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Molecular Control of Cardiomyocyte Mitophagy by the RhoGAP GRAF1

RhoGAP GRAF1 对心肌细胞线粒体自噬的分子控制

基本信息

批准号：
10521295
负责人：
Joan M Taylor
金额：
$ 43.29万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-12-15 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10521295
关键词：
Actins Affect Apoptosis Autophagocytosis Autophagosome Award Binding Biogenesis Cardiac Cardiac Myocytes Cardiopulmonary Cells Cessation of life Coupled Data Defect Disease Exhibits Extravasation Foundations Generations Genetic Genomics Heart Heart Diseases Homeostasis Impairment Injury Ischemia Knockout Mice Learning Link Lipid Binding Lipids Mediating Membrane Metabolic Metabolism Mitochondria Modeling Molecular Mus Muscle Cells Myocardial Infarction Myocardial Ischemia Myocardium Necrosis Operative Surgical Procedures Organelles Outcome Outer Mitochondrial Membrane Oxidative Stress Oxygen PH Domain Parkin Pathologic Pathway interactions Patients Phospholipids Phosphorylation Phosphotransferases Play Process Production Protein-Serine-Threonine Kinases Proteins Quality Control Reperfusion Therapy Research Role SH3 Domains Stress Structure Time cardiogenesis cardioprotection depolymerization experimental study fatty acid oxidation fetal fluorophore gain of function heart cell loss of function metabolomics mitochondrial dysfunction mitochondrial membrane novel novel strategies novel therapeutics oxidative damage postnatal rational design receptor receptor binding recruit response rho GTPase-activating protein ubiquitin-protein ligase

项目摘要

Summary Because mitochondrial dysfunction affects ATP production and promotes oxidative damage, these organelles are turned over every 2-3 weeks in healthy cardiomyocytes by a process that involves autophagy. Defects in mitochondrial quality control also enhance the progression of cardiac disease making it critical to identify the mechanisms that regulate this process. Several major pathways have been implicated that involve binding of the cytosolic E3 ubiquitin ligase, Parkin, to the outer mitochondrial membrane followed by the subsequent recruitment of mitochondria into double-membraned autophagosomes. This process is facilitated by the recruitment of the autophagosomal protein, LC3, by LC3-binding receptors that accumulate on damaged mitochondria providing a critical link between the cargo to be degraded and the autophagosome. Although evidence suggests that certain autophagy receptors promote the clearance of specific organelles or organelle components, little is known about the precise LC3-receptors that mediate cardiomyocyte mitophagy. Having previously identified GRAF1 as a critical regulator of cardiac form and function, our current data indicate that GRAF1 plays an important role in regulating cardiomyocyte mitochondrial clearance and metabolism. GRAF1 is expressed at high levels in the heart from E17 onwards and is poised to co-regulate actin- and lipid- dynamics by virtue of its multi-domain structure that includes a lipid binding/bending BAR domain, a phospholipid binding PH domain, a Rho-GAP domain, and a protein-interaction SH3 domain. Importantly, GRAF1 depletion in primary cardiomyocytes led to impaired mitochondrial OXPHOX-mediated ATP generation, mitochondrial membrane depolarization, increased mitochondrial-associated ROS, and increased ischemia/reperfusion-dependent myocyte death. GRAF1 depletion in cultured cardiomyocytes reduced LC3 mediated autophagic flux and led to the accumulation of mitochondria, and we observed similar effects in hearts from genetically modified GRAF1-deficient mice. Mechanistically, we showed that GRAF1 facilitates Parkin-dependent mitophagy by serving as a novel LC3 receptor. Our aims for this award are three-fold: In aim1, we will undertake a step-wise approach using sophisticated pH sensitive fluorophores to identify the precise mechanisms by which GRAF1 regulates cardiomyocyte mitochondrial homeostasis. In aim 2, we will use our newly developed cardiac-restricted GRAF1 knock-out mice to assess GRAF1’s contributions to cardiac metabolic reprogramming and in aim 3 we will use these mice to evaluate a Role of GRAF1-mediated mitophagy in cardioprotection. Results from the experiments proposed herein will provide the scientific foundation for the rational design of strategies to control cardiomyocyte mitophagy and could lead to novel approaches to treat ischemic heart disease.

