Evidence that SbcB and RecF pathway functions contribute to RecBCD-dependent transductional recombination

Abstract

A role for the RecF, RecJ, and SbcB proteins in the RecBCD-dependent recombination pathway is suggested on the basis of the effect of null recF, recJ, and sbcB mutations in Salmonella typhimurium on a "short-homology" P22 transduction assay. The assay requires recombination within short (ϳ3-kb) sequences that flank the selected marker and lie at the ends of the transduced fragment. Since these ends are subject to exonucleolytic degradation, the assay may demand rapid recombination by requiring that the exchange be completed before the essential recombining sequences are degraded. In this assay, recF, recJ, and sbcB null mutations, tested individually, cause a small decrease in recombinant recovery but all pairwise combinations of these mutations cause a 10-to 30-fold reduction. In a recD mutant recipient, which shows increased recombination, these pairwise mutation combinations cause a 100-fold reduction in recombinant recovery. In a standard transduction assay (about 20 kb of flanking sequence), recF, recJ, and sbcB mutations have a very small effect on recombinant frequency. We suggest that these three proteins promote a rate-limiting step in the RecBCdependent recombination process. The above results were obtained with a lysogenic recipient strain which represses expression of superinfecting phage genomes and minimizes the contribution of phage recombination functions. When a nonlysogenic recipient strain is used, coinfecting phage genomes express functions that alter the genetic requirements for recombination in the short-homology assay.

A broadening view of recombinational DNA repair in bacteria

“…1 are not necessarily as distinct as shown. For example, some evidence exists that the RecF pathway functions may participate in RecBCDmediated recombination pathways under at least some conditions (Miesel & Roth 1996). Second, the resolution of the recombination crossover in either pathway can occur in two ways.…”

Single-Strand-Specific Exonucleases Prevent Frameshift Mutagenesis by Suppressing SOS Induction and the Action of DinB/DNA Polymerase IV in Growing Cells

“…First, defects in recombination are seen when two or three are defective (30,35,46). Second, we show here that lack of even two can be mutagenic via increased SOS induction and PolIV up-regulation.…”

“…Single-strand-specific exonucleases of either 5Ј or 3Ј polarity are required for normal levels of homologous recombination (30,35,46) and presumably recombinational repair. In their absence, recombination of linear substrates is reduced (30,35,46), suggesting that processing of double-strand ends to yield either 3Ј or 5Ј singlestrand overhangs promotes recombination (35,37,40).In the E. coli methyl-directed mismatch repair system, single-strand-specific exonucleases are required for degradation of the nicked, error-containing strand in vitro (44). The polarity of the exonuclease required depends on the location of the incision relative to the mismatch, and any of the four semiredundant single-strand-specific exonucleases is sufficient (44).…”

“…Despite their differing polarities, these exonucleases are wholly or partially redundant for their functions in vivo. Single-strand-specific exonucleases of either 5Ј or 3Ј polarity are required for normal levels of homologous recombination (30,35,46) and presumably recombinational repair. In their absence, recombination of linear substrates is reduced (30,35,46), suggesting that processing of double-strand ends to yield either 3Ј or 5Ј singlestrand overhangs promotes recombination (35,37,40).…”

Role of the RecBCD Recombination Pathway in Salmonella Virulence

“…This result suggests that either exonuclease V or IX is required for systemic infection and hence that RecJ can participate in RecBCD-mediated recombinational repair in the absence of the RecD subunit. Previous observations have suggested that RecD and RecJ provide alternative functions for recombinational repair of DNA damage in both Salmonella (11,23,27) and E. coli (39). Function redundancy may thus explain why RecD Ϫ and RecJ Ϫ single mutants are virulent (Table 2; see also reference 40).…”