RpoHI and RpoHII regulons in Rhodobacter sphaeroides 2.4.1
Ontology highlight
ABSTRACT: This SuperSeries is composed of the following subset Series: GSE39711: RpoHI and RpoHII regulons in Rhodobacter sphaeroides 2.4.1 from chromatin immuno-precipitation GSE39712: RpoHI and RpoHII regulons in Rhodobacter sphaeroides 2.4.1 from gene expression profiling Refer to individual Series
Project description:In this study, we performed a ChIP-chip experiment to determine the respective regulons of RpoHI and RpoHII in Rhodobacter sphaeroides. We grew R. sphaeroides in aerobic conditions and induced either proteins ectopically and immuno-precipitated the regions of the genomic DNA interacting with the sigma factors. DNA immunoprecipitated with antibodies against RpoHI or RpoHII was labelled with Cy5 and hybridized on two-color tilling arrays (triplicates for each) with genomic DNA as an input control labelled with Cy3.
Project description:We determined the genomic locations of the SigR binding sites by chromatin immuno-precipitation of cross-linked DNA and hybridization on a tilling oligonucleotide array after 20 minutes of diamide treatment on cell cultures. Three independent biological samples were prepared separately from the wild-type cells and pooled before hybridization on a single array. Three more independent biological samples were prepared from sigR deletion mutant cells, pooled, and hybridized on another single array. After normalization, the signal from the mutant cells was subtracted from the signal from the wild-type cells to recover SigR specific signal.
Project description:In Rhodobacter sphaeroides a transcriptional response to the reactive oxygen species singlet oxygen is controlled by the group IV sigma factor RpoE and the anti-sigma factor ChrR. In this study, we integrated various large datasets to identify genes within the singlet oxygen stress response that contain RpoE-dependent promoters within R. sphaeroides. Transcript pattern clustering and a RpoE-binding sequence model were used to predict candidate promoters that respond to singlet oxygen stress in R. sphaeroides. These candidate promoters were experimentally validated to nine R. sphaeroides RpoE-dependent promoters that control the transcription of 15 genes activated by singlet oxygen. DNA immunoprecipitated with polyclonal antibodies against RpoE or the Beta' subunit of RNA polymerase was labelled with Cy5 and hybridized on two-color tilling arrays (triplicates for each) with genomic DNA as an input control labelled with Cy3.
Project description:In this study, we performed a ChIP-chip experiment to determine the regulon of FnrL in Rhodobacter sphaeroides. We grew R. sphaeroides under anaerobic photosnthetic conditions, in which FnrL is expected to bind DNA and contol gene expression, and immuno-precipitated FnrL, but also sigma70 and the Beta' subunits of RNA polymerase to determine transcription activity. DNA immunoprecipitated with polyclonal antibodies against FnrL, sigma70, or the Beta' subunit of RNA polymerase was labelled with Cy5 and hybridized on two-color tilling arrays (triplicates for each) with genomic DNA as an input control labelled with Cy3.
Project description:Mapping the occupancy of ArcA throughout the genome of Escherchia coli MG1655 K-12 using an affinity purified antibody under anaerobic and aerobic growth conditions. As a control, we also performed ChIP-chip onArcA in a M-bM-^HM-^FarcA mutant strain of Escherchia coli MG1655 K-12. Described in the manuscript The response regulator ArcA uses a diverse binding site architechture to globally regulate carbon oxidation in E. coli Mapping of occupancy of ArcA in the genome of Escherchia coli MG1655 K-12 during anaerobic fermentation and aerobic respiration. Immunoprecipitated DNA compared to INPUT for each sample.
Project description:Mapping the occupancy of FNR, HNS, and IHF throughout the genome of Escherchia coli MG1655 K-12 using an affinity purified antibody under anerobic growth conditions. We also mapped the binding of the M-CM-^_ subunit of RNA Polymerase under both aerobic and anaerobic growth conditions. As a control, we also performed ChIP-chip on FNR in a M-bM-^HM-^Ffnr mutant strain of Escherchia coli MG1655 K-12. We also examined FNR immunoprecipitation at various FNR concentrations using IPTG and Ptac::fnr (PK8263). The M-bM-^HM-^Fhns/M-bM-^HM-^FstpA strains were also used. Descirbed in the manuscript Genome-scale Analysis of E. coli FNR Reveals the Complexity of Bacterial Regulon Structure Mapping of occupancy of FNR, NNS, IHF and M-CM-^_ of RNAP in the genome of Escherchia coli MG1655 K-12 under aerobic or anaerobic growth conditions. Immunoprecipitated DNA compared to INPUT for each sample.
Project description:Despite the prevalence of antisense transcripts in bacterial transcriptomes, little is known about how their synthesis is controlled. We report that a major function of the Escherichia coli termination factor Rho and its co-factor NusG is suppression of ubiquitous antisense transcription genome-wide. Rho binds C-rich unstructured nascent RNA (high C/G ratio) prior to its ATP-dependent dissociation of transcription complexes. NusG is required for efficient termination at minority subsets (~20%) of both antisense and sense Rho-dependent terminators with lower C/G ratio sequences. In contrast, a widely studied nusA deletion proposed to compromise Rho-dependent termination had no effect on antisense or sense Rho-dependent terminators in vivo. Global co-localization of the nucleoid-associated protein H-NS with Rho-dependent terminators and genetic interactions between hns and rho suggest that H-NS aids Rho in suppression of antisense transcription. The combined actions of Rho, NusG, and H-NS appear to be analogous to the Sen1-Nrd1-Nab3 and nucleosome systems that suppress antisense transcription in eukaryotes. Chromatin immunoprecipitation (ChIP) experiments were performed using antibodies against RNA polymerase (RNAP; Beta subunit) in wild-type cells or cells deleted for hns, nusG, or a partial deletion of nusA. Differentially labeled ChIP DNA and genomic DNA were competitively hybridized to an E. coli K-12 MG1655 tiling array with overlapping probes at ~12bp spacing across the entire genome. The series contains 12 datasets.
Project description:Among its many roles in development, retinoic acid determines the anterior-posterior identity of differentiating motor neurons by activating Retinoic Acid Receptor (RAR)-mediated transcription. RAR is thought to bind the genome constitutively, and only induce transcription in the presence of the retinoid ligand. However, little is known about where RAR binds to the genome or how it selects target sites. We tested the constitutive RAR binding model using the retinoic acid-driven differentiation of mouse embryonic stem cells into differentiated motor neurons. We find that retinoic acid treatment results in widespread changes in RAR genomic binding, including novel binding to genes directly responsible for anterior-posterior specification, as well as the subsequent recruitment of the basal polymerase machinery. Finally, we discovered that the binding of transcription factors at the embryonic stem cell stage can accurately predict where in the genome RAR binds after initial differentiation. We have characterized a ligand-dependent shift in RAR genomic occupancy at the initiation of neurogenesis. Our data also suggests that enhancers active in pluripotent embryonic stem cells may be preselecting regions that will be activated by RAR during neuronal differentiation. The differentiation of ventral motor neurons is induced by treating embryonic stem cell cultures with retinoic acid. Here, ChIP-seq is used to profile the genome-wide occupancy of RAR isofroms both immediately prior to and during exposure of the cells to retinoic acid. ChIP-seq is also used to profile the genomic occupancy of Pol2 with phosphorylated serine 5 (Pol2-S5P) and phosphorylated serine 2 (Pol2-S2P) after exposure to retinoic acid.