Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
Mol Microbiol. 2022 Oct;118(4):426-442. doi: 10.1111/mmi.14976. Epub 2022 Sep 11.
DNA replication is highly regulated and primarily controlled at the step of initiation. In bacteria, the replication initiator DnaA and the origin of replication oriC are the primary targets of regulation. Perturbations that increase or decrease replication initiation can cause a decrease in cell fitness. We found that multiple mechanisms, including an increase in replication elongation and a decrease in replication initiation, can compensate for lethal over-initiation. We found that in Bacillus subtilis, under conditions of rapid growth, loss of yabA, a negative regulator of replication initiation, caused a synthetic lethal phenotype when combined with the dnaA1 mutation that also causes replication over-initiation. We isolated several classes of suppressors that restored viability to dnaA1 ∆yabA double mutants. Some suppressors (relA, nrdR) stimulated replication elongation. Others (dnaC, cshA) caused a decrease in replication initiation. One class of suppressors decreased replication initiation in the dnaA1 ∆yabA mutant by causing a decrease in the amount of the replicative helicase, DnaC. We found that decreased levels of helicase in otherwise wild-type cells were sufficient to decrease replication initiation during rapid growth, indicating that the replicative helicase is limiting for replication initiation. Our results highlight the multiple mechanisms cells use to regulate DNA replication.
DNA 复制受到高度调控，主要在起始阶段进行控制。在细菌中，复制起始因子 DnaA 和复制起点 oriC 是主要的调控靶标。增加或减少复制起始的干扰会导致细胞适应性降低。我们发现，多种机制，包括增加复制延伸和减少复制起始，可以补偿致死性的过度起始。我们发现，在枯草芽孢杆菌中，在快速生长的条件下，当与 dnaA1 突变（也导致复制过度起始）结合时，复制起始负调控因子 yabA 的缺失会导致合成致死表型。我们分离出几类能够恢复 dnaA1 ∆yabA 双突变体活力的抑制子。一些抑制子（relA、nrdR）刺激复制延伸。其他抑制子（dnaC、cshA）则减少复制起始。一类抑制子通过减少复制酶 DnaC 的量来减少 dnaA1 ∆yabA 突变体中的复制起始。我们发现，在其他方面为野生型的细胞中降低解旋酶的水平足以在快速生长期间减少复制起始，表明复制解旋酶对复制起始有限制作用。我们的结果强调了细胞用于调节 DNA 复制的多种机制。
