EXPRESSION OF THE MULTIDRUG-RESISTANCE OPERON MEXA-MEXB-OPRM IN PSEUDOMONAS-AERUGINOSA - MEXR ENCODES A REGULATOR OF OPERON EXPRESSION

The region upstream of the multiple antibiotic resistance efflux opero n mexA-mexB-oprM in Pseudomonas aeruginosa was sequenced, and a gene, mexR, was identified. The predicted MexR product contains 147 amino ac ids with a molecular mass of 16,964 Da, which is consistent with the o bserved size of the overexpressed mexR gene product. MexR was homologo us to MarR, the repressor of MarA-dependent multidrug resistance in Es cherichia coli, and other repressors of the MarR family. A mexR knocko ut mutant showed a twofold increase in expression of both plasmid-born e and chromosomal mexA-reporter gene fusions compared with the MexR(+) parent strain, indicating that the mexR gene product negatively regul ates expression of the mexA-mexB-oprM operon. Furthermore, the cloned mexR gene product reduced expression of a plasmid-borne mexA-lacZ fusi on in E. coli, indicating that MexR represses mexA-mexB-oprM expressio n directly, Consistent with the increased expression of the efflux ope ron in the mexR mutant, the mutant showed an increase (relative to its MexR(+) parent) in resistance to several antimicrobial agents. Expres sion of a mexR-lacZ fusion increased threefold in a mexR knockout muta nt, indicating that mexR is negatively autoregulated, OCR1, a nalB mul tidrug-resistant mutant which overproduces OprM, exhibited a greater t han sevenfold increase in expression of a chromosomal mexA-phoA fusion compared with its parent. Introduction of a mexR knockout mutation in strain OCR1 eliminated this increase in efflux gene expression and, a s expected, increased the susceptibility of the strain to a variety of antibiotics, The nucleotide sequences of the mexR genes of OCR1 and i ts parental strain revealed a single base substitution in the former w hich would cause a predicted substitution of Trp for Arg at position 6 9 of its mexR product, These data suggest that MexR possesses both rep ressor and activator function in vivo, the activator form being favore d in nalB multidrug-resistant strains.