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，阐明在脊椎动物神经系统中对雪旺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，雪旺细胞形成髓鞘，髓鞘包裹轴突，并允许动作电位的快速传递。髓磷脂的破坏会导致周围神经病变，这种使人衰弱的疾病影响着美国150万人。对控制髓磷脂形成的信号的理解的空白阻碍了髓鞘轴突修复治疗的发展。在破坏髓鞘形成的突变的基因筛选中，我们在10个不同的基因中发现了13个突变，这些基因在髓鞘轴突的发育中具有特定的功能。细胞和分子研究表明，这些突变决定了在髓鞘轴突发育的许多步骤中起作用的基因，包括胶质命运规范、雪旺细胞迁移、Ranvier节点的组织、髓鞘胶质内特定髓磷脂mrna的运输以及雪旺细胞对髓鞘形成的承诺。在这个应用中，我们关注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)在之前的研究中，我们已经发现了编码神经调节蛋白（Nrg）信号异质受体成分的erbb2和erbb3基因的突变，并表明ErbB信号传导对雪旺细胞的定向迁移至关重要。在目前的应用中，我们建议识别操作信号，并确定它们如何指导雪旺细胞沿生长的周围神经轴突迁移。我们建议确定哪些Nrg亚型引导雪旺细胞迁移，确定这些信号在发育中的神经中的表达位置，并确定Nrg信号是否足以引导雪旺细胞迁移到异位位置。(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。我们建议确定该蛋白是否作为信号或受体，或两者兼而有之，并测试该基因在神经元中指导雪旺细胞启动髓鞘形成的假设。此外，我们将测试这种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基因是雪旺细胞髓鞘形成所必需的，我们正在通过定位克隆进行鉴定。我们的研究表明，对st64突变的分析将定义一个在雪旺细胞发育中起重要作用的新基因的功能。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。公共卫生相关性：髓鞘的破坏导致周围神经病变，使人衰弱的疾病，影响150万人在美国。该项目的长期目标是利用在斑马鱼中可用的强大的遗传和细胞方法，发现在髓磷脂形成中具有重要功能的新基因。新基因的发现和新动物模型的建立将是髓磷脂修复治疗周围神经病变和其他髓磷脂疾病的重要一步。

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. PUBLIC HEALTH RELEVANCE: Disruption of the myelin sheath causes peripheral neuropathies, debilitating diseases that affect 1.5 million people in the United States. The long-term goal of this project is to discover new genes with essential functions in myelin formation using the powerful genetic and cellular approaches available in the zebrafish. The discovery of new genes and the development of new animal models will be an important step toward myelin repair therapies for peripheral neuropathies and other diseases of myelin.