Project description:Quorum sensing controls the expression of multiple virulence factors. PA14 genes lasR and rhlR are necessary for quorum sensing via homoserine lactones. A PA14 lasR rhlR deficient mutant exhibits a reduced oxidative stress response. Here we conducted a microarray to determine oxidative stress response gene regulation mediated by the homoserine lactone quorum sensing circuits. A PA14 lasR rhlR deficient mutant was compared to the wild-type with and without H2O2 stress.
Project description:Quorum sensing controls the expression of multiple virulence factors. PA14 genes lasR and rhlR are necessary for quorum sensing via homoserine lactones. A PA14 lasR rhlR deficient mutant exhibits a reduced oxidative stress response. Here we conducted a microarray to determine oxidative stress response gene regulation mediated by the homoserine lactone quorum sensing circuits.
Project description:RNA-seq of the previously uncharacterized QslA and QteE regulons revealed overlapping, yet distinct groups of differentially expressed genes. Simultaneous inactivation of qteE and qslA had the largest effect on gene expression with 999 genes induced and 798 genes repressed in the double mutant vs. wild-type. We found that LasR and RhlR-activated QS genes formed a subset of the genes induced in the qteE, qslA, and double mutant. The activation of almost all of these QS genes was advanced from stationary phase to log phase in the qteE qslA double mutant.
Project description:To better understand the role of QscR in P. aeruginosa gene regulation and to better understand the relationship between QscR, LasR and RhlR control of gene expression we used transcription profiling to identify a QscR-dependent regulon. Our analysis revealed that QscR activates some genes and represses others. Some of the repressed genes are not regulated by the LasR-I or RhlR-I systems while others are. The LasI-generated 3-oxododecanoyl-homoserine lactone serves as a signal molecule for QscR. Thus QscR appears to be an integral component of the P. aeruginosa quorum sensing circuitry. QscR uses the LasI-generated acyl-homoserine lactone signal and controls a specific regulon that overlaps with the already overlapping LasR and RhlR-dependent regulons. Keywords: Quorum sensing regulon, Direct activation
Project description:Pseudomonas aeruginosa is a troublesome opportunistic pathogen isolated from diverse environmental sources. An arsenal of degrading enzymes and antagonistic factors contribute to P. aeruginosa persistence and damage of a susceptible host. Largely through density-dependent regulation referred to as quorum sensing, the LasR, RhlR, and PqsR transcription factors collectively modulate hundreds of genes, including the expression of several virulence factors, in response to diffusible signals called autoinducers. LasR loss-of-function (LasR-) strains are commonly isolated from clinical samples and produce fewer toxins in monoculture, yet these strains are associated with worse clinical outcomes. We show that in co-culture with P. aeruginosa wild type where LasR loss-of-function strains are often found in vivo, ∆lasR hyperproduces RhlR/I dependent antagonistic factors. Specifically, we present a cyclic model of interaction between wild type and ∆lasR wherein the iron-scavenging siderophore pyochelin produced by the lasR mutant induces citrate release and cross-feeding from the wild type to ∆lasR to stimulate production of antagonistic factors with native functions involved in iron acquisition. Co-culture specific behaviors mediated by altered metabolite secretion and metabolism may explain complications associated with LasR loss-of-function strains. More broadly, this report illustrates how heterogenous behaviors within a mono-species community can promote antagonism associated with carbon and metal assimilation.
Project description:The PqsE enzyme plays a vital role in quorum sensing and virulence in Pseudomonas aeruginosa, yet its enzymatic function is unknown. Here, we identify the protein interaction network of PqsE as well as that of a catalytically dead variant, PqsE(D73A) in P. aeruginosa PA14. Our analyses identify proteins that interact with PqsE that are independent of and that depend on PqsE catalytic function. One such catalysis-independent interaction is with the quorum-sensing regulator, RhlR, consistent with our previous work. We also characterize the PqsE interaction network in a delta rhlR P. aeruginosa PA14 strain and identify additional proteins as PqsE-interactors.
