的重要性已经在CMS中清楚地确定。 这是该项目的目的是证明替代自噬的功能意义 消除对相关应力的响应并阐明潜在的，消除心脏中的线粒体受损 分子机制。假设1：针对心肌缺血，心脏激活ATG7- 独立/ULK1依赖性替代自噬，在介导清除率中起着至关重要的作用 线粒体受损，并保护心脏免受心肌缺血。假设2：ULK1磷酸化 在SER555充当诱导自噬体形成和磷酸化的Rab9相互作用的支架 Ser616的DRP1用于线粒体裂变，这两者在介导线粒体自噬中很重要 对心肌缺血的反应。我们将：目标1：证明与ATG7无关和ULK1依赖性 替代自噬被心肌缺血激活。评估替代自噬是否保护 心肌缺血期间的心脏。为此，我们将使用心脏特异性的ATG7-和ULK1敲除小鼠， 电子显微镜，替代自噬和线粒体自噬的特定报道以及功能 线粒体分析。我们将证明损坏的线粒体主要通过替代性降解 心肌缺血期间的自噬。目标2：评估Ser55上ULK1的磷酸化是否播放 通过在心肌缺血期间介导替代自噬和心脏保护的重要作用 刺激与Rab9和Ser616磷酸化DRP1的相互作用。为此，我们将使用功能丧失 以及型鼠标模型以及替代自噬和溶酶体的独特可靠记者 线粒体的降解。从这个目标获得的知识应导致特定的发展 在心肌缺血期间调节线粒体的干预措施。总而言之，我们的研究将证明 替代自噬是一种新颖而主要的线粒体降解机制，这是必不可少的 为了维持缺血期间心脏中心的线粒体质量。