Role of UCH-L1 Oxidative Modification in Parkinson's Disease Pathogenesis

UCH-L1 氧化修饰在帕金森病发病机制中的作用

• 批准号: 8009813
• 负责人: JEANNE Elizabeth MCKEON
• 金额: $ 3.05万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2012-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8009813
• 关键词: Affect; Alanine; Aspartate; Autophagocytosis; Autopsy; Biochemical; Biological Assay; Bradykinesia; Brain; Brain region; Cell Death; Cell Survival; Cell physiology; Cells; Cysteine; Degradation Pathway; Deposition; Deubiquitinating Enzyme; Deubiquitination; Development; Disease; Dopaminergic Cell; Exposure to; Family; Functional disorder; Generations; Human; Hydrolase; Impairment; In Vitro; Individual; Isoleucine; Lewy Bodies; Link; Lysine; Mass Spectrum Analysis; Mediating; Methionine; Modification; Molecular Chaperones; Motor; Mus; Mutate; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Oxidative Stress; Parkinson Disease; Pathogenesis; Pathway interactions; Patients; Polyacrylamide Gel Electrophoresis; Predisposition; Property; Proteins; Reactive Oxygen Species; Reporting; Rest Tremor; Risk Factors; Role; Rotenone; Severities; Substantia nigra structure; System; Testing; Tissues; Toxic Environmental Substances; Toxicant exposure; Ubiquitin; Western Blotting; axonal degeneration; dinitrophenylhydrazine; disorder risk; immunocytochemistry; inhibition of autophagy; motor learning; multicatalytic endopeptidase complex; mutant; novel therapeutics; overexpression; oxidation; prevent; protein degradation; research study; small hairpin RNA; two-dimensional

DESCRIPTION (provided by applicant): Parkinson's disease (PD) is the most common neurodegenerative disorder affecting the motor system, and is characterized clinically by bradykinesia and resting tremor, and pathologically by dopaminergic neurodegeneration and intracellular Lewy body deposition. Ubiquitin carboxyl-terminal-hydrolase LI (UCH- L1) is an abundant neuronal deubiquitinating enzyme, and recently the I93M mutation of UCH-L1 was described in a family with autosomal dominant PD. UCH-L1 is implicated in both the ubiquitin proteasome system (UPS) and chaperone mediated autophagy (CMA) protein degradation pathways. PD-linked I93M mutant UCH-L1 demonstrates significantly reduced deubiquitinating activity and additionally, I93M mutant UCH-L1 overexpression is associated with dopaminergic neurodegeneration and inhibition of the CMA pathway. Mice lacking functional UCH-L1 display axonal degeneration, reduced free monomeric ubiquitin level, and motor and learning deficits. Decreased expression of UCH-L1 is observed in cortex of sporadic PD patients, as well as extensive modification by carbonylation and oxidation at methionine and cysteine residues. Together these observations suggest that UCH-L1 dysfunction is detrimental to neurons, and may contribute to PD pathogenesis. Exposure to environmental toxicants which are linked to increased reactive oxygen species (ROS) generation has been identified as a PD risk factor. The mechanism by which environmental toxicants contribute to PD pathogenesis remains unclear, but one possibility is that increased ROS induce oxidative modification of particular target proteins, such as UCH-L1, leading to pathological alteration of their cellular function. This proposal will test the hypothesis that environmental-toxicant induced oxidative stress causes oxidative modification of UCH-L1, thereby disrupting the role of UCH-L1 in the UPS and CMA degradation pathways, which leads to toxic protein accumulation and dopaminergic cell death. The specific aims are to 1) determine the severity of UCH-L1 oxidative modification in the predominantly affected brain region in PD, the substantia nigra, and to evaluate UCH-L1 as a target of oxidative modification caused by environmental toxicant exposures; 2) characterize the effects of oxidative modification of UCH-L1 on enzymatic activity and interactions with CMA machinery; and 3) determine the effects of UCH-L1 oxidative modification on the UPS and CMA protein degradation pathways and cell viability. Understanding the role of UCH-L1 oxidative modification and subsequent alterations of its function on protein degradation and cell viability will contribute to the understanding of PD pathogenesis and to the development of novel therapeutics or treatments for this disease.

描述（由申请人提供）：帕金森病（PD）是最常见的影响运动系统的神经退行性疾病，临床表现为运动迟缓和静息性震颤，病理表现为多巴胺能神经变性和细胞内路易体沉积。泛素羧基末端水解酶LI （Ubiquitin carboxyl-terminal-hydrolase LI, UCH-L1）是一种丰富的神经元去泛素化酶，最近在一个常染色体显性PD家族中发现了UCH-L1的I93M突变。UCH-L1参与泛素蛋白酶体系统（UPS）和伴侣介导的自噬（CMA）蛋白降解途径。pd相关的I93M突变体UCH-L1显示去泛素化活性显著降低，此外，I93M突变体UCH-L1过表达与多巴胺能神经变性和CMA通路抑制有关。缺乏功能性UCH-L1的小鼠表现为轴突变性，游离单体泛素水平降低，运动和学习缺陷。在散发性PD患者的皮层中观察到UCH-L1的表达降低，并且在蛋氨酸和半胱氨酸残基上被羰基化和氧化广泛修饰。综上所述，这些观察结果表明UCH-L1功能障碍对神经元有害，并可能导致帕金森病的发病。暴露于与活性氧（ROS）生成增加有关的环境毒物已被确定为帕金森病的危险因素。环境毒物促进PD发病的机制尚不清楚，但一种可能性是ROS增加诱导特定靶蛋白（如UCH-L1）的氧化修饰，导致其细胞功能的病理改变。该提案将验证环境毒物诱导的氧化应激导致UCH-L1氧化修饰的假设，从而破坏UCH-L1在UPS和CMA降解途径中的作用，从而导致有毒蛋白积累和多巴胺能细胞死亡。具体目的是：1)确定PD患者大脑主要受影响区域黑质中UCH-L1氧化修饰的严重程度，并评估UCH-L1作为环境毒物暴露引起的氧化修饰的靶点；2)表征UCH-L1氧化修饰对酶活性的影响及其与CMA机制的相互作用；3)确定UCH-L1氧化修饰对UPS和CMA蛋白降解途径及细胞活力的影响。了解UCH-L1氧化修饰及其随后对蛋白质降解和细胞活力的功能改变的作用，将有助于了解PD的发病机制，并有助于开发新的治疗方法或治疗方法。