Role of p62 in Protein Aggregation and Neurodegeneration in ALS

p62 在 ALS 蛋白质聚集和神经变性中的作用

• 批准号: 7676126
• 负责人: Haining Zhu
• 金额: $ 18.3万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7676126
• 关键词: Address; Amyotrophic Lateral Sclerosis; Animal Model; Autophagocytosis; Autophagosome; Binding; Cessation of life; Collaborations; Cultured Cells; Data; Disease Progression; Electrons; Etiology; Familial Amyotrophic Lateral Sclerosis; Goals; Homeostasis; In Vitro; Kentucky; Knockout Mice; Length; Link; Lysosomes; Mediating; Microscopic; Molecular; Motor; Motor Neurons; Mus; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Onset of illness; Pathway interactions; Polyubiquitin; Precipitation; Process; Proteins; Reporting; Role; Scientist; Series; Signal Transduction; Spinal; System; Techniques; Testing; Time; Tissues; Toxic effect; Transgenic Animals; Transgenic Mice; Ubiquitin; Ubiquitination; Universities; Western Blotting; age related; copper zinc superoxide dismutase; design; domain mapping; in vivo; innovation; insight; motor neuron degeneration; multicatalytic endopeptidase complex; mutant; neuronal survival; new therapeutic target; overexpression; protein aggregate; protein aggregation; protein degradation; protein misfolding

DESCRIPTION (provided by applicant): Protein aggregates containing mutant copper-zinc superoxide dismutase (SOD1) are a hallmark of familial amyotrophic lateral sclerosis (ALS, Lou Gehrig's disease), an age-related neurodegenerative disease. Conformational differences between wild-type (WT) and mutant SOD1 proteins have been demonstrated by many in vitro studies using a variety of biophysical techniques, but in vivo details of the aggregation process remain to be defined. It is also unclear whether protein aggregates are toxic or beneficial. P62/Sequestome 1 (referred as p62 in this proposal) is a multifunctional protein involved in both of the two major protein degradation mechanisms, ubiquitin-proteasome system (UPS) and autophagy-lysosome pathway. We recently reported that the endogenous mouse p62 levels significantly increased prior to the disease onset in G93A SOD1 transgenic mice. In addition, p62 was co-localized with mutant SOD1 and ubiquitin in the protein aggregates in spinal motor neurons in G93A SOD1 transgenic mice. We have established collaboration with Dr. Marie Wooten at Auburn University who is the leading scientist studying p62 in signaling cascades and who will provide the p62 knockout mice for the study. The long term goal of this collaborative project is to understand the role of p62 in protein aggregation caused by ALS-linked SOD1 mutants and motor neuron degeneration in ALS. The innovative aspect of this project is that the role of p62 in the ALS etiology is largely undefined. The central hypothesis to be tested in the project is that p62 can recognize misfolded mutant SOD1 and that p62 can ameliorate the mutant SOD1 induced toxicity by shuttling such misfolded proteins to the ubiquitin-proteasome system (UPS) and/or autophagy. Our preliminary studies demonstrate that p62 specifically recognizes mutant SOD1 and the ubiquitin-association (UBA) domain of p62 is not essential to such interaction. Our results also show that p62 enhances the formation of aggresome-like inclusion of mutant SOD1 but had little effect on WT SOD1. Recent results from other groups showed that p62 can directly bind to autophagy effector protein Atg8/LC3 and that the formation of autophagy was increased in the G93A SOD1 transgenic mice. These data strongly support the central hypothesis. Three specific aims are designed to determine the mechanisms by which p62 recognizes mutant SOD1, the downstream pathways activated by such interaction, and the functional role of p62 in neuronal degeneration. Aim 1 is to map the domains of p62 essential for recognizing and interacting with mutant SOD1. Aim 2 is to determine whether and how p62 mediates the autophagy activation induced by mutant SOD1. Aim 3 is to study how p62 influences protein aggregation and ALS disease progression in vivo using p62 KO mice. The results from this study will provide invaluable insights into the role of p62 in protein aggregation and neurodegeneration in ALS, which will result in better understanding of ALS etiology and potential discovery of new therapeutic target for ALS treatment. Project Narrative This collaborative project between Dr. Zhu at University of Kentucky and Dr. Wooten at Auburn University is to understand the role of p62 in mutant SOD1 induced protein aggregation and motor neuron degeneration in amyotrophic lateral sclerosis (ALS), an age-related neurodegenerative disease. The innovative aspect of this project is that the role of p62 in the ALS etiology is largely undefined. P62 is a multifunctional protein involved in both of the two major protein degradation mechanisms, ubiquitin-proteasome system (UPS) and autophagy- lysosome pathway. The central hypotheses to be tested in the project are that p62 can recognize misfolded mutant SOD1 and that p62 can ameliorate the mutant SOD1 induced toxicity by shuttling such misfolded proteins to UPS and/or autophagy. We have obtained substantial amount of preliminary data that strongly support these hypotheses. Three specific aims are designed to further characterize the molecular details regarding the role of p62 in protein homeostasis (degradation and aggregation) and neuronal survival in ALS. Aim 1 is to map the domains of p62 essential for recognizing and interacting with mutant SOD1. Aim 2 is to determine whether and how p62 mediates the autophagy activation induced by mutant SOD1. Aim 3 is to study how p62 influences protein aggregation and ALS disease progression in vivo using p62 KO mice. The results from this study will provide invaluable insights into the role of p62 in protein aggregation and neurodegeneration in ALS, which will result in better understanding of ALS etiology and potential discovery of new therapeutic target for ALS treatment.

