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.

核蛋白的多聚ADP-核糖基化修饰是 细胞对化学物质造成的DNA损伤的反应。 聚ADP-核糖 聚合酶（PADPRP）催化活性完全依赖于DNA， 它的活性与细胞中链断裂的数量直接相关， DNA. 因此，当细胞暴露于化学致癌物时， 细胞NAD立即下降并伴随快速和短暂增加 在多聚ADP核糖基化核蛋白中。 该项目的最新进展涉及克隆、测序和 这种酶转染基因在人和 鼠系统。 使用瞬时转染的初步数据表明， 高表达PADPRP的细胞DNA修复的速度加快。 我们 我们也能够在这两个基因的精确染色体定位， 人和小鼠。 在更新期间，我们打算利用分子和 获得的生化数据，以更好地阐明和表征的作用， DNA修复机制中的ADP-核糖基化。 目标I中的方法利用 包括反义核酸和核酶在内的多种分子方法 表达，以实验性地调节细胞中的ADP-核糖基化潜力 进行DNA链断裂，并评估其影响，利用 精确测量优先基因修复的新方法， 重组酶活性的测定。 ADP核糖基化的核靶点是Aim II的重点。 新 分子信息将在此用于关联PADPRP的结构（和 其核蛋白受体）发挥作用。 例如，c-fos是一种 PADPRP的核受体，也被链- 破坏化学品 由反义调节的PADPRP活性将用于 解决Fos的PADPRP的生物学功能，并作为一个例子， 其他核蛋白受体的方法 更新期。