Elucidation of Sigma Factor-Associated Networks in Pseudomonas aeruginosa Reveals a Modular Architecture with Limited and Function-Specific Crosstalk

Abstract

Sigma factors are essential global regulators of transcription initiation in bacteria which confer promoter recognition specificity to the RNA polymerase core enzyme. They provide effective mechanisms for simultaneously regulating expression of large numbers of genes in response to challenging conditions, and their presence has been linked to bacterial virulence and pathogenicity. In this study, we constructed nine his-tagged sigma factor expressing and/or deletion mutant strains in the opportunistic pathogen Pseudomonas aeruginosa. To uncover the direct and indirect sigma factor regulons, we performed mRNA profiling, as well as chromatin immunoprecipitation coupled to high-throughput sequencing. We furthermore elucidated the de novo binding motif of each sigma factor, and validated the RNA- and ChIP-seq results by global motif searches in the proximity of transcriptional start sites (TSS). Our integrated approach revealed a highly modular network architecture which is composed of insulated functional sigma factor modules. Analysis of the interconnectivity of the various sigma factor networks uncovered a limited, but highly function-specific, crosstalk which orchestrates complex cellular processes. Our data indicate that the modular structure of sigma factor networks enables P. aeruginosa to function adequately in its environment and at the same time is exploited to build up higher-level functions by specific interconnections that are dominated by a participation of RpoN.

Toxic Electrophiles Induce Expression of the Multidrug Efflux Pump MexEF-OprN in Pseudomonas aeruginosa through a Novel Transcriptional Regulator, CmrA

“…Our results confirmed that the transcript of PA2048 begins 315 nucleotides upstream from ATG PA2048 as determined in the PA14 sigmulome (19). This long untranslated region (UTR) contains two inverted repeat sequences of 10 nucleotides with a spacer of 91 nucleotides (Fig.…”

“…2B) successfully generated an nfxC2 resistance phenotype, while complementation with a shorter fragment lacking the RpoN/RpoS SFBMs (Ϫ68 bp upstream from the TSS) had no effect (data not shown). These data provide evidence that the RpoN/RpoS region is necessary to activate the expression of cmrA (19). Whether cmrA is under the control of RpoN and/or RpoS remains to be determined.…”

“…Analysis of the putative sigma factor binding motifs (SFBMs) of PA14 (sigmulome) (19) proved to be useful to predict the location of the cmrA promoter (P cmrA ). Hence, comparison of the DNA region upstream from cmrA TSS with the PA14 sigmulome Homologs in strain PAO1 are indicated in brackets.…”

Pseudomonas aeruginosa AlgU Contributes to Posttranscriptional Activity by Increasing rsmA Expression in a mucA22 Strain

“…There are conflicting data regarding RpoS control of rsmA expression (9,27). The potential RpoS-binding site upstream of rsmA implicated RpoS in the control of rsmA (Fig.…”

“…Currently, only a single rsmA promoter region, which resembles a 70 (RpoD) promoter, has been identified in P. aeruginosa (24)(25)(26). A recent chromatin immunoprecipitation sequencing study showed that both RpoD and RpoS likely control rsmA expression (27). However, another study found similar RsmA levels in both an rpoS mutant and the wildtype strain (9).…”