Studies on Eukaryotic Replication

真核复制研究

• 批准号: 7903851
• 负责人: Jerard Hurwitz
• 金额: $ 30.99万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-31 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7903851
• 关键词: Binding; Cell Cycle; Cell Death; Cells; Complex; Coupled; DNA; DNA Primase; DNA biosynthesis; DNA-Directed DNA Polymerase; Event; Goals; Grant; Hereditary Disease; Human; Individual; Investigation; Laboratories; Lead; Malignant Neoplasms; Phosphotransferases; Polymerase; Pre-Replication Complex; Process; Property; Proteins; Reaction; Research; Role; Work; cohesin; helicase; in vivo; novel; protein complex

DESCRIPTION (provided by applicant): The goal of our research is to identify and characterize the machinery involved in eukaryotic DNA replication. We focus on the isolation of the individual proteins and protein complexes required for DNA synthesis and define their role in this critical macromolecular process. All growing cells must duplicate their DNA accurately and perform this feat once and only once per cell cycle. Events that interfere with these requirements can lead to cell death or altered proliferation that result in cancers. At present, many steps involved in supporting replication and its control are either unknown or poorly characterized. We plan to extend our previous work on various key replication proteins and their complexes that support the activation of initiation complexes required for the elongation of DNA chains. For this purpose, we will study: a) the role of a complex, containing Cdc45, the MCM2-7 complex and GINS (CMG) which contains DNA helicase activity and may contribute important to the unwinding reaction at the replication fork; b) the interaction between GINS and the Pol 1-primase complex which markedly stimulates its DNA polymerase activity; c) the properties of cloned Pol5 and its interaction with PCNA required for its processive elongation of primed DNA templates. We have also observed functional interactions of GINS with Pol5 as well as Pol4. We plan to examine these effects further and evaluate whether the CMG complex does the same. Coupled replication and unwinding reactions will be carried out which may contribute to strand selection; d) We plan to explore the role of the Cdc7 kinase on the activity and formation of the CMG complex. In vivo, this kinase was shown to influence the interaction of Cdc45 with the MCM2-7 complex. We will further characterize a novel interaction recently detected between the Cdc7 kinase complex and the cohesin loader complex Scc2/Scc4. The requirements for this interaction and its consequences will be investigated. All cells must make identical copies of their DNA to survive. Our studies are directed at how these copies are made and regulated. We plan to isolate the proteins involved in copying DNA and determine how they function. Results from these studies will have a direct impact on genetic diseases and cancer.

描述（由申请人提供）：我们研究的目标是识别和表征真核 DNA 复制所涉及的机制。我们专注于 DNA 合成所需的单个蛋白质和蛋白质复合物的分离，并确定它们在这一关键大分子过程中的作用。所有生长的细胞都必须准确地复制它们的 DNA，并且每个细胞周期只执行一次这一壮举。干扰这些要求的事件可能导致细胞死亡或增殖改变，从而导致癌症。目前，支持复制及其控制所涉及的许多步骤要么是未知的，要么是描述不明确。 我们计划扩展我们之前对各种关键复制蛋白及其复合物的研究，这些复制蛋白及其复合物支持 DNA 链延长所需的起始复合物的激活。为此，我们将研究：a) 包含 Cdc45、MCM2-7 复合物和 GINS (CMG) 的复合物的作用，该复合物含有 DNA 解旋酶活性，可能对复制叉处的解旋反应发挥重要作用； b) GINS 和 Pol 1-引物酶复合物之间的相互作用，显着刺激其 DNA 聚合酶活性； c) 克隆的 Pol5 的特性及其与引发的 DNA 模板持续延伸所需的 PCNA 相互作用。我们还观察到 GINS 与 Pol5 以及 Pol4 的功能相互作用。我们计划进一步研究这些影响并评估 CMG 复合物是否具有相同的作用。将进行耦合复制和解旋反应，这可能有助于链选择； d) 我们计划探索Cdc7激酶对CMG复合物的活性和形成的作用。在体内，该激酶被证明会影响 Cdc45 与 MCM2-7 复合物的相互作用。我们将进一步描述最近检测到的 Cdc7 激酶复合体和粘连蛋白装载复合体 Scc2/Scc4 之间的新型相互作用。将调查这种相互作用的要求及其后果。所有细胞都必须复制相同的 DNA 才能生存。我们的研究针对的是这些副本是如何制作和监管的。我们计划分离出参与 DNA 复制的蛋白质并确定它们的功能。这些研究的结果将对遗传疾病和癌症产生直接影响。