In response to DNA replication stress, DNA replication checkpoint kinase Mec1 phosphorylates Mrc1, which in turn activates Rad53 to prevent the generation of deleterious single-stranded DNA, a process that remains poorly understood. We previously reported that lagging-strand DNA synthesis proceeds farther than leading strand in rad53-1 mutant cells defective in replication checkpoint under replication stress, resulting in the exposure of long stretches of the leading-strand templates. Here, we show that asymmetric DNA synthesis is also observed in mec1-100 and mrc1-AQ cells defective in replication checkpoint but, surprisingly, not in mrc1∆ cells in which both DNA replication and checkpoint functions of Mrc1 are missing. Furthermore, depletion of either Mrc1 or its partner, Tof1, suppresses the asymmetric DNA synthesis in rad53-1 mutant cells. Thus, the DNA replication checkpoint pathway couples leading- and lagging-strand DNA synthesis by attenuating the replication function of Mrc1-Tof1 under replication stress.

为应对 DNA 复制压力，DNA 复制检查点激酶 Mec1 磷酸化 Mrc1，进而激活 Rad53 以防止有害单链 DNA 的产生，这一过程仍然知之甚少。我们之前报道过，在复制压力下，在复制检查点缺陷的rad53-1突变细胞中，滞后链 DNA 合成比前导链进行得更远，导致前导链模板的长链暴露。在这里，我们表明在复制检查点有缺陷的mec1-100和mrc1-AQ细胞中也观察到不对称 DNA 合成，但令人惊讶的是，在mrc1Δ中没有缺少 Mrc1 的 DNA 复制和检查点功能的细胞。此外，Mrc1 或其伙伴 Tof1 的消耗抑制了rad53-1突变细胞中的不对称 DNA 合成。因此，DNA 复制检查点途径通过在复制压力下减弱 Mrc1-Tof1 的复制功能来耦合前导链和滞后链 DNA 合成。