Genetic Mechanisms of Myelination in Zebrafish

描述（由申请人提供）：该项目将利用斑马鱼细胞和遗传方法的力量来阐明对脊椎动物神经系统中施万细胞发育和髓鞘形成至关重要的新基因的功能。在周围神经系统中，许旺细胞形成包裹轴突的髓鞘，并允许动作电位的快速传输。髓磷脂的破坏导致周围神经病变，这是一种使美国150万人衰弱的疾病。对控制髓鞘形成的信号的理解上的差距阻碍了有髓鞘轴突修复疗法的发展。在对破坏髓鞘形成的突变进行基因筛选时，我们在10个不同的基因中发现了13个突变，这些基因在有髓鞘轴突的发育中具有特定功能。细胞和分子研究表明，这些突变定义了在有髓轴突发育的许多步骤中发挥作用的基因，包括胶质细胞命运的指定、许旺细胞的迁移、朗维尔结的组织、特定髓鞘mRNA在髓鞘形成胶质细胞内的转运以及许旺细胞对髓鞘形成的承诺。在这个应用程序中，我们专注于一组在雪旺细胞发育和髓鞘形成中具有关键功能的基因。(1)在以前的研究中，我们已经确定了erbb2和erbb3基因的突变，这些基因编码神经调节蛋白（Nrg）信号的异聚体受体的成分，并表明ErbB信号传导对于雪旺细胞的定向迁移是必不可少的。在本申请中，我们提出识别操作信号并确定当雪旺细胞沿生长的外周神经的轴突沿着迁移时它们如何引导雪旺细胞。我们建议确定哪些Nrg亚型引导雪旺细胞迁移，以确定这些信号在发育中的神经中表达的位置，并确定Nrg信号是否足以引导迁移的雪旺细胞到异位位置。(2)在最近的工作中，我们发现我们的两个突变破坏了一种新的跨膜蛋白。我们建议确定蛋白质是否作为一个信号或受体，或两者兼而有之，并测试的假设，该基因在神经元中的行为，以指导雪旺细胞启动髓鞘。此外，我们将测试这一新的跨膜蛋白通过激活雪旺细胞中的krox20来触发髓鞘形成的假设。(3)我们的初步研究表明，st64基因，我们正在努力确定的定位克隆，是必需的许旺细胞髓鞘。我们的表征表明，分析的st64突变将定义一个新的基因在雪旺细胞发育中的重要作用的功能。我们建议通过突变体的表型分析和突变基因的分子分析来确定st64基因的细胞和生化功能。

DESCRIPTION (provided by applicant): This project will exploit the power of cellular and genetic approaches in zebrafish to illuminate the functions of new genes that are essential for Schwann cell development and myelination in the vertebrate nervous system. In the peripheral nervous system, Schwann cells form the myelin sheath that wraps axons and allows for the rapid transmission of action potentials. Disruption of myelin causes peripheral neuropathies, debilitating diseases that affect 1.5 million people in the United States. Gaps in the understanding of the signals that govern the formation of myelin have hindered the development of therapies for the repair of myelinated axons. In genetic screens for mutations that disrupt myelination, we identified 13 mutations in 10 different genes with specific functions in the development of myelinated axons. Cellular and molecular studies demonstrate that the mutations define genes that function at many steps of the development of myelinated axons, including glial fate specification, Schwann cell migration, organization of the nodes of Ranvier, transport of specific myelin mRNAs within myelinating glia, and commitment of Schwann cells to myelination. In this application, we focus on a group of genes that have key functions in Schwann cell development and myelination. (1) In previous studies, we have identified mutations in erbb2 and erbb3, genes that encode components of a heteromeric receptor for Neuregulin (Nrg) signals, and showed that ErbB signaling is essential for directed migration of Schwann cells. In the present application, we propose to identify the operative signals and determine how they direct Schwann cells as they migrate along axons of growing peripheral nerves. We propose to determine which Nrg isoforms guide Schwann cell migration, to determine where these signals are expressed in developing nerves, and to determine whether Nrg signals are sufficient to guide migrating Schwann cells to ectopic locations. (2) In recent work, we found that two of our mutations disrupt a novel transmembrane protein. We propose to determine whether the protein acts as a signal or a receptor, or both, and to test the hypothesis that the gene acts in neurons to instruct Schwann cells to initiate myelination. In addition, we will test the hypothesis that this new transmembrane protein triggers myelination by activating krox20 in Schwann cells. (3) Our preliminary studies show that the st64 gene, which we are working to identify by positional cloning, is required for Schwann cell myelination. Our characterization suggests that analysis of the st64 mutation will define the function of a novel gene with an essential role in Schwann cell development. We propose to define the cellular and biochemical functions of the st64 gene by phenotypic analysis of the mutants and molecular analysis of the mutated gene.