Chaperone-mediated Autophagy and Synaptic Dysfunction in Parkinson's Disease

帕金森病（PD）是第二常见的神经退行性疾病，其特征在于： 黑质延髓部多巴胺能（DA）神经元的退行性丢失。研究细胞和 动物模型和死后PD患者的大脑显示，环境因素引发的突触功能障碍 遗传应激是PD发病的早期事件。然而，精确和关键的机制， PD中潜在的突触功能障碍仍有待确定。液泡（H+）-ATP酶（V-ATP酶）是一种ATP- 依赖性质子泵参与酸化突触囊泡，并在多个步骤中发挥关键作用， 突触囊泡生命周期，如与突触前膜融合和神经递质释放/重新加载。 我们的初步研究表明，突触V-ATP酶可能是由伴侣介导的自噬控制的 (CMA)这是一种基于溶酶体的过程，专门处理氧化或受损的蛋白质， 在压力下维持细胞功能。值得注意的是，CMA受到与以下相关的多种应激条件的抑制： PD包括衰老、神经毒性应激、氧化应激、ER应激和遗传应激。然而，无论和 细胞应激信号和CMA如何参与V-ATP酶调节突触功能仍然是未知的。在 目前的项目，通过利用多个模型系统，包括尖端的人类诱导多能干细胞 （iPSC）模型、体内啮齿动物模型（啮齿动物脑的ER和遗传应激）和死后PD患者 我们的目标是确定CMA是否直接降解突触V-ATP酶的受损成分， 维持突触囊泡功能的压力下，以及是否损失足够的CMA活动损害突触 在PD中发挥作用。首先，我们将确定生物化学，细胞生物学和电生理学，如果CMA 直接调节源自人iPSC的DA神经元中的突触V-ATP酶和突触功能。第二、 我们将测试与PD相关的多种应激条件是否通过CMA调节突触V-ATP酶和功能， 人iPSC衍生的DA神经元。最后，我们将评估CMA-V-ATP酶-突触的调节和作用， 在多种PD动物模型和来自PD患者的死后脑中，我们提出的研究 将大大推进我们对压力如何调节突触囊泡的理解，揭示一个关键的致病因素， PD中突触失败的潜在机制，并为开发 诊断/生物标志物和更有效的疾病预防和治疗策略。

Parkinson's disease (PD) is the second most common neurodegenerative disease characterized by the degenerative loss of dopaminergic (DA) neurons in the substantia nigra pars compacta. Studies of cellular and animal models and postmortem PD patient brains reveal that synaptic dysfunction triggered by environmental and genetic stress is an early event in PD pathogenesis. However, the precise and key mechanisms that underlie synaptic dysfunction in PD remain to be defined. The vacuolar (H+)-ATPase (V-ATPase) is an ATP- dependent proton pump involved in acidifying synaptic vesicles and plays a critical role in multiple steps of synaptic vesicle life cycle such as fusion with pre-synaptic membrane and neurotransmitter release/reloading. Our preliminary studies show that synaptic V-ATPase may be controlled by chaperone-mediated autophagy (CMA), a lysosome-based process specialized in disposing oxidized or damaged proteins for degradation to maintain cellular function under stress. Notably, CMA is inhibited by multiple stress conditions associated with PD including aging, neurotoxic stress, oxidative stress, ER stress, and genetic stress. However, whether and how cellular stress signals and CMA engage V-ATPase to modulate synaptic function is still unknown. In the current project, by utilizing multiple model systems, including cutting-edge human induced pluripotent stem cell (iPSC) model, in vivo rodent models (ER and genetic stress of rodent brains), and postmortem PD patient brains, we aim to determine whether CMA directly degrades damaged components of synaptic V-ATPase to maintain synaptic vesicle function under stress, and whether loss of adequate CMA activity impairs synaptic function in PD. First, we will determine biochemically, cell biologically, and electro-physiologically if CMA directly regulates synaptic V-ATPase and synaptic function in DA neurons derived from human iPSCs. Second, we will test if multiple stress conditions related to PD regulate synaptic V-ATPase and function via CMA in human iPSC-derived DA neurons. Last, we will assess the regulation and role of CMA–V-ATPase–synaptic vesicle pathway in multiple PD animal models and in postmortem brains from PD patients. Our proposed study will significantly advance our understanding of how stress regulates synaptic vesicle, reveal a key pathogenic mechanism underlying synaptic failure in PD, and offer new opportunities for developing diagnostics/biomarkers and more effective prevention and treatment strategies for the disease.