ISOPRENYLATION OF RAS AND OTHER PROTEINS IN YEAST

酵母中 RAS 和其他蛋白质的异戊二烯化

• 批准号: 2625435
• 负责人: Fuyuhiko Tamanoi
• 金额: $ 30.95万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-06-01 至 2003-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2625435
• 关键词: G protein; Saccharomyces cerevisiae; Schizosaccharomyces pombe; actins; alkyltransferase; cytoskeleton; enzyme activity; farnesyl compound; fungal genetics; gene expression; guanine nucleotide binding protein; membrane fusion; mutant; oncoproteins; polymerase chain reaction; posttranslational modifications; protein localization; protein structure function; yeast two hybrid system

Modification of proteins by the addition of a farnesyl groups has been shown to be critical for the function of proteins such as Ras. This facilitates membrane association of Ras which is required for its transforming activity. The modification is catalyzed by protein farnesyltransferase which recognizes the C-terminal CAAX motif of farnesylated proteins. These studies culminated in the development of farnesyltransferase inhibitors which exhibit remarkable ability to block the growth of human tumor cells. Characterization of the effects of these inhibitors, however, has raised the possibility that proteins other than Ras are targets of the inhibitors. In addition, a growing number of G-proteins such as Rheb, RhoB and RhoE have been shown to be farnesylated. These observations lead to a question how many proteins are farnesylated and what their physiological functions are. Our long term objective is to address these questions. Recent determination of the entire sequence of the yeast genome provides a unique opportunity to explore these issues. For the first time in the study of farnesylation, we can obtain a complete picture of farnesylated proteins in an eukaryotic organism. By searching for potentially farnesylated proteins using the yeast protein database, we identified 22 proteins ending with the CAAX motif. Of these, 14 are newly identified proteins or partially characterized proteins. We will take a targeted approach to characterize these proteins. First, we will focus on two G-proteins, Rheb and Rho3. Rheb is of particular interest, since its human counterpart is upregulated in a variety of human tumor cells, and has recently been shown to be farnesylated. We will carry out gene disruption as well as expression of mutant proteins to gain insights into the function of this G-protein. The other is Rho3 which plays a critical role in the polarized growth of yeast cells, apparently through its action on actin cytoskeleton and membrane fusions. We will investigate the significance of farnesylation on the function of Rho3. Finally, a systematic gene disruption study will be carried out to identify new farnesylated proteins which are essential. Our study will be the first full scale attempt to characterize farnesylated proteins in an eukaryotic organism. The results obtained should dramatically widen our understanding of protein farnesylation.

通过添加法呢基团对蛋白质进行修饰已被 已被证明对 Ras 等蛋白质的功能至关重要。 这 促进 Ras 的膜结合，这是其必需的 转化活动。 修饰是由蛋白质催化的 法呢基转移酶可识别 C 端 CAAX 基序 法尼基化蛋白质。 这些研究最终导致了 法尼基转移酶抑制剂具有显着的阻断能力 人类肿瘤细胞的生长。 影响的表征 然而，这些抑制剂提出了蛋白质的可能性 除了 Ras 之外，其他都是抑制剂的靶点。 此外，日益增长的 一些 G 蛋白，如 Rheb、RhoB 和 RhoE 已被证明 法尼基化。 这些观察结果引出了一个问题：有多少蛋白质 是法尼基化的以及它们的生理功能是什么。 我们的长 术语目标就是解决这些问题。 最近确定 酵母基因组的整个序列提供了独特的机会 来探讨这些问题。 第一次在研究中 法尼基化，我们可以获得法尼基化蛋白质的全貌 在真核生物中。 通过寻找潜在的法呢基化 蛋白质 使用酵母蛋白质数据库，我们鉴定了 22 种蛋白质 以 CAAX 主题结束。 其中，14种是新发现的蛋白质 或部分表征的蛋白质。 我们将采取有针对性的方法 来表征这些蛋白质。 首先，我们将关注两种 G 蛋白， Rheb 和 Rho3。 Rheb 特别令人感兴趣，因为它是人类的 对应物在多种人类肿瘤细胞中上调，并且具有 最近被证明是法尼基化的。 我们将进行基因 突变蛋白的破坏和表达以获得见解 进入该 G 蛋白的功能。 另一个是 Rho3，它的作用是 在酵母细胞的极化生长中起着关键作用，显然是通过 它对肌动蛋白细胞骨架和膜融合的作用。我们将 研究法尼基化对 Rho3 功能的意义。 最后，将进行系统的基因破坏研究 识别重要的新法呢基化蛋白质。 我们的研究将 首次全面尝试表征法呢基化蛋白质 在真核生物中。获得的结果应该显着 拓宽我们对蛋白质法呢基化的理解。