Probing Post-Transcriptional Control by Small Regulatory RNAs in Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen, whose large regulatory network is able to coordinate the expression of genes in response to the changing environments. Small regulatory RNAs (sRNAs) represent an important component of this regulatory network. To characterize the post-transcriptional regulation by sRNAs, I used a modified in vivo RNA proximity ligation method and probed global sRNA interactions with their targets in Pseudomonas aeruginosa. I validated the method by showing that known targets of an iron-regulated sRNA PrrF1 were recovered by the global in vivo RNA proximity ligation experiment under iron depletion condition. I identified two sRNAs (Sr0161 and ErsA) that negatively regulate OprD, the major porin for the uptake of basic amino acids, basic peptides and carbapenem antibiotics. Both sRNAs inhibit translation initiation by base-pairing with the 5’ UTR of oprD, leading to increased resistance to meropenem. Further proximity ligation experiments and enrichment for Sr0161 identified additional Sr0161 targets, including the regulator of type III secretion system. Another sRNA, Sr006, positively regulates the expression of PagL, a lipid A deacylase that is implicated in modulating the immunogenic property and polymyxin resistance of bacteria. The up-regulation of PagL by Sr006 is independent of the known RNA chaperone Hfq. Collectively, our work shows that an analysis of global sRNA-mRNA interactions can lead to discoveries of novel regulatory mechanisms involved in bacterial adaptation to different environments.