CARCINOGENS AND CHROMATIN STRUCTURE AND FUNCTION

致癌物和染色质结构与功能

• 批准号: 2087359
• 负责人: MARK E SMULSON
• 金额: $ 19.58万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1979
• 资助国家: 美国
• 起止时间: 1979-06-01 至 1996-02-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2087359
• 关键词: ADP ribosylation; DNA directed DNA polymerase; DNA repair; DNA replication; RNA splicing; antisense nucleic acid; chemical carcinogen; enzyme mechanism; enzyme structure; flow cytometry; genetic transcription; human tissue; molecular cloning; nicotinamide adenine dinucleotide; nucleoproteins; nucleosomes; plasmids; protein structure function; ribosomal RNA; ribozymes; tissue /cell culture; transfection; transposon /insertion element

The poly ADP-ribosylation modification of nuclear proteins is the initial cellular response to DNA damage caused by chemicals. Poly(ADP-ribose) polymerase (PADPRP) catalytic activity is absolutely dependent upon DNA and its activity is directly correlated with the number of strand breaks in DNA. Thus, when cells, are exposed to chemical carcinogens, there is an immediate drop in cellular NAD and concomitant rapid and transient increase in poly ADP-ribosylated nuclear proteins. Recent progress on this project involved the cloning, sequencing, and hyperexpression of transfected genes for this enzyme in both human and murine systems. Preliminary data using transient transfection indicated that cells hyperexpressing PADPRP have increased rates of DNA repair. We also were able to map the precise chromosomal loci for this gene in both human and mouse. During the renewal period, we intent to utilize the molecular and biochemical data obtained to better clarify and characterize the role of ADP-ribosylation in DNA repair mechanisms. Methods in Aim I utilize a variety of molecular approaches including antisense and ribozyme expression, to experimentally modulate ADP-ribosylation potential in cells subjected to DNA strand breaks and assess the effects of this by utilizing new methods for precisely measuring preferential gene repair and recombinase activity using novel plasmid assays. Nuclear targets for ADP ribosylation are the focus of Aim II. The new molecular information will be used here to relate structure of PADPRP (and its nuclear protein acceptors) to function. For example, c-fos is a nuclear acceptor for PADPRP and also transcriptionally activated by strand- breaking chemicals. PADPRP activity modulated by antisense will be used to address the biological function of the PADPRP of fos, and as an example of approaches to be used for other nuclear protein acceptors during the renewal period.

核蛋白的聚ADP-核糖基化修饰是初始 细胞对化学物质引起的DNA损伤的反应。 聚（ADP-核糖） 聚合酶（PADPRP）催化活性绝对取决于DNA和 它的活性与链断的数量直接相关 脱氧核糖核酸。 因此，当细胞暴露于化学致癌物时，有一个 立即下降细胞NAD和伴随的快速和瞬态增加 在聚ADP-核糖基化的核蛋白中。 该项目的最新进展涉及克隆，测序和 该酶在人和该酶 鼠系统。 使用瞬态转染的初步数据指示 该细胞过度使用PADPRP的DNA修复速率增加。 我们 还能够为这两个基因绘制精确的染色体基因座 人和老鼠。 在续签期间，我们意图利用分子和 获得的生化数据是为了更好地阐明和表征 DNA修复机制中的ADP-核糖基化。 AIM中的方法我利用 各种分子方法包括反义和核酶 表达，实验调节细胞中的ADP-核糖基势 通过利用DNA链断裂并评估其影响 精确测量优先基因修复和的新方法 使用新型质粒测定的重组酶活性。 ADP核糖基化的核靶标是AIM II的重点。 新的 分子信息将在这里用于关联PADPRP的结构（和 它的核蛋白受体）发挥作用。 例如，c-fos是 PADPRP的核受体，也被链的转录激活 破坏化学物质。 反义调节的PADPRP活动将用于 解决了FOS PADPRP的生物学功能，作为一个例子 在此期间用于其他核蛋白受体的方法 续期。