描述（由申请人提供）：含有突变型铜锌超氧化物歧化酶（SOD 1）的蛋白质聚集体是家族性肌萎缩侧索硬化症（ALS，Lou Gehrig病）的标志，这是一种年龄相关的神经退行性疾病。野生型（WT）和突变体SOD 1蛋白之间的构象差异已被证明了许多体外研究使用各种生物物理技术，但在体内的聚集过程的细节仍有待确定。目前还不清楚蛋白质聚集体是有毒还是有益的。P62/Sequestome 1（以下简称p62）是一种多功能蛋白，参与泛素-蛋白酶体系统（UPS）和自噬-溶酶体途径两种主要的蛋白质降解机制。我们最近报道，内源性小鼠p62水平显着增加之前，在G93 A SOD 1转基因小鼠的疾病发作。此外，在G93 A SOD 1转基因小鼠脊髓运动神经元中，p62与突变型SOD 1和泛素共定位于蛋白聚集体中。我们已经与奥本大学的玛丽伍滕博士建立了合作关系，她是研究p62信号级联的主要科学家，并将为这项研究提供p62基因敲除小鼠。该合作项目的长期目标是了解p62在ALS相关SOD 1突变体引起的蛋白质聚集和ALS运动神经元变性中的作用。该项目的创新之处在于p62在ALS病因学中的作用在很大程度上是不确定的。在该项目中要测试的中心假设是，p62可以识别错误折叠的突变体SOD 1，并且p62可以通过将这种错误折叠的蛋白质穿梭到泛素-蛋白酶体系统（UPS）和/或自噬来改善突变体SOD 1诱导的毒性。我们的初步研究表明，p62特异性识别突变SOD 1和泛素相关（乌巴）结构域的p62是不是必不可少的这种相互作用。我们的研究结果还表明，p62增强了突变体SOD 1的侵袭体样包涵体的形成，但对WT SOD 1几乎没有影响。其他研究小组的最新结果表明，p62可以直接结合自噬效应蛋白Atg 8/LC 3，并且在G93 A SOD 1转基因小鼠中自噬的形成增加。这些数据有力地支持了中心假设。三个具体的目的是确定p62识别突变SOD 1的机制，这种相互作用激活的下游通路，以及p62在神经元变性中的功能作用。目的1是定位p62识别突变SOD 1并与其相互作用所必需的结构域。目的2是确定p62是否以及如何介导突变SOD 1诱导的自噬激活。目的3是使用p62 KO小鼠研究p62如何影响体内蛋白聚集和ALS疾病进展。这项研究的结果将为p62在ALS蛋白聚集和神经变性中的作用提供宝贵的见解，这将导致更好地了解ALS病因学和潜在发现ALS治疗的新治疗靶点。肯塔基州的朱博士和奥本大学的Wooten博士的合作项目是了解p62在肌萎缩侧索硬化症（ALS）中突变SOD 1诱导的蛋白质聚集和运动神经元变性中的作用，ALS是一种与年龄相关的神经退行性疾病。该项目的创新之处在于p62在ALS病因学中的作用在很大程度上是不确定的。P62是一种多功能蛋白，参与泛素-蛋白酶体系统（UPS）和自噬-溶酶体途径两种主要的蛋白质降解机制。在该项目中要测试的中心假设是p62可以识别错误折叠的突变体SOD 1，并且p62可以通过将这种错误折叠的蛋白质穿梭到UPS和/或自噬来改善突变体SOD 1诱导的毒性。我们已经获得了大量的初步数据，有力地支持这些假设。三个具体的目的是为了进一步表征p62在蛋白质稳态（降解和聚集）和ALS神经元存活中的作用的分子细节。目的1是定位p62识别突变SOD 1并与其相互作用所必需的结构域。目的2是确定p62是否以及如何介导突变SOD 1诱导的自噬激活。目的3是使用p62 KO小鼠研究p62如何影响体内蛋白聚集和ALS疾病进展。这项研究的结果将为p62在ALS蛋白聚集和神经变性中的作用提供宝贵的见解，这将导致更好地了解ALS病因学和潜在发现ALS治疗的新治疗靶点。