Probing the ArcA Regulon Under Aerobic/ROS Conditions in Salmonella Enterica Serovar Typhimurium

Background: Hydrogen peroxide (H₂O₂) is a reactive oxygen species (ROS), which is part of the oxidative burst encountered upon internalization of Salmonella enterica serovar Typhimurium (S. Typhimurium) by phagocytic cells. It has previously been established that, the ArcAB two-component system plays a critical role in ROS resistance, but the genes regulated by the system remained undetermined to date. We therefore investigated the ArcA regulon in aerobically growing S. Typhimurium before and after exposure to H₂O₂ by querying gene expression and other physiological changes in wild type and ΔarcA strains.

Results: In the ΔarcA strain, expression of 292 genes showed direct or indirect regulation by ArcA in response to H₂O₂, of which 141were also regulated in aerobiosis, but in the opposite direction. Gene set enrichment analysis (GSEA) of the expression data from WT and ΔarcA strains, revealed that, in response to H₂O₂ challenge in aerobically grown cells, ArcA down regulated multiple PEP-PTS and ABC transporters, while up regulating genes involved in glutathione and glycerolipid metabolism and nucleotide transport. Further biochemical analysis guided by GSEA results showed that deletion of arcA during aerobic growth lead to increased reactive oxygen species (ROS) production which was concomitant with an increased NADH/NAD+ ratio. In absence of ArcA under aerobic conditions, H₂O₂ exposure resulted in lower levels of glutathione reductase activity, leading to a decreased GSH (reduced glutathione)/GSSG (oxidized glutathione) ratio.

Conclusion: The ArcA regulon was defined in 2 conditions, aerobic growth and the combination of peroxide treatment and aerobic growth in S. Typhimurium. ArcA coordinates a response that involves multiple aspects of the carbon flux through central metabolism, which ultimately modulates the reducing potential of the cell.