Hfq‐dependent alterations of the transcriptome profile and effects on quorum sensing in Pseudomonas aeruginosa

The Pseudomonas aeruginosa quorum‐sensing (QS) systems, Las and Rhl, control the production of several virulence factors and other proteins, which are important to sustain adverse conditions. A comparative transcriptome analysis of a rpoS ⁻ and a rpoS⁻hfq ⁻ strain indicated that the Sm‐like RNA‐binding protein Hfq affects approximately 5% of the P. aeruginosa O1 transcripts. Among these transcripts 72 were identified to be QS regulated. Expression studies revealed that Hfq does not control the master regulators of the Las system, LasR and LasI. Upon entry into stationary phase, Hfq exerted a moderate stimulatory effect on translation of the rhlR gene and on the qscR gene, encoding a LasR/RhlR homologue. However, Hfq considerably stimulated translation of the rhlI gene, encoding the synthetase of the autoinducer N‐Butyryl‐homoserine lactone (C4‐HSL). Correspondingly, the C4‐HSL levels were reduced in a hfq⁻ strain. To elucidate the stimulatory effect of Hfq on rhlI expression we asked whether Hfq affects the stability of the regulatory RNAs RsmY and RsmZ, which have been implicated in sequestration of the translational repressor RsmA, which in turn is known to negatively regulate RhlI synthesis. We demonstrate that Hfq binds to and stabilizes the regulatory RNA RsmY, which is further shown to bind to the regulatory protein RsmA. A model for the Hfq regulatory network is presented, wherein an alleviation of the negative effect of RsmA accounts for the observed stimulation of rhlI expression by Hfq. The model is corroborated by the observation that a rsmY⁻ mutant mimics the hfq ⁻ phenotype with regard to rhlI expression.