The Recombination Genes addAB Are Not Restricted to Gram-Positive Bacteria: Genetic Analysis of the Recombination Initiation Enzymes RecF and AddAB in Rhizobium etli

Abstract

Single-strand gaps (SSGs) and double-strand breaks (DSBs) are the major initiation sites for recombination. In bacteria, the SSGs are repaired by RecFOR, while the DSBs are processed by RecBCD in gramnegative bacteria and AddAB in gram-positive bacteria. Unexpectedly, instead of recBCD genes, the addAB genes were found in members of the ␣-proteobacteria group (gram negative). Taking Rhizobium etli as a model, the role of recF and addAB genes in homologous recombination and repair of damaged DNA was evaluated. Inactivation of either recF or addA provoked strong sensitivity to UV radiation and mitomycin C, while an additive effect was observed in the recF-addA mutant. The DSBs generated by nalidixic acid caused low viability only in the addA mutant. The recombination frequency of large and small plasmids was reduced in the recF mutant (24-and 36-fold, respectively), whereas a slight decrease (threefold) in the addA mutant was observed. Moreover, an additive effect (47-and 90-fold, respectively) was observed in the double mutant, but it was not as dramatic as that in a recA mutant. Interestingly, the frequency of deletion and Campbell-type recombination was slightly affected in either single or double mutants. These results suggest that another pathway exists that allows plasmid and Campbell-type recombination in the absence of recF and addA genes.

Genome Sequence of Lactobacillus helveticus , an Organism Distinguished by Selective Gene Loss and Insertion Sequence Element Expansion

“…Alternatively, a missing or malfunctioning recombination enzyme(s) encoded by the DPC 4571 bacterium may have limited intragenomic rearrangements without affecting chromosomal segregation and repair of damaged DNA. The L. helveticus genome encodes a complete set of enzymes with significant similarity to the recognized bacterial recombination pathways in gram-positive bacteria (50). The DPC 4571 genome encodes predicted AddAB (Lhv1273-Lhv1274) and RecFOR (Lhv4, Lhv1300, and Lhv402, respectively) pathway proteins, in addition to orthologues of RecG (Lhv1394), RecJ (Lhv1329), RecQ (Lhv1098), RecN (Lhv1410), RecU (Lhv1266), RecX (Lhv678), and RuvAB (Lhv429-Lhv430).…”

Gene Conversion Tracts Associated with Crossovers in Rhizobium etli

“…To evaluate the association of crossover formation to gene conversion, cointegrates between pMC11 (harboring eight different RFLPs) and pSym were selected, as described in Materials and Methods. In R. etli, formation of cointegrates of this kind is strictly dependent on recA (50). Interestingly, when plasmid pMC11 was used as a donor, integrants were obtained at a very low frequency (1.63 ϫ 10 Ϫ7 Ϯ 0.52 ϫ 10 Ϫ7 ).…”

RecBCD Enzyme and the Repair of Double-Stranded DNA Breaks

“…An alternative class of enzymes, the AddAB family (also referred to as RexAB), was originally thought to fulfill the same break-processing function in a restricted niche of gram-positive bacteria (57,59,60,101,118). However, AddAB-type complexes are now appreciated to be more prevalent, having also been found widely in the proteobacteria (3,203,234,328). Like RecBCD, the AddAB enzyme is implicated in recombinational repair, and the inactivation of either subunit renders cells sensitive to DNA damage and reduces viability.…”