Genome-scale mapping reveals complex regulatory activities of RpoN inYersinia pseudotuberculosis

Abstract

RpoN, an alternative sigma factor commonly known as sigma 54, is implicated in persistent stages of Yersinia pseudotuberculosis infections in which genes associated with this regulator are upregulated. We here combined phenotypic and genomic assays to provide insight into its role and function in this pathogen. RpoN was found essential for Y. pseudotuberculosis virulence in mice, and in vitro functional assays showed that it controls biofilm formation and motility. Mapping genome-wide associations of Y. pseudotuberculosis RpoN using chromatin immunoprecipitation coupled with next-generation sequencing identified an RpoN-binding motif located at 103 inter- and intragenic sites on both sense and anti-sense strands. Deletion of rpoN had a large impact on gene expression, including down-regulation of genes encoding proteins involved in flagellar assembly, chemotaxis, and quorum sensing. There were also clear indications of cross talk with other sigma factors, together with indirect effects due to altered expression of other regulators. Matching differential gene expression with locations of the binding sites implicated around 130 genes or operons potentially activated or repressed by RpoN. Mutagenesis of selected intergenic binding sites confirmed both positive and negative regulatory effects of RpoN binding. Corresponding mutations of intragenic sense sites had less impact on associated gene expression. Surprisingly, mutating intragenic sites on the anti-sense strand commonly reduced expression of genes encoded by the corresponding sense strand

Diversified repertoire of phage resistance inKlebsiella pneumoniaeand bidirectional steering effects impacting antibiotic susceptibility

“…Our results indicate that mutation of mnmE can substantially delay phage replication; however, the exact mechanism warrants further investigation. For RpoN, an RNA polymerase factor sigma-54 which regulates the transcription of genes across diverse cellular functions 25, 26, 27, 28 , a previous study reported its association with phage resistance in Pseudomonas aeruginosa 29 . With the diverse regulatory roles played by RpoN, the specific mechanism for its association with phage infection is unknown.…”