Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Chromatin immunoprecipitation of human cell lines with oestrogen receptor (ER), the E2F transcription factor 4 (E2F4),

ABSTRACT: The Goal of the experiment was to validate the predicted regulatory modules identified by an algorithm.For this we took advantage of the the genome-wide location analysis technique (or ?ChIP/chip?). We selected modules predicted to be bound by the estrogen receptor (ER), the E2F transcription factor 4 (E2F4), the signal transducer and activator of transcription 3 (STAT3), and the hypoxia-inducible factor 1 HIF1), to print a DNA microarray. In the current study, the microarray was then probed by ChIP/chip for ER and E2F4.

INSTRUMENT(S): Axon- GenePix4000B

ORGANISM(S): Homo sapiens

SUBMITTER: Dominique Bergeron

PROVIDER: E-MEXP-471 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Genome-wide computational prediction of transcriptional regulatory modules reveals new insights into human gene expression.

Blanchette Mathieu M Bataille Alain R AR Chen Xiaoyu X Poitras Christian C Laganière Josée J Lefèbvre Céline C Deblois Geneviève G Giguère Vincent V Ferretti Vincent V Bergeron Dominique D Coulombe Benoit B Robert François F

Genome research 20060410 5

The identification of regulatory regions is one of the most important and challenging problems toward the functional annotation of the human genome. In higher eukaryotes, transcription-factor (TF) binding sites are often organized in clusters called cis-regulatory modules (CRM). While the prediction of individual TF-binding sites is a notoriously difficult problem, CRM prediction has proven to be somewhat more reliable. Starting from a set of predicted binding sites for more than 200 TF families ...[more]

PMID: 16606704

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Project description:Transcription of the several hundred of mouse and human Ribosomal RNA (rRNA) genes accounts for the majority of RNA synthesis in the cell nucleus and is the determinant of cytoplasmic ribosome abundance, a key factor in regulating gene expression. The rRNA genes, referred to globally as the rDNA, are clustered as direct repeats at the Nucleolar Organiser Regions, NORs, of several chromosomes, and in many cells the active repeats are transcribed at near saturation levels. The rDNA is also a hotspot of recombination and chromosome breakage, and hence understanding its control has broad importance. Despite the need for a high level of rDNA transcription, typically only a fraction of the rDNA is transcriptionally active, and some NORs are permanently silenced by CpG methylation. Various chromatin-remodelling complexes have been implicated in counteracting silencing to maintain rDNA activity. However, the chromatin structure of the active rDNA fraction is still far from clear. Here we have combined a high-resolution ChIP-Seq protocol with conditional inactivation of key basal factors to better understand what determines active rDNA chromatin. The data resolve questions concerning the interdependence of the basal transcription factors, show that preinitiation complex formation is driven by the architectural factor UBF (UBTF) independently of transcription, and exclude a significant role for termination by a torpedomechanism. They further reveal the existence of an asymmetric Boundary Complex formed by CTCF, Cohesin and three phased nucleosomes lying adjacent to the rDNA Enhancer and an arrested RNA Polymerase I complex. We find that this complex is the only site of active histone modification in the whole 45kbp rDNA repeat. Strikingly, the Enhancer Boundary Complex not only delimits each functional rRNA gene, but also is stably maintained after gene inactivation and the re-establishment of surrounding repressive chromatin. Our data define the poised state of rDNA chromatin and place the Enhancer Boundary Complex as the likely entry point for the chromatin remodelling complexes.

Project description:Cross-talk between DNA methylation and histone modifications drives the establishment of composite epigenetic signatures and is traditionally studied using correlative rather than direct approaches. Here we present sequential ChIP-bisulfite-sequencing (ChIP- BS-seq) as an approach to quantitatively assess DNA methylation patterns associated with chromatin modifications or chromatin-associated factors directly. A chromatin- immunoprecipitation (ChIP)-capturing step is used to obtain a restricted representation of the genome occupied by the epigenetic feature of interest, for which a single-base resolution DNA methylation map is then generated. When applied to H3 lysine 27 tri- methylation (H3K27me3), we found that H3K27me3 and DNA methylation are compatible throughout most of the genome, except for CpG islands, where these two marks are mutually exclusive. Further ChIP-BS-seq-based analysis in Dnmt triple- knock-out (TKO) embryonic stem cells revealed that total loss of CpG methylation is associated with alteration of H3K27me3 levels throughout the genome: H3K27me3 in localized peaks is decreased while broad local enrichments (BLOCs) of H3K27me3 are formed. At an even broader scale, these BLOCs correspond to regions of high DNA methylation in wild-type ES cells, suggesting that DNA methylation prevents H3K27me3 deposition locally and at megabase scale. Our strategy provides an unique way of investigating global interdependencies between DNA methylation and other chromatin features. ChIP (chromatin immunoprecipitation) is followed by bisulfite conversion and deep sequencing to directly assess DNA methylation levels in captured chromatin fragments (ChIP-BS-seq). We used ChIP-BS-seq to study the potential global cross-talk between H3K27me3 and DNA methylation, which are both linked to repression. First, we used capturing of methylated DNA, followed by bisulfite-deep sequencing (MethylCap-BS-seq). Genomic DNA isolated from normal and tumor colon tissues was used for MethylCap-BS-seq as well as for conventional MethylCap-seq experiments. Second, we performed ChIP-BS-seq on H3K27me3, using HCT116 colon carcinoma cells. Third, to further study the relevance of the observations, we generated genome-wide profiles for H3K27me3 and DNA methylation by conventional ChIP-seq and MethylCap-seq, and RNA-seq, respectively. Finally, we performed H3K27me3-ChIP-BS-seq and MethylCap-seq on wild-type mouse ES cells as well as Dnmt-triple-knockout (TKO) mouse ES cells.

Dataset Information

Chromatin immunoprecipitation of human cell lines with oestrogen receptor (ER), the E2F transcription factor 4 (E2F4),

Publications

Genome-wide computational prediction of transcriptional regulatory modules reveals new insights into human gene expression.

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