Chloride Homeostasis in Lysosomal Function and Parkinson's Disease

溶酶体功能和帕金森病中的氯稳态

• 批准号: 10515961
• 负责人: ZIXU MAO
• 金额: $ 60.46万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10515961
• 关键词: Affect; Albers-Schonberg disease; Animal Model; Animals; Anions; Automobile Driving; Autophagocytosis; Autophagosome; Autopsy; Binding; Biochemical; Biological Markers; Biological Process; Biology; Brain; Brain Diseases; CLC Gene; Cathepsin C; Cell physiology; Cells; Chloride Channels; Chloride Ion; Chlorides; Complex; Data; Degenerative Disorder; Degradation Pathway; Disease; Environmental Risk Factor; Event; Family; Functional disorder; GTP-Binding Protein alpha Subunits, Gs; Genes; Genetic; Genetic Determinism; Genetic study; Health; Homeostasis; Human; Hydrolase; Impairment; Induced pluripotent stem cell derived neurons; Intracellular Space; LRRK2 gene; Link; Lysosomal Storage Diseases; Lysosomes; Modeling; Molecular; Mus; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Organelles; Oxidative Stress; Parkinson Disease; Pathogenesis; Pathogenicity; Pathologic; Pathology; Pathway interactions; Patients; Play; Process; Proteins; Regulation; Reporting; Research; Rodent Model; Role; Signal Transduction; Site; Stress; Substantia nigra structure; Testing; base; design; detection of nutrient; dopaminergic neuron; experimental study; human model; induced pluripotent stem cell; innovation; member; mutant; nervous system disorder; new therapeutic target; pars compacta; response; targeted biomarker

Parkinson's disease (PD) is a progressive and degenerative disorder of the brain. It is pathologically characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta (SNc). The key events driving the pathogenesis in PD are not completely understood. The long-term objective of my research is to understand the molecular and cellular processes by which neurons respond to stress and how dysfunction of these responsive mechanisms contributes to neurodegenerative process. I propose to investigate a new molecular regulatory process of lysosomal chloride and its role in PD pathogenesis. Chloride ion is the most abundant anion in both extra- and intracellular spaces of animal cells. No longer regarded as an inert anion, chloride is found to play discrete roles in cells and its homeostasis needs to be tightly regulated at a subcellular organelle level. Lysosomal chloride is important for the function of acidic hydrolases. In addition to this general role, lysosomal chloride itself also has specific roles affecting lysosomal functions such as binding directly to cathepsin C to regulate its activity. Lysosomal chloride is mainly regulated by chloride channel 7 (CLC-7) in complex with its beta subunit Ostm-1. Loss of either protein severely comprises lysosomal chloride homeostasis, reduces lysosomal degradation capacity, and causes accumulation of lysosomal storage materials and autophagosomes, leading to diseases in human and rodent models including neuronal damages and degeneration. How CLC-7 is regulated remains largely unknown and no studies have ever reported its involvement in PD pathogenesis. We discovered unexpectedly a regulatory link between leucine-rich repeat kinase 2 (LRRK2), one of the most common genetic determinants associated with PD, and CLC-7. Our new preliminary data show a direct interaction between LRRK2 and CLC-7. This interaction is pathogenically enhanced by LRRK2 G2019S mutation and by oxidative stress, leading to aberrantly high level of lysosomal chloride and reduced lysosomal activities. We hypothesize that LRRK2 interacts with CLC-7 to modulate lysosomal chloride homeostasis and pathogenic mutant LRRK2G2019S dysregulates this process and impairs lysosomal functions. We propose to assess the molecular effects of LRRK2 on CLC-7 function in DA neurons derived from iPSCs of PD patients (aim I), assess the cellular effects of aberrant LRRK2-CLCL-7 interaction under genetic and oxidative stress on lysosomal functions in DA neurons derived from control and PD patient iPSCs (aim II); assess the molecular and cellular effects of LRRK2 on lysosomal CLC-7, chloride, and functions in animal models of PD (aim III), and assess the lysosomal LRRK2, CLC-7, and chloride in postmortem brains of PD patients (aim IV). The study will identify the key process that controls lysosomal chloride, establish its role in PD cytopathogenesis, and possibly reveal new therapeutic targets and biomarkers for the disease.

帕金森病（Parkinson's disease，PD）是一种进行性和退行性的脑部疾病。这是病理学上的 其特征在于黑质腹侧部（SNc）中多巴胺能神经元的损失。关键 在PD中驱动发病机制的事件尚未完全理解。我研究的长期目标是 是了解神经元对压力的反应的分子和细胞过程，以及功能障碍是如何发生的。 这些反应机制的作用导致神经退行性过程。我提议调查一个新的 溶酶体氯的分子调控过程及其在PD发病机制中的作用氯离子是最 在动物细胞的细胞外和细胞内都有丰富的阴离子。不再被视为惰性阴离子， 氯离子在细胞中起着不同的作用，它的稳态需要在亚细胞水平上受到严格的调节。 细胞器水平。溶酶体氯化物对酸性水解酶的功能很重要。除此之外，一般 作用，溶酶体氯化物本身也具有影响溶酶体功能的特定作用，例如直接结合 组织蛋白酶C调节其活性。溶酶体氯主要由氯离子通道7（CLC-7）调节， 与其β亚基Ostm-1的复合物。任一蛋白质的丢失严重地包括溶酶体氯化物 体内平衡，降低溶酶体降解能力，并导致溶酶体蓄积 材料和自噬体，导致人类和啮齿动物模型中的疾病，包括神经元损伤 和退化CLC-7是如何调节的在很大程度上仍然是未知的，也没有研究报道过它的作用。 参与PD发病机制。我们意外地发现富含亮氨酸的重复序列之间存在调节联系 激酶2（LRRK 2），与PD相关的最常见的遗传决定因素之一，和CLC-7。我们的新 初步数据显示LRRK 2和CLC-7之间的直接相互作用。这种相互作用在病理学上是 通过LRRK 2 G2019 S突变和氧化应激增强，导致异常高水平的溶酶体 氯化物和降低的溶酶体活性。我们假设LRRK 2与CLC-7相互作用， 溶酶体氯稳态和致病性突变体LRRK 2G 2019 S失调这一过程， 溶酶体功能我们建议评估LRRK 2对DA神经元中CLC-7功能的分子作用 来源于PD患者的iPSC（目的I），评估异常LRRK 2-CLCL-7的细胞效应 遗传和氧化应激对多巴胺神经元溶酶体功能的影响 对照组和PD患者iPSC（目的II）;评估LRRK 2对溶酶体的分子和细胞影响 CLC-7，氯化物和功能在PD动物模型中的作用（目的III），并评估溶酶体LRRK 2，CLC-7，和 PD患者死后脑中的氯化物（目的IV）。该研究将确定控制的关键过程 溶酶体氯化物，建立其在PD细胞病变中的作用，并可能揭示新的治疗靶点， 疾病的生物标志物。