RecA-dependent and independent recombination in Pseudomonas aeruginosa and Escherichia coli

MORO CAMILA; BORGOGNO MARÍA VICTORIA; MONTI MARIELA R; ARGARAÑA CARLOS E.

Congreso; Reunión Conjunta de Sociedades de Biociencias; 2017

Genetic recombinationparticipates in multiple physiological pathways that are crucial for themaintenance and diversification of the genome. The recombinase RecA is acentral factor in this process, mediating the exchange between DNA moleculescontaining perfect homology (homologous, HO) or very low divergence(homeologous, HE). At present multiple mechanisms are known to mediate genetic recombination,both dependent and independent of RecA. In order to study the recombinationprocess in Gram-negative bacteria, we used a LacZ based system to determineboth HO and HE recombination. Using this system, we determined therecombination rates in wild-type (WT) and RecA deficient (ΔrecA) strains of Pseudomonas aeruginosa and Escherichiacoli.Moreover, the recombination process creates a functional copy of the lacZ gene and thus, therecombinant clones can be detected by their β galactosidase activity.In both bacteria, HO recombination rates were approximately 60-200-fold higherthan HE recombination rates. In the RecA-deficient E. coli strain, both HO and HE recombination rateswere 60-fold lower than that obtained for the WT strain. The recA deletionmutant of P.aeruginosa showedthat HO and HE recombination rates decreased 15 and 3-fold respectively,compared with the WT strain. When the β galactosidase activityof recombined clones was determined, we found that all E. coli clones showed enzymaticactivity asit was expected. In P. aeruginosa however, many clones showed no β galactosidase activity. Furthermore, the molecularanalysis of therecombined regions of these clones showed the presence of point mutationsand also deletions of as much as 800bp. These results indicate the existence ofa significantly mutagenic recombination mechanism, independent of RecA in P. aeruginosa, at difference of E. coli.