Nonperiodic activity of the human anaphase-promoting complex-Cdh1 ubiquitin ligase results in continuous DNA synthesis uncoupled from mitosis
Nonperiodic activity of the human anaphase-promoting complex-Cdh1 ubiquitin ligase results in continuous DNA synthesis uncoupled from mitosis
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DOI：
10.1128/mcb.20.20.7613-7623.2000
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发表时间：
2000-10-01
影响因子：
5.3
通讯作者：
Lukas, J
中科院分区：
文献类型：
--
作者：
Sorensen, CS;Lukas, C;Lukas, J
泛素-蛋白酶体介导的限速蛋白的破坏是通过主要细胞周期转换及时进展所必需的。由Cdh 1亚基周期性激活的后期促进复合物（APC）是主要的细胞泛素连接酶之一，在酿酒酵母和果蝇属中，触发退出有丝分裂和在G（1）阻止非计划DNA复制。在这项研究中，我们调查的APC-Cdh 1活性的周期性振荡的重要性，在人类细胞的细胞周期进程。我们发现，在G（1）/S转换APC-Cdh 1解离的条件干扰导致无法积累一个令人惊讶的范围广泛的关键有丝分裂调节，包括细胞周期蛋白B1，细胞周期蛋白A，Plk 1，Pds 1，有丝分裂素（CENP-F），Aim 1，和Cdc 20。出乎意料的是，尽管组成型组装的APC-Cdh 1也延迟了G（1）/S转换，降低了S期DNA合成的速率，但DNA复制所必需的一些活性明显增强，这主要是由于E2 F依赖性细胞周期蛋白E转录的进行性增加和p27（Kip 1）细胞周期蛋白依赖性激酶抑制剂的快速周转。因此，未能使APC-Cdh 1在G（1）/S转换之后停止，不仅抑制了生产性细胞分裂，而且支持缓慢但不间断的DNA复制，排除了S期退出并导致基因组的大规模过度复制。我们的数据表明，APC-Cdh 1泛素连接酶活性的及时振荡是协调DNA复制与细胞分裂的重要步骤，并且调控APC与Cdh 1活化亚基的关联的机制的失败可以破坏哺乳动物细胞中的基因组稳定性。
Ubiquitin-proteasome-mediated destruction of rate-limiting proteins is required for timely progression through the main cell cycle transitions. The anaphase-promoting complex (APC), periodically activated by the Cdh1 subunit, represents one of the major cellular ubiquitin ligases which, in Saccharomyces cerevisiae and Drosophila spp., triggers exit from mitosis and during G(1) prevents unscheduled DNA replication. In this study we investigated the importance of periodic oscillation of the APC-Cdh1 activity for the cell cycle progression in human cells. We show that conditional interference with the APC-Cdh1 dissociation at the G(1)/S transition resulted in an inability to accumulate a surprisingly broad range of critical mitotic regulators including cyclin B1, cyclin A, Plk1, Pds1, mitosin (CENP-F), Aim1, and Cdc20. Unexpectedly, although constitutively assembled APC-Cdh1 also delayed G(1)/S transition and lowered the rate of DNA synthesis during S phase, some of the activities essential for DNA replication became markedly amplified, mainly due to a progressive increase of E2F-dependent cyclin E transcription and a rapid turnover of the p27(Kip1) cyclin-dependent kinase inhibitor. Consequently, failure to inactivate APC-Cdh1 beyond the G(1)/S transition not only inhibited productive cell division but also supported slow but uninterrupted DNA replication, precluding S-phase exit and causing massive overreplication of the genome. Our data suggest that timely oscillation of the APC-Cdh1 ubiquitin ligase activity represents an essential step in coordinating DNA replication with cell division and that failure of mechanisms regulating association of APC with the Cdh1 activating subunit can undermine genomic stability in mammalian cells.