Molecular Characterization of Axon-Glial Interactions

描述（由申请人提供）：多种衰弱的人类神经系统疾病是由神经胶质相互作用的功能障碍引起的，如多发性硬化症等疾病。神经元和神经胶质之间复杂的相互作用构成了跨物种轴突嵌套和轴突胶质分离连接形成的潜在基础。我们之前对果蝇和脊椎动物的研究已经证实，存在于有髓鞘轴突和果蝇周围神经系统中的AGSJs的分子组成依赖于三个关键的同源蛋白：CASPR[果蝇Neurexin IV (NRX IV)]、Contactin （CONT）和有髓鞘胶质神经束蛋白（NF155）。我们最近的研究表明，NRX IV在胚胎中枢神经系统的轴突-胶质相互作用和轴突嵌套中起重要作用。在这里，NRX IV与两种含有中线胶质（MLG）特异性免疫球蛋白（Ig）结构域的蛋白，Wrapper （WRAP）和KIRRE（脊椎动物Nephrins的同源物）反式相互作用，以建立神经元-胶质支架和AGSJs。我们已经生成了一个令人兴奋的新工具，使我们能够以一种前所未有的方式可视化MLG的形态学和解剖学特征。这些形态学和解剖学特征与脊椎动物少突胶质细胞相似。尽管它们作为引导轴突的信号中心具有重要意义，但令人惊讶的是，人们对MLG在轴突嵌套中的作用知之甚少，也不知道MLG如何作为一个简单而强大的模型来理解少突胶质细胞生物学。为了进一步推动果蝇MLG和脊椎动物少突胶质细胞可能使用共同的遗传程序和分子原理的观点，我们进行了第一次向前遗传筛选，以确定控制MLG形态发生的新的额外分子参与者，并可能控制中枢神经系统中的轴突嵌套。我们在整个果蝇基因组中筛选了约550个个体染色体缺陷系，并鉴定了一些可能在MLG形态发生和轴突鞘鞘中起作用的新基因。这些基因中有许多在少突胶质细胞中表达的脊椎动物同源物，但没有在任何系统中对这些基因进行详细的体内功能分析。该提案旨在利用果蝇中可用的强大的遗传，分子和生化技术，以及我们已经确定的新基因和我们已经生成的工具，来解决与神经胶质生物学和轴突嵌套相关的关键问题。本研究的具体目的是：(1)果蝇中枢神经系统中MLG形态发生、轴突-胶质相互作用和轴突嵌套的解剖学和分子基础是什么？(2)控制MLG形态发生的遗传和分子途径是什么？它们与脊椎动物少突胶质细胞形态发生的关系如何？我们期望我们的工作将为体内调节神经胶质形态发生和神经元-神经胶质相互作用的分子和细胞机制提供令人兴奋的新见解，并将告诉我们在脊椎动物髓鞘胶质细胞分化中运作的类似机制。

DESCRIPTION (provided by applicant): A diverse array of debilitating human neurological conditions result from dysfunction in neuron-glial interactions, as seen in diseases like Multiple Sclerosis. The intricate interactions between neurons and glia form the underlying basis of axonal ensheathment and formation of axo-glial septate junctions (AGSJs) across species. Our previous studies using Drosophila and vertebrates have established that the molecular make up of AGSJs, present in myelinated axons and fly peripheral nervous system are dependent on three key homologous proteins: CASPR [Neurexin IV (NRX IV) in flies], Contactin (CONT) and myelinating glial Neurofascin (NF155). Our more recent studies reveal that NRX IV plays an important role in axon-glial interactions and axonal ensheathment in the embryonic CNS. Here NRX IV interacts in trans with two midline glia (MLG)-specific immunoglobulin (Ig) domain containing proteins, Wrapper (WRAP) and KIRRE (a homolog of vertebrate Nephrins) to establish neuron-glial scaffolds and AGSJs. We have generated an exciting new tool that allows us to visualize the morphological and anatomical features of MLG in a way not seen before. These morphological and anatomical characteristics of MLG resemble those displayed by vertebrate oligodendrocytes. Despite their fundamental importance as signaling centers for guiding axons, surprisingly little is known about the role of MLG in ensheathment of axons and how MLG might serve as a simple, yet powerful model for understanding oligodendrocyte biology. In order to further provide impetus to the idea that Drosophila MLG and vertebrate oligodendrocytes might employ common genetic programs and molecular principles, we carried out the first forward genetic screen to identify new additional molecular players that control MLG morphogenesis and, potentially, axonal ensheathment in the CNS. We screened ~550 individual chromosomal deficiency lines across the entire Drosophila genome and identified a number of novel genes that potentially function in MLG morphogenesis and axonal ensheathment. Many of these genes have vertebrate homologs that are expressed in oligodendrocytes, but no detailed in vivo functional analysis of these genes has been carried out in any system. This proposal aims to use the powerful array of genetic, molecular and biochemical techniques available in Drosophila, together with the new genes that we have identified and tools we have generated, to address key questions related to glial biology and axonal ensheathment. The specific aims of this proposal are: (1) what are the anatomical and molecular bases of MLG morphogenesis, axon-glial interactions and axonal ensheathment in the Drosophila CNS? and (2) what are the genetic and molecular pathways that control MLG morphogenesis and how they relate to vertebrate oligodendrocyte morphogenesis? We expect that our work will provide exciting new insights into the molecular and cellular mechanisms regulating glial morphogenesis and neuron-glial interactions in vivo, and will inform us about similar mechanisms that operate in the differentiation of vertebrate myelinating glial cells. PUBLIC HEALTH RELEVANCE: Genetic and molecular mechanisms that govern glial morphogenesis, axonal ensheathment, establishment of neuro-glial scaffolds and organization of specialized electron-dense structures, the axo-glial septate junctions (AGSJs) are of utmost importance to human neuronal function. AGSJs coordinate organization of axonal domains and allow myelinated axons to propagate nerve impulses in a saltatory manner. Better understanding of the fundamental mechanisms that underlie axonal ensheathment and organization of axonal domains is critical in designing future therapeutic strategies to myelin-related diseases or demyelination disorders like for example multiple sclerosis (MS) where remyelination is required and the axonal domain structure must be preserved to allow action potential propagation and neuronal function.