LRRK2 and oxidative stress in Parkinson’s disease

LRRK2 与帕金森病的氧化应激

基本信息

批准号：
10799999
负责人：
J Timothy Greenamyre
金额：
$ 55.65万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-06-01 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10799999
关键词：
4 hydroxynonenal Accounting Address Alanine Automobile Driving Brain Pathology Cells Chloroquine Complex Consensus Curcumin Cysteine Disease Dopamine Genetic Genetic Risk Human Hydrogen Peroxide Idiopathic Parkinson Disease Ionophores Knock-out LRRK2 gene Link Microglia Mitochondria Mitochondrial Proteins Modeling Monovalent Cations Morphology Mutate Mutation Nerve Degeneration Neurons Oxidation-Reduction Oxidative Stress Oxidative Stress Induction Paraquat Parkinson Disease Pathogenesis Pathogenicity Patients Phosphotransferases Play Prevention Process Production Protein Import Proteins Rattus Regulation Reporting Research Personnel Risk Factors Rodent Model Role Rotenone Signal Transduction Specificity Stress Substantia nigra structure Toxic effect Trichloroethylene Variant autosome brain tissue design dihydroethidium dopaminergic neuron enzyme activity experimental study in vivo kinase inhibitor mitochondrial dysfunction mutant neurotoxicity non-genetic novel novel therapeutic intervention oxidation pharmacologic physiologic stressor prevent receptor response risk variant stressor tool toxicant trafficking

项目摘要

Mutations in LRRK2 are the most common cause of autosomal dominant Parkinson's disease (PD) and it appears that all such pathogenic mutations are associated with aberrantly enhanced LRRK2 kinase activity. Independent of mutations, however, there is also evidence that increased LRRK2 kinase activity contributes to the pathogenesis of idiopathic PD (iPD). The current project is designed to elucidate how wildtype (non-mutated) LRRK2 kinase activity is stimulated, where in the cell this occurs, and what the downstream consequences are. In keeping with the well-known association of iPD with oxidative stress and mitochondrial dysfunction, we focus on these processes in relation to LRRK2. Specifically, this project examines (i) the oxidative activation of LRRK2, (ii) the translocation of LRRK2 to mitochondria under conditions of mitochondrial stress, and (iii) LRRK2 kinase activity-dependent oxidative stress. The overarching premise of this proposal is that oxidative stress and LRRK2 kinase activity are intimately and bidirectionally intertwined in PD pathogenesis. The project has 3 broad Specific Aims to address these issues: Aim 1: To elucidate the oxidative activation of LRRK2 kinase, we will (a) assess/compare WT LRRK2 activation by physiological stressors (H2O2 and DA) to the PD-associated toxicants: rotenone, paraquat (PQ) and trichloroethylene (TCE); (b) Determine whether monensin & chloroquine activate LRRK2 via oxidative mechanisms; (c) Examine the role of cysteine residues C2024/5 in oxidative LRRK2 activation. Aim 2: To examine translocation of LRRRK2 to mitochondria, we will (a) Assess mitochondrial localization of LRRK2 in response to various mitochondrial stressors; (b) See if translocation requires (i) mitochondrial ROS, (ii) cytosolic ROS, (iii) LRRK2 activity or (iv) Cys2024/5; (c) Determine whether mitochondrial translocation of LRRK2 occurs in vivo in rat models of PD; (d) Evaluate whether LRRK2 association with mitochondria is aberrantly enhanced in human iPD brain tissue. Aim 3: To elucidate the role of LRRK2 kinase in oxidative stress and its downstream consequences, we will (a) Use genetic and pharmacological approaches to see if pathogenic LRRK2 mutations cause oxidative stress and whether blocking LRRK2 kinase activity prevents oxidative stress; (b) Examine the potential role of Cys2024/5 in driving LRRK2 kinase activity-dependent oxidative stress; (c) Determine if rotenone-induced mitochondrial ROS production is reduced by LRRK2 inhibition/knockout; (d) Determine if rotenone-induced cytosolic NOX2 activation is reduced by LRRK2 inhibition/knockout; (e) Assess ROS production in patient-derived healthy control, iPD and G2019S mutant lymphoblastoid cells. Together, these experiments will begin to elucidate the intimate and bidirectional relationship between oxidative stress and LRRK2 in PD pathogenesis. By doing so, new therapeutic strategies are likely to emerge.

LRRK2 突变是常染色体显性帕金森病 (PD) 的最常见原因，而且似乎所有这些致病突变都与异常增强的 LRRK2 激酶活性有关。独立的然而，也有证据表明 LRRK2 激酶活性的增加有助于特发性PD（iPD）的发病机制。当前的项目旨在阐明野生型（非突变）如何 LRRK2 激酶活性被刺激，这种情况发生在细胞的哪个部位，以及下游的后果是什么。为了与众所周知的 iPD 与氧化应激和线粒体功能障碍相关，我们重点关注与 LRRK2 相关的这些过程。具体而言，该项目检查 (i) LRRK2 的氧化激活， (ii) LRRK2 在线粒体应激条件下易位至线粒体，以及 (iii) LRRK2 激酶活动依赖性氧化应激。该提案的首要前提是氧化应激和 LRRK2 激酶活性在 PD 发病机制中密切且双向地交织在一起。该项目有 3 个广泛的具体目标来解决这些问题：目标 1：为了阐明 LRRK2 激酶的氧化激活，我们将 (a) 评估/比较 WT LRRK2 激活生理应激源（H2O2 和 DA）对 PD 相关毒物：鱼藤酮、百草枯 (PQ) 和三氯乙烯（TCE）； (b) 确定莫能菌素和氯喹是否通过氧化激活 LRRK2 机制； (c) 检查半胱氨酸残基 C2024/5 在氧化 LRRK2 激活中的作用。目标 2：为了检查 LRRRK2 向线粒体的易位，我们将 (a) 评估 LRRK2 响应各种线粒体应激源； (b) 看看易位是否需要 (i) 线粒体 ROS， (ii) 胞质 ROS，(iii) LRRK2 活性或 (iv) Cys2024/5； (c) 确定线粒体是否易位 LRRK2 存在于 PD 大鼠模型体内； (d) 评估 LRRK2 与线粒体的关联是否人类 iPD 脑组织中异常增强。目标 3：为了阐明 LRRK2 激酶在氧化应激及其下游后果中的作用，我们将 (a) 使用遗传和药理学方法来观察致病性 LRRK2 突变是否会导致氧化应激和阻断 LRRK2 激酶活性是否可以预防氧化应激； (b) 审查 Cys2024/5 的潜在作用驱动 LRRK2 激酶活性依赖性氧化应激； (c) 确定鱼藤酮是否诱导线粒体 LRRK2 抑制/敲除可减少 ROS 产生； (d) 确定鱼藤酮是否诱导胞质 NOX2 LRRK2 抑制/敲除可减少激活； (e) 评估源自患者的健康体内 ROS 的产生对照、iPD 和 G2019S 突变淋巴母细胞。总之，这些实验将开始阐明氧化之间的密切和双向关系。 PD发病机制中的应激和LRRK2。通过这样做，新的治疗策略可能会出现。