Project description:To better understand the role of QscR in P. aeruginosa gene regulation and to better understand the relationship between QscR, LasR and RhlR control of gene expression we used transcription profiling to identify a QscR-dependent regulon. Our analysis revealed that QscR activates some genes and represses others. Some of the repressed genes are not regulated by the LasR-I or RhlR-I systems while others are. The LasI-generated 3-oxododecanoyl-homoserine lactone serves as a signal molecule for QscR. Thus QscR appears to be an integral component of the P. aeruginosa quorum sensing circuitry. QscR uses the LasI-generated acyl-homoserine lactone signal and controls a specific regulon that overlaps with the already overlapping LasR and RhlR-dependent regulons. Experiment Overall Design: First comparisaon Experiment Overall Design: We first compared transcriptomes of the qscR mutant P. aeruginosa PAO-R3 and the parental strain PAO1 at several points during growth (OD600nm 0.5, 0.8, 1.4, 2.0 and 3.5). To identify those genes with expression significantly different between PAO1 and PAO-R3 at different culture densities we used CYBER-T. The Bayesian prior estimate was 10 and the sliding window size was 101. The p-value threshold was 0.001, the posterior probability of differential expression >0.95, and fold change was >2.5. Experiment Overall Design: Second comparison Experiment Overall Design: We selected genes that were at least 3-fold differentially expressed in the strain containing the L-arabinose promoter-driven qscR allele vs the strain containing L-arabinose promoter-driven qscR-Ddbd allele, and were also at least 3-fold differentially expressed in the parent strain PAO1 as compared to the strain containing the qscR null mutation. RNA were extracted from cultures at OD 0.5, 0.8, 1.4, 2.0 and 3.5
Project description:Early stages of host microbe adaptations involve 'system status changes' (rewiring of pre-existing cellular signaling networks and components) of the host and microbe. We posited that under certain environmental conditions these changes leads to maladaptations and favor emergence of new infectious diseases, and these adaptations will have characteristic signatures representative of the adaptation. Here using Arabidopsis seedlings in a submerged environment treated with P. aerugionsa, we show one such rewired regulation where the master two-component regulator GacA (previously shown to act upstream of quorum sensing, including the regulator LasR, that in turn controls a subset of virulence factors) is completely dispensable. The gacA mutant behaves similar to wild type P. aeruginosa (strain PA14) by a number of read-outs. Consistent with that, the gene expression data here indicates that the transcriptome pattern of the host is identical when treated with wild type PA14 or PA14-gacA mutant. Single time point (10 day old Arabidopsis seedlings infected with wild type PA14 or mutant bacteria PA14DgacA, and analyzed 24h after infection) with two independent experimenal replicates per treatment
Project description:Early stages of host microbe adaptations involve 'system status changes' (rewiring of pre-existing cellular signaling networks and components) of the host and microbe. We posited that under certain environmental conditions these changes leads to maladaptations and favor emergence of new infectious diseases, and these adaptations will have characteristic signatures representative of the adaptation. Here using Arabidopsis seedlings in a submerged environment treated with P. aerugionsa, we show one such rewired regulation where the master two-component regulator GacA (previously shown to act upstream of quorum sensing, including the regulator LasR, that in turn controls a subset of virulence factors) is completely dispensable. The gacA mutant behaves similar to wild type P. aeruginosa (strain PA14) by a number of read-outs. Consistent with that, the gene expression data here indicates that the transcriptome pattern of the host is identical when treated with wild type PA14 or PA14-gacA mutant.
Project description:Gene regulation via transcription factors influences the metabolic, adaptive and pathogenic capabilities of the organism. We report the transcriptomes of the mutants of six major P. aeruginosa PA14 trancription factors - RhlR, LasR, Anr, GacA, FleQ and CbrB.