FLAGGING OF DNA DAMAGE SITES BY RNA POLYMERASE II

通过 RNA 聚合酶 II 标记 DNA 损伤位点

• 批准号: 2748917
• 负责人: DAVID B BREGMAN
• 金额: $ 9.13万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-08-01 至 1998-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2748917
• 关键词: ADP ribosylation; DNA damage; DNA directed RNA polymerase; DNA repair; HeLa cells; biological signal transduction; crosslink; enzyme activity; enzyme complex; genetic transcription; nucleic acid sequence; site directed mutagenesis; transfection; ubiquitin

Certain proteins essential for DNA repair are also required for transcriptional initiation by RNA polymerase II, the multisubunit enzyme that transcribes protein endoding genes. In addition, DNA lesions located on the transcribed strand of active genes are repaired more rapidly than other DNA lesions. It has therefore been proposed that RNA polymerase Ii may function to identify such lesions as it "needs" the DNA. Nevertheless, a critical question remains: how does RNA polymerase II signal to the rest of the DNA repair system that it has encountered damaged DNA and that the machinery must switch from "transcription mode" to "repair mode"? This investigator has identified novel, upper molecular weight forms of RNA polymerase II's largest subunit (Pol II LS) which are induced shortly after cells are exposed to DNA damaging agents. Preliminary studies indicate that two covalent modifications already implicated in DNA repair, ubiquitination the ADP-ribosylation, may be involved in generating the upper molecular weight forms." In vitro and in vivo approaches will be utilized to (1) determine whether the upper molecular weight forms of Pol II LS are the result of ubiquitination and/or ADP-ribosylation. (2) determine whether the modification maps tot he regulatory C-terminal domain of Pol II and (3) determine whether the subset of RNA Polymerase II molecules which get modified represent elongating molecules which have stalled at DNA lesions. Because maintaining the accuracy of DNA is a sine qua non of normal cell function and because defective DNA repair contributes to the development of cancer this topic impacts profoundly on the study of human disease.

DNA 修复所必需的某些蛋白质也是 由多亚基酶 RNA 聚合酶 II 启动转录 转录蛋白质内含基因。 此外，DNA 损伤位于 活性基因转录链上的修复速度比 其他 DNA 损伤。 因此有人提出 RNA 聚合酶 Ii 可能会发挥识别此类病变的作用，因为它“需要”DNA。 尽管如此， 一个关键问题仍然存在：RNA 聚合酶 II 如何向其他酶发出信号 DNA 修复系统遇到了受损的 DNA，并且 机械必须从“转录模式”切换到“修复模式”？ 这 研究人员发现了新的、高分子量的 RNA 形式 聚合酶 II 最大的亚基 (Pol II LS) 在不久后被诱导 细胞暴露于 DNA 损伤剂。 初步研究表明 两个共价修饰已经与 DNA 修复有关， ADP-核糖基化的泛素化，可能参与生成 高分子量形式。” 将利用体外和体内方法来（1）确定是否 Pol II LS 的高分子量形式是以下结果 泛素化和/或 ADP-核糖基化。 (2)判断是否 修饰映射到 Pol II 的 C 末端调控域和 (3) 确定 RNA 聚合酶 II 分子的子集是否得到 修饰代表在 DNA 损伤处停滞的伸长分子。 因为维持DNA的准确性是正常细胞的必要条件 功能，并且因为有缺陷的 DNA 修复有助于发展 癌症这个话题对人类疾病的研究影响深远。