A quantitative framework for understanding endosomal trafficking networks in Alzheimer's disease

了解阿尔茨海默氏病内体运输网络的定量框架

• 批准号: 10241471
• 负责人: JEFFREY W HARPER
• 金额: $ 45.94万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10241471
• 关键词: Abeta clearance; Abeta synthesis; Affect; Alzheimer' s Disease; Amyloid beta-Protein; Architecture; Autophagocytosis; Biochemical; Biological; Biological Models; Biotinylation; CRISPR screen; Candidate Disease Gene; Cell Line; Cell membrane; Cells; Defect; Degradation Pathway; Dependence; Disease; Event; Excision; Genes; Genetic; Goals; Golgi Apparatus; Image; Individual; Laboratories; Link; Lysosomes; Maps; Membrane Proteins; Methodology; Mitochondria; Molecular; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Organelles; PINK1 gene; Pathogenesis; Pathway Analysis; Pathway interactions; Predisposition; Prevention; Process; Proteins; Proteomics; Quality Control; Recycling; Reporter; Role; Sorting - Cell Movement; Specificity; System; Testing; Therapeutic; Time; Toxic effect; Ubiquitin; Validation; Work; abeta accumulation; abeta toxicity; arm; base; cell type; design; experimental study; gain of function mutation; genetic analysis; human embryonic stem cell; insight; late endosome; multicatalytic endopeptidase complex; mutant; neuropathology; novel; protein degradation; protein misfolding; protein transport; proteostasis; repaired; screening; tissue/cell culture; trafficking

PROJECT SUMMARY Accumulating evidence suggest that neurodegenerative diseases including Alzheimer’s disease (AD) are frequently the result of alterations in endosomal trafficking and proteostasis pathways, one consequence of which can be the accumulation of aggregation-prone proteins. Numerous familial and sporadic loss or gain-of- function mutations have been identified in such pathways, illuminating potential drivers of disease pathogenesis. In addition, excess protein mis-folding due to altered trafficking could affect proteostasis pathways by blocking protein turnover or trafficking. Protein and organelle trafficking within cells is a highly dynamic and interconnected process, and defects in one arm of the system can affect other aspects of the network in unpredictable ways including reduced flux in turnover pathways. Indeed, it is conceivable that many seemingly unrelated mutations across the trafficking landscape in various neurodegenerative diseases reveal common mechanistic vulnerabilities downstream but with distinct cell-type sensitivities reflective of the identity of mis-trafficked proteins. As such, understanding the global architecture of trafficking systems and the key machinery that controls the directionality and efficiency of trafficking, particularly of aggregation-prone neurodegenerative proteins such as APP and its aggregation-prone form A, represents a central goal of the field. A aggregation as a toxic driver of AD neuropathology has been a dominant hypothesis in the field. However, thus far therapeutics directed at aggregate prevention or removal have not been successful, and alternative hypotheses including alterations in intracellular trafficking as an important event in neuropathology have emerged. Here, we seek to combine powerful genetic and proteomic approaches to develop a quantitative framework for understanding how disruption of major endosomal trafficking systems – retromer and retriever, found defective in neurodegenerative diseases – alter global membrane protein trafficking, and specific trafficking and processing of APP proteoforms. These studies make use of an extensive tool-kit of mutant tissue culture cell lines and induced neurons derived from human embryonic stem cell (hESC), in combination with targeted and unbiased proteomics of individual organelles linked with endosomal trafficking, to assemble a global map of cargo and trafficking dependencies. In parallel, we will employ novel flux-based screening strategies to search for genes controlling APP/A trafficking to the lysosome and the plasma membrane, and will examine the extent to which A accumulation within the endo-lysosomal system alters selective autophagic flux using new cargo-specific reporters. The central hypothesis being tested is that specific defects in protein trafficking networks underlies the susceptibility of neurons to A and other aggregation prone proteins and that these defects can be molecularly unmasked through systematic network and genetic analysis.

项目总结 越来越多的证据表明，包括阿尔茨海默病(AD)在内的神经退行性疾病 通常是内体运输和蛋白平衡途径改变的结果，其中一个后果是 这可能是聚集倾向的蛋白质的积累。许多家族性和零星的损失或收益 已经在这些通路中发现了功能突变，从而阐明了潜在的疾病驱动因素 发病机制。此外，由于运输改变而导致的过量蛋白质错误折叠可能会影响蛋白平衡。 通过阻止蛋白质周转或运输的途径。蛋白质和细胞器在细胞内的运输是一种高度 动态的和相互关联的过程，系统的一个臂中的缺陷可能会影响到 网络以不可预测的方式出现，包括周转路径上的流量减少。事实上，可以想象，许多人 各种神经退行性疾病的贩运格局中看似无关的突变揭示 下游常见的机械漏洞，但具有反映身份的不同细胞类型敏感度 被非法贩卖的蛋白质。因此，了解贩运系统的全球架构和关键 控制贩运的方向性和效率的机制，特别是易于聚集的贩运 神经退行性蛋白，如APP及其易于聚集的A，代表了 菲尔德。聚集作为AD神经病理的有毒驱动因素一直是该领域的主导假说。 然而，到目前为止，针对集体预防或清除的治疗尚未成功，并且 替代假说，包括细胞内转运的改变是神经病理学中的一个重要事件 已经出现了。在这里，我们寻求结合强大的遗传学和蛋白质组学方法来开发一种 理解主要内体转运系统如何被破坏的定量框架--回溯 和寻回犬，被发现患有神经退行性疾病-改变全球膜蛋白运输，以及 APP蛋白形式的具体贩运和加工。这些研究利用了一个广泛的工具包 人胚胎干细胞(HESC)突变型组织培养细胞系及诱导神经元 结合与内体贩运有关的单个细胞器的定向和无偏倚蛋白质组学， 汇编一张货物和贩运依赖关系的全球地图。同时，我们将采用新的基于助熔剂的 寻找控制APP/A转运到溶酶体和血浆的基因的筛选策略 膜，并将检查在内溶酶体系统内积累的改变程度 使用新的货物专用记者的选择性自噬通量。正在测试的中心假设是 蛋白质运输网络中的特定缺陷是神经元对A和其他 容易聚集的蛋白质，这些缺陷可以通过系统网络在分子上揭示出来 和基因分析。