Mec1 is activated at the onset of normal S phase by low dNTP pools impeding DNA replication [microarray]

The Mec1 and Rad53 kinases play a central role during acute replication stress in budding yeast. They are also essential for viability in normal growth conditions, but the signal that activates the Mec1–Rad53 pathway in the absence of exogenous insults is currently unknown. Here, we show that this pathway is active at the onset of normal S phase because dNTP levels present in G1 phase are not sufficient to support processive DNA synthesis and impede DNA replication. This activation can be suppressed experimentally by increasing dNTP levels in G1 phase. Moreover, we show that unchallenged cells entering S phase in the absence of Rad53 undergo irreversible fork collapse and mitotic catastrophe. Together, these data indicate that cells use dNTP shortage to detect the onset of DNA replication and activate the Mec1–Rad53 pathway, which in turn maintains functional forks and triggers dNTP synthesis, allowing the completion of DNA replication.Mec1与Rad53激酶在出芽酵母（budding yeast）的急性复制应激过程中发挥核心作用。二者在正常生长条件下对细胞存活亦不可或缺，但目前尚不清楚在无外源胁迫的情况下，激活Mec1–Rad53通路的信号分子为何。本研究证实，该通路在正常S期起始阶段便已激活，这是由于G1期的脱氧核苷三磷酸（deoxynucleoside triphosphate, dNTP）水平无法满足持续性DNA合成的需求，进而阻碍DNA复制进程。通过实验手段提升G1期的dNTP水平，可抑制该通路的激活。此外，本研究发现，缺失Rad53的未受胁迫细胞进入S期后，会发生不可逆的复制叉崩解与有丝分裂灾难。综合上述实验结果可知，细胞通过感知dNTP不足来检测DNA复制的起始，并激活Mec1–Rad53通路；该通路可维持功能性复制叉的稳定，并触发dNTP合成，从而保障DNA复制顺利完成。