Regulation of cell cycle checkpoints that monitor DNA damage and DNA replication block.

Chkl调节哺乳动物中的DNA损伤和复制检查点。虽然Chk 1对胚胎细胞的存活至关重要，但其在正常细胞周期中的功能仍不清楚。使用Cre/lox条件小鼠系，我们在这里证明了Chk 1在调节胚胎细胞周期的S期进展和预防有丝分裂起始中的重要和不可或缺的作用。激光扫描细胞仪分析发现，Chk 1缺陷导致细胞有丝分裂提前，细胞周期蛋白B-Cdc 2异常激活导致染色体提前浓缩，组蛋白H3在丝氨酸10位磷酸化，细胞色素c释放导致caspase 3和caspase 9激活。这种缺陷与磷酸化组蛋白H2 AX（gH 2AX）形成病灶有关，特别是在S期。Cdc 2AF是一种不能在抑制位点磷酸化的突变体，其异位表达也诱导ES细胞和HCT 116细胞在S期由gH 2AX引起的过早有丝分裂和病灶形成。用咖啡因抑制ATM和ATR可防止过早有丝分裂引起的细胞死亡。p53缺陷细胞对过早有丝分裂的致死性也具有高度抗性。因此，我们的结果表明，ATM/ATR-p53是有丝分裂灾难所必需的，它消除了逃避Chk 1依赖性有丝分裂调节的细胞。该功能的丧失可能在哺乳动物肿瘤发生中是重要的。

Chkl regulates both DNA damage and replication checkpoints in mammals. Although Chk1 is crucial for embryonic cell survival, its function in normal cell cycle remains largely unknown. With the use of a Cre/lox-conditional mouse line, we demonstrate here the essential and indispensable role of Chk1 in regulation of S-phase progression and prevention of mitotic initiation in embryonic cell cycle. An analysis using laser scanning cytometer revealed that Chk1 deficiency resulted in premature mitosis with the premature chromosome condensation and the phosphorylation of histone H3 at serine10 due to abnormal activation of cyclin B-Cdc2, and led to the activation of caspases 3 and 9 triggered by cytoplasmic release of cytochrome c. This deficiency was associated with foci formation by the phosphorylated histone, H2AX (gH2AX), specifically at S phase. Ectopic expression of Cdc2AF, a mutant that cannot be phosphorylated at inhibitory sites, also induced premature mitosis and foci formation by gH2AX at S phase in both ES cells and HCT116 cells. Inhibition of ATM and ATR with caffeine protected against cell death from premature mitosis. p53-deficient cells were highly resistant to lethality from premature mitosis as well. Our results therefore suggest that ATM/ATR-p53 is required for mitotic catastrophe that eliminates cells escaping Chk1-dependent mitotic regulation. Loss of this function might be important in mammalian tumorigenesis.