GSE137738
Overexpression of the small RNA PA0805.1 modulates the expression of a large set of genes and proteins, resulting in altered motility, cytotoxicity and tobramycin resistance
Purpose:  Pseudomonas aeruginosa is a motile species that initiates swarming motility in response to specific environmental cues, i.e., a semisolid surface with amino acids as a nitrogen source (relevant to the human lung). Swarming is an intricately regulated process, but to date post-transcriptional regulation has not been extensively investigated. Small non-coding RNAs (sRNA) are hypothesized to play post-transcriptional regulatory roles, largely through suppression of translation, and we previously demonstrated 20 sRNA species that were dysregulated under swarming conditions. Overexpressing these revealed one sRNA, PA0805.1,  that when cloned, transformed into PAO1 WT and overexpressed led to broad phenotypic changes. Methods: The sRNA PA0805.1 was cloned and overexpressed in PAO1 WT. Phenotypic screens including motility, cytotoxicity and adherence assays were performed. RNA-Seq and proteomics were performed under swarming conditions compared to the empty vector strain to discover which genes were influenced by the overexpression of PA0805.1 Results:  Phenotypic screens revealed that PA0805.1 resulted in reduced swarming, swimming and twitching motility, as well as increased adherence, cytotoxicity, and tobramycin resistance. A deletion mutant ?PA0805.1 was supersusceptible to tobramycin under swarming conditions. The strain overexpressing PA0805.1 was compared to the empty vector strain by RNA-Seq and proteomics under swarming conditions to determine sRNA targets. Broad transcriptional and proteomic profiles were revealed, encompassing 1121 differentially expressed genes and 258 proteins with significantly different abundance. Importantly, these included 106 transcriptional regulators, two-component regulatory systems, sigma and anti-sigma factors. Downstream of these regulators were found downregulated type IV pilus genes, many upregulated adherence and virulence factors, and two multidrug efflux systems, mexXY and mexGHI-opmD. Conclusions:  The sRNA PA0805.1 appears to regulate diverse bacterial lifestyles, most likely through a regulatory cascade. Overall design: Five biological replicates of the wildtype strain overexpressing PA0805.1 were compared to five biological replicates of the wildtype with the empty vector as a control