Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Division of Genome Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet and Science for Life Laboratory, Stockholm, Sweden.
Genome Damage and Stability Centre, University of Sussex, Brighton, UK.
Mol Cell. 2018 Jul 5;71(1):117-128.e3. doi: 10.1016/j.molcel.2018.05.026. Epub 2018 Jun 28.
To maintain genome stability, cells need to replicate their DNA before dividing. Upon completion of bulk DNA synthesis, the mitotic kinases CDK1 and PLK1 become active and drive entry into mitosis. Here, we have tested the hypothesis that DNA replication determines the timing of mitotic kinase activation. Using an optimized double-degron system, together with kinase inhibitors to enforce tight inhibition of key proteins, we find that human cells unable to initiate DNA replication prematurely enter mitosis. Preventing DNA replication licensing and/or firing causes prompt activation of CDK1 and PLK1 in S phase. In the presence of DNA replication, inhibition of CHK1 and p38 leads to premature activation of mitotic kinases, which induces severe replication stress. Our results demonstrate that, rather than merely a cell cycle output, DNA replication is an integral signaling component that restricts activation of mitotic kinases. DNA replication thus functions as a brake that determines cell cycle duration.
为了维持基因组的稳定性，细胞在分裂前需要复制其 DNA。在大量 DNA 合成完成后，有丝分裂激酶 CDK1 和 PLK1 变得活跃，并驱动细胞进入有丝分裂。在这里，我们检验了这样一个假设，即 DNA 复制决定了有丝分裂激酶激活的时间。我们使用优化的双 degron 系统，以及激酶抑制剂来严格抑制关键蛋白，发现无法起始 DNA 复制的人类细胞会过早地进入有丝分裂。阻止 DNA 复制的许可和/或启动会导致 CDK1 和 PLK1 在 S 期迅速激活。在存在 DNA 复制的情况下，抑制 CHK1 和 p38 会导致有丝分裂激酶过早激活，这会诱导严重的复制应激。我们的结果表明，DNA 复制不是细胞周期的输出，而是一个完整的信号成分，限制了有丝分裂激酶的激活。因此，DNA 复制作为一个刹车，决定了细胞周期的持续时间。
