Genetic analysis of transductional recombination in Escherichia coli reveals differences in the postsynaptic stages of RecBCD and RecFOR pathways

Pub Date : 2023-05-05 DOI:10.18054/pb.v124i3-4.23604
K. Zahradka, J. Repar, Damir Đermić, D. Zahradka
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Abstract

Background and purpose: Homologous recombination in Escherichia coli proceeds via two pathways, RecBCD and RecFOR, which use different enzymes for DNA end resection and loading of RecA recombinase. The postsynaptic reactions following RecA-mediated homologous pairing have mostly been studied within the RecBCD pathway. They involve RuvABC helicase/resolvase complex, RecG and RadA helicases that process recombination intermediates to produce recombinant DNA molecules. Also, RecG functionally interacts with the PriA protein in initiation of recombination-dependent replication. Here, we studied the individual and combined effects of ruvABC, recG and radA null mutations on transductional recombination in both pathways. The effect of the priA300 mutation, which acts as a suppressor of the recG mutation, was also tested. The goal was to characterize the postsynaptic stage of transductional recombination in more details, especially in the RecFOR pathway, which is less well-studied. Materials and methods: Phage P1vir-mediated transduction was used to measure recombination efficiency in a series of recombination mutants. The proA+ marker was used for selection in transductional crosses with various ΔproA recipients. Results: The ruvABC mutation moderately decreased recombination in both recombination pathways, while radA had no effect. The recG mutation reduced recombination in the RecBCD pathway but not in the RecFOR pathway. The strong recombination defect of recG radA double mutants in both pathways was completely suppressed by the priA300 mutation, and this suppression depended on the functional RuvABC complex. Conclusions: RecG and RadA proteins have a redundant role in transductional recombination via RecFOR pathway. In both recombination pathways, RecG and RadA functionally interact with PriA, probably during initiation of recombination-dependent replication.
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大肠杆菌转导重组的遗传分析揭示了RecBCD和RecFOR途径突触后阶段的差异
背景和目的:大肠杆菌中的同源重组通过RecBCD和RecFOR两种途径进行,这两种途径使用不同的酶进行DNA末端切除和装载RecA重组酶。RecA介导的同源配对后的突触后反应主要在RecBCD途径中进行研究。它们涉及RuvABC解旋酶/再分解酶复合物、RecG和RadA解旋酶,它们处理重组中间体以产生重组DNA分子。此外,RecG在重组依赖性复制的启动过程中与PriA蛋白功能性相互作用。在这里,我们研究了ruvABC、recG和radA-null突变对两种途径中转导重组的单独和联合影响。还测试了作为recG突变抑制剂的priA300突变的效果。目的是更详细地描述转导重组的突触后阶段,特别是在研究较少的RecFOR途径中。材料和方法:Phage P1vir介导的转导用于测量一系列重组突变体的重组效率。proA+标记用于与不同ΔproA受体的转导杂交中的选择。结果:ruvABC突变适度降低了两种重组途径中的重组,而radA没有影响。recG突变减少了RecBCD途径中的重组,但没有减少RecFOR途径中的复合。两种途径中recG-radA双突变体的强重组缺陷被priA300突变完全抑制,这种抑制依赖于功能性RuvABC复合物。结论:RecG和RadA蛋白在通过RecFOR途径进行转导重组中具有冗余作用。在两种重组途径中,RecG和RadA可能在重组依赖性复制的起始过程中与PriA发生功能性相互作用。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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