The role of metals in hypothiocyanite resistance in Escherichia coli

The innate immune system employs a variety of antimicrobial oxidants to control and kill host-associated bacteria. Hypothiocyanite/hypothiocyanous acid (-OSCN/HOSCN) is one such antimicrobial oxidant that is synthesized by lactoperoxidase, myeloperoxidase, and eosinophil peroxidase at sites throughout the human body. HOSCN has potent antibacterial activity while being largely non-toxic towards human cells. The molecular mechanisms by which bacteria sense and defend themselves against HOSCN have only recently begun to be elaborated, notably by the discovery of bacterial HOSCN reductase (RclA), an HOSCN-degrading enzyme widely conserved among bacteria that live on epithelial surfaces. In this paper, I show that Ni2+ sensitizes Escherichia coli to HOSCN by inhibiting glutathione reductase, and that inorganic polyphosphate protects E. coli against this effect, probably by chelating Ni2+ ions. I also found that RclA is very sensitive to inhibition by Cu2+ and Zn2+, metals that are accumulated to high levels by innate immune cells, and that, surprisingly, thioredoxin and thioredoxin reductase are not involved in HOSCN stress resistance in E. coli. These results advance our understanding of the contribution of different oxidative stress response and redox buffering pathways to HOSCN resistance in E. coli and illustrate important interactions between metal ions and the enzymes bacteria use to defend themselves against oxidative stress. IMPORTANCEHypothiocyanite (HOSCN) is an antimicrobial oxidant produced by the innate immune system. The molecular mechanisms by which host-associated bacteria defend themselves against HOSCN have only recently begun to be understood. The results in this paper are significant because they show that the redox buffer glutathione and enzyme glutathione reductase are critical components of the Escherichia coli HOSCN response, working by a mechanism distinct from that of the HOSCN-specific defenses provided by the RclA, RclB, and RclC proteins, and that metal ions (including nickel, copper, and zinc) may impact the ability of bacteria to resist HOSCN by inhibiting specific defensive enzymes (e.g. glutathione reductase or RclA).