Pseudomonas aureofaciens strain 30-84 is a biological control bacterium that utilizes a two-component GacS/GacA regulatory system interconnected with the PhzR/PhzI quorum sensing system to positively regulate biosynthesis of phenazine antibiotics that contribute to its association with plant hosts. To date, no negative regulators of phenazine production have been identified, nor has the role of repression been studied. Here we describe a novel repressor of secondary metabolism in P. aureofaciens strain 30-84, RpeA, whose deduced amino acid sequence is similar to those of a group of putative two-component regulatory systems of unknown function found in several animal and plant-pathogenic bacteria. In minimal medium where phenazine production is very low, inactivation of the rpeA gene enhanced phenazine biosynthetic gene expression and increased phenazine production but did not increase quorum sensing signal accumulation. Furthermore, RpeA functioned to block phenazine biosynthetic gene transcription in minimal medium even when quorum-sensing signals were at a level that was sufficient for induction of phenazine gene expression in rich medium. Additionally, in the absence of rpeA, the quorum sensor PhzR was not required for phenazine production. Although repression plays a critical role in phenazine regulation, the rpeA mutation could not bypass the requirement for a functional GacS/GacA system, demonstrating that activation is required even in the absence of the RpeA repressor. This study reinforces that multiple signals, including nutrition and population density, are integrated to control the appropriate expression of phenazine antibiotics.
