Obesity-associated Mitophagy Resistance

肥胖相关的线粒体自噬抵抗

• 批准号: 9750720
• 负责人: Michael J. Jurczak
• 金额: $ 35.21万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-25 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9750720
• 关键词: Acetylation; Affect; Biochemical; Biology; Cell Respiration; Cells; Citric Acid Cycle; Confocal Microscopy; Data; Development; Diet; Discipline; Disease; Event; Excision; Fatty Liver; Free Radicals; Generations; Genetic; Health; Hepatic; Homeostasis; Impairment; In Vitro; Insulin Resistance; Knowledge; Lipids; Liver; Liver Mitochondria; Lysine; Mass Spectrum Analysis; Mediating; Metabolic; Metabolic Diseases; Metabolic stress; Mitochondria; Mitochondrial Proteins; Modeling; Molecular; Mus; Mutagenesis; Non-Insulin-Dependent Diabetes Mellitus; Nutritional; Obese Mice; Obesity; Obesity associated liver disease; Outer Mitochondrial Membrane; Oxidants; Oxidative Phosphorylation; PINK1 gene; Pathogenesis; Pathology; Pathway interactions; Phosphorylation; Polyubiquitin; Post-Translational Protein Processing; Predisposition; Production; Protein Acetylation; Protein-Serine-Threonine Kinases; Proteins; Proteomics; Quality Control; Reactive Oxygen Species; Reporting; Research Design; Resistance; Signal Pathway; Signal Transduction; Stress; Testing; Therapeutic; Thinness; Ubiquitin; Ubiquitination; base; design; improved; in vivo; insight; insulin sensitivity; insulin signaling; lifestyle intervention; mitochondrial dysfunction; novel; obesity development; recruit; stable cell line; targeted treatment; ubiquitin-protein ligase

Project summary/Abstract Abnormal mitochondrial function is proposed to contribute to the hepatic insulin resistance and steatosis associated with obesity and type 2 diabetes. Recently, diet-induced obesity in mice was found to be associated with reduced rates of hepatic mitophagy, a mitochondrial quality control pathway that regulates selective removal of damaged mitochondria from the cell. Mitophagy signaling and targeting of damaged mitochondria relies on generation of a phosphorylated ubiquitin signaling motif on the outer mitochondrial membrane that is produced through the coordinated activation of the ubiquitin E3 ligase PARKIN and the serine/threonine kinase PINK1. This proposal will test the hypothesis that obesity-associated liver metabolic disease arises in part due to a loss of mitochondrial homeostasis resulting from impaired PINK1-PARKIN-mediated signaling and subsequently reduced mitophagy. The approach to test this hypothesis consists of characterizing a novel mouse line with liver-specific deletion of PARKIN at baseline and during diet-induced obesity, and will include a breadth of in vivo and ex vivo approaches designed to evaluate hepatic mitochondrial oxidative metabolism, reactive oxygen species production, hepatic insulin sensitivity, rates of mitophagic flux, and changes in hepatic insulin signaling and stress signaling pathways. These studies will define how genetic inhibition of PARKIN- mediated mitophagy in liver impacts mitochondrial biology and the pathogenesis of obesity-associated liver metabolic disease. Additionally, key aspects of PINK1-PARKIN-mediated mitophagy signaling will be evaluated in lean and obese mice using a combination of approaches that includes quantitative mass spectrometry- based proteomics, confocal microscopy and mutagenesis followed by in vivo and in vitro functional characterization using primary and stable cell lines and cell free models. These studies will define how nutritional stress in the obese liver contributes to post-translational modifications of mitochondrial proteins that then interfere with recently defined phosphorylation and ubiquitination signaling events that activate the mitophagy pathway. They will provide further insight into how PINK1 and PARKIN dynamics are affected in the obese liver and may account for the reduced mitophagy observed in liver from diet-induced obese mice. Overall, these studies will explore a novel pathway by which nutritional stress in liver may contribute to the pathogenesis of hepatic steatosis, insulin resistance and mitochondrial dysfunction. This new knowledge will ultimately improve our understanding of the biochemical and molecular events that contribute to these diseases, which may lend themselves to the development of effective life style interventions or targeted therapeutics to treat obesity-associated liver disease.

项目概要/摘要 线粒体功能异常被认为导致肝脏胰岛素抵抗和脂肪变性 与肥胖和2型糖尿病有关。最近，发现饮食引起的小鼠肥胖与 肝线粒体自噬率降低，这是一种调节选择性的线粒体质量控制途径 从细胞中去除受损的线粒体。线粒体自噬信号传导和针对受损线粒体的靶向 依赖于线粒体外膜上磷酸化泛素信号基序的生成，即 通过泛素 E3 连接酶 PARKIN 和丝氨酸/苏氨酸激酶的协同激活产生 粉色1。该提案将检验以下假设：肥胖相关的肝脏代谢疾病的发生部分是由于 由于 PINK1-PARKIN 介导的信号传导受损而导致线粒体稳态丧失 随后减少线粒体自噬。检验这一假设的方法包括描述一部小说的特征 在基线和饮食诱导肥胖期间肝脏特异性缺失 PARKIN 的小鼠品系，将包括 旨在评估肝线粒体氧化代谢的体内和离体方法的广度， 活性氧的产生、肝脏胰岛素敏感性、线粒体自噬通量率以及肝脏的变化 胰岛素信号传导和应激信号传导途径。这些研究将确定 PARKIN- 的基因抑制如何 肝脏介导的线粒体自噬影响线粒体生物学和肥胖相关肝脏的发病机制 代谢性疾病。此外，还将评估 PINK1-PARKIN 介导的线粒体自噬信号传导的关键方面 在瘦小鼠和肥胖小鼠中使用包括定量质谱在内的组合方法 基于蛋白质组学、共聚焦显微镜和诱变，然后进行体内和体外功能 使用原代细胞系和稳定细胞系以及无细胞模型进行表征。这些研究将定义如何 肥胖肝脏中的营养应激导致线粒体蛋白的翻译后修饰， 然后干扰最近定义的磷酸化和泛素化信号事件，激活 线粒体自噬途径。他们将进一步深入了解 PINK1 和 PARKIN 动态在 肥胖的肝脏可能是饮食诱导的肥胖小鼠肝脏中观察到的线粒体自噬减少的原因。 总的来说，这些研究将探索一种新的途径，肝脏中的营养应激可能会导致 肝脂肪变性、胰岛素抵抗和线粒体功能障碍的发病机制。这些新知识将 最终提高我们对促成这些的生化和分子事件的理解 疾病，这可能有助于制定有效的生活方式干预措施或有针对性的治疗 治疗肥胖相关肝病的疗法。