ChIP-seq study of sigma factors in Pseudomonas aeruginosa
Ontology highlight
ABSTRACT:
ORGANISM(S): Pseudomonas aeruginosa
PROVIDER: GSE54997 | GEO | 2015/02/12
SECONDARY ACCESSION(S): PRJNA238228
REPOSITORIES: GEO
Similar Datasets
Project description:Sigma factors are master regulators of bacterial transcription which direct gene expression of specific subsets of genes. In particular, alternative sigma factors are well-known to be key players of bacterial adaptation to changing environments. To elucidate the regulatory network of sigma factors in P. aeruginosa, an integrative approach including sigma factor-dependent mRNA profiling was performed to define the primary regulon of each sigma factor.
2015-02-12 | GSE54998 | GEO
Project description:Sigma factors are master regulators of bacterial transcription which direct gene expression of specific subsets of genes. In particular, alternative sigma factors are well-known to be key players of bacterial adaptation to changing environments. To elucidate the regulatory network of sigma factors in P. aeruginosa, an integrative approach including ChIP-seq of 11 polyhistidine-tag sigma factors was performed to define the primary regulon of each sigma factor. Sigma factor genes were fused to a polyhistidine-tag and provided in trans. Under optimal conditions regarding sigma factor activity and induction of sigma factor expression, DNA-sigma factor interactions were conserved by formaldehyde treatment. Upon DNA fragmentation by sonication, the complexes were specifically immunoprecipitated by polyclonal anti-6X His-tag antibodies and the purified DNA was analyzed by Illumina sequencing. DNA enrichment to a control strain was calculated and used for peak calling within promoter region (-500,+100) to identify directly regulated genes/operons.
2015-02-12 | E-GEOD-54997 | biostudies-arrayexpress
Project description:Sigma factors are master regulators of bacterial transcription which direct gene expression of specific subsets of genes. In particular, alternative sigma factors are well-known to be key players of bacterial adaptation to changing environments. To elucidate the regulatory network of sigma factors in P. aeruginosa, an integrative approach including sigma factor-dependent mRNA profiling was performed to define the primary regulon of each sigma factor. Sigma factor hyper-expressing strains harboring the sigma factor gene in trans under control of the araBAD promoter and sigma factor deletion mutants were constructed. Under optimal conditions regarding sigma factor activity and optional induction of sigma factor expression, bacteria were harvested and total RNA was extracted. Upon mRNA enrichment, RNA was fragmented and ligated to specific RNA-adapters containing a hexameric barcode sequence for multiplexing. These RNA-libraries were reverse transcribed and amplified resulting in cDNA libraries which were sequenced on Illumina platforms. Sequence reads were separated according to their barcodes and barcode sequences were removed. The short reads were mapped to the genome sequence of the reference strain P. aeruginosa PA14 wild-type using stampy with default settings. The R package DESeq was used for differential gene expression analysis.
2015-02-12 | E-GEOD-54998 | biostudies-arrayexpress
Project description:Next to the two-component and quorum sensing systems, cell-surface signaling (CSS) has been recently identified as an important regulatory system in Pseudomonas aeruginosa. CSS senses signals from outside the cell and transmits them into the cytoplasm. It consists of a TonB-dependent outer membrane receptor, a cytoplasmic membrane-localized sigma factor regulator (or anti-sigma factor), and an extracytoplasmic function (ECF) sigma factor. Upon perception of the extracellular signal by the receptor the ECF sigma factor is activated and promotes the transcription of a specific set of gene(s). Although most P. aeruginosa ECF sigma factors are involved in the regulation of iron uptake, we have identified a novel ECF sigma factor (PA0675) involved in the regulation of virulence. By microarray analysis of cells overexpressing PA0675 from the pMUM3 plasmid we have identified the genes regulated by this sigma factor.
