Hypochlorous acid and hydrogen peroxide-induced negative regulation of Salmonella enterica serovar Typhimurium ompW by the response regulator ArcA

H2O2 and HOCl influx was determined both in vitro and in vivo. A S. Typhimurium ompW mutant strain (？ompW) exposed to sub-lethal levels of H2O2 and HOCl showed a decreased influx of both compounds as compared to a wild type strain. Further evidence of H2O2 and HOCl diffusion through OmpW was obtained by using reconstituted proteoliposomes. We hypothesized that ompW expression should be negatively regulated upon exposure to H2O2 and HOCl to better exclude these compounds from the cell. As expected, qRT-PCR showed a negative regulation in a wild type strain treated with sub-lethal concentrations of these compounds. A bioinformatic analysis in search for potential negative regulators predicted the presence of three ArcA binding sites at the ompW promoter region. By electrophoretic mobility shift assay (EMSA) and using transcriptional fusions we demonstrated an interaction between ArcA and one site at the ompW promoter region. Moreover, qRT-PCR showed that the negative regulation observed in the wild type strain was lost in an arcA and in arcB mutant strains.OmpW allows the influx of H2O2 and HOCl and is negatively regulated by ArcA by direct interaction with the ompW promoter region upon exposure to both toxic compounds.Hydrogen peroxide (H2O2) and hypochlorous acid (HOCl) are reactive oxygen species that are part of the oxidative burst encountered by S. Typhimurium upon internalization by phagocytic cells. Under acidic conditions, such as those found inside the phagosome, H2O2 is generated spontaneously by the reaction of two superoxide anion (O2？) molecules [1]. Moreover, S. Typhimurium encodes both periplasmic and cytoplasmic superoxide dismutases that catalyze O2？ dismutation to generate H2O2 and molecular oxygen [2-4]. HOCl is produced by the action of myeloperoxidase (MPO) in a reaction that depends on H2O2, Cl？and acidic conditions [5,6]. Taken together, H2O2 and HOCl react with thiol and heme groups, copper and iron salts generating the reactive hydroxyl radica