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(PD)是一种进行性和退行性的脑部疾病。这是病理性的 以黑质致密部(SNC)多巴胺能神经元丢失为特征。钥匙 帕金森病的发病机制尚不完全清楚。我研究的长期目标是 是了解神经元对应激做出反应的分子和细胞过程，以及功能障碍 这些反应机制中的一种参与了神经退行性变过程。我提议调查一项新的 氯化溶酶体的分子调控过程及其在帕金森病发病中的作用。氯离子是最多的 在动物细胞的胞外和胞内空间都有丰富的阴离子。不再被视为惰性负离子， 氯被发现在细胞中扮演着离散的角色，它的动态平衡需要在亚细胞中受到严格的调控 细胞器水平。溶酶体氯化物对酸性水解酶的功能很重要。除了这位将军 作用，溶酶体氯本身也有影响溶酶体功能的特定作用，如直接结合 组织蛋白酶C调节其活性。溶酶体氯主要受氯通道7(ClC-7)调节。 具有其β亚基OSTM-1的复合体。任何一种蛋白质的丢失都严重地包括溶酶体氯 动态平衡，降低溶酶体的降解能力，并导致溶酶体储存的积累 材料和自噬小体，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是如何被调控的在很大程度上仍然是未知的，也没有研究报道它 参与帕金森病的发病机制。我们意外地发现了富含亮氨酸的重复序列之间的调节联系 激酶2(LRRK2)，与帕金森病相关的最常见的遗传决定因素之一，以及ClC-7。我们的新产品 初步数据显示，LRRK2和ClC-7之间存在直接相互作用。这种相互作用是病理性的 LRRK2 G2019S突变和氧化应激增强，导致溶酶体异常高水平 氯化物和降低的溶酶体活性。我们假设LRRK2与ClC-7相互作用调节 溶酶体氯动态平衡和致病突变体LRRK2G2019S失调这一过程并损害 溶酶体的功能。我们建议评估LRRK2对DA神经元ClC-7功能的分子效应 帕金森病患者IPSCs来源(目标I)，评估LRRK2-CLCL-7异常对细胞的影响 遗传和氧化应激对下丘脑多巴胺能神经元溶酶体功能的影响 对照组和帕金森病患者IPSCs(AIM II)；评估LRRK2对溶酶体的分子和细胞效应 ClC-7、ClC-7和PD动物模型的功能(Aim III)，并评估溶酶体LRRK2、ClC-7和 帕金森病患者死后脑中的氯化物(目标四)。这项研究将确定控制 溶酶体氯，确定其在帕金森病细胞病变中的作用，并可能揭示新的治疗靶点和 疾病的生物标志物。