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

基本信息

批准号：
9686111
负责人：
JEFFREY W HARPER
金额：
$ 48.23万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
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/9686111
关键词：
Abeta clearance Abeta synthesis Affect Alzheimer&apos 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 tool trafficking

项目摘要

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.

项目概要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是通常是内体运输和蛋白质稳态途径改变的结果，这是以下因素的结果之一：这可能是许多家族性和零星性损失或增加的易聚集蛋白质的积累。在这些途径中已经发现了功能突变，阐明了疾病的潜在驱动因素此外，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质稳态。通过阻断细胞内蛋白质和细胞器运输的途径是高度有效的。动态且相互关联的过程，系统某一部分的缺陷可能会影响系统的其他方面网络以不可预测的方式，包括周转路径的流量减少，确实，可以想象，许多。各种神经退行性疾病中贩运景观中看似无关的突变揭示了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性因此，了解贩运系统的全球架构和关键。控制贩运方向和效率的机制，特别是易于聚集的贩运方向和效率神经退行性蛋白，例如 APP 及其易于聚集的形式 A，代表了该研究的中心目标 A 聚集作为 AD 神经病理学的毒性驱动因素一直是该领域的主导假设。然而，迄今为止，针对总体预防或消除的治疗尚未成功，并且其他假设，包括细胞内运输的改变作为神经病理学的重要事件在这里，我们寻求结合强大的遗传和蛋白质组学方法来开发一种新的方法。用于了解主要内体运输系统如何被破坏的定量框架 – Retromer 和猎犬，被发现存在神经退行性疾病缺陷——改变全球膜蛋白运输，以及这些研究利用了广泛的工具包。源自人胚胎干细胞 (hESC) 的突变组织培养细胞系和诱导神经元，与与内体运输相关的单个细胞器的有针对性和公正的蛋白质组学相结合，同时，我们将采用基于通量的新型方法来构建全球货物和贩运依赖性地图。寻找控制 APP/A 运输至溶酶体和血浆的基因的筛选策略膜，并将检查 A 在内溶酶体系统内的积累改变的程度使用新的货物特异性生产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 和其他物质的易感性的基础易于聚集的蛋白质，并且这些缺陷可以通过系统网络从分子上揭示和遗传分析。