H-NS Family Members MvaT and MvaU Regulate the Pseudomonas aeruginosa Type III Secretion System.

Pseudomonas aeruginosa is an opportunistic Gram-negative pathogen capable of causing severe disease in immunocompromised individuals. A major P. aeruginosa virulence factor is the type III secretion system (T3SS). The T3SS is used to translocate effector proteins into host cells, causing cytotoxicity. The T3SS is under the transcriptional control of the master regulator ExsA. ExsA is encoded in the exsCEBA operon and autoregulates transcription via the P exsC promoter. There is also a Vfr-dependent promoter (P exsA ) located in the intergenic region between exsB and exsA A previous chromatin immunoprecipitation (ChIP)-on-chip experiment identified strong binding signatures for MvaT and MvaU in the intergenic region containing the P exsA promoter. MvaT and MvaU are DNA-binding histone-like nucleoid-structuring proteins that can repress gene expression. As predicted from the previous ChIP data, purified MvaT specifically bound to the P exsA promoter region in electrophoretic mobility shift assays. Whereas disruption of mvaT or mvaU by either transposon insertion or clustered regularly interspaced short palindromic repeat interference (CRISPRi) derepressed P exsA promoter activity and T3SS gene expression, overexpression of MvaT or MvaU inhibited P exsA promoter activity. Disruption of mvaT, however, did not suppress the Vfr requirement for P exsA promoter activity. Mutated MvaT/MvaU defective in transcriptional silencing exhibited dominant negative activity, resulting in a significant increase in P exsA promoter activity. Because no effect of MvaT or MvaU on Vfr expression was detected, we propose a model in which the primary effect of MvaT/MvaU on T3SS gene expression is through direct silencing of the P exsA promoter.IMPORTANCE Global regulatory systems play a prominent role in controlling the P. aeruginosa T3SS and include the Gac/RsmA, c-di-GMP, and Vfr-cAMP signaling pathways. Many of these pathways appear to directly or indirectly influence exsA transcription or translation. In this study, the histone-like proteins MvaT and MvaU are added to the growing list of global regulators that control the T3SS. MvaT and MvaU bind AT-rich regions in the genome and silence xenogeneic genes, including pathogenicity islands. The T3SS gene cluster has been horizontally transmitted among many Gram-negative pathogens. Control by MvaT/MvaU may reflect a residual effect that has persisted since the initial acquisition of the gene cluster, subsequently imposing a requirement for active regulatory mechanisms to override MvaT/MvaU-mediated silencing.

Fitting Pieces into the Puzzle of Pseudomonas aeruginosa Type III Secretion System Gene Expression.

Type III secretion systems (T3SS) are widely distributed in Gram-negative microorganisms and critical for host-pathogen and host-symbiont interactions with plants and animals. Central features of the T3SS are a highly conserved set of secretion and translocation genes and contact dependence wherein host-pathogen interactions trigger effector protein delivery and serve as an inducing signal for T3SS gene expression. In addition to these conserved features, there are pathogen-specific properties that include a unique repertoire of effector genes and mechanisms to control T3SS gene expression. The Pseudomonas aeruginosa T3SS serves as a model system to understand transcriptional and posttranscriptional mechanisms involved in the control of T3SS gene expression. The central regulatory feature is a partner-switching system that controls the DNA-binding activity of ExsA, the primary regulator of T3SS gene expression. Superimposed upon the partner-switching mechanism are cyclic AMP and cyclic di-GMP signaling systems, two-component systems, global regulators, and RNA-binding proteins that have positive and negative effects on ExsA transcription and/or synthesis. In the present review, we discuss advances in our understanding of how these regulatory systems orchestrate the activation of T3SS gene expression in the context of acute infections and repression of the T3SS as P. aeruginosa adapts to and colonizes the cystic fibrosis airways.