CENTRAL REGULATORY ROLE FOR THE RPOS SIGMA-FACTOR IN EXPRESSION OF SALMONELLA-DUBLIN PLASMID VIRULENCE GENES

The plasmid virulence genes spvABCD of Salmonella spp. are regulated by SpvR and the stationary-phase sigma factor RpoS. The transcription of spy genes is induced during the post exponential phase of bacterial growth in vitro. We sought to investigate the relationship between growth phase and RpoS in spy regulation. rpoS insertion mutations were constructed in S. dublin Lane and plasmid-cured LD842 strains, and the mutants were found to be attenuated for virulence and deficient in spy gene expression. We utilized the plasmid pBAD::rpoS to express rpoS independent of the growth phase under the control of the arabinose-inducible araBAD promoter. SpvA expression was induced within 2 h after the addition of 0.1% arabinose, even when bacteria were actively growing. This suggested that the level of RpoS, instead of the growth phase itself, controls induction of the spy genes. However, RpoS did not activate transcription of spvA in the absence of SpvR protein. Using a constitutive tet promoter to express spvR, we found that the spvA gene can be partially expressed in the rpoS mutant, suggesting that RpoS is required for SpvR synthesis. We confirmed that spvR is poorly expressed in the absence of RpoS. With an intact rpoS gene, spvR expression is not dependent on an intact spvR gene but is enhanced by spvR supplied in trans. We propose a model for Salmonella spy gene regulation in which both RpoS and SpvR are required for maximal expression at the spvR and spvA promoters.