2009-04-21 | GSE15697 | GEO
Project description:We analyzed a deletion mutant of the ECF σ factor SigX and applied mRNA profiling to define the SigX dependent regulon in P. aeruginosa in response to low osmolarity medium conditions. Furthermore, the combination of transcriptional data with chromatin immunoprecipitation (ChIP) followed by high-throughput sequencing led to the identification of the DNA binding motif of SigX. Genome-wide mapping of SigX-binding regions revealed enrichment of downstream genes involved in fatty acid biosynthesis, type III secretion, swarming and c-di-GMP signaling. In accordance, a sigX deletion mutant exhibited an altered fatty acid composition of the cell membrane, reduced cytotoxicity, impaired swarming activity, elevated c-di-GMP levels and increased biofilm formation. In conclusion, a combination of ChIP-seq with transcriptional profiling and bioinformatic approaches to define consensus DNA binding sequences proved to be effective for the elucidation of the regulon of the alternative σ factor SigX revealing its role in complex virulence-associated phenotypes in P. aeruginosa.
2013-11-11 | GSE50937 | GEO
Project description:Analysis of a SigX knockout mutant of Pseudomonas aeruginosa H103 strain in minimal medium with glucose as carbon source (M9G). SigX, one of the 19 extra-cytoplasmic function sigma factors of P. aeruginosa, was only known to be involved in transcription of the gene encoding the major outer membrane protein OprF in Pseudomonas aeruginosa. Deletion of the ECF sigma factor sigX gene provide insights into the SigX role in several virulence and biofilm- related phenotypes in Pseudomonas aeruginosa.
2014-01-01 | GSE51076 | GEO
Project description:Next to the two-component and quorum sensing systems, cell-surface signaling (CSS) has been recently identified as an important regulatory system in Pseudomonas aeruginosa. CSS senses signals from outside the cell and transmits them into the cytoplasm. It consists of a TonB-dependent outer membrane receptor, a cytoplasmic membrane-localized sigma factor regulator (or anti-sigma factor), and an extracytoplasmic function (ECF) sigma factor. Upon perception of the extracellular signal by the receptor the ECF sigma factor is activated and promotes the transcription of a specific set of gene(s). Although most P. aeruginosa ECF sigma factors are involved in the regulation of iron uptake, we have identified a novel ECF sigma factor (PA0675) involved in the regulation of virulence. By microarray analysis of cells overexpressing PA0675 from the pMUM3 plasmid we have identified the genes regulated by this sigma factor. Two different samples are analyzed namely P. aeruginosa (pMMB67EH) (control/reference sample) and P. aeruginosa (pMUM3) (overexpressing the PA0675 ECF sigma factor). Two different replicates per sample are included.
2009-04-21 | E-GEOD-15697 | biostudies-arrayexpress
Project description:Bacterial gene expression is controlled by modifying the promoter affinity of the RNA polymerase (RNAP). This control occurs through the substitution of the RNAP σ subunit and by the interaction with transcription factors. The pathogen Pseudomonas aeruginosa contains several σ factors, including σVreI. This protein belongs to the extracytoplasmic function group of the σ70 family. Expression and activity of σECFs are tightly regulated and only occur in response to specific signals. σVreI is encoded within the vreAIR gene cluster. The expression of this cluster occurs in response to inorganic phosphate (Pi) limitation. σVreI activity is modulated by VreR anti-sigma factor, which keeps σVreI inactive. In response to a still unidentified host signal, VreR is proteolytically degraded and σVreI released and activated. σVreI interacts then with the RNAP and targets expression of the σVreI regulon, which includes virulence genes that increase P. aeruginosa pathogenicity. In this work, we compared the transcriptome of the P. aeruginosa PAO1 wild-type strain with that of the ΔvreR mutant upon bacterial growth in Pi starvation. Forty-six transcripts were more abundant in the ΔvreR mutant than in the PAO1 wild-type strain, and nine were less abundant, including the vreR transcript.
