Dynamics of DNA replication initiation complex in fission yeast during cell cycle

细胞周期中裂殖酵母 DNA 复制起始复合物的动态

• 批准号: 10480193
• 负责人: MURAKAMI Yota
• 金额: $ 2.37万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10480193/
• 关键词: Fission yeast; chromosomal Replication; ORC; Cell cycle; check point control; 複製開始

The origin recognition complex (ORC) has been identified as a protein complex of six subunits which binds to an replication origin in S. cerevisiae. The role of ORC is considered as a landing pad for replication factors. The subsequent studies suggest the involvement of ORC in several processes other than DNA replication including silencing, checkpoint control, and mitosis. However, it had been unclear to what extent ORC is involved in the regulation of these processes. We characterized a fifth subunit of ORC in S. pombe, Orp5 (ORC related protein 5). The orp5ィイD1+ィエD1 gene was essential for cell viability, and its depletion resulted in incomplete S-phase and inability to enter mitosis. Further analysis of three different alleles of orp5 temperature sensitive mutants elucidated the multiple functions of Orp5 which were separable : the initiation of DNA replication, S/M-checkpoint, S-phase progression, S-phase checkpoint, and mitosis. The orp5-H19 mutant was defective in DNA replication initiation and checkpoint control which prevent mitosis until S-phase is complete. The other mutant, orp5-K37 appeared to exit G1-phase, but the S-phase was stalled. The orp5-H37 has a deficiency entering M-phase while DNA synthesis itself is complete. In addition, the orp5-H37 was defective in DNA-damage checkpoint in S-phase which are regulated by rad3-cds1 pathway. Taken together, we propose that ORC is a chromatin component which constitutes a base for these events and manage cells to proceed the cell cycle properly.

起点识别复合物（ORC）已被鉴定为由六个亚基组成的蛋白质复合物，其与酿酒酵母中的复制起点结合。 ORC 的作用被认为是复制因子的着陆平台。随后的研究表明 ORC 参与 DNA 复制以外的几个过程，包括沉默、检查点控制和有丝分裂。然而，目前尚不清楚 ORC 在多大程度上参与了这些过程的监管。我们鉴定了粟酒裂殖酵母中 ORC 的第五个亚基 Orp5（ORC 相关蛋白 5）。 orp5ィイD1+ィエD1基因对于细胞活力至关重要，其缺失会导致S期不完整且无法进入有丝分裂。对orp5温度敏感突变体的三个不同等位基因的进一步分析阐明了Orp5可分离的多种功能：DNA复制的起始、S/M检查点、S期进展、S期检查点和有丝分裂。 orp5-H19突变体在DNA复制起始和检查点控制方面存在缺陷，这会阻止有丝分裂直到S期完成。另一个突变体 orp5-K37 似乎退出 G1 期，但 S 期停滞。 orp5-H37 有缺陷进入 M 期，而 DNA 合成本身已完成。此外，orp5-H37在S期DNA损伤检查点存在缺陷，该检查点受rad3-cds1途径调节。综上所述，我们认为 ORC 是一种染色质成分，它构成这些事件的基础并管理细胞以正确地进行细胞周期。