The core and accessory Hfq interactomes across Pseudomonas aeruginosa lineages.

The major RNA-binding protein Hfq interacts with mRNAs, either alone or together with regulatory small noncoding RNAs (sRNAs), affecting mRNA translation and degradation in bacteria. However, studies tend to focus on single reference strains and assume that the findings may apply to the entire species, despite the important intra-species genetic diversity known to exist. Here, we use RIP-seq to identify Hfq-interacting RNAs in three strains representing the major phylogenetic lineages of Pseudomonas aeruginosa. We find that most interactions are in fact not conserved among the different strains. We identify growth phase-specific and strain-specific Hfq targets, including previously undescribed sRNAs. Strain-specific interactions are due to different accessory gene sets, RNA abundances, or potential context- or sequence- dependent regulatory mechanisms. The accessory Hfq interactome includes most mRNAs encoding Type III Secretion System (T3SS) components and secreted toxins in two strains, as well as a cluster of CRISPR guide RNAs in one strain. Conserved Hfq targets include the global virulence regulator Vfr and metabolic pathways involved in the transition from fast to slow growth. Furthermore, we use rGRIL-seq to show that RhlS, a quorum sensing sRNA, activates Vfr translation, thus revealing a link between quorum sensing and virulence regulation. Overall, our work highlights the important intra-species diversity in post-transcriptional regulatory networks in Pseudomonas aeruginosa.