ISOLATION OF POTENTIAL ONCOGENES FROM TELEOST TUMORS

从最古老的肿瘤中分离潜在的癌基因

• 批准号: 3916923
• 负责人: D K WATSON
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3916923
• 关键词: calcium phosphate; chemical carcinogen; clone cells; fish; genetic mapping; mesothelioma; natural gene amplification; neoplastic cell culture for noncancer research; neoplastic transformation; nitrosamines; nucleic acid hybridization; nucleic acid probes; nucleic acid sequence; oncogenes; transfection

Previous studies have suggested that fish have oncogene sequences homologous to those found in mammalian and avian species. We were the first to confirm the presence of fish oncogenes by isolating and sequencing the c-myc gene from rainbow trout (Van Beneden et al., 1986). Other fish genes homologous to known mammalian oncogenes were identified by Southern blot hybridization. In order to examine the role of fish oncogenes, Southern blots were prepared using DNA digests from tumor and normal tissue. Hybridization to known oncogene sequences did not detect rearrangements or gene amplifications. In order to detect other oncogenes, we developed a transfection system in which fish DNA in the presence of calcium phosphate was transfected into NIH 3T3 cells. The transforming ability of fish tumor DNA was examined by standard focus assay, nude mouse assay, and colony selection assay. DNA from diethylnitrosamine-induced mesothelioma in medaka was the most efficient in transformation of NIH 3T3 cells. Secondary transfectants caused formation of tumors in nude mice of greater than 20mm in 1-1/2 weeks following injection. Southern blot analysis of these transfectant DNAs hybridized to medaka genomic DNA probe showed bands present in tumor-induced transfectants. No bands were present in DNA from NIH 3T3 controls and cells transfected with non-tumorigenic medaka DNA. This suggests the presence of specific fish sequences in transformants. These sequences do not appear to be homologous to K-ras, H-ras, N-ras, c-myc m-met or v-erbB. This study will continue to identify, clone, and sequence the transforming gene from fish in order to examine its role in tumor formation.

先前的研究表明鱼类有致癌基因序列 与在哺乳动物和鸟类中发现的那些同源。 我们 第一个证实鱼类致癌基因存在的人， 并对虹鳟鱼的c-myc基因进行测序（货车Beneden et 例如，1986年）。 与已知哺乳动物同源的其他鱼类基因 通过Southern印迹杂交鉴定癌基因。 为了 为了检测鱼类致癌基因的作用，制备了Southern印迹， 使用来自肿瘤和正常组织的DNA引物。 杂交 已知的癌基因序列没有检测到重排或基因 扩增。 为了检测其他致癌基因，我们开发了 一种转染系统，其中鱼DNA在钙的存在下 磷酸盐转染NIH 3 T3细胞。 转化 通过标准病灶分析检测鱼肿瘤DNA的能力， 裸鼠试验和集落选择试验。 的DNA 二乙基亚硝胺诱发的青少年间皮瘤最多， 转化NIH 3 T3细胞。 二次 转染子在裸鼠中引起肿瘤的形成， 在注射后1-1/2周内超过20 mm。 Southern印迹 分析这些与青鳉基因组杂交的转染DNA DNA探针显示在肿瘤诱导的转染子中存在条带。 没有 NIH 3 T3对照和细胞的DNA中存在条带 用非致瘤性青鳉DNA转染。 这表明 转化体中存在特异性FISH序列。 这些 序列似乎与K-ras，H-ras，N-ras， c-myc m-met或v-erbB。 这项研究将继续确定， 从鱼类中克隆并测序转化基因， 研究它在肿瘤形成中的作用。