LAMININ AND FIBRONECTIN IN PERIPHERAL NERVE DEVELOPMENT

拟议研究的长期目标是了解 细胞外基质（ECM）蛋白质纤维连接蛋白（FN）和层粘连蛋白（LAM） 周围神经系统（PNS）的发展。 ECM分子调节 胚胎发生过程中的细胞形状、分化和运动。 在 PNS、ECM分子似乎参与神经嵴迁移， 神经肌肉接头的发育和再生。 生长锥 从神经管或周围神经节迁移的神经纤维穿过 也可能含有粘合剂分子的底物途径。 的 粘附糖蛋白FN和LAM是介导 PNS细胞迁移和分化的特定阶段，由于其 在胚胎中的位置和与体外PNS细胞的相互作用。 首先， FN和LAM的功能区介导神经元-基底层粘附， 神经突延伸 新生力量的两个地区开展了这些活动， 揭示了与FN分子的神经元类型特异性相互作用。 LAM的功能区域不太明确，但也可能促进细胞凋亡。 特定类型的行为。 将使用FN和LAM的蛋白水解片段， 结合抗体，以评估1）功能差异 FN与细胞相互作用的区域之间，2）细胞表面是否 硫酸乙酰肝素介导这些相互作用中的某些，以及3）每种相互作用如何 结构域有助于介导的细胞行为的完整的FN 分子。 此外，生长锥粘附在神经突中的作用 将研究FN片段上的延伸。 第二，FN和LAM（以及 特定区域的每一个）所使用的细胞在发展PNS？ 方法 将是1）免疫细胞化学，以揭示时间和空间关联 2）免疫标记TEM和SEM， 鉴定产生LAM和FN的细胞及其与生长的接近程度 锥体，和3）抑制，与抗体，神经突延伸上 天然存在的基质和体内，以探测 每个分子 第三，FN和LAM的定义区域如何有助于 神经嵴细胞的迁移和分化 做嵴细胞 亚群附着在这些区域的某些区域上和/或在这些区域上移动， 在特定的分化阶段的分子？ 相反，定义 ECM分子区域对嵴细胞发挥分化作用 亚群？ (To与詹姆斯韦斯顿博士的 实验室）。 凭借我们在细胞-ECM相互作用方面的背景以及现在的工具， 我们将能够准确地评估FN的作用， PNS开发中的LAM。

The long-range goal of the proposed research is to understand the roles of the extra-cellular matrix (ECM) proteins fibronectin (FN) and laminin (LAM) in peripheral nervous system (PNS) development. ECM molecules regulate cell shape, differentiation, and movement during embryogenesis. In the PNS, ECM molecules appear to be involved in neural crest migration and development and regeneration of neuromuscular junctions. Growth cones of nerve fibers migrating from the neural tube or peripheral ganglia traverse substrate pathways that are also likely to contain adhesive molecules. The adhesive glyco-proteins FN and LAM are good candidates for mediating particular stages of PNS cell migration and differentiation, due to their locations in embryos and interactions with PNS cells in vitro. First, what functional regions of FN and LAM mediate neuron-substratum sdhesion and neurite extension? Two regions of FN have these activities and have revealed neuronal type-specific interactions with the FN molecule. Functional regions of LAM are less well-defined, but may also promote cell type-specific behavior. Proteolytic fragments of FN and LAM will be used, in conjuction with antibodies, to evaluate 1) the functional distinctions between the regions of FN that interact with cells, 2) whether cell surface heparan sulfate mediates certain of these interactions, and 3) how each domain contributes to mediation of cell behavior by the intact FN molecule. In addition, the role of growth cone adhesion in neurite extension on FN fragments will be studied. Second, how are FN and LAM (and specific regions of each) used by cells in the developing PNS? Approaches will be 1) immunocytochemistry to reveal temporal and spatial associations between cells and ECM molecules, 2) immunolabeling for TEM and SEM to identify the cells producing LAM and FN and their proximity to growth cones, and 3) inhibition, with antibodies, of neurite extension on naturally-occurring matrices and in vivo to probe the relative roles of each molecule. Third, how do defined regions of FN and LAM contribute to migration and differentiation of neural crest cells? Do crest cell subpopulations attach to, and/or move upon, certain regions of these molecules at particular stages of differentiation? Conversely, do defined regions of ECM molecules exert differentiative effects on crest cell subpopulations? (To be studied in collaboration with Dr. James Weston's lab). With our background in cell-ECM interactions and the tools now available to us, we will be able to evaluate precisely the roles of FN and LAM in PNS development.