SV 40 DNA REPLICATION

SV 40 DNA 复制

• 批准号: 6299962
• 负责人: BRUCE S STILLMAN
• 金额: $ 37.41万
• 依托单位: COLD SPRING HARBOR LABORATORY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-01-01 至 2000-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6299962
• 关键词: DNA replication; DNA replication origin; Escherichia coli; bovine papillomavirus; cell cycle; cell transformation; chromatin; flow cytometry; guanine nucleotide binding protein; immunoprecipitation; laboratory mouse; laboratory rabbit; molecular cloning; molecular oncology; nucleic acid metabolism; oncoprotein p21; proliferating cell nuclear antigen; protein purification; protein structure function; simian virus 40; tumor antigens; virus genetics; virus replication; western blottings

Understanding the mechanism of DNA replication in human cells is a fundamental problem in biology and has direct relevance to cancer research. Towards this goal, the mechanism of DNA replication from the Simian Virus 40 (SV40) and the bovine papillomavirus (BPV) origins of DNA replication will be investigated. In addition, the mechanism and regulation of DNA replication-dependent chromatin assembly will also be studies. Using cell-free extracts prepared from human cells and either purified SV40 T antigen or the BPV E1 and E2 proteins, factors required for virus DNA replication and their mechanism of action will be studies. A cellular protein essential for BVP DNA replication, but not for SV40 DNA replication, will be purified and its functions characterized. The mechanism of DNA polymerase switching involving Replication Factor C and the proliferating cell nuclear antigen (PCNA) will also be investigated, as will the interaction of these proteins with the cell division cycle regulator, p21. These proteins also appear to function in repair of radiation damaged DNA and therefore, these studies will provide insight into the controls that coordinated DNA replication, DNA repair and cell cycle progression. Finally, the functions of chromatin assembly factor 1 (CAF1) and its role in DNA replication- dependent chromatin assembly will be determined. Other chromatin assembly factors cooperate with CAF1 to assemble nucleosomes onto replication DNA and these will be purified with the goal of establishing the replication-dependent assembling in vitro with purified proteins. The proposed studies will determine the mechanism of initiation and elongation of DNA replication and replication-coupled chromatin assembly, all important aspects of DNA metabolism in proliferating cells. These studies are directly relevant to the overall goals of the program project.

了解人体细胞中 DNA 复制的机制是 生物学中的一个基本问题，与癌症直接相关 研究。 为了实现这一目标，DNA复制机制 来自猿猴病毒 40 (SV40) 和牛乳头瘤病毒 将研究 DNA 复制的 (BPV) 起源。 此外， DNA复制依赖的机制和调控 染色质组装也将被研究。 使用无细胞提取物 由人类细胞和纯化的 SV40 T 抗原或 BPV E1 和 E2 蛋白，病毒 DNA 复制所需的因子 并对其作用机制进行研究。 一种细胞蛋白 对于 BVP DNA 复制至关重要，但对于 SV40 DNA 则不然 复制，将被纯化并表征其功能。 这 涉及复制的 DNA 聚合酶转换机制 C 因子和增殖细胞核抗原 (PCNA) 也会 研究这些蛋白质与细胞的相互作用 除法周期调节器，第 21 页。 这些蛋白质似乎也发挥作用 修复辐射损伤的 DNA，因此，这些研究将 提供对协调 DNA 复制的控制的深入了解， DNA 修复和细胞周期进展。 最后，功能 染色质组装因子 1 (CAF1) 及其在 DNA 复制中的作用 - 将确定依赖的染色质组装。 其他染色质 组装因子与 CAF1 配合将核小体组装到 复制DNA，这些将被纯化，目的是 建立体外复制依赖性组装 纯化的蛋白质。 拟议的研究将确定 DNA复制的起始和延伸机制 复制耦合染色质组装，所有重要方面 增殖细胞中的 DNA 代谢。 这些研究直接 与计划项目的总体目标相关。