The heme-responsive PrrH sRNA regulatesPseudomonas aeruginosapyochelin gene expression

Abstract

Pseudomonas aeruginosais an opportunistic pathogen that requires iron for growth and virulence, yet this nutrient is sequestered by the innate immune system during infection. When iron is limiting,P. aeruginosaexpresses the PrrF1 and PrrF2 small regulatory RNAs (sRNAs), which post-transcriptionally repress expression of non-essential iron-containing proteins thus sparing this nutrient for more critical processes. The genes for the PrrF1 and PrrF2 sRNAs are arranged in tandem on the chromosome, allowing for the transcription of a longer heme-responsive sRNA, termed PrrH. While the functions of PrrF1 and PrrF2 have been studied extensively, the role of PrrH inP. aeruginosaphysiology and virulence is not well understood. In this study, we performed transcriptomic and proteomic studies to identify the PrrH regulon. In shaking cultures, the pyochelin synthesis proteins were increased in two distinctprrHmutants compared to wild type, while the mRNAs for these proteins were not affected byprrHmutation. We identified complementarity between the PrrH sRNA and sequence upstream of thepchEmRNA, suggesting potential for PrrH to directly regulate expression of genes for pyochelin synthesis. We further showed thatpchEmRNA levels were increased in theprrHmutants when grown in static but not shaking conditions. Moreover, we discovered controlling for the presence of light was critical for examining the impact of PrrH onpchEexpression. As such, our study reports on the first likely target of the PrrH sRNA and highlights key environmental variables that will allow for future characterization of PrrH function.

Pseudomonas aeruginosa heme metabolites biliverdin IXβ and IXδ are integral to lifestyle adaptations associated with chronic infection

“…For static cultures, cells were removed from the plate with a sterile loop, and the total pellet weight was recorded in pre-weighed Eppendorf tubes. Sample preparation and quantitative label-free proteomics analyses were performed as previously described ( 76 – 79 ). In brief, cells were harvested by centrifugation at 2,000 rpm for 30 s at 4°C and then subsequently lysed in 4% sodium deoxycholate, reduced, alkylated, and trypsinolyzed on filter as previously described ( 80 ).…”