Antibiotic Inducibility of the mexXY Multidrug Efflux Operon of Pseudomonas aeruginosa Involvement of the MexZ Anti-Repressor ArmZ.docVIP

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Antibiotic Inducibility of the mexXY Multidrug Efflux Operon of Pseudomonas aeruginosa Involvement of the MexZ Anti-Repressor ArmZ.doc

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Antibiotic Inducibility of the mexXY Multidrug Efflux Operon of Pseudomonas aeruginosa Involvement of the MexZ Anti-Repressor ArmZ

AntibioticInducibilityofthemexXYMultidrugEfflux OperonofPseudomonasaeruginosa:Involvementofthe MexZAnti-RepressorArmZ ThomasHay,SebastienFraud,CalvinHo-FungLau,ChristieGilmour,KeithPoole* DepartmentofBiomedicalandMolecularSciences,BotterellHall,Queen’sUniversity,Kingston,Ontario,Canada Abstract Expression of the mexXY multidrug efflux operon in wild type Pseudomonas aeruginosa is substantially enhanced by the ribosome-targeting antimicrobial spectinomycin (18-fold) andthis is whollydependentupon theproduct of thePA5471 gene.InamutantstrainlackingthemexZgeneencodingarepressorofmexXYgeneexpression,expressionoftheefflux operon increases modestly (5-fold) and is still responsive (18-fold) to spectinomycin. Spectinomycin induction of mexXY expressioninthemexZmutantis,however,independentofPA5471suggestingthatPA5471functionsasananti-repressor (dubbedArmZforanti-repressorMexZ)thatservesonlytomodulateMexZ’srepressoractivity,withadditionalgene(s)/gene product(s) providing for the bulk of the antimicrobial-inducible mexXY expression. Consistent with PA5471/ArmZ functioning as a MexZ anti-repressor, an interaction between MexZ and ArmZ was confirmed using a bacterial 2-hybrid assay.Mutationscompromisingthisinteraction(P68S,G76S,R216C,R221W,R221Q,G231DandG252S)wereidentifiedand localized to one region of an ArmZ structural model that may represent a MexZ-interacting domain. Introduction of representative mutations into the chromosome of P. aeruginosa reduced (P68S, G76S) or obviated (R216C, R2211W) antimicrobial induction of mexXY gene expression, rendering the mutants pan-aminoglycoside-susceptible. These data confirm the importance of an ArmZ-MexZ interaction for antimicrobial-inducible mexXY expression and intrinsic aminoglycosideresistanceinP.aeruginosa. Citation: Hay T, Fraud S, Lau CH-F, Gilmour C, Poole K (2013) Antibiotic Inducibility of the mexXY Multidrug Efflux Operon of Pseudomonas aeruginosa: InvolvementoftheMexZAnti-RepressorArmZ.PLoSONE8(2):e56858.doi:10.1371