Copper distribution networks in Pseudomonas aeruginosa

Bacterial Cu + homeostasis is based on protein networks that mobilize copper to cellular targets, enabling both metallation of cuproproteins and maintenance of non‐toxic metal levels. Here, we use genome wide expression analysis (RNA‐Seq) to characterize the response of Pseudomonas aeruginosa to external 0.5 mM CuSO 4 , a condition where cell viability is uncompromised. Analysis of copR and cueR mutant strains revealed that compartmental Cu levels are independently controlled; the cytoplasmic Cu sensor CueR regulates cytoplasmic chaperones and plasma membrane transporters, whereas the two component system CopR/S shows a putative DNA recognition sequence, forming a regulon composed of genes involved in periplasmic Cu‐homeostasis. Response to Cu‐imbalance resulted in distinct transcriptomic landscapes: a pre‐steady‐state (5 min), where general metabolism is slowed down, and a steady‐state (2 h), where general metabolism is restored. Specific Cu homeostasis genes were strongly regulated in both conditions. Our system‐wide analysis revealed induction of three Cu‐efflux systems (a P 1B ‐ATPase, a porin, and a resistance‐nodulation‐division (RND) system), a putative Cu‐binding periplasmic chaperone and the unusual presence of two cytoplasmic CopZ proteins. Importantly, novel transmembrane transporters probably mediating Cu‐influx were among those largely repressed genes upon Cu‐stress. Three proteins were identified among these: OprC, an outer membrane porin; PA3789, an inner membrane protein; and PA5030, a Met/His rich protein, member of the major facilitator superfamily (MFS) of transporters. Alternatively, genes encoding previously proposed Cu‐importers ccoA and hmtA did not change their expression levels. The occurrence of cell influx transporters is a prerequisite for homeostasis of a given cellular constituent. However, deletion of the putative Cu‐importers OprC and PA1530 does not affect the Cu content of whole cells, suggesting that Pseudomonas has alternative ways to acquire Cu. As Pseudomonas secrete two major siderophores (pyochelin and pyoverdin) to scavenge metals, we hypothesize that those siderophores will constitute an alternative way to import copper. Interestingly, genes involved in both, pyochelin and pyoverdin pathways, are overexpressed when the outer membrane porin is absent, but only pyochelin‐related genes are up‐regulated when the putative inner‐membrane transporters are deleted. This suggests that pyochelin can be the alternative Cu‐import system that Pseudomonas uses under copper‐starving conditions. Summarizing, our studies show a system‐wide network of sensors/regulators, soluble chaperones, and influx/efflux transporters that control the Cu levels in P. aeruginosa compartments. Support or Funding Information This work was supported by NIH Grant R01 GM114949 to J.M.A. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .