MOLECULAR STUDIES OF A NOVEL OLIGODENDROCYTE

描述：少突胶质细胞特异性蛋白(OSP)是最近分离到的一种 中枢神经系统髓鞘特异蛋白，其功能与调控表达 申请人建议解决的中枢神经系统发育和神经胶质肿瘤的发生。 OSP在大脑中的发育表达将使用 原位杂交，以确定少突胶质细胞处于哪个阶段 分化的蛋白质很可能起作用。基因的启动子区域 将分离OSP基因并对其进行特征分析，以确定影响因素 来调节这个基因的细胞特异性表达。这一分析将 在细胞培养和转基因小鼠中进行。几行 有证据支持OSP在调节少突胶质细胞增殖中的作用。 因此，OSP基因剂量对少突胶质细胞生长和生长的影响 将检查髓鞘形成情况。少突胶质细胞前体细胞将 将含有OSP基因的逆转录病毒表达载体 OSP反义cDNA以确定上下效应 OSP在细胞生长和分化中的表达转基因小鼠将 由髓鞘碱性蛋白启动子驱动的OSP转基因创造 为了促进OSP的过度表达，将创建OSP“基因敲除”小鼠以 确定OSP低表达的影响。这些突变的小鼠将是 通过免疫组织化学、原位杂交和电子显微镜进行分析 显微镜，以揭示形态、结构和生化效应 改变OSP基因剂量。申请人和他的同事也将 利用酵母鉴定OSP与之相互作用的蛋白质 双杂交系统。最后，他们将检查OSP的表达和 OSP基因在一些少突胶质细胞瘤中的拷贝数，试图 研究其在肿瘤发生中的潜在作用。

DESCRIPTION: Oligodendrocyte-specific protein (OSP) is a recently isolated CNS myelin-specific protein, whose function and regulated expression during CNS development and glial tumorigenesis the applicant proposes to address. OSP's developmental expression in the brain will be characterized using in situ hybridization, to determine at which stage in oligodendrocyte differentiation the protein is likely to act. The promoter region of the OSP gene will be isolated and characterized in order to identify factors that regulate the cell-specific expression of this gene. This analysis will be performed in cell culture and in transgenic mice. Several lines of evidence support a role of OSP in regulating oligodendrocyte proliferation. Therefore, the effect of OSP gene dosage on oligodendroglial growth and myelination will be examined. Oligodendrocyte precursors will be transfected with a retroviral expression vector containing the OSP cDNA and OSP anti-sense cDNAs in order to determine the effect of over and under expression of OSP on cell growth and differentiation. Transgenic mice will be created with an OSP transgene driven by the myelin basic protein promoter to promote OSP overexpression, and OSP "knock-out" mice will be created to determine the effect of OSP underexpression. These mutant mice will be analyzed by immunohistochemistry, in situ hybridization, and electron microscopy, to reveal the morphological, structural, and biochemical effects of alterred OSP gene dosage. The applicant and his colleagues will also identify the proteins with which OSP interacts by exploiting the yeast two-hybrid system. Finally, they will examine expression of OSP and the copy number of the OSP gene in a number oligodendrogliomas, in an attempt to investigate its potential role in tumorigenesis.