Sandler S J
Department of Microbiology, University of Massachusetts, Amherst 01003, USA.
Genetics. 2000 Jun;155(2):487-97. doi: 10.1093/genetics/155.2.487.
In Escherichia coli, the primosome assembly proteins, PriA, PriB, PriC, DnaT, DnaC, DnaB, and DnaG, are thought to help to restart DNA replication forks at recombinational intermediates. Redundant functions between priB and priC and synthetic lethality between priA2::kan and rep3 mutations raise the possibility that there may be multiple pathways for restarting replication forks in vivo. Herein, it is shown that priA2::kan causes synthetic lethality when placed in combination with either Deltarep::kan or priC303:kan. These determinations were made using a nonselective P1 transduction-based viability assay. Two different priA2::kan suppressors (both dnaC alleles) were tested for their ability to rescue the priA-priC and priA-rep double mutant lethality. Only dnaC809,820 (and not dnaC809) could rescue the lethality in each case. Additionally, it was shown that the absence of the 3'-5' helicase activity of both PriA and Rep is not the critical missing function that causes the synthetic lethality in the rep-priA double mutant. One model proposes that replication restart at recombinational intermediates occurs by both PriA-dependent and PriA-independent pathways. The PriA-dependent pathways require at least priA and priB or priC, and the PriA-independent pathway requires at least priC and rep. It is further hypothesized that the dnaC809 suppression of priA2::kan requires priC and rep, whereas dnaC809,820 suppression of priA2::kan does not.
在大肠杆菌中，引发体组装蛋白PriA、PriB、PriC、DnaT、DnaC、DnaB和DnaG被认为有助于在重组中间体处重新启动DNA复制叉。PriB和PriC之间的冗余功能以及priA2::kan与rep3突变之间的合成致死性增加了体内可能存在多种重新启动复制叉途径的可能性。在此表明，priA2::kan与Deltarep::kan或priC303::kan组合时会导致合成致死性。这些测定是使用基于非选择性P1转导的活力测定法进行的。测试了两种不同的priA2::kan抑制子（均为dnaC等位基因）挽救priA-priC和priA-rep双突变致死性的能力。在每种情况下，只有dnaC809,820（而不是dnaC809）能够挽救致死性。此外，还表明PriA和Rep的3'-5'解旋酶活性的缺失不是导致rep-priA双突变体合成致死性的关键缺失功能。一种模型提出，在重组中间体处的复制重新启动通过PriA依赖性和PriA非依赖性途径发生。PriA依赖性途径至少需要priA和priB或priC，而PriA非依赖性途径至少需要priC和rep。进一步假设，dnaC809对priA2::kan的抑制需要priC和rep，而dnaC809,820对priA2::kan的抑制则不需要。
