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ChIP-chip from dissected Drosophila egg chambers using antibody recognizing RNAPII

ABSTRACT: Chromatin immunoprecipitation of RNAPII followed by hybridization to genome-wide tiling microarrays demonstrated that RNAPII localizes to all six Drosophila amplicons in follicle cells (DAFC) as well as other non-amplified genomic loci Localization of RNAPII in stage 10 egg chambers with the nurse cells manually removed

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Terry Orr-Weaver

PROVIDER: E-GEOD-29518 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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ORC2, RNAPII, and tetra-acetylated histone H4 ChIP-chip egg chambers and RNA-Seq 16C ovarian follicle cells

Project description:This SuperSeries is composed of the following subset Series: GSE29517: ChIP-chip from Drosophila egg chambers using ORC2 antibody GSE29518: ChIP-chip from dissected Drosophila egg chambers using antibody recognizing RNAPII GSE29520: ChIP-chip from Drosophila egg chambers using antibody recognizing tetra-acetylated histone H4 GSE29526: Expression profile of 16C ovarian follicle cells Refer to individual Series

2011-05-26 | E-GEOD-29527 | biostudies-arrayexpress

ChIP-chip from Drosophila egg chambers using ORC2 antibody

Project description:Chromatin immunoprecipitation of the Origin Recognition Complex (ORC) followed by hybridization to genome-wide tiling microarrays demonstrated that ORC localizes to the six Drosophila amplicons in follicle cells (DAFC) as well as other non-amplified loci Comparison of ORC2 ChIP sample compared to input DNA for two different Drosophila strains and two developmental stage samples

2011-05-27 | E-GEOD-29517 | biostudies-arrayexpress

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2011-09-06 | E-GEOD-31889 | biostudies-arrayexpress

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Project description:This SuperSeries is composed of the following subset Series: GSE31889: ChIP-chip from Drosophila egg chambers using MCM2-7 antibody GSE31890: ChIP-chip from Drosophila egg chambers using ORC2 antibody Refer to individual Series

2011-09-06 | E-GEOD-31891 | biostudies-arrayexpress

ChIP-chip from Drosophila egg chambers using ORC2 antibody

Project description:Chromatin immunoprecipitation of the Origin Recognition Complex (ORC) followed by hybridization to genome-wide tiling microarrays demonstrated that ORC does not localize to DAFC-34B after stage 10, despite a second round of replication initiation Comparison of ORC2 ChIP sample compared to input DNA for Drosophila egg chambers

2011-09-06 | E-GEOD-31890 | biostudies-arrayexpress

gammaH2Av ChIP-seq in Drosophila stage 10B and 13 follicle cells

Project description:We analyzed gamaH2Av ChIP-seq from hand dissected stage 10B and 13 follicle cell nuclei. Egg chambers were dissected from wild-type (OrR) or H2Av[ΔCT] ovaries to assess binding at the Drosophila Follicle Cell Amplicons and across the genome. ChIP-seq of gammaH2Av bound to follicle cell DNA from stage 10B and 13 egg chambers, collected from wild-type (OrR) and H2Av[ΔCT] Drosophila ovaries. Sequences analyzed by Illumina sequencing. Two replicates are included for each ChIP reaction.

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Kin28 Regulates the Transient Association of Mediator with Core Promoters

Project description:Mediator is an essential, broadly utilized eukaryotic transcriptional co-activator. How and what it communicates from activators to RNA polymerase II remains an open question. Here we performed genome-wide location profiling of yeast Mediator subunits. Mediator is not found at core promoters but rather occupies the upstream activating sequence (UAS), upstream of the pre-initiation complex. In the absence of Kin28/CDK7 kinase activity, or in cells where the CTD is mutated to replace Ser5 with alanines, however, Mediator accumulates at core promoters together with RNAPII. We propose that Mediator is quickly released from promoters upon Ser5 phosphorylation by Kin28/CDK7, which also allows for RNAPII to escape from the promoter. We took a systematic approach to examine the genome-wide distribution (using ChIP-chip) of the various Mediator subunits. Mediator occupancy was also assayed in mutants for most of the CTD kinases and in strains where some CTD serines had been replaced by alanines. Mediator ChIPs were performed with Myc-tagged subunits, except in some preliminary experiments where polyclonal antibodies were used. Most ChIPs (in Cy5) were hybridyzed against a control ChIP sample from an isogenic non-tagged strain (in Cy3). In some of the preliminary experiments, non immunoprecipitated DNA (input) was used as the control. In addition to Mediator ChIPs, the project includes TFIIB and RNAPII (Rpb3) ChIP-chip datasets. All ChIP-chip experiments were done in duplicates except for the preliminary experiments that were done in monoplicat for the most part. Each microarray was normalized using the Lima Loess and replicates were combined using a weighted average method as previously described (Pokholok et al., 2005).

2014-03-24 | E-GEOD-55402 | biostudies-arrayexpress

ChIP-seq of RNA Polymerase II (Rpb3) from wild-type and Rpb7-QHF cells

Project description:Our structural and biochemical studies show that S. cerevisiae RNA Polymerase II (RNAPII) homodimerizes through the stalk domain (formed by the Rpb4-Rpb7 subunits). To explore the biological impact of disrupting this interaction we introduced a triple point mutation at the RNAPII dimerization interface in Rpb7 (Q96A, H97A, F109K). To test the impact of the dimerziation mutant on RNAPII occupancy, we performed ChIP-seq using a monoclonal antibody against the Rpb3 subunit (1Y26, abcam) of RNAPII, in WT and Rpb7-QHF cells.

2023-12-31 | E-MTAB-13369 | biostudies-arrayexpress

DSIF and RNA polymerase II CTD Phosphorylation Coordinate the Recruitment of Rpd3S to Actively Transcribed Genes

Project description:Histone deacetylase Rpd3 is part of two distinct complexes: the large (Rpd3L) and small (Rpd3S) complexes. While Rpd3L targets specific promoters for gene repression, Rpd3S is recruited to ORFs to deacetylate histones in the wake of RNA polymerase II, to prevent cryptic initiation within genes. Methylation of histone H3 at lysine 36 by the Set2 methyltransferase is thought to mediate the recruitment of Rpd3S. Here, we confirm by ChIP-Chip that Rpd3S binds active ORFs. Surprisingly, however, Rpd3S is not recruited to all active genes, and its recruitment is Set2-independent. However, Rpd3S complexes recruited in the absence of H3K36 methylation appear to be inactive. Finally, we present evidence implicating the yeast DSIF complex (Spt4/5) and RNA polymerase II phosphorylation by Kin28 and Ctk1 in the recruitment of Rpd3S to active genes. Taken together, our data support a model where Set2-dependent histone H3 methylation is required for the activation of Rpd3S following its recruitment to the RNA polymerase II C-terminal domain. In order to examine the genome-wide localization of RNAPII and Spt6 in Saccharomyces cerevisiae, RNAPII and Spt6 along with associated DNA sequences were immunoprecipitated using anti-8WG16 and anti-HA antibodies, respectively. The RNAPII and Spt6 chromatin immunoprecipitation was performed in duplicate from WT cells as described below. The extracted DNA was hybridized to a DNA microarray containing an average of 4 probes per kilobase across the whole yeast genome. The combined datasets are available in the supplemental files of the related publication.

2010-11-28 | E-GEOD-22636 | biostudies-arrayexpress

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