Ivanov E L, Sugawara N, Fishman-Lobell J, Haber J E
Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, Massachusetts 02254-9110, USA.
Genetics. 1996 Mar;142(3):693-704. doi: 10.1093/genetics/142.3.693.
HO endonuclease-induced double-strand breaks (DSBs) within a direct duplication of Escherichia coli lacZ genes are repaired either by gene conversion or by single-strand annealing (SSA), with > 80% being SSA. Previously it was demonstrated that the RAD52 gene is required for DSB-induced SSA. In the present study, the effects of other genes belonging to the RAD52 epistasis group were analyzed. We show that RAD51, RAD54, RAD55, and RAD57 genes are not required for SSA irrespective of whether recombination occurred in plasmid or chromosomal DNA. In both plasmid and chromosomal constructs with homologous sequences in direct orientation, the proportion of SSA events over gene conversion was significantly elevated in the mutant strains. However, gene conversion was not affected when the two lacZ sequences were in inverted orientation. These results suggest that there is a competition between SSA and gene conversion processes that favors SSA in the absence of RAD51, RAD54, RAD55 and RAD57. Mutations in RAD50 and XRS2 genes do not prevent the completion, but markedly retard the kinetics, of DSB repair by both mechanisms in the lacZ direct repeat plasmid, a result resembling the effects of these genes during mating-type (MAT) switching.
在大肠杆菌lacZ基因的直接重复序列内，HO核酸内切酶诱导的双链断裂（DSB）可通过基因转换或单链退火（SSA）进行修复，其中超过80%是通过SSA修复。此前已证明，DSB诱导的SSA需要RAD52基因。在本研究中，分析了属于RAD52上位性组的其他基因的作用。我们发现，无论重组发生在质粒还是染色体DNA中，RAD51、RAD54、RAD55和RAD57基因对于SSA都不是必需的。在具有直接同向同源序列的质粒和染色体构建体中，突变菌株中SSA事件相对于基因转换的比例显著升高。然而，当两个lacZ序列反向排列时，基因转换不受影响。这些结果表明，在缺乏RAD51、RAD54、RAD55和RAD57的情况下，SSA和基因转换过程之间存在竞争，这种竞争有利于SSA。RAD50和XRS2基因的突变并不妨碍在lacZ直接重复质粒中通过这两种机制进行的DSB修复的完成，但会显著延缓其动力学，这一结果类似于这些基因在交配型（MAT）转换过程中的作用。
