FUNCTION OF THE CPH ONCOGENE IN MAMMALIAN CARCINOGENESIS

本延续申请的主要目的是确定 新的cph癌基因和原癌基因的功能， 在肿瘤发生过程中激活的癌基因。 cph癌基因是 在PI的实验室中从化学引发的肿瘤性 叙利亚仓鼠胚胎细胞，一种与人类相关的致癌模型 肿瘤形成我们以前的研究结果表明， 在鼠成纤维细胞的转化中与ras协同作用。 在本资助期内取得的结果显示， 从酵母到人类细胞都是保守的， 组织，这表明它在细胞中起着重要作用， 调控 此外，我们还发现：i）cph是由 点突变缺失导致截短的 蛋白产物，ii）CPH蛋白具有显著的全局同源性 iii）当过表达时，cph使细胞 更能抵抗各种形式的压力。 我们假设 激活的cph癌基因是一种失调的，过度活跃的核苷 二磷酸酶（NDP 3），其为细胞提供上级应激- 存活功能和增殖优势。 这一假设将 可以通过几种方法进行测试：1）结构决定因素分析 (gain和/或调节序列的丢失）。 2)通过以下方法证明cph蛋白的NDP 3活性： GDPase-null酵母突变体的互补和纯化， 天然哺乳动物蛋白的微测序和活性测定。 第三章 研究了cph转化的机理， NDV活性是必需的，cph癌蛋白是否 错误定位或与异常细胞基质相互作用。 四、 建立cph参与细胞应激反应的机制 途径。这些研究将扩大我们对 癌基因在癌变过程中的作用 NDR 4对恶性肿瘤的作用。 长期目标是 建议将获得的知识扩展到人类细胞，并应用于 研究cph在人类肿瘤形成中的可能作用。

The central objective of this continuation application is to determine the function of the novel cph oncogene and proto-oncogene, and the role of the activated oncogene in tumorigenesis. The cph oncogene was isolated in the PI's laboratory from chemically-initiated, neoplastic Syrian hamster embryo cells, e carcinogenesis model relevant to human neoplasia. Our previous results demonstrated that cph acts synergistically with ras in the transformation of murine fibroblasts. Results obtained during the present funding period have shown that cph is conserved from yeast to human cells and is expressed in most adult tissues, suggesting that it plays an important role in cellular regulation. In addition we have found that: i) cph is activated by point mutational deletions which result in the synthesis of truncated protein products, ii) the cph proteins have significant global homology to the yeast GDPase, and iii) when overexpressed, cph makes the cells more resistant to various forms of stress. We hypothesize that the activated cph oncogene is a disregulated, hyperactive nucleoside diphosphatase (NDPase) which provides the cells with superior stress- survival functions and proliferative advantage. This hypothesis will be tested by several approaches: 1) Analysis of structural determinants (gain and/or loss of regulatory sequences) of cph transforming activity. 2) Demonstration of the NDPase activity of the cph proteins, by complementation of GDPase-null yeast mutants and by purification, microsequencing and activity assays of native mammalian proteins. 3) Study the mechanism of cph transformation with regard to whether the NDPase activity is required and whether the cph oncoprotein is either mislocalized or interacts with abnormal cellular substrates. 4) Establish the participation of cph in the cellular stress-response pathways. These studies will expand our knowledge on the role of oncogenes in the carcinogenesis process by showing for the first time the contribution of a NDPase to malignancy. The long term goal of this proposal is to extend the knowledge gained to human cells, and to apply it to study the possible role of cph in human neoplasia.