Institute of Cell Biology, University of Edinburgh, Edinburgh, United Kingdom.
PLoS One. 2009 Aug 6;4(8):e6542. doi: 10.1371/journal.pone.0006542.
DNA double-strand breaks can be repaired by homologous recombination involving the formation and resolution of Holliday junctions. In Escherichia coli, the RuvABC resolvasome and the RecG branch-migration enzyme have been proposed to act in alternative pathways for the resolution of Holliday junctions. Here, we have studied the requirements for RuvABC and RecG in DNA double-strand break repair after cleavage of the E. coli chromosome by the EcoKI restriction enzyme. We show an asymmetry in the ability of RuvABC and RecG to deal with joint molecules in vivo. We detect linear DNA products compatible with the cleavage-ligation of Holliday junctions by the RuvABC pathway but not by the RecG pathway. Nevertheless we show that the XerCD-mediated pathway of chromosome dimer resolution is required for survival regardless of whether the RuvABC or the RecG pathway is active, suggesting that crossing-over is a common outcome irrespective of the pathway utilised. This poses a problem. How can cells resolve joint molecules, such as Holliday junctions, to generate crossover products without cleavage-ligation? We suggest that the mechanism of bacterial DNA replication provides an answer to this question and that RecG can facilitate replication through Holliday junctions.
DNA 双链断裂可以通过同源重组修复，涉及 Holliday 连接点的形成和解决。在大肠杆菌中，RuvABC 解旋酶和 RecG 分支迁移酶被提出在 Holliday 连接点的解决中发挥替代途径的作用。在这里，我们研究了 RuvABC 和 RecG 在 EcoKI 限制酶切割大肠杆菌染色体后的 DNA 双链断裂修复中的要求。我们显示了 RuvABC 和 RecG 在体内处理连接分子的能力存在不对称性。我们检测到线性 DNA 产物与 RuvABC 途径的 Holliday 连接点的切割-连接兼容，但与 RecG 途径不兼容。然而，我们表明，无论 RuvABC 或 RecG 途径是否活跃，XerCD 介导的染色体二聚体解析途径的生存都是必需的，这表明交叉是一种常见的结果，无论所使用的途径如何。这带来了一个问题。细胞如何解决连接分子，如 Holliday 连接点，以产生交叉产物而不进行切割-连接？我们认为细菌 DNA 复制的机制为这个问题提供了答案，并且 RecG 可以通过 Holliday 连接点促进复制。
