Differential expression from two iron-responsive promoters in Salmonella enterica serovar Typhimurium reveals the presence of iron in macrophage-phagosomes

The metal status of macrophage-phagosomes during Salmonella infection is largely unknown. In this study, we have precisely calibrated the metal-specificities of two metal-responsive promoters, P_iroBCDE and P_sodB, from Salmonella enterica serovar Typhimurium and used these to directly monitor iron-levels in Salmonella-containing macrophage-phagosomes. Expression from the P_iroBCDE promoter is highly elevated in metal-depleted media but low in media supplemented with iron or cobalt, and to a lesser extent manganese. In contrast, P_sodB shows low levels of expression in metal-depleted media but is induced in media supplemented with iron but no other metals at maximum permissive concentrations. In both cases, iron-responsive expression corresponds to changes in the number of iron atoms per bacterial cell and is unaffected by pH or the presence of reactive oxygen species (hydrogen peroxide and superoxide). Importantly, expression from P_iroBCDE remained low while expression from P_sodB was elevated during infection of both Nramp1^+/+ and Nramp1^−/− macrophages. Expression from a control promoter, P_polA, unaffected by metal ions, remained unchanged. These findings are therefore consistent with the presence of iron within Salmonella-containing macrophage-phagosomes and support a model in which the toxic potential of iron may be exploited as a component of the respiratory burst killing mechanism.