BIOSYNTHESIS OF SCHWANN CELL EXTRACELLULAR MATRIX

这个项目的目标是了解细胞外基质的作用 在周围神经系统的发展中起着重要作用。 我们专注于 雪旺细胞（SC）产生的基底膜及其与细胞增殖的关系 影响SC终末分化。 之前的实验表明 1）SC是结构组件的主要来源，如果不是唯一来源的话 这些包括胶原蛋白（主要是I型和IV型），层粘连蛋白 和硫酸乙酰肝素蛋白聚糖; 2）SC终末分化（即， 鞘化和髓鞘形成）依赖于与该基质的接触; 3）SC基质组装受神经细胞控制。 工作提出的 下一个资助期的目标是阐明 参与调节基质生产和发育的影响， SC上的矩阵。 作为实现我们概述的这些目标的步骤， 以下目标。 1)来识别和描述地下室 膜和SC蛋白介导的细胞基质相互作用，我们将 利用免疫学方法来利用观察结果， 重组基底膜诱导培养的正常分化 SC;将针对该基质和SC制备特异性抗体 膜蛋白，并测试它们阻断SC扩散的能力 与母体接触引起的。 2)使其结构和 SC质膜蛋白聚糖的功能特性， 我们将试图描绘的结构域结构的细胞骨架， 这种分子，并研究，使用体外结合试验，其模式， 与基底膜和细胞骨架结合;我们也会尝试 纯化这种分子并产生特异性抗体。 3)阐明本 神经细胞调控SC基底膜形成的机制; 我们将确定是否神经细胞接触或可溶性因子释放 由神经细胞介导这种现象;我们还将研究是否 调节特定SC基质蛋白水平的神经细胞， 这是通过改变合成、降解或利用的速率实现的 这些蛋白质。 4)描述基底膜诱导 伴随形态学变化的SC细胞骨架的改变 这是终端分化所需的;我们将研究 细胞骨架蛋白代谢的变化（合成速率， 降解、翻译后修饰和聚合状态）， 生物化学方法;细胞骨架丝的结构改变将 通过免疫荧光与特异性抗体和电子 显微镜

The goal of this project is to understand the role of extracellular matrix in the development of the peripheral nervous system. We have focused our studies on the basement membrane produced by Schwann cells (SCs) and its effects on SC terminal differentiation. Previous experiments have shown that 1) SCs are the major, if not sole, source of the structural components of this matrix; these include collagen (primarily types I and IV), laminin and heparan sulfate proteoglycan; 2) SC terminal differentiation (i.e., ensheathment and myelination) is dependent upon contact with this matrix; and 3) SC matrix assembly is controlled by nerve cells. The work proposed for the next grant period is aimed at elucidating the molecular mechanisms involved in regulating matrix production and the developmental effect of the matrix on SCs. As steps toward achieving these goals we have outlined the following objectives. 1) To identify and characterize the basement membrane and SC proteins which mediate the cell-matrix interaction; we will utilize an immunological approach to exploit the observation that simple, reconstituted basement membrane induces normal differentiation in cultured SCs; specific antibodies will be prepared against this matrix and SC membrane proteins and tested for their ability to block the SC spreading induced by contact with the matrix. 2) To charcterize the structure and functional properties of a SC plasma membrane proteoglycan that is anchored to the cytoskeleton; we will attempt to delineate the domain structure of this molecule, and to study, using in vitro binding assays, its mode of binding to basement membrane and the cytoskeleton; we will also at tempt to purify this molecule and produce specific antibodies. 3) To elucidate the mechanism of regulation by nerve cells of SC basement membrane formation; we will determine whether nerve cell contact or a soluble factor released by nerve cells mediate this phenomenon; we will also investigate whether nerve cells, which modulate the levels of specific SC matrix proteins, do this through changes in the rates of synthesis, degradation or utilization of these proteins. 4) To describe the basement membrane induced alterations in the SC cytoskeleton that accompany the morphological changes which are required for terminal differentiation; we will investigate changes in cytoskeletal protein metabolsim (rates of synthesis, degradation, post-translational modification, and polymerization state) by biochemical methods; structural alterations in cytoskeletal filaments will be studied by immunofluorescence with specific antibodies and electron microscopy.