PINK1 in the Regulation of Macroautophagy and Parkinsonian Neurodegeneration.

PINK1 在巨自噬和帕金森神经变性的调节中的作用。

• 批准号: 8071041
• 负责人: Salvatore James Cherra
• 金额: $ 1.25万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2011-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8071041
• 关键词: 1-Methyl-4-phenylpyridinium; Abbreviations; Affect; Alzheimer' s Disease; Apoptotic; Autophagocytosis; Axon; Biochemical; Brain; Catabolism; Cell Line; Cells; Child; Clinical; Cultured Cells; Data; Dendrites; Disease; Dopamine; Drug Delivery Systems; Elderly; Functional disorder; Gene Mutation; Genes; Green Fluorescent Proteins; Health; High temperature of physical object; Huntington Disease; Immunofluorescence Microscopy; Impairment; In Vitro; Individual; Induced Mutation; Injury; Knockout Mice; LRRK2 gene; Lead; Length; Life; Light; Link; Maintenance; Mass Spectrum Analysis; Measures; Mediating; Mediator of activation protein; Microscopy; Microtubule-Associated Proteins; Mitochondria; Morphology; Mus; Mutation; Nerve Degeneration; Neurites; Neurodegenerative Disorders; Neuronal Injury; Neuronal Plasticity; Neurons; Neurotoxins; Organelles; Oxidopamine; PINK1 gene; PTEN gene; Parkinson Disease; Parkinsonian Disorders; Pathogenesis; Pathway interactions; Patients; Phosphoproteins; Phosphotransferases; Play; Proteins; RNA Interference; RNAi vector; Regulation; Relative (related person); Research Proposals; Role; Sampling; Signal Pathway; Signal Transduction; Synapses; Synaptic plasticity; Toxic effect; Toxin; Training; Wild Type Mouse; design; effective therapy; inhibitor/antagonist; insight; leucine-rich repeat kinase 2; loss of function mutation; mitochondrial autophagy; mitochondrial dysfunction; mutant; neuroblastoma cell; neuroprotection; overexpression; pre-doctoral; prevent; protective effect; protein profiling; reuptake; synaptic function; therapeutic target; vector

DESCRIPTION (provided by applicant): Neurodegenerative diseases afflict millions of individuals ranging from children to the elderly. Without known causes or effective therapies, this class of diseases outstrips the brain's capacity for compensatory neuronal plasticity with debilitating results. Two pathways involved with neurodegeneration are mitochondrial dysfunction and macroautophagy. This research proposal focuses on the interplay between these two pathways as regulators of axonal and dendritic degeneration (neurite degeneration). Macroautophagy (hereafter, autophagy) is the bulk catabolism of long-lived proteins and organelles, including mitochondria. While basal levels of autophagy are required for health, dysregulation has been implicated as a cause for neurite degeneration. Mitochondria are ubiquitous organelles that play key roles in the proper function and plasticity of neurons. Dysregulated macroautophagy and mitochondrial dysfunction/loss have been implicated in the pathogenesis of Parkinson's disease (PD). Recent studies have illustrated the neuroprotective activity of the PTEN-induced kinase 1 (PINK1). Additionally, PINK1 has been linked to autosomal recessive PD, presumably through loss of function mutations. I hypothesize that PINK1 signaling regulates neurite degeneration through its effects on autophagy/mitophagy. The role of PINK1 in regulating neurite degeneration, autophagy, and mitochondrial loss will be measured in vitro by biochemical and microscopy studies. Aim 1 will determine whether levels of PINK1 expression regulate autophagy and protect against neuronal injuries that cause neurite degeneration. Aim 2 will determine whether PINK1 knockdown or disease-associated mutations induce autophagy and neurite degeneration. I will also determine whether PINK1 knockdown or mutants require autophagy for neurite degeneration. Aim 3 will use a non-biased proteomies approach to identify downstream mitochondrial targets of PINK1. Phosphorylated protein profiles will be compared between PINK1 deficient mice or cells and wild type mice or cells, respectively. Potential downstream PINK1 targets will be identified as phosphoproteins in wild-type samples that are reduced in PINK1 deficient samples. Using RNAi, I will determine if the putative downstream PINK1 mediators are required for the PINK1 neuroprotective effects. By identifying potential pathways that regulate PINK1-mediated neuroprotection and autophagic "selfdigestion", the results of this study will provide insight into the mechanisms of neurite degeneration in Parkinson's and related diseases. In addition to providing predoctoral training to the applicant, this research proposal is designed to uncover promising drug targets for the treatment of neurodegenerative diseases.

描述(申请人提供)：神经退行性疾病困扰着数以百万计的人，从儿童到老年人。在没有已知的病因或有效的治疗方法的情况下，这类疾病超过了大脑的代偿性神经元可塑性，导致虚弱的结果。参与神经退行性变的两条途径是线粒体功能障碍和巨大的自噬。这项研究计划集中在这两条通路之间的相互作用，作为轴突和树突变性(轴突变性)的调节者。巨自噬(以下简称自噬)是指长寿命蛋白质和细胞器(包括线粒体)的大量分解代谢。虽然基础水平的自噬是健康所必需的，但调节失调已被认为是轴突退化的原因之一。线粒体是无处不在的细胞器，对神经元的正常功能和可塑性起着关键作用。巨噬细胞功能失调和线粒体功能障碍/缺失与帕金森病(PD)的发病有关。最近的研究表明，PTEN诱导的激酶1(PINK1)具有神经保护作用。此外，PINK1可能通过功能突变的丧失与常染色体隐性遗传性帕金森病相关联。我推测PINK1信号通过对自噬/有丝分裂吞噬的影响来调节轴突变性。PINK1在调节轴突变性、自噬和线粒体丢失中的作用将在体外通过生化和显微镜研究来衡量。目的1将确定PINK1的表达水平是否调节自噬并保护神经元免受导致轴突变性的神经元损伤。目标2将确定PINK1基因敲除或疾病相关突变是否会导致自噬和轴突变性。我还将确定PINK1基因敲除或突变是否需要对轴突变性进行自噬。AIM 3将使用一种无偏见的蛋白质组方法来识别PINK1的下游线粒体靶点。PINK1缺陷小鼠或细胞与野生型小鼠或细胞之间的磷酸化蛋白质图谱将分别进行比较。在野生型样本中，潜在的下游PINK1靶标将被鉴定为磷蛋白，而在PINK1缺失的样本中，野生型样本中的磷蛋白被减少。利用RNAi，我将确定PINK1下游介体是否对PINK1的神经保护作用是必需的。通过识别调节PINK1介导的神经保护和自噬“自我消化”的潜在途径，这项研究的结果将为帕金森病和相关疾病的轴突变性机制提供深入的认识。除了为申请者提供博士前培训外，这项研究提案还旨在发现治疗神经退行性疾病的有希望的药物靶点。