BIN1-interactome in Alzheimer's disease pathophysiology

BIN1-相互作用组在阿尔茨海默病病理生理学中的作用

基本信息

批准号：
10677190
负责人：
Joseph McMillan
金额：
$ 3.79万
依托单位：
UNIVERSITY OF SOUTH FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-14 至 2025-08-13
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10677190
关键词：
Address Adult Affect Age Alternative Splicing Alzheimer&apos s Disease Alzheimer&apos s disease model Alzheimer&apos s disease pathology Alzheimer&apos s disease patient Alzheimer&apos s disease risk American Amyloidosis Bioinformatics Biological Process Biotin Biotinylation Brain Brain Diseases Cellular biology Clathrin Cluster Analysis Complement Core Facility Coupled Data Analyses Development Disease Dynamin Elderly Endocytosis Environment Enzymes Experimental Designs Functional disorder Future Genes Genetic Gliosis Health Human Immunofluorescence Immunologic Immunoprecipitation In Vitro Interdisciplinary Study Investigation Kinetics Knowledge Label Laboratories Late Onset Alzheimer Disease Learning Ligase Link Mass Spectrum Analysis Mediating Mentors Methods Mus Muscle Development Nature Neighborhoods Neuroblastoma Neurodegenerative Disorders Neurons Pathogenesis Pathogenicity Pathology Pathway interactions Protein Isoforms Proteins Proteome Proteomics Radial Recycling Research Research Training Retrieval Risk Risk Factors SYNJ1 gene Severities Skeletal Muscle Synapses Synapsins Synaptic Vesicles System Therapeutic Translating Translations Tumor Suppressor Proteins Wild Type Mouse Work age related neurodegeneration anticancer research apolipoprotein E-4 brain tissue detection method experimental study genetic risk factor genome wide association study grasp gray matter human old age (65+)in vivo innovation interest lifestyle factors molecular pathology mouse model neuroblastoma cell neuron loss novel pre-doctoral promoter protein protein interaction protein purification superresolution microscopy tau Proteins tool vesicular release

项目摘要

PROJECT SUMMARY Alzheimer’s disease (AD) is a devastating neurodegenerative disorder with no definitive treatment that reverses the course of the disease, and we still lack a firm grasp on how older people develop AD. Bridging Integrator 1 (BIN1) is the second-largest genetic risk factor for late-onset AD. At least 12 different alternatively spliced BIN1 isoforms are expressed in the brain, including the neuron-specific BIN1 isoform 1 (BIN1iso1) and the ubiquitously expressed BIN1 isoform 9 (BIN1iso9). In the brain gray matter of patients with AD, there is a decrease in neuronal BIN1iso1 and an increase in BIN1iso9 compared to healthy controls. Thus far, evidence has shown that neuronal BIN1 isoforms participate in clathrin-mediated endocytosis, endocytic recycling, and synaptic vesicle release and retrieval. However, the mechanisms by which BIN1 contributes to these functions and the neighborhood of proteins that BIN1 interacts with to accomplish these cellular tasks remain largely undefined. Therefore, a fundamental gap in the field is an unbiased characterization of BIN1iso1 interacting proteins and proximal neighbors. Closing this gap will help define BIN1’s biological functions in healthy and diseased brain neurons. Utilizing the highly innovative proximity biotin ligase, TurboID, fused to BIN1iso1, will allow the identification of all proteins within a 10-nm radius. TurboID-based proximity labeling coupled with the most recent advanced quantitative mass spectrometry and data analysis methods represents a powerful strategy for discovery research. My preliminary in vitro studies using this approach in mouse N2a neuroblastoma cells resulted in the discovery of 234 proteins as BIN1-associated (proximal) or interacting proteins. These results identified several known BIN1 interactors such as tau, dynamin, synaptojanin, and many previously unknown proximal proteins. The following Specific Aims will translate these findings in vivo and dramatically advance the field. Aim 1. Identify neuronal BIN1iso1 interacting proteins in vivo using wild-type mice under homeostatic conditions. Aim 2. Establish neuronal BIN1iso interactome in mouse models of AD (5XFAD and PS19) before and after the onset of pathology. This project will not only advance the field by providing those studying AD and BIN1 with a list of BIN1iso1 proximal proteins to generate novel hypotheses but will also support the applicant’s pre-doctoral research training in AD pathophysiology, advanced methods such as in vivo AAV transduction, proximity-based labeling in the context of AD pathology, large-scale proteomics data analysis, and bioinformatics analysis of BIN1 functional pathways. The dynamic and highly collaborative research environment at the USF Health Byrd Alzheimer’s Institute will enhance the learning opportunities in cell biology and molecular pathology within an established AD laboratory led by a committed mentor. Furthermore, research training in proteomics approaches offered by the collaborator’s lab and the Proteomics Core facilities will complement the applicant’s interdisciplinary research training in age-related neurodegenerative disease.

项目摘要阿尔茨海默氏病（AD）是一种毁灭性的神经退行性疾病，没有明确治疗疾病的过程，我们仍然对老年人如何发展广告缺乏坚定的掌握。桥接整合器1 （BIN1）是晚发广告的第二大遗传危险因素。至少12个不同的剪接Bin1 同工型在大脑中表达，包括神经特异性BIN1同工型1（BIN1ISO1）和无处不在的表达的BIN1同工型9（BIN1ISO9）。在AD患者的脑灰质中，神经元降低与健康对照相比，BIN1ISO1和BIN1ISO9的增加。远的证据表明神经元参与网格蛋白介导的内吞作用，内吞回收和突触囊泡释放的BIN1同工型检索。但是，BIN1促进这些功能的机制以及 BIN1与完成这些细胞任务相互作用的蛋白质在很大程度上不确定。因此，该领域的基本差距是BIN1ISO1相互作用蛋白和近端邻居。缩小此差距将有助于定义BIN1在健康和患病的大脑中的生物学功能神经元。利用高度创新的接近生物素连接酶，涡轮增压，融合到bin1iso1，将允许鉴定10 nm半径内的所有蛋白质。基于涡轮的接近标签以及最新的高级定量质谱和数据分析方法代表了一种强大的策略发现研究。我在小鼠N2A神经母细胞中使用这种方法的初步体外研究导致发现234种蛋白作为BIN1相关（近端）或相互作用的蛋白质。这些结果确定了几个已知的BIN1相互作用者，例如Tau，Dynamin，Synaptojanin，以及许多以前未知的近端蛋白质。以下特定目标将在体内转化这些发现，并急剧地转化推进领域。 AIM 1。使用野生型小鼠在体内识别Neuronal Bin1ISO1相互作用的蛋白稳态条件。 AIM 2。在AD的小鼠模型中建立神经元bin1iso Interactome（5XFAD和 PS19）病理发作前后。该项目不仅将通过提供这些项目来推进该领域使用BIN1ISO1近端蛋白列表来研究AD和BIN1，以产生新的假设，但也将支持申请人在AD病理生理学的博士前研究培训，例如In Vivo AAV等高级方法在AD病理学，大规模蛋白质组学数据分析和 BIN1功能途径的生物信息学分析。动态和高度协作的研究环境阿尔茨海默氏症研究所在USF健康，将增强细胞生物学和分子的学习机会由坚定的心理领导的既定广告实验室内的病理。此外，研究培训合作者实验室提供的蛋白质组学方法和蛋白质组学核心设施将完成申请人对年龄相关的神经退行性疾病的跨学科研究培训。