Rapid cycling and precocious termination of G1 phase in cells expressing CDK1AF
Rapid cycling and precocious termination of G1 phase in cells expressing CDK1AF
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DOI：
10.1091/mbc.e08-02-0172
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发表时间：
2008-08-01
影响因子：
3.3
通讯作者：
Ferrell, James E., Jr.
中科院分区：
文献类型：
--
作者：
Pomerening, Joseph R.;Ubersax, Jeffrey A.;Ferrell, James E., Jr.
在非洲爪蟾胚胎中，细胞周期由一个自主的生化振荡器驱动，该振荡器控制周期蛋白B1-CDK 1的周期性激活和失活。振荡器电路包括三个相互连接的正反馈回路和双负反馈回路（CDK 1-> Cdc 25-> CDK 1; CDK 1-竖线Wee 1-竖线CDK 1;以及CDK 1-竖线Myt 1-竖线CDK 1）的系统，这些回路共同用作触发器。先前的工作确定，这种触发器对于早期胚胎细胞周期中的CDK 1振荡至关重要。在这里，我们评估触发器在体细胞周期中的重要性，检查点和其他调控机制可能会使其无效。我们的方法是在HeLa细胞中表达磷酸化位点突变体CDK 1AF，该突变体使反馈回路短路，并通过活细胞荧光显微镜监测细胞周期进程。我们发现，表达CDK 1AF的细胞进行相对正常的第一次有丝分裂，但随后经历周期蛋白B1积累和破坏的快速周期，间隔3-6小时。在这些周期中，细胞进入和退出M期样状态，而不进行胞质分裂或核分裂。在Wee 1敲低细胞中观察到表型相似的快速周期。这些研究结果表明，CDK 1，Wee 1/Myt 1，和Cdc 25之间的相互作用是必需的G1期的建立，为正常的类似20小时的细胞周期期间，和开关样振荡的周期蛋白B1丰度特征的体细胞周期。我们建议，HeLa细胞周期是建立在一个不可靠的负反馈振荡器和正常的高可靠性，缓慢的步伐和开关样的字符周期是由一个负反馈CDK 1/Wee 1/Myt 1/Cdc 25系统。
In Xenopus embryos, the cell cycle is driven by an autonomous biochemical oscillator that controls the periodic activation and inactivation of cyclin B1-CDK1. The oscillator circuit includes a system of three interlinked positive and double-negative feedback loops (CDK1 -> Cdc25 -> CDK1; CDK1 -vertical bar Wee1 -vertical bar CDK1; and CDK1 -vertical bar Myt1 -vertical bar CDK1) that collectively function as a bistable trigger. Previous work established that this bistable trigger is essential for CDK1 oscillations in the early embryonic cell cycle. Here, we assess the importance of the trigger in the somatic cell cycle, where checkpoints and additional regulatory mechanisms could render it dispensable. Our approach was to express the phosphorylation site mutant CDK1AF, which short-circuits the feedback loops, in HeLa cells, and to monitor cell cycle progression by live cell fluorescence microscopy. We found that CDK1AF-expressing cells carry out a relatively normal first mitosis, but then undergo rapid cycles of cyclin B1 accumulation and destruction at intervals of 3-6 h. During these cycles, the cells enter and exit M phase-like states without carrying out cytokinesis or karyokinesis. Phenotypically similar rapid cycles were seen in Wee1 knockdown cells. These findings show that the interplay between CDK1, Wee1/Myt1, and Cdc25 is required for the establishment of G1 phase, for the normal similar to 20-h cell cycle period, and for the switch-like oscillations in cyclin B1 abundance characteristic of the somatic cell cycle. We propose that the HeLa cell cycle is built upon an unreliable negative feedback oscillator and that the normal high reliability, slow pace and switch-like character of the cycle is imposed by a bistable CDK1/Wee1/Myt1/Cdc25 system.