Dataset Information

Identification of the PA1113 Gene Product as an ABC Transporter Involved in the Uptake of Carbenicillin in Pseudomonas aeruginosa PAO1.

ABSTRACT: The resistance of Pseudomonas aeruginosa to antibiotics is multi factorial and complex. Whereas efflux pumps such as MexAB-OprM have been thought to predominate, here we show that a novel ATP Binding Cassette (ABC) transporter that mediates influx of carbenicillin from the periplasm to the cytoplasm and away from its cell wall target plays an important role in the resistance of P. aeruginosa to this antibiotic. Treatment of P. aeruginosa with verapamil, an inhibitor of ABC transporters in eukaryotic cells, increases its sensitivity to carbenicillin. Using amino acid sequence homology with known verapamil protein targets as a probe, we determined that the PA1113 gene product, an ABC transporter, mediates carbenicillin uptake into the bacterial cytoplasm. Docking and pharmacological analyses showed that verapamil and carbenicillin compete for the same site on the PA1113 gene protein, explaining the inhibitory effect of verapamil on carbenicillin uptake, and furthermore suggest that the PA1113 ABC transporter accounts for about 30% of P. aeruginosa carbenicillin resistance. Our findings demonstrate that the PA1113 gene product helps mediate carbenicillin resistance by transporting it away from its cell wall target and represents a promising new therapeutic target.

SUBMITTER: Hulen C

PROVIDER: S-EPMC7559151 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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Identification of the PA1113 Gene Product as an ABC Transporter Involved in the Uptake of Carbenicillin in Pseudomonas aeruginosa PAO1.

Hulen Christian C Racine Pierre-Jean PJ Chevalier Sylvie S Feuilloley Marc M Lomri Nour-Eddine NE

Antibiotics (Basel, Switzerland) 20200911 9

The resistance of Pseudomonas aeruginosa to antibiotics is multi factorial and complex. Whereas efflux pumps such as MexAB-OprM have been thought to predominate, here we show that a novel ATP Binding Cassette (ABC) transporter that mediates influx of carbenicillin from the periplasm to the cytoplasm and away from its cell wall target plays an important role in the resistance of P. aeruginosa to this antibiotic. Treatment of P. aeruginosa with verapamil, an inhibitor of ABC t ...[more]

PMID: 32933058

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Project description:DNA methylation is widespread among prokaryotes, and most DNA methylation reactions are catalyzed by adenine DNA methyltransferases, which are part of restriction-modification (R-M) systems. R-M systems are known for their role in the defense against foreign DNA; however, DNA methyltransferases also play functional roles in gene regulation. In this study, we used single-molecule real-time (SMRT) sequencing to uncover the genome-wide DNA methylation pattern in the opportunistic pathogen Pseudomonas aeruginosa PAO1. We identified a conserved sequence motif targeted by an adenine methyltransferase of a type I R-M system and quantified the presence of N6-methyladenine using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Changes in the PAO1 methylation status were dependent on growth conditions and affected P. aeruginosa pathogenicity in a Galleria mellonella infection model. Furthermore, we found that methylated motifs in promoter regions led to shifts in sense and antisense gene expression, emphasizing the role of enzymatic DNA methylation as an epigenetic control of phenotypic traits in P. aeruginosa Since the DNA methylation enzymes are not encoded in the core genome, our findings illustrate how the acquisition of accessory genes can shape the global P. aeruginosa transcriptome and thus may facilitate adaptation to new and challenging habitats.IMPORTANCE With the introduction of advanced technologies, epigenetic regulation by DNA methyltransferases in bacteria has become a subject of intense studies. Here we identified an adenosine DNA methyltransferase in the opportunistic pathogen Pseudomonas aeruginosa PAO1, which is responsible for DNA methylation of a conserved sequence motif. The methylation level of all target sequences throughout the PAO1 genome was approximated to be in the range of 65 to 85% and was dependent on growth conditions. Inactivation of the methyltransferase revealed an attenuated-virulence phenotype in the Galleria mellonella infection model. Furthermore, differential expression of more than 90 genes was detected, including the small regulatory RNA prrF1, which contributes to a global iron-sparing response via the repression of a set of gene targets. Our finding of a methylation-dependent repression of the antisense transcript of the prrF1 small regulatory RNA significantly expands our understanding of the regulatory mechanisms underlying active DNA methylation in bacteria.

Dataset Information

Identification of the PA1113 Gene Product as an ABC Transporter Involved in the Uptake of Carbenicillin in Pseudomonas aeruginosa PAO1.

Publications

Identification of the PA1113 Gene Product as an ABC Transporter Involved in the Uptake of Carbenicillin in <i>Pseudomonas aeruginosa</i> PAO1.

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