Endolysosomal defects in secretory autophagy and microglial toxicity in FTD

FTD 中分泌性自噬和小胶质细胞毒性的内溶酶体缺陷

基本信息

批准号：
10623233
负责人：
Jayanta Debnath
金额：
$ 71.39万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-09-15 至 2027-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10623233
关键词：
Affect Amyloid beta-Protein Autophagocytosis Autophagosome Brain Catabolism Cell secretion Coculture Techniques Defect Degradation Pathway Disease Equilibrium Exocytosis Follow-Up Studies Frontotemporal Dementia Functional disorder Funding Genetic Gliosis Homeostasis Inflammation Mediators Inflammatory Response Intracellular Transport Laboratories Lipids Lysosomes MYO5A gene Mediating Microglia Modeling Mus Myosin ATPase Nerve Degeneration Neurodegenerative Disorders Neuroglia Neurons Organelles PGRN gene Pathogenesis Pathology Pathway interactions Phenotype Phospholipids Process Proteins Proteomics RNA-Binding Proteins Regulation Role Signal Transduction Testing Toxic effect Up-Regulation Vesicle Work age related age related neurodegeneration aging brain cell type extracellular extracellular vesicles glial activation hyperphosphorylated tau insight lipid metabolism lipid transport loss of function misfolded protein nanoparticle neuron loss neurotoxic neurotoxicity particle protein TDP-43 proteostasis receptor trafficking transcriptomics

项目摘要

PROJECT SUMMARY Glial pathology is increasingly recognized as a key feature in neurodegenerative diseases. However, the exact mechanisms that promotes glial pathology remain unclear. Several studies implicate the autophagy machinery, a highly conserved lysosomal degradation pathway, in maintaining protein homeostasis in neurons during brain aging and neurodegeneration. In addition, autophagy regulates the check-and-balance of microglia-mediated inflammatory response to misfolded proteins, including β-amyloid peptides, hyperphosphorylated Tau, and TDP- 43, which accumulate during neurodegeneration. Collectively, these results underscore the critical and cell type- specific role of the autophagy machinery in maintaining neuron-glia homeostasis by regulating the trafficking of intracellular organelles and cargoes, such as misfolded proteins. In the previous funding period, we uncovered new autophagy-dependent secretory processes in non-neuronal and glial cells, including most notably, LC3- dependent extracellular vesicle loading and secretion (LDELS). LDELS targets include RNA binding proteins and extracellular vesicle (EV) cargoes that have been implicated in the pathogenesis of frontotemporal dementia (FTD). In addition, our results showed that loss of function in FTD gene Progranulin (Grn) leads to age-dependent endolysosomal defects and hypersecretory phenotypes in microglia that promotes neuronal loss and abnormal accumulation of the RNA binding protein TDP-43 in Grn-/- mouse brain. Together, these results underscore the important roles of autophagy and endolysosomal trafficking in microglial activation in FTD. In follow up studies, we have further expanded the scope of our work and identified a highly regulated mechanism, Secretory Autophagy during Lysosome Inhibition (SALI), which is critically dependent on multiple ATGs for autophagosome formation and on Rab27a for vesicle exocytosis. Using TMT-based quantitative proteomics, we uncovered that loss of Progranulin leads to a marked increase in unconventional myosin Myo5a, a known Rab27a interacting partner. Given the well-documented role of Rab27a and Myo5a in the transport of intracellular organelles, these results broach the hypothesis that blockage of endolysosomal trafficking disrupts microglial homeostasis and promotes secretory autophagy and neurodegeneration in Grn-/- mice. To test this hypothesis, we will: 1) Delineate how FTD-associated endolysosomal defects influence secretory autophagy and dissect the effects of SALI on intracellular homeostasis; 2) Scrutinize how secretory autophagy and endolysosomal defects in microglia impact lipid metabolism and microglial toxicity; and 3) Determine whether genetic autophagy loss mitigates microglial toxicity and neurodegeneration in Progranulin-deficient mice. Together, the studies proposed in this renewal harness the unique expertise in Drs. Huang and Debnath’s laboratories and provide critical insights on how SALI controls the delicate balance of secretion and intracellular trafficking of disease-relevant proteins in microglia to promote neurodegeneration in FTD.

项目摘要胶质病理越来越多地被认为是神经退行性疾病中的关键特征。但是，确切的促进神经胶质病理学的机制尚不清楚。几项研究暗示了自噬机械，高度保守的溶酶体降解途径，在脑中维持神经元中的蛋白质稳态衰老和神经变性。此外，自噬调节小胶质细胞介导的检查和余额对错误折叠蛋白的炎症反应，包括β-淀粉样胡椒，高磷酸化的tau和TDP- 43，在神经退行性过程中积累。总的来说，这些结果强调了关键和细胞类型自噬机械在维持神经胶质体内平衡中的具体作用，通过调节贩运细胞内细胞器和货物，例如错误折叠的蛋白质。在上一个资金期间，我们发现了非神经元和神经胶质细胞中的新自噬依赖性分泌过程，包括最著名的是LC3- 依赖的细胞外囊泡负荷和分泌（LDELS）。 LDELS靶标包括RNA结合蛋白在额颞痴呆的发病机理中隐含的细胞外囊泡（EV）货物（FTD）。此外，我们的结果表明，FTD基因前植物（GRN）的功能丧失导致年龄依赖性促进神经元丧失和异常的小胶质细胞中的内溶性缺陷和性表型 RNA结合蛋白TDP-43在GRN - / - 小鼠脑中的积累。这些结果在一起强调了自噬和内溶性贩运在FTD中的小胶质细胞激活中的重要作用。在后续研究中，我们进一步扩大了工作范围，并确定了高度监管的机制，秘书溶酶体抑制期间的自噬（SALI），这是自噬小体的多个ATG 形成和Rab27a蔬菜胞吐作用。使用基于TMT的定量蛋白质组学，我们发现了失去尿素的损失导致非常规肌球蛋白肌球蛋白肌球蛋白肌球蛋白显着增加，这是一种已知的RAB27A相互作用伙伴。鉴于Rab27a和Myo5a在细胞内细胞器中的运输中有据可查的作用，这些结果提出了以下假设：内溶性贩运的阻塞破坏了小胶质体内稳态和促进GRN - / - 小鼠的秘书自噬和神经退行性。为了检验这一假设，我们将：1）描述 FTD相关的内溶性缺陷如何影响秘书自噬并剖析Sali对细胞内稳态； 2）仔细检查小胶质细胞影响的秘书自噬和内溶性缺陷脂质代谢和小胶质毒性； 3）确定遗传自噬损失是否减轻小胶质细胞缺陷小鼠的毒性和神经退行性。共同提出的研究合计利用博士的独特专业知识。黄和黛布纳斯的实验室，并就萨利的方式提供关键见解控制小胶质细胞中与疾病相关蛋白的分泌和细胞内运输的微妙平衡在FTD中促进神经变性。