Ruvolo Michael V, Mach Kathleen E, Burkholder William F
Department of Biological Sciences, Stanford University, 371 Serra Mall, Stanford, CA 94305-5020, USA.
Mol Microbiol. 2006 Jun;60(6):1490-508. doi: 10.1111/j.1365-2958.2006.05167.x.
Cells of Bacillus subtilis actively co-ordinate the initiation of sporulation with DNA replication and repair. Conditions that perturb replication initiation or replication elongation induce expression of a small protein, Sda, that specifically inhibits the histidine kinases required to initiate spore development. Previously, the role of Sda has been studied during chronic blocks to DNA replication. Here we show that induction of Sda is required to delay the initiation of sporulation when replication elongation is transiently blocked or after UV irradiation. During the recovery phase, cells efficiently sporulated, but this required the proteolysis of Sda. The rapid proteolysis of Sda required the ClpXP protease and the uncharged C-terminal sequence of Sda. Replacing the last two residues of Sda, both serines, with aspartic acids markedly stabilized Sda. Strains expressing sdaDD from the endogenous sda locus were unable to efficiently initiate sporulation after transient replication stress. We conclude that the Sda replication checkpoint is required to delay the initiation of sporulation when DNA replication is transiently perturbed, and that the intrinsic instability of Sda contributes to shutting off the pathway. The Sda checkpoint thus co-ordinates early events of spore development, including the polar cell division, with successful completion of chromosome replication.
枯草芽孢杆菌的细胞积极地将芽孢形成的起始与DNA复制和修复协调起来。干扰复制起始或复制延伸的条件会诱导一种小蛋白Sda的表达，该蛋白特异性抑制启动孢子发育所需的组氨酸激酶。此前，Sda的作用已在DNA复制的慢性阻滞过程中得到研究。在此我们表明，当复制延伸被短暂阻断或在紫外线照射后，Sda的诱导对于延迟芽孢形成的起始是必需的。在恢复阶段，细胞能够高效地形成芽孢，但这需要Sda的蛋白水解。Sda的快速蛋白水解需要ClpXP蛋白酶和Sda的不带电荷的C末端序列。将Sda的最后两个残基（均为丝氨酸）替换为天冬氨酸可显著稳定Sda。在内源性sda基因座表达sdaDD的菌株在短暂的复制应激后无法有效地起始芽孢形成。我们得出结论，当DNA复制被短暂干扰时，Sda复制检查点对于延迟芽孢形成的起始是必需的，并且Sda的内在不稳定性有助于关闭该途径。因此，Sda检查点将孢子发育的早期事件（包括极性细胞分裂）与染色体复制的成功完成协调起来。
