Small RNA-mediated wiring of quorum sensing systems in Pseudomonas aeruginosa

Many of the numerous virulence factors produced by Pseudomonas aeruginosa are under the control of quorum sensing (QS), which uses diffusible autoinducer molecules as a way to monitor cell density. Specific genes are thus activated when bacterial cell population density, and hence autoinducer concentration, exceeds a threshold. P. aeruginosa has at least three QS systems, with distinct autoinducer molecules, that are hierarchically arranged. The las system is the first to become activated, and in turn it stimulates the other systems known as rhl and pqs, which additionally regulate each other. In this study, the novel P. aeruginosa small RNA ReaL has been characterized. Our results indicate that ReaL is embedded in the P. aeruginosa QS playing the role of wiring las to pqs systems. In fact, we show that ReaL responds to the las autoinducer receptor LasR and impacts positively the synthesis of the pqs quinolone autoinducer PQS. Our results suggest that the stimulation of PQS synthesis is mediated by a positive post-transcriptional effect of ReaL on the pqsC gene belonging to the pqsABCD cluster involved in the biosynthesis of the PQS-precursor HHQ. We suggest that a fine balancing between the already known regulatory effects of LasR on PQS synthesis and those mediated by ReaL can influence the timing and magnitude of expression of QS-regulated virulence factors. This view is consistent with the evidence that perturbations of ReaL levels affect pyocyanin synthesis, biofilm formation and swarming motility, processes that are known to be influenced by PQS synthesis. Besides being regulated by LasR, ReaL responds to infection relevant cues that P. aeruginosa can experience in mammalian hosts such as temperature and oxygen availability. Furthermore, ReaL shows a growth phase-dependent pattern of expression, being up-regulated in stationary phase. Together, these ReaL regulations are expected to contribute to the fine co-modulation of PQS synthesis.