MpaR-driven expression of an orphan terminal oxidase subunit supports Pseudomonas aeruginosa biofilm respiration and development during cyanogenesis

Pseudomonas aeruginosa is a common, biofilm-forming pathogen that exhibits complex pathways of redox metabolism. It produces four different types of terminal oxidases for aerobic respiration, and for one of these, the cbb3-type terminal oxidases, it has the capacity to produce at least 16 isoforms encoded by partially redundant operons. It also produces small-molecule virulence factors that interact with the respiratory chain, including the poison cyanide. Previous studies had indicated a role for cyanide in activating expression of an orphan terminal oxidase subunit gene called ccoN4 and that the product contributes to P. aeruginosa cyanide resistance, fitness in biofilms, and virulence, but the mechanisms underlying this process had not been elucidated. Here, we show that the regulatory protein MpaR, which is predicted to be a pyridoxal phosphate-binding transcription factor and is encoded just upstream of ccoN4, controls ccoN4 expression in response to endogenous cyanide. Paradoxically, we find that cyanide production is required to support CcoN4's contribution to respiration in biofilms. We identify a palindromic motif required for cyanide- and MpaR-dependent expression of ccoN4 and co-expressed adjacent loci. We also characterize the regulatory logic of this region of the chromosome. Finally, we identify residues in the putative cofactor-binding pocket of MpaR, which are required for ccoN4 expression. Together, our findings illustrate a novel scenario in which the respiratory toxin cyanide acts as a signal to control gene expression in a bacterium that produces the compound endogenously.

Automated summary

Pseudomonas aeruginosa is a biofilm-forming pathogen with complex pathways of redox metabolism. This study reveals the role of cyanide in activating the expression of a gene in Pseudomonas aeruginosa, which is involved in cyanide resistance, fitness in biofilms, and virulence. The regulatory protein MpaR is identified as the key protein controlling the expression of this gene in response to endogenous cyanide. The findings demonstrate a novel scenario where cyanide acts as a signal to control gene expression in a bacterium that produces it endogenously.