1192. H-NS-like Proteins in Pseudomonas aeruginosa Coordinately Silence Intragenic and Antisense Transcription

Abstract

BackgroundThe H-NS-like proteins MvaT and MvaU act coordinately as global repressors in Pseudomonas aeruginosa by binding to AT-rich regions of the chromosome, which include horizontally acquired genes and numerous virulence factors. Although cells can tolerate the loss of either protein, identifying their combined regulatory effects has been challenging because the loss of both proteins is lethal due to induction of the prophage Pf4 and subsequent superinfection of the cell. Although in other bacteria, H-NS promotes cellular fitness by inhibiting intragenic transcription from AT-rich target regions, preventing them from sequestering RNA polymerase, a role for MvaT and MvaU in repressing transcription from intragenic promoters has not been demonstrated.MethodsHere we utilize a parental strain that cannot be infected by Pf4 phage to identify the combined MvaT and MvaU regulon. RNA-seq was utilized to identify genes differentially expressed in cells lacking MvaU or both MvaU and MvaT. ChIP-seq was utilized to identify genes directly regulated by MvaT and MvaU in wild-type cells. Further, ChIP-seq was performed in cells of the parental strain and cells lacking both MvaT and MvaU to map genome-wide σ 70-dependent promoters that were active in the presence or absence of both H-NS-like proteins.ResultsWe demonstrate that the loss of both MvaT and MvaU, but not MvaU alone, leads to increased sense, antisense, and intragenic transcription from loci directly controlled by these proteins. We further show that the loss of MvaT and MvaU leads to a striking redistribution of RNA polymerase containing σ 70 to those genomic regions vacated by these proteins.Loss of MvaT and MvaU causes global changes in gene expression Loss of MvaT and MvaU results in increased sense and antisense transcription Loss of both MvaT and MvaU results in genome-wide redistribution of RNA polymerase ConclusionOur findings suggest that the ability of H-NS-like proteins to repress intragenic transcription is a universal function of these proteins and describe a second mechanism by which MvaT and MvaU may contribute to the growth of P. aeruginosa.Disclosures All Authors: No reported disclosures