Influence of the pqs genes on Pseudomonas quorum sensing
The opportunistic pathogen Pseudomonas aeruginosa has complex quorum sensing (QS) circuitry, which involves two acylhomoserine lactone (AHL) systems, the LasI AHL synthase and LasR AHL-dependent transcriptional activator system and the RhlI AHL synthase-RhlR AHL-responsive transcriptional activator. There is also a quinoline signaling system (the Pseudomonas quinolone signal, PQS, system). Although there is a core set of genes regulated by the AHL circuits, there is substantial strain-to-strain variation in the non-core QS regulated genes. Reductive evolution of the QS regulon, and variation in specific genes activated by QS occurs in laboratory evolution experiments with the model strain PAO1. We used a transcriptomics approach to test the hypothesis that reductive evolution in the PAO1 QS regulon can in large part be explained by a simple null mutation in pqsR, the gene encoding the transcriptional activator of the pqs operon. We found that PqsR had very little influence on the AHL QS regulon. This was a surprising finding because the last gene in the PqsR-dependent pqs operon, pqsE, codes for a protein, which physically interacts with RhlR and this interaction is required for full RhlR-dependent activation of some genes. We used comparative transcriptomics to examine the influence of a pqsE mutation on the QS regulon and identified only three transcripts, which were strictly dependent on PqsE. By using reporter constructs we showed that the PqsE influence on other genes was dependent on experimental conditions and we have gained some insight about those conditions. This work adds to our understanding of the plasticity of the P. aeruginosa QS regulon and to the role PqsE plays in RhlR-dependent gene activation. Overall design: We used comparative transcriptomics to contrast the quorum sensing (QS) regulons of the lab strain PAO1, PAO1 ?pqsR and PAO1 ?pqsE grown in buffered LB to better understand the effect of PqsE on acylhomoserine lactone (AHL) QS-regulated genes. The AHL-degrading enzyme AiiA lactonase (100 ug/ml) was added (or not)  to our growth medium to prevent activation of the LasR/I and RhlR/I circuits by their respective signals, 3OC12-HSL and C4-HSL. Lactonase does not degrade the third QS signal, pseudomonas quinolone signal (PQS), the activator of the pqs circuit. Using RNAseq we compared the transcripts from cells collected at OD 2.0 with and without AHL QS-activation to determine the core genes under QS control in each strain. We then compared the QS regulons of the wildtype, PqsR and PqsE mutants.