Molecular and genetic decoding of neuron-glial interactions

项目摘要 正常的神经系统功能取决于神经元和非神经元细胞之间的适当通信。 在人脑中，大多数非神经元细胞被称为神经胶质细胞，占脑细胞总数的近一半。 尽管几十年来的工作都集中在神经元和神经胶质上，但它们如何相互交流仍然存在。 不太了解。这部分是由于缺乏方法来全面分析分子， 在动态细胞-细胞相互作用期间在神经元-神经胶质界面富集。该建议旨在制定一项 基因引导的蛋白质组学工具箱，用于时空分析神经胶质细胞富集的关键分子 接口在体内，允许神经元-胶质细胞相互作用的分子和遗传解剖。我们将首先设计 并生成具有时空精度的胶质细胞特异性细胞表面邻近标记探针。我们将应用 少突胶质细胞是中枢神经系统（CNS）中唯一的髓鞘生成细胞， 确定少突胶质细胞轴突鞘化起始的分子机制。我们将 利用这种新方法，沿着单细胞RNA测序，全基因组CRISPR筛选， 转基因小鼠品系，询问一个神秘的细胞阶段（前髓鞘少突胶质细胞）， 发育和适应性髓鞘形成。为了扩展神经胶质表面邻近标记工具包，我们将开发一个 神经元-胶质细胞互补邻近标记系统，其允许瞬时神经元-胶质细胞的可视化 相互作用和捕获的分子仅在体内神经元-神经胶质界面富集。使用这个系统，我们将 解决了控制少突胶质细胞亚群之间髓鞘形成选择性的分子密码， 功能不同的神经元亚群。这项拟议的工作将填补髓鞘生物学的知识空白， 并将在理解神经元-神经胶质和神经胶质-神经胶质相互作用机制方面具有更广泛的意义。 本工作建立的方法和试剂也将极大地丰富研究多种细胞的方法， 细胞通信，包括神经免疫和癌症免疫相互作用。

Project Summary Normal nervous system function depends on proper communications between neurons and non-neuronal cells. In the human brain, the majority of non-neuronal cells are called glia that consist of nearly half of total brain cells. Despite decades of work focusing on neurons and glia alone, how they communicate with each other remains poorly understood. This is partly due to a dearth of methods to comprehensively profile molecules that are enriched at the neuron-glial interface during dynamic cell-cell interactions. This proposal aims to develop a genetically-guided proteomic toolbox to spatiotemporally profile critical molecules enriched at neuron-glial interface in vivo, allowing for molecular and genetic dissection of neuron-glial interactions. We will first design and generate glial-specific cell surface proximity-labeling probes with the spatiotemporal precision. We will apply this platform in the oligodendrocytes, the sole myelin-producing cells in the central nervous system (CNS), to determine the molecular mechanisms governing the initiation of oligodendrocyte-axon ensheathment. We will leverage this new method, along with single-cell RNA sequencing, genome-wide CRISPR screens, and novel transgenic mouse strains, to interrogate a mysterious cell stage (the pre-myelinating oligodendrocytes) during developmental and adaptive myelination. To extend the glial surface proximity labeling toolkit, we will develop a neuron-glial complementary proximity labeling system allowing for visualization of transient neuron-glial interactions and capture of molecules only enriched at neuron-glial interface in vivo. Using this system we will address the molecular codes governing myelination selectivity between subsets of oligodendrocytes and functionally distinct neuronal subpopulations. This proposed work will fulfill the knowledge gap in myelin biology, and will have broader implications in understanding neuron-glial and glia-glial interaction mechanisms in general. The methods and reagents established by the work will also greatly enrich the methods in studying diverse cell- cell communications, including neuro-immune and cancer-immune interactions.