Autophagy-dependent exosome loading and biogenesis in AD and FTD

AD 和 FTD 中自噬依赖性外泌体负载和生物发生

• 批准号: 9919472
• 负责人: Jayanta Debnath
• 金额: $ 65.02万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-15 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9919472
• 关键词: Address; Affect; Alzheimer' s Disease; Amyloid; Amyloid beta-Protein Precursor; Astrocytes; Autophagocytosis; Autophagosome; Binding Proteins; Biochemical; Biogenesis; Bioinformatics; Biotinylation; Cell Death; Cell Survival; Cells; Cellular biology; Cleaved cell; Cytoplasm; Defect; Disease; Equilibrium; Estrogen receptor positive; Frontotemporal Dementia; Functional disorder; Goals; Lead; Lipids; Lysosomes; Mediating; Membrane; Microglia; Microtubules; Modeling; Molecular; Multivesicular Body; Mus; N-terminal; Nerve Degeneration; Neurodegenerative Disorders; Neurofibrillary Tangles; Neuroglia; Neurons; Oligodendroglia; PGRN gene; Pathogenesis; Pathologic; Pathway interactions; Peptide Signal Sequences; Peptides; Population; Process; Protein Isoforms; Protein Secretion; Proteins; Proteomics; RNA; RNA-Binding Proteins; Role; Senile Plaques; Specific qualifier value; Sphingomyelinase; Stress; Synapses; Tauopathies; Testing; To specify; Vesicle; amyloid peptide; base; exosome; glial activation; hyperphosphorylated tau; misfolded protein; novel; protein TDP-43; protein aggregation; protein degradation; protein misfolding; proteostasis; recruit; stress granule; tau Proteins; trafficking

PROJECT SUMMARY Alzheimer's Disease (AD) is characterized by the progressive accumulation of abnormally cleaved A amyloid peptides and hyperphosphorylated tau proteins, which lead to amyloid plaques and neurofibrillary tangles, respectively. While it remains unclear what triggers these proteinopathies, several lines of evidence indicate that defects in intracellular trafficking may regulate AD pathogenesis. Importantly, emerging evidence suggests that AD-related proteins, including tau, amyloid precursor protein (APP) and A amyloid peptides are secreted via exosomes. Despite these findings, it remains unclear what regulates the formation and packaging of exosomes, whether exosome biogenesis is functionally connected to intracellular trafficking of disease-related proteins, whether neurons and glia develop different mechanism to process these proteins, and if so, how abnormal proteostasis in neurons and glia cooperatively promotes neurodegeneration. We have discovered a new pathway in which the autophagy machinery specifies packaging and secretion of proteins within exosomes. Traditionally studied as an autodigestive pathway that promotes cell survival during stress, autophagy also promotes the unconventional secretion of proteins lacking N-terminal signal sequences. Using a proximity-based biotinylation (BioID) proteomics strategy, we have uncovered ~90 novel putative targets of autophagy-dependent secretion, including numerous proteins released within exosomes. These proteins biochemically interact with MAP1LC3B, a mammalian ATG8 isoform and autophagy regulator crucial for cargo sequestration. Based on these results, we hypothesize that the autophagy machinery mediates the LC3-dependent recruitment and packaging of specific intracellular cargo for their secretion via exosomes. Furthermore, we hypothesize that autophagy controls a delicate balance of secretion and intracellular trafficking of disease-relevant proteins in neurons and glia to promote neurodegeneration in AD and FTD. To test these predictions, we will: 1) Dissect whether and how autophagy specifies exosome packaging and secretion in normal and Alzheimer neural cell populations; and 2) Delineate how lysosomal dysfunction in AD and FTD impacts LC3-dependent exosome packaging and proteostasis. These studies are uniquely poised to define new functions for the autophagy machinery in the biogenesis and secretion of exosomes and to delineate its contributions to AD pathogenesis. This multi-PI R01 application synergistically merges the unique expertise of Dr. Jayanta Debnath in the cell biology of autophagy and Dr. Eric Huang in the molecular mechanisms of neurodegenerative diseases to address the goals of RFA-AG-17-051 by uncovering new machineries directing exosome biogenesis and the secretion of exosomal cargo molecules in AD.

项目概要 阿尔茨海默病 (AD) 的特点是异常裂解的 A 淀粉样蛋白逐渐积累 肽和过度磷酸化的 tau 蛋白，导致淀粉样蛋白斑和神经原纤维缠结， 分别。虽然目前尚不清楚是什么引发了这些蛋白质病，但多项证据表明 细胞内运输缺陷可能调节 AD 发病机制。重要的是，新出现的证据表明 AD 相关蛋白，包括 tau、淀粉样前体蛋白 (APP) 和 A 淀粉样肽被分泌 通过外泌体。尽管有这些发现，但仍不清楚是什么调节了形成和包装 外泌体，外泌体生物发生是否与疾病相关的细胞内运输有功能相关 蛋白质，神经元和神经胶质细胞是否发展出不同的机制来处理这些蛋白质，如果是的话，如何 神经元和神经胶质细胞的异常蛋白质稳态共同促进神经退行性变。 我们发现了一条新的途径，自噬机制指定包装和分泌 外泌体中的蛋白质。传统上研究作为促进细胞存活的自消化途径 在压力下，自噬还会促进缺乏 N 末端信号序列的蛋白质的非常规分泌。 使用基于邻近的生物素化 (BioID) 蛋白质组学策略，我们发现了约 90 个新的推定蛋白 自噬依赖性分泌的目标，包括外泌体中释放的许多蛋白质。这些 蛋白质与 MAP1LC3B 发生生化相互作用，MAP1LC3B 是一种哺乳动物 ATG8 亚型，也是自噬调节因子的关键 用于货物扣押。基于这些结果，我们假设自噬机制介导 依赖于 LC3 的特定细胞内货物的募集和包装，以通过外泌体进行分泌。 此外，我们假设自噬控制着分泌和细胞内的微妙平衡。 神经元和神经胶质细胞中疾病相关蛋白的运输促进 AD 和 FTD 的神经变性。到 为了测试这些预测，我们将：1）剖析自噬是否以及如何指定外泌体包装和 正常和阿尔茨海默神经细胞群的分泌； 2) 描述 AD 中的溶酶体功能障碍如何 FTD 影响 LC3 依赖性外泌体包装和蛋白质稳态。这些研究独特地旨在 定义自噬机制在外泌体生物发生和分泌中的新功能 描述其对 AD 发病机制的贡献。该多 PI R01 应用程序协同融合了独特的 Jayanta Debnath 博士在自噬细胞生物学方面的专业知识和 Eric Huang 博士在分子生物学方面的专业知识 通过发现新的神经退行性疾病机制来实现 RFA-AG-17-051 的目标 AD 中指导外泌体生物合成和外泌体货物分子分泌的机制。