ISOLATION AND ANALYSIS OF PROTEINS WHICH REGULATE RAS

哺乳动物细胞分裂的调控是一个复杂的过程； 协调跨细胞质传递的促生长信号 膜由生长因子启动，信号由元素启动 监测细胞的营养状态。细胞的发现 癌基因为研究生物化学提供了一种手段，至少是部分的。 控制细胞分裂的机制。其中一组癌基因，即ras 基因被发现在大量和广泛的突变中被激活 各种各样的人类肿瘤。Ras基因在进化中高度保守， 这表明它们在细胞的生物学中扮演着重要的角色。两者都有 Ras蛋白的功能及如何调控ras蛋白 对于哺乳动物细胞来说，这仍然是一个谜。酵母RAS基因的发现 使研究RAS蛋白功能的系统方法成为可能，因为 在这种有机体中可能进行的基因操纵。这项建议是 目的是研究酵母中的CDC25蛋白，它不仅可以调节 酵母Ras蛋白起作用，但人的H-ras蛋白也起作用。这 调节可能是由于蛋白质与蛋白质之间的直接相互作用。 CDC25和RAS蛋白。这种蛋白质-蛋白质相互作用的研究和 它的保存和进化提供了一种独特的手段来识别 调节ras蛋白功能的哺乳动物分子。我们将发展 编码或识别结构域的DNA探针和免疫试剂 RAS激活所必需的CDC25蛋白。这些试剂将是 用于克隆与CDC25同源的编码结构域的cDNA。此外，我们还将 使用基因筛查来识别人类cDNA，它可以补充缺失的 CDC25在酵母中发挥功能。黄曲霉毒素的分子机制鉴定 人类ras蛋白的作用可能使分子设计成为可能 逆转突变ras蛋白致瘤作用的合理药物。

The regulation of cell division in mammalian cells is a complex process; coordinating growth promoting signals transduced across the cytoplasmic membrane initiated by growth factors, with signals initiated by elements monitoring the nutritional status of the cells. The discovery of cellular oncogenes provide a means to study, at least in part, the biochemical mechanisms controlling cell division. One group of these oncogenes, the ras genes, are found to be mutationally activated in a large number and wide variety of human tumors. The ras genes are highly conserved in evolution, suggesting they play a fundamental role in the biology of the cell. Both the function of the ras proteins and how the ras proteins are regulated remains a mystery for mammalian cells. The discovery of the yeast RAS genes made possible a systematic approach to study RAS protein function because of the genetic manipulation possible in that organism. This proposal is aimed at studying the CDC25 proteins in yeast which can regulate, not only yeast RAS protein function but the human H-ras protein as well. This regulation is likely due to a direct protein-protein interaction between CDC25 and RAS proteins. The study of this protein-protein interaction and its conservation t rough evolution provides a unique means to identify the mammalian molecule which regulates ras protein function. We will develop DNA probes and immunological reagents which encode or recognize the domain of the CDC25 protein essential for RAS activation. These reagents will be used to clone cDNAs encoding domains homologous to CDC25. Further, we will use a genetic screen to identify human cDNAs which can complement loss of CDC25 function in yeast. The identification of the molecular mechanism of action of the human ras proteins may allow the design of molecularly rational drugs to reverse the tumorigenic effect of mutant ras proteins.