ISOPRENYLATION OF RAS AND OTHER PROTEINS IN YEAST

通过添加法尼基基团对蛋白质进行修饰已经被广泛应用。 对Ras等蛋白质的功能至关重要。 这 促进Ras的膜结合，这是其 转化活动。 这种修饰是由蛋白质催化的 法尼基转移酶，其识别 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显示出显著的阻断 人类肿瘤细胞的生长。 影响的特征 然而，这些抑制剂提高了蛋白质 是抑制剂的靶点。 此外，日益增长的 许多G蛋白如Rheb、RhoB和RhoE已被证明是 法尼基化的 这些观察结果引出了一个问题， 以及它们的生理功能。 我们漫长 长期目标是解决这些问题。 最近确定 酵母基因组的整个序列提供了一个独特的机会 来探讨这些问题。 在研究中， 法尼基化，我们可以获得法尼基化蛋白质的完整图像 在真核生物中。 通过寻找潜在的法尼基化 利用酵母蛋白质数据库，我们鉴定了22种蛋白质 以CAAX主题结尾。 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Rho 3. Rheb特别令人感兴趣，因为它的人类 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。 我们将执行基因 破坏以及突变蛋白的表达， 这个G蛋白的功能 另一个是Rho3，它扮演一个 在酵母细胞的极化生长的关键作用，显然通过 它对肌动蛋白细胞骨架和膜融合作用。我们将 探讨法尼基化对Rho3功能的影响。 最后，将进行系统的基因中断研究， 鉴定新的必需的法尼基化蛋白质。 我们的研究将 是第一次全面尝试表征法尼基化蛋白质 在真核生物中。所获得的结果应显着 拓宽我们对蛋白质法尼基化的理解。

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.