Identification of recG genetic interactions in Escherichia coli by transposon sequencing

Maintaining the integrity of the genome is of utmost importance for cell division and propagation. In Escherichia coli, the RecG protein has been implicated in processing branched recombination intermediates during DNA repair processes, but the primary cellular role(s) of RecG and the repair pathways in which it acts in have been difficult to define. To gain additional insight into RecG function, we employed transposon sequencing (Tn-seq) to identify recG genetic interactions and reveal complementary or redundant functions. The strongest hits from the screen were the dam, uvrD, rnhA, radA, and rep genes. The conditional importance of these five genes in cells lacking recG was confirmed using a plasmid-based assay, revealing synthetic lethal interactions for most double deletion strains. Several of the synthetic lethal gene combinations were suppressed by deletion of recF or recO, indicating that their genetic relationships involved roles in ssDNA gap repair. Additionally, loss of the RecG/SSB interaction phenocopied a recG deletion when combined with several of the top screen hits. The results support a role for RecG as a general genome guardian, where RecG alleviates toxic DNA intermediates resulting from recombination reactions.