JPND - Intraneuronal transport-related pathways across neurodegenerative diseases

该方案的总体目标是确定Abeta1-42、Tau、α -突触核蛋白或HTTExon 1不同聚集体扩散的途径/蛋白质网络，这些聚集体分别与阿尔茨海默病、帕金森病和亨廷顿病有关。病变的扩散是所有这些疾病的一个共同特征，但引人注目的是，每种病理在不同的脑次区域开始和发展。根据最近的观察，不同的聚集构象可能导致不同的临床亚型。在这里，我们旨在揭示Abeta1-42、Tau、α -突触核蛋白和HTTExon 1不同的纤维蛋白多态性被神经元摄取和运输的具体机制。我们将使用在原代培养中维持的不同类型的小鼠神经元，通过分析它们在细胞外应用后的结合、摄取和细胞运输，来建立不同致病蛋白组装及其构象的结构-功能关系。通过这样做，我们将解决：1 .不同的致病性原纤维蛋白多态性如何与神经元细胞结合，以及与哪些膜组分结合；2 .外源性致病原纤维蛋白如何被摄取和运输；3 .致病性原纤维蛋白如何通过识别在细胞内运输；4 .致病性原纤维蛋白的转运与神经元活动的关系；5 .解释不同疾病的蛋白质网络；6 .遗传危险因素如何影响不同致病性原纤维蛋白多态性的相互作用组和转运机制；共同途径的识别作为新的治疗靶点。更具体地说，我们将量化这些不同致病性纤维蛋白多态性与神经元质膜的结合；*识别它们的“受体”或膜蛋白伴侣；*评估内吞作用；*识别细胞内的货物；*定量细胞内运输及其在顺行或逆行运输后在细胞外介质中的输出/释放，以确定导致不同疾病的共同和不同的蛋白质网络和特征。选择了该项目的不同合作伙伴，因为他们的专业知识涵盖了这些具体任务。* LAC/ENS Paris-Saclay团队将应用其最近的神经元内运输测定，依靠荧光纳米金刚石跟踪。* neuropsi和CNCR团队在相互作用蛋白质组学方面具有专业知识，可以揭示在不同运输步骤中具有功能相似性和特异性的蛋白质。*该联盟还将利用FP7-HEALTH AgedBrainSYSBIO联盟（由LAC/ENS Paris-Saclay协调）最近获得并验证的新型小鼠转基因LOAD模型、PD模型和halo标记敲入模型，以确定与神经元内运输相关的共同潜在机制。*这些读数将用于建立总量贩运的定量模型，并利用商业智能合作伙伴和MSSM作为外部合作伙伴的专业知识确定共同途径。

The overall goal of this proposal is to identify pathways/protein networks underlying the spreading of distinct aggregates of Abeta1-42, Tau, alpha-synuclein or HTTExon 1 involved in Alzheimer disease, Parkinson's disease, and Huntington disease, respectively. The spread of the lesions is a common feature of all these diseases, but strikingly starts and progresses in different brain sub-regions for each pathology. According to recent observations distinct aggregate conformers could be responsible for different clinical subtypes. Here, we aim at revealing the specific mechanisms by which distinct fibrillary protein polymorphs of Abeta1-42, Tau, alpha-synuclein and HTTExon 1, are taken up and transported by neurons. We will use different types of mouse neurons maintained in primary culture to establish a structure-function relationship for distinct pathogenic protein assemblies and their conformers by analyzing their binding, uptake and cellular trafficking after extracellular application. By doing so we will address:By doing so we will address:1. How distinct pathogenic fibrillary protein polymorphs bind to neuronal cells and to which membrane components;2. How exogenous pathogenic fibrillary proteins are taken up and transported;3. How pathogenic fibrillary proteins traffic within cells by identifying;4. The relationship between the traffic of pathogenic fibrillary proteins and neuronal activity;5. The protein networks that account for distinct diseases;6. How genetic risk factors affect the interactomes and transport mechanisms of distinct pathogenic fibrillary proteins polymorphs;7. The identification of common pathways involved as novel therapeutic targets.More specifically, we will* quantify the binding of these distinct pathogenic fibrillary proteins polymorphs to the neuronal plasma membrane;* identify their "receptors" or membrane protein partners;* assess endocytosis;* identify their cargoes within the cells;* quantify intracellular transport and their export/release in the extracellular medium after anterograde or retrograde transport to identify common and divergent protein networks and characteristics that account for distinct diseases.The different partners of this project have been selected for their expertise to cover these specific tasks.* The LAC/ENS Paris-Saclay team will apply its recent intraneuronal transport assay, relying on fluorescent nanodiamond tracking.* Neuro-PSI and CNCR teams have expertise in interaction proteomics to reveal proteins with functional similarities and specificities in the different trafficking steps.* The consortium will also take advantage of novel mouse transgenic LOAD models, PD models and Halo-tagged-knockin models recently obtained and validated by the FP7-HEALTH AgedBrainSYSBIO consortium (coordinated by LAC/ENS Paris-Saclay) to identify common underlying mechanisms linked to intraneuronal transport.* These readouts will be used to build quantitative models of aggregates trafficking and identify common pathways, using the expertise of the BI partner and of the MSSM as external partner.