Characterization of a PecS Family Transcriptional Regulator in Vibrio vulnificus

Abstract

Vibrio vulnificus, marine bacteria, are serious human pathogens. Virulent strains can cause food poisoning upon consumption of uncooked seafood and they can cause wound infections in humans. A genomic locus found in V. vulnificus consists of a gene, which encodes a homologue of the multiple antibiotic resistance regulator (MarR), and a divergently oriented gene encoding a transmembrane protein, predicted to function as an efflux pump. The encoded proteins are predicted to be homologues of PecS and PecM from plant pathogens where PecS controls virulence gene expression and PecM exports antioxidants. The predicted V. vulnificus PecS was cloned from V. vulnificus YJ016 genomic DNA, overexpressed in E. coli and purified via affinity chromatography. Electrophoretic Mobility Shift Assays revealed specific binding of PecS to multiple sites in the intergenic region between pecS and pecM genes with a Kd of 0.6 ±0.1 nM. Further analysis showed attenuation of PecS binding to intergenic DNA in the presence of urate and xanthine, but not with hypoxanthine, allantonin, guanosine, or GMP. The attenuation of complex formation in presence of ligands is expected to correlate with de-repression of pecS and pecM expression. This may enable the bacteria to efflux antioxidants, increasing bacterial resistance to reactive oxygen species produced by the host and promote virulence. Since urate and xanthine are expected to be produced as part of the oxidative burst during host defenses, we propose that these products of the host purine degradation pathway function to elevate bacterial virulence. Supported by NSF (DBI-1051610).