概括由于线粒体功能障碍会影响 ATP 的产生并促进氧化损伤，这些细胞器在健康心肌细胞中，通过涉及自噬的过程每 2-3 周更新一次。缺陷于线粒体质量控制还会促进心脏病的进展，因此确定线粒体质量控制至关重要调节这一过程的机制。涉及的几个主要途径涉及结合胞质 E3 泛素连接酶 Parkin 连接至线粒体外膜，然后是后续的连接酶将线粒体募集到双膜自噬体中。这个过程是由通过在受损细胞上积累的 LC3 结合受体募集自噬体蛋白 LC3 线粒体在待降解的货物和自噬体之间提供了关键的联系。虽然有证据表明某些自噬受体促进特定细胞器或细胞器的清除但对于介导心肌细胞线粒体自噬的精确 LC3 受体知之甚少。拥有先前确定 GRAF1 是心脏形态和功能的关键调节因子，我们当前的数据表明 GRAF1在调节心肌细胞线粒体清除和代谢中发挥重要作用。格拉夫1 从 E17 起在心脏中高水平表达，并准备共同调节肌动蛋白和脂质凭借其多域结构实现动力学，其中包括脂质结合/弯曲 BAR 域、磷脂结合 PH 结构域、Rho-GAP 结构域和蛋白质相互作用 SH3 结构域。重要的是，原代心肌细胞中 GRAF1 的缺失导致线粒体 OXPHOX 介导的 ATP 受损产生，线粒体膜去极化，线粒体相关活性氧增加，以及增加缺血/再灌注依赖性心肌细胞死亡。培养心肌细胞中 GRAF1 的缺失降低了 LC3 介导自噬流并导致线粒体积累，我们在来自转基因 GRAF1 缺陷小鼠的心脏。从机制上讲，我们表明 GRAF1 促进 Parkin 依赖性线粒体自噬作为新型 LC3 受体。我们获得该奖项的目标有三个：目标1，我们将采取逐步的方法，使用复杂的 pH 敏感荧光团来识别 GRAF1 调节心肌细胞线粒体稳态的精确机制。在目标 2 中，我们将使用我们新开发的心脏受限 GRAF1 敲除小鼠来评估 GRAF1 对心脏的贡献代谢重编程，在目标 3 中，我们将使用这些小鼠来评估 GRAF1 介导的作用线粒体自噬在心脏保护中的作用。本文提出的实验结果将提供科学依据为合理设计控制心肌细胞线粒体自噬的策略奠定了基础，并可能导致新的研究治疗缺血性心脏病的方法。

项目成果

期刊论文数量（5）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Reply to 'Lactate as a major myokine and exerkine'.

回复“乳酸作为主要的肌动因子和运动因子”。

DOI：
10.1038/s41574-022-00726-y
发表时间：
2022
期刊：
Nature reviews. Endocrinology
影响因子：
0
作者：
Chow,LisaS;Gerszten,RobertE;Taylor,JoanM;Pedersen,BenteK;vanPraag,Henriette;Trappe,Scott;Febbraio,MarkA;Galis,ZorinaS;Gao,Yunling;Haus,JacobM;Lanza,IanR;Lavie,CarlJ;Lee,Chih-Hao;Lucia,Alejandro;Moro,Cedric;Pandey,A
通讯作者：
Pandey,A

Down-regulation of Beclin1 promotes direct cardiac reprogramming.

DOI：
10.1126/scitranslmed.aay7856
发表时间：
2020-10-21
期刊：
Science translational medicine
影响因子：
17.1
作者：
Wang L;Ma H;Huang P;Xie Y;Near D;Wang H;Xu J;Yang Y;Xu Y;Garbutt T;Zhou Y;Liu Z;Yin C;Bressan M;Taylor JM;Liu J;Qian L
通讯作者：
Qian L

The actin depolymerizing factor destrin serves as a negative feedback inhibitor of smooth muscle cell differentiation.

肌动蛋白解聚因子 destrin 充当平滑肌细胞分化的负反馈抑制剂。