BIOCHEMICAL ANALYSIS OF MITOTIC CHROMOSOME CONDENSATION

有丝分裂染色体凝聚的生化分析

• 批准号: 6636164
• 负责人: TATSUYA HIRANO
• 金额: $ 36.2万
• 依托单位: COLD SPRING HARBOR LABORATORY
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-05-01 至 2004-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6636164
• 关键词: DNA binding protein; Xenopus oocyte; adenosinetriphosphatase; cell cycle; cell cycle proteins; chromatin; chromosome movement; enzyme complex; laboratory mouse; protein purification; protein reconstitution; protein structure function; ultracentrifugation; ultraviolet spectrometry

Chromosome condensation is an essential cellular process that ensures the faithful segregation of genetic information during mitosis and meiosis. The long-term goal of this project is to understand how this dynamic and highly orchestrated process of DNA-protein assembly is achieved, and how it is regulated precisely during the eukaryotic cell cycle. A major emphasis will be made on the structural and functional characterization of 13S condensin, a recently identified protein complex that plays a key role in this process. 13S condensin is a DNA-stimulated ATPase that consists of two SMC (structural maintenance of chromosomes) subunits and three non-SMC subunits. The complex has an ability to reconfigure DNA by introducing global positive writhe in an ATP-dependent manner. In this proposal, (1) the five-subunit complex will be dissected biochemically to determine how individual subunits and subcomplexes contribute to condensin functions. (2) A cell cycle-dependent interaction between 13S condensin and chromatin will be reconstituted in vitro and the molecular basis of mitosis-specific condensation will be determined. (3) Structural, cell biological and biophysical approaches will be combined to understand when, where and how the condensin complex works in the context of higher-order chromosome structure. (4) New structural components that cooperate with 13S condensin to build up mitotic chromosomes will be identified and characterized. The information obtained from this work will ultimately contribute to a better understanding of human health because chromosome anomalies, such as aneuploidy and translocations, are tightly associated with tumor development or birth defects.

染色体凝聚是一个重要的细胞过程，在有丝分裂和减数分裂过程中确保遗传信息的忠实分离。这个项目的长期目标是了解这个动态的和高度协调的DNA-蛋白质组装过程是如何实现的，以及它如何在真核细胞周期中受到精确调控。重点将放在13S凝集素的结构和功能特征上，这是最近发现的一种在这一过程中发挥关键作用的蛋白质复合体。13S凝集素是一种DNA刺激的ATPase，由两个SMC亚基和三个非SMC亚基组成。该复合体具有通过以依赖于ATP的方式引入全球正扭曲来重新配置DNA的能力。在这项建议中，(1)将对五个亚基复合体进行生化解剖，以确定单个亚基和亚复合体如何对凝集素功能做出贡献。(2)13S凝集素与染色质之间依赖于细胞周期的相互作用将在体外重建，并将确定有丝分裂特异性凝聚的分子基础。(3)结构、细胞生物学和生物物理方法将结合起来，以了解凝集素复合体在高阶染色体结构中何时、何地和如何发挥作用。(4)与13S凝集素协同构建有丝分裂染色体的新结构成分将被鉴定和鉴定。从这项工作中获得的信息最终将有助于更好地了解人类健康，因为染色体异常，如非整倍体和易位，与肿瘤发展或出生缺陷密切相关。