CONICET | Buscador de Institutos y Recursos Humanos

Copper resistance in Gram-negative bacteria is principally controlled by the cue regulon, which in Escherichia coli is composed by the Cu(I) sensor CueR, a MerR-like transcriptional regulator, that modulates the expression of copA and cueO, coding for an integral inner-membrane Cu-transporting P-type ATPase, and a periplasmic oxygen-dependent multicopper-Cu(I) oxidase, respectively. E. coli also relies on the cus system to increase copper resistance in anaerobiosis. Salmonella harbors the components of the cue regulon, but lacks the cus system, and still it can resist higher copper concentrations than E. coli under anaerobic conditions. We have recently uncovered a novel, Salmonella-specific, CueR-regulated gene, cueP, whose product is responsible for this increased copper resistance. Atomic absorption spectrometry, as well as UV-visible spectroscopy demonstrated that CueP is able to bind two Cu equivalents per monomer. CueP homolog proteins were identified in bacterial species that lack the cus system. The alignment of these homologues highlighted the presence of conserved amino acid residues predicted to participate in metal recognition. We generated site-directed mutants in these conserved residues and analyzed their contribution to copper resistance. CueP mutant proteins in conserved histidine and cysteine residues were affected in their ability to interact with copper. Furthermore, these mutants were unable to restore copper resistance both in aerobic and anaerobic conditions, and, unlike the wild-type CueP, could not rescue the smooth colony morphology of the otherwise mucoid phenotype of the cuiD mutant strain. Overall, these results establish the physiological function of Salmonella CueP in copper resistance and highlight the role of the conserved amino acid residues in this function.