Study of the involvement of PolIV, ImuB and DnaE2 error-prone DNA Polymerases in UV-induced and stationary-phase mutagenesis in Pseudomonas aeruginosa

FELIZIANI S; LUJÁN AM; MOYANO AJ; SMANIA AM

San Miguel de Tucumán

Congreso; XLV Reunión Anual de la SAIB; 2009

Here, we studied the role of PolIV, ImuB and DnaE2 in Pseudomonas aeruginosa (PA) stress-induced mutagenesis. We constructed PAO1 dinB, imuB and dnaE2 mutants and analyzed their role in UV-induced mutagenesis. After UV irradiation, PAO1 showed a 2.6 fold increase (P=0.038) in the mutation frequency confirming that PA displays a subtle UV-induced mutator phenotype. PolIV deficiency resulted in levels of UV-induced mutagenesis similar to the wild type strain (ratio 3.6; P=0.019). In contrast, imuB (ratio 0.83, P=0.09) and dnaE2 (ratio 0.72, P=0.2) showed no increase in the mutation frequencies after induction, indicating that UV-induced mutagenesis in PA might depend on ImuB and DnaE2 but not on PolIV. We further investigated the role of these DNA polymerases in stationary-phase mutagenesis using a plasmid-based test system that allows the detection of -1 deletions and substitutions. These plasmids contain the reporter gene pheA harboring loss-of-function mutations that produce phenol monooxygenase inactivation, but whose reversion allows bacteria to grow using phenol as the sole carbon source. Results obtained from the incubation of PAO1 carrying the plasmids on phenol selective plates showed that PA displays stationary-phase mutagenesis. We are currently analyizing dinB, imuB and dnaE2 strains in order to study the role of these DNA polymerases in stationary-phase mutagenesis.