PmtA Regulates Pyocyanin Expression and Biofilm Formation in Pseudomonas aeruginosa

Abstract

The opportunistic pathogen <i>Pseudomonas aeruginosa</i> expresses a small molecular weight, cysteine-rich protein (PmtA), identified as a metallothionein (MT) protein family member. The MT family proteins have been well-characterized in eukaryotes as essential for zinc and copper homeostasis, protection against oxidative stress, and the ability to modify a variety of immune activities. Bacterial MTs share sequence homology, antioxidant chemistry, and heavy metal-binding capacity with eukaryotic MTs, however, the impact of bacterial MTs on virulence and infection have not been well-studied. In the present study, we investigated the role of PmtA in <i>P. aeruginosa</i> PAO1 using a PmtA-deficient strain (Δ<i>pmtA</i>). Here we demonstrated the virulence factor, pyocyanin, relies on the expression of PmtA. We showed that PmtA may be protective against oxidative stress, as an alternative antioxidant, glutathione, can rescue pyocyanin expression. Furthermore, the expression of <i>phzM</i>, which encodes a pyocyanin precursor enzyme, was decreased in the Δ<i>pmtA</i> mutant during early stationary phase. Upregulated <i>pmtA</i> expression was previously detected in confluent biofilms, which are essential for chronic infection, and we observed that the Δ<i>pmtA</i> mutant was disrupted for biofilm formation. As biofilms also modulate antibiotic susceptibility, we examined the Δ<i>pmtA</i> mutant susceptibility to antibiotics and found that the Δ<i>pmtA</i> mutant is more susceptible to cefepime and ciprofloxacin than the wild-type strain. Finally, we observed that the deletion of <i>pmtA</i> results in decreased virulence in a waxworm model. Taken together, our results support the conclusion that PmtA is necessary for the full virulence of <i>P. aeruginosa</i> and may represent a potential target for therapeutic intervention.