DinB (DNA polymerase IV), ImuBC and RpoS contribute to the generation of ciprofloxacin-resistance mutations in Pseudomonas aeruginosa.

We investigated the role(s) of the damage-inducible SOS response dinB and imuBC gene products in the generation of ciprofloxacin-resistance mutations in the important human opportunistic bacterial pathogen, Pseudomonas aeruginosa. We found that the overall numbers of ciprofloxacin resistant (Cip) mutants able to be recovered under conditions of selection were significantly reduced when the bacterial cells concerned carried a defective dinB gene, but could be elevated to levels approaching wild-type when these cells were supplied with the dinB gene on a plasmid vector; in turn, firmly establishing a role for the dinB gene product, error-prone DNA polymerase IV, in the generation of Cip mutations in P. aeruginosa. Further, we report that products of the SOS-regulated imuABC gene cassette of this organism, ImuB and the error-prone ImuC DNA polymerase, are also involved in generating Cip mutations in this organism, since the yields of Cip mutations were substantially decreased in imuB- or imuC-defective cells compared to wild-type. Intriguingly, we found that the mutability of a dinB-defective strain could not be rescued by overexpression of the imuBC genes. And similarly, overexpression of the dinB gene either only modestly or else failed to restore Cip mutations in imuB- or imuC-defective cells, respectively. Combined, these results indicated that the products of the dinB and imuBC genes were acting in the same pathway leading to the generation of Cip mutations in P. aeruginosa. In addition, we provide evidence indicating that the general stress response sigma factor σ, RpoS, is required for mutagenesis in this organism and is in part at least modulating the dinB (DNA polymerase IV)-dependent mutational process. Altogether, these data provide further insight into the complexity and multifaceted control of the mutational mechanism(s) contributing to the generation of ciprofloxacin-resistance mutations in P. aeruginosa.