BIOCHEMICAL ANALYSIS OF MITOTIC CHROMOSOME CONDENSATION

有丝分裂染色体凝聚的生化分析

• 批准号: 6386253
• 负责人: TATSUYA HIRANO
• 金额: $ 34.17万
• 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/6386253
• 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，确保在有丝分裂和减数分裂期间遗传信息的忠实分离。 该项目的长期目标是了解这种动态和高度协调的DNA-蛋白质组装过程是如何实现的，以及它在真核细胞周期中是如何精确调控的。 一个主要的重点将在13 S凝聚素，最近确定的蛋白质复合物，在这一过程中发挥了关键作用的结构和功能特性。 13 S凝聚素是一种DNA刺激的ATP酶，由两个SMC亚基和三个非SMC亚基组成。 该复合物具有通过以ATP依赖的方式引入全局正扭来重构DNA的能力。 在这个提议中，（1）五亚基复合物将被生物化学解剖，以确定单个亚基和亚复合物如何有助于凝聚素功能。 (2)将在体外重建13 S凝聚素和染色质之间的细胞周期依赖性相互作用，并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学的方法将结合起来，以了解何时，何地以及如何在高阶染色体结构的背景下，凝聚素复合物的工作。 (4)与13 S凝聚素合作建立有丝分裂染色体的新结构组分将被鉴定和表征。 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。