EFFECTS OF CHEMICAL CARCINOGENS ON TRANSFORMING DNA SEQUENCES AND EXPRESSION

该项目的目标是识别、表征和克隆这些基因 它推动了肿瘤的发展，其恶性潜能 这是化学致癌物引起的变化的结果。(1)识别 苯并[a]芘(BP)激活的转化基因，来自三个国家的DNA 对BP转化的BALB/3T3细胞进行DNA转移和 NIH/3T3系统中的焦点形成。所有三个测试品系都显示出 转换彼此不同以及与RAS不同的活动 癌基因的限制性内切酶敏感性和MspI定位。这些 通过同胞选择，可能正在克隆新的转化基因。 来自Charon 4A噬菌体基因组文库的协议。三个协议周期 已进行选择，阳性池未显示存在 Ras癌基因。(2)探讨化学物质的作用机制。 致癌物可能激活原癌基因，黄曲霉毒素B1的分布 (AFB)加合物的基因分析使用纯化的肝细胞核和 试管中的微粒体组。AFB加合物优先定位于DNase I 基因组的过敏区。(3)阐明遗传事件 在小鼠皮肤肿瘤促进机制的基础上，新基因 称为PRO 1和PRO 2，指定对诱导的敏感性 TPA在JB-6细胞中的转化，通过同源选择克隆 对启动子敏感的克隆细胞的大小选择DNA文库。通过 限制性内切酶图谱、异源双链分析和直接杂交 原基因与癌基因或其他已知的基因不同，也不相关 基因。这两个前基因都已经测序，它们的功能正在研究中 调查过了。人类前基因的同源物已经从 人鼻咽癌细胞系CNE2。(4)使用NIH/3T3 Focus 在人前列腺癌细胞中检测到一种转化基因 并正在克隆中；这些细胞显然不包含 突变的ras癌基因。

The aim of this project is to identify, characterize, and clone those genes that drive the development of neoplasia and whose malignant potential results from changes caused by chemical carcinogens. (1) To identify transforming genes activated by benzo[a]pyrene (BP), DNA from three BALB/3T3 cell lines transformed by BP was analyzed by DNA transfer and focus formation in the NIH/3T3 system. All three tested lines showed transforming activity that differed from each other and from the ras oncogenes by restriction endonuclease sensitivity and MspI mapping. These possibly new transforming genes are now being cloned by the sib selection protocol from Charon 4A phage genomic libraries. Three protocol cycles of selections have been carried out and the positive pools show no presence of the ras oncogenes. (2) To explore the mechanisms by which chemical carcinogens may activate proto-oncogenes, the distribution of aflatoxin Bl (AFB) adducts on genes was analyzed using purified liver nuclei and microsomes in vitro. AFB adducts were preferentially located in DNAse I hypersensitive regions of the genome. (3) To elucidate the genetic events underlying the mechanisms of tumor promotion in mouse skin, novel genes termed pro 1 and pro 2, specifying sensitivity to induction of transformation by TPA in JB-6 cells, were cloned by sib selection from a size-selected DNA library of clonal cells sensitive to promotion. By restriction mapping, heteroduplex analyses and direct hybridization, the pro genes are different from and unrelated to oncogenes or other known genes. Both pro genes have been sequenced and their functions are being investigated. Human homologs of pro genes have been isolated from the human nasopharyngeal carcinoma cell line CNE2. (4) Using the NIH/3T3 focus assay, a transforming gene was detected in human prostate carcinoma cell line PC-3 and is being cloned; these cells apparently do not contain altered ras oncogens.