Defective lysosome acidification in dystrophic neurites in Alzheimer's disease contributes to failure of autophagy

阿尔茨海默病营养不良的神经突中溶酶体酸化缺陷导致自噬失败

基本信息

批准号：
10670489
负责人：
Matthew Schrag
金额：
$ 73.88万
依托单位：
VANDERBILT UNIVERSITY MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10670489
关键词：
3-Dimensional Adult Alzheimer&apos s Disease Alzheimer&apos s disease brain Alzheimer&apos s disease model Alzheimer&apos s disease related dementia Alzheimer&apos s disease risk Amyloid Amyloid beta-Protein Animal Model Autophagocytosis Axon Biological Models Brain Catalytic Domain Cells Clinical Clinical Research Cognition Cognitive Cognitive aging Complex Crowding Custom Cytosol Data Data Set Defect Disease Drug Targeting Embryo Ensure Enzymes Epidemic Evaluation Failure Functional disorder Future Genes Genetic Human Hydrogels Image Impaired cognition Impairment In Vitro Individual Ions Lead Link Lysosomes Measures Mediating Membrane Memory Metabolic Methods Microscopy Molecular Molecular Target Neurites Neurobiology Neurons Organelles Outcome Pathologic Pathology Pathway interactions Pattern Peptides Point Mutation Production Proteins Proteomics RNA-Binding Proteins Regulation Research Risk Factors Role Senile Plaques Slice Structure Techniques Ubiquitination Variant Work base cell injury data streams design drug development drug discovery druggable target effective therapy genetic risk factor golgin human model in vitro Model induced pluripotent stem cell loss of function molecular drug target mouse model neuropathology next generation novel pre-clinical protein aggregation proteostasis religious order study risk variant tau Proteins tau aggregation transcriptome sequencing transcriptomics vacuolar H+-ATPase

项目摘要

Alzheimer’s disease is a national crisis and the burden of this terrible disease is rising. After decades of failure to produce a disease modifying treatment for Alzheimer’s disease, it is critical that we identify drug targets that are linked to causal mechanisms underlying the diverse neuropathological substrates of Alzheimer’s disease. Our research focuses on the function of lysosomes within an axonal structure called a dystrophic neurite. Dystrophic neurites form when axon segments pass near a β-amyloid plaque and become distended and clogged with lysosomes and related organelles. Tau aggregates are also present in some dystrophic neurites, so this pathology seems to link the major pathological hallmarks of Alzheimer’s disease. Because lysosomes are critical for protein homeostasis, we hypothesize that loss of function of these lysosomes is a key step in the formation of toxic protein aggregates. We demonstrated a pattern of changes in the function this pool of lysosomes which could be explained by failure of acidification, so in this proposal we will evaluate 1) whether lysosomes in dystrophic neurites are able to acidify and what molecular mechanisms may underlie failed acidification, 2) whether loss of lysosome acidification results in defective autophagy and accumulation of pathologic autophagic intermediates within dystrophic neurites and 3) the degree to which a novel Alzheimer’s disease genetic risk factor, RBFOX1, contributes to altered autophagy and pH regulation in neuronal lysosomes. For each of these aims, we leverage advanced in vitro methods including organotypic brain slice cultures from adult model animals and three-dimensional human neuronal cultures derived from induced pluripotent stem cells grown in a custom designed hydrogel matrix. Using live-imaging, we will quantitatively measure the pH in lysosomes in dystrophic neurites. We will conduct detailed neuropathological studies simultaneously in human and mouse model systems to identify altered distribution of vacuolar ATPase subunits, ARL1 and Golgin A4 which are key regulators of autophagy, and RBFOX1. Additionally, we want to understand the cognitive and neuropathological associations of each component of the molecular pathways that contribute to failed acidification, defective autophagy and tau aggregation, so we will leverage RNAseq and proteomic data from large clinical studies, in particularly the Religious Orders Study/Memory and Alzheimer’s Project, to ensure these pathways are functionally important. These synergistic streams of data will converge to robustly establish the mechanism and consequence of loss of lysosome acidification in dystrophic neurites and identify molecular targets for future drug discovery efforts.

阿尔茨海默病是一场全国性的危机，这种可怕疾病的负担正在不断增加。几十年来未能研制出治疗阿尔茨海默病的疾病治疗方法，这一点至关重要我们确定与多种潜在因果机制相关的药物靶点我们的研究重点是阿尔茨海默病的神经病理学底物的功能。轴突结构内的溶酶体称为营养不良性神经突，当营养不良性神经突形成时。轴突片段经过 β-淀粉样斑块附近并变得遥远并被溶酶体和相关细胞器也存在于一些营养不良者中。神经突，因此这种病理学似乎与阿尔茨海默病的主要病理特征有关由于溶酶体对于蛋白质稳态至关重要，因此我们考虑了溶酶体的损失。这些溶酶体的功能是形成有毒蛋白质聚集体的关键步骤。证明了该溶酶体库的功能变化模式，这可能是解释为酸化失败，因此在本提案中，我们将评估 1）溶酶体是否营养不良的神经突能够酸化以及酸化失败的分子机制是什么酸化，2）溶酶体酸化的丧失是否会导致自噬缺陷和营养不良的神经突内病理性自噬中间产物的积累，3) 一种新的阿尔茨海默病遗传风险因素 RBFOX1 在多大程度上导致了阿尔茨海默病的改变神经元溶酶体中的自噬和 pH 调节对于每个目标，我们都加以利用。先进的体外方法，包括成年模型动物的器官型脑切片培养以及源自诱导多能干细胞的三维人类神经培养物在定制设计的水凝胶基质中生长，我们将使用实时成像进行定量测量。我们将进行详细的神经病理学研究。同时在人类和小鼠模型系统中识别液泡的分布 ATP 酶亚基、ARL1 和高尔金 A4（它们是自噬的关键调节因子）以及 RBFOX1。此外，我们想了解每种疾病的认知和神经病理学关联导致酸化失败、自噬缺陷的分子途径的组成部分和 tau 聚合，因此我们将利用来自大型临床研究的 RNAseq 和蛋白质组数据，特别是宗教秩序研究/记忆和阿尔茨海默氏症项目，以确保这些这些协同数据流在功能上非常重要。建立营养不良患者溶酶体酸化丧失的机制和后果神经突并确定未来药物发现工作的分子靶标。