Project description:The Pseudomonas aeruginosa quorum-sensing (QS) systems contribute to bacterial homeostasis and pathogenicity. Although many regulators have been characterized to control the production of virulence factors and QS signaling molecules, its detailed regulatory mechanisms still remain elusive. Here, we performed chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) on 10 key QS regulators. The direct regulation of these genes by corresponding regulator has been confirmed by Electrophoretic mobility shift assays (EMSAs) and quantitative real-time polymerase chain reactions (qRT-PCR). Binding motifs are found by using MEME suite and verified by footprint assays in vitro. Collectively, this work provides new cues to better understand the detailed regulatory networks of QS systems. ChIP-seq of 10 QS regulators in Pseudomonas aeruginosa
Project description:Analysis of Pseudomonas aeruginosa PAO1 treated with 200 µM sphingomyelin. Results provide insight into the response to sphingomyelin in P. aeruginosa.
Project description:Immune responses in plants are triggered in part by conserved microbe-associated molecular pattern (MAMP) molecules such as bacterial flagellin. Upon MAMP perception, plants rapidly turn on the induction of numerous defense-related genes. We have identified a novel type of plant innate immunity elicitor, protease IV from Pseudomonas aeruginosa. Genome-wide transcriptomic profiles obtained with Affymetrix Arabidopsis ATH1 GeneChips® of 10-day old Arabidopsis seedlings treated with 20 nM purified protease IV for 1 hour were compared to published bacterial flagellin- and oligogalacturonide-triggered responses. We used microarrays to characterize the global transcriptomic changes in Arabidopsis seedlings upon protease IV treatment.
Project description:Oxygenated unsaturated fatty acids, known as oxylipins, are signaling molecules commonly used for cell-to-cell communication in eukaryotes. However, a role for oxylipins in mediating communication in prokaryotes has not previously been described. Bacteria mainly communicate via quorum sensing (QS) , which involves the production and detection of diverse small molecules termed autoinducers. We showed that oleic acid-derived oxylipins 10-HOME and 7,10-DiHOME produced by Pseudomonas aeruginosa function as autoinducers of a novel quorum sensing system termed Oxylipin-Dependent QS Sytem (ODS). This experiment was designed to determine the genes whose expression is altered by these P. aeruginosa oxylipins. We found that the ODS system controls the cell density-dependent expression of a P. aeruginosa gene subset through the mediation of 10-HOME and 7,10-DiHOME oxylipins.
Project description:Airway epithelium is the initial point of host-pathogen interaction in Pseudomonas aeruginosa infection, an important pathogen in cystic fibrosis and nosocomial pneumonia. We used global gene expression analysis to determine airway epithelial transcriptional responses dependent on matrilysin (MMP-7) and stromelysin-2 (MMP-10), two matrix metalloproteinases induced by acute P. aeruginosa pulmonary infection. Extraction of Differential Gene Expression (EDGE) analysis of gene expression changes in P. aeruginosa infected organotypic tracheal epithelial cell cultures from wildtype, Mmp7-/-, and Mmp10-/- mice identified 2,089 matrilysin-dependent and 1,628 stromelysin-2-dependent genes that were differentially expressed. Key node network analysis showed that these MMPs controlled distinct gene expression programs involved in proliferation, cell death, immune responses, and signal transduction, among other host defense processes. Our results demonstrate discrete roles for these MMPs in regulating epithelial responses to pseudomonas infection and show that a global genomics strategy can be used to assess MMP function. Keywords: time course
Project description:Purpose: To determine effects of arsenic on gene expression in polarized primary human bronchial epithelial (HBE) cells and impact on transcriptional response to Pseudomonas aeruginosa infection Methods: mRNA profiles of HBE cells from 6 donors exposed to 0, 5, 10 or 50 ug/L total arsenic +/- Pseudomonas aeruginosa (48 samples) were generated using Illumina sequencing, aligned in CLC Genomics workbench and analyzed for DE in EdgeR Findings: 20-30 million reads were mapped per sample and transcripts were identifed that were significantly differentially expressed in response to arsenic and Pseudomonas aeruginosa
Project description:The Pseudomonas aeruginosa quorum-sensing (QS) systems contribute to bacterial homeostasis and pathogenicity. Although many regulators have been characterized to control the production of virulence factors and QS signaling molecules, its detailed regulatory mechanisms still remain elusive. Here, we performed chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) on 10 key QS regulators. The direct regulation of these genes by corresponding regulator has been confirmed by Electrophoretic mobility shift assays (EMSAs) and quantitative real-time polymerase chain reactions (qRT-PCR). Binding motifs are found by using MEME suite and verified by footprint assays in vitro. Collectively, this work provides new cues to better understand the detailed regulatory networks of QS systems.
Project description:As production of virulence factors by Pseudomonas aeruginosa is influenced by the host environment, we examined the effect of 10% adult bovine serum (ABS) on the global transcription with P. aeruginoas PAO1 at different phases of growth. At early exponential phase (4 h), serum significantly enhanced expression of 138 genes, most of which are repressed by iron and carry binding sequences for the ferric uptake regulator or the Fur-regulated extracytoplasmic function sigma factor PvdS. The expression of another 40 genes was reduced at 4h. However, the expression of only a few genes was significantly altered (reduced) at the early stationary phase of growth (8 h). Transcriptional fusion analyses confirmed serum enhances the expression of toxA, regA, and their regulatory genes regA and pvdS, yet does not interfere with the repression of these genes by iron. The P. aeruginosa strain PAO1 was grown in the iron-limited medium TSB-DC with or without 10% (v/v) ABS. The presence of 10% ABS reduced PAO1 growth, but despite this reduction, the cultures reached early exponential phase at 4 h and early stationary phase at 8 h in both media. Accordingly, we determined the PAO1 transcriptome in both media at these two specific time points. To standardize the comprison, we adjusted the cultures to the same optical density at 600 nm (biomass) by diluting with fresh TSB-DC PAO1 grown in TSB-DC to the same OD as PAO1 grown in TSB-DC/ABS. We obtained the transcriptome profile of 5,552 genes representing the complete P. aeruginosa PAO1 genome.