SummaryWe report here the results of an analysis of the regulatory range of the GacS/GacA two-component system in Pseudomonas aeruginosa. Using microarrays, we identified a large number of genes that are regulated by the system, and detected a near complete overlap of these genes with those regulated by two small RNAs (sRNAs), RsmY and RsmZ, suggesting that the expression of all GacA-regulated genes is RsmY/Z-dependent. Using genome-wide DNA-protein interaction analyses, we identified only two genomic regions that associated specifically with GacA, located upstream of the rsmY and rsmZ genes. These results demonstrate that in P. aeruginosa, the GacS/ GacA system transduces the regulatory signals to downstream genes exclusively by directly controlling the expression of only two genes rsmY and rsmZ. These two sRNAs serve as intermediates between the input signals and the output at the level of mRNA stability, although additional regulatory inputs can influence the levels of these two riboregulators. We show that the A+T-rich DNA segment upstream of rsmZ is bound and silenced by MvaT and MvaU, the global gene regulators of the H-NS family. This work highlights the importance of post-transcriptional mechanisms involving sRNAs in controlling gene expression during bacterial adaptation to different environments.

Type III secretion systems are toxin delivery systems that are present in a large number of pathogens. A hallmark of all type III secretion systems studied to date is that expression of one or more of their components is induced upon cell contact. It has been proposed that this induction is controlled by a negative regulator that is itself secreted by means of the type III secretion machinery. Although candidate proteins for this negative regulator have been proposed in a number of systems, for the most part, a direct demonstration of their role in regulation is lacking. Here, we report the discovery of ExsE, a negative regulator of type III secretion gene expression in Pseudomonas aeruginosa. Deletion of exsE deregulates expression of the type III secretion genes. We provide evidence that ExsE is itself secreted by means of the type III secretion machinery and physically interacts with ExsC, a positive regulator of the type III secretion regulon. Taken together, these data demonstrate that ExsE is the secreted negative regulator that couples triggering of the type III secretion machinery to induction of the type III secretion genes.contact-dependent secretion ͉ pathogenesis ͉ virulence

The histone-like nucleoid structuring protein, H-NS, is a prominent global regulator of gene expression. Many Gram-negative bacteria contain multiple members of the H-NS family of proteins. Thus, a key question is whether H-NS family members have overlapping or distinct functions. To address this question we performed genomewide location analyses with MvaT and MvaU, the two H-NS family members present in Pseudomonas aeruginosa. We show that MvaT and MvaU bind the same chromosomal regions, coregulating the expression of Ϸ350 target genes. We show further that like H-NS in enteric bacteria, which functions as a transcriptional silencer of foreign DNA by binding to AT-rich elements, MvaT and MvaU bind preferentially to AT-rich regions of the chromosome. Our findings establish that H-NS paralogs can function coordinately to regulate expression of the same set of target genes, and suggest that MvaT and MvaU are involved in silencing foreign DNA elements in P. aeruginosa.MvaT ͉ MvaU ͉ Pseudomonas aeruginosa