2019-03-13 | GSE122253 | GEO
Project description:The bacterial heat-shock response is regulated by the alternative sigma factor sigma 32 (RpoH), which responds to misfolded protein stress and directs the RNA polymerase to the promoterss for genes required for protein refolding or degradation. In P. aeruginosa, RpoH is essential for viability under laboratory growth conditions. Here, we used a transcriptomics approach to identify the genes of the RpoH regulon, including RpoH-regulated genes that are essential for P. aeruginosa. We placed the rpoH gene under control of the arabinose inducible PBAD promoter, then deleted the chromosomal rpoH allele. This allowed transcriptomic analysis of the RpoH regulon following a short up-shift in the cellular concentration of RpoH by arabinose addition, in the absence of a sudden increase in temperature. The P. aeruginosa ∆rpoH (PBAD-rpoH) strain grew in the absence of arabinose, indicating that some rpoH expression occurs without arabinose-induction. When arabinose was added, the rpoH mRNA abundance of P. aeruginosa ∆rpoH (PBAD-rpoH) measured by RT-qPCR increased fivefold within 15 min of arabinose addition. Whole genome transcriptome results showed that P. aeruginosa genes required for protein repair or degradation are induced by increased RpoH levels, and that many of the genes induced by RpoH are essential for P. aeruginosa growth. Other stress response genes induced by RpoH are involved in nucleic acid damage and repair and in amino acid metabolism. Annotation of the hypothetical proteins under RpoH control included proteins that may play a role in antibiotic resistances and in non-ribosomal peptide synthesis. The P. aeruginosa ∆rpoH (PBAD-rpoH) strain is impaired in its ability to survive during starvation compared to the wild-type strain. P. aeruginosa ∆rpoH (PBAD-rpoH) also has increased sensitivity to aminoglycoside antibiotics, but not to other classes of antibiotics, whether cultured planktonically or in biofilms. The enhanced aminoglycoside resistance of the mutant strain may be due to indirect effects, such as the build-up of toxic misfolded proteins, or to the direct effect of genes such as aminoglycoside acetyl transferases that are regulated by RpoH. Overall, the results demonstrate that RpoH regulates genes that are essential for viability of P. aeruginosa, that it protects P. aeruginosa from damage from aminoglycoside antibiotics, and that it is required for survival during nutrient limiting conditions. We used Affymetrix microarrays to characterize the RpoH regulon in P. aeruginosa. Using the P. aeruginosa ∆rpoH strain with rpoH under control of the PBAD promoter, we were able to perform transcriptomic analysis of genes induced by a sudden increase (15 min) in the cellular concentration of RpoH, independent from a sudden increase in temperature.
2022-11-06 | GSE217157 | GEO
Project description:Alginate overproduction by P. aeruginosa, also known as mucoidy is associated with chronic endobronchial infections in cystic fibrosis (CF). Alginate biosynthesis in this bacterium is initiated by the extracytoplasmic function sigma factor (σ22, AlgU/T). In the wild type (wt) nonmucoid strains, such as PAO1, AlgU is sequestered by the anti-sigma factor MucA that inhibits alginate production. However, the degradation of MucA by activated intramembrane proteases AlgW and/or MucP can lead to the conversion from nonmucoid strains to mucoid. Previously we reported that the absence of the sensor kinase KinB in PAO1 causes the initiation of AlgW-dependent proteolysis of MucA resulting in alginate overproduction. In the kinB mutant this activation requires alternate sigma factor RpoN (σ54). To determine the RpoN-dependent KinB regulon, microarray and proteomic analyses were performed on a mucoid kinB mutant and an isogenic nonmucoid kinB rpoN double mutant. In the kinB mutant, RpoN controlled the expression of approximately 20% of the genome. Besides alginate biosynthesis and regulator genes such as AlgW, KinB, in concert with RpoN, also control a large number of genes including: those involved in carbohydrate metabolism, quorum sensing, iron regulation, rhamnolipid production, and motility. In an acute pneumonia murine infection model, mice exhibited better survival when challenged with the kinB mutant than wt PAO1. Together, these data strongly suggest that KinB controls virulence factors important for acute pneumonia and conversion to mucoidy.
2012-03-01 | GSE35248 | GEO