Molecular profiling of activated neurons by phosphorylated ribosome capture
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Zachary Knight
PROVIDER: E-GEOD-40995 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:We report that phosphorylated ribosomes can be immunoprecipitated from mouse brain homogenates, enriching for mRNAs that were selectively expressed in activated neurons Mice were given an injection of concentrated salt solution (2M NaCl, 325 uL) or PBS, and then sacrificed 2h later and hypothalami dissected. Tissue homogenates were prepared from hypothalamus and phosphorylated ribosomes immunoprecipitated using pS6 244/247 antibodies. The immunoprecipitated RNA and total RNA from both control and salt treated animals were then purified and analyzed by Illumina Microarray.
2012-09-20 | E-GEOD-40994 | biostudies-arrayexpress
Project description:We report that phosphorylated ribosomes can be immunoprecipitated from mouse brain homogenates, resulting in enrichment of transcripts expressed in activated neurons. Mice were either injected with a concentrated salt solution or vehicle, hypothalami dissected, and phosphorylated ribosomes immunoprecipitated. RNA was sequenced from the input and IP for each condition (4 samples total).
2012-09-20 | E-GEOD-40969 | biostudies-arrayexpress
Project description:When C. elegans larvae hatch in the absence of food they persist in a stress resistant, developmentally arrested state (L1 arrest) for weeks or until food becomes available. We characterized growth, mRNA expression, and RNA Polymerase II activity genome-wide during L1 arrest and recovery. RNA Pol II binding data resulting from ChIP-Seq experiments using the Illumina Genome Analyzer are included in this GEO submission. Complementary mRNA expression data from the Affymetrix microarray platform can be found at GEO accession# GSE11055. The goals of the Pol II ChIP-Seq compononent of this project were to use Pol II antibodies 1) to investigate patterns of transcription in developmentally arrested larvae, when mRNA expression levels have reached steady state; 2) to investigate patterns of transcription in immediate response to feeding, when mRNA expression levels change dramatically; and 3) to investigate accumulation of Pol II at promoters during arrest and recovery. We started our ChIP studies with the S2 antibody (Abcam ab5095) since it was raised against a Ser2 phosphorylated peptide from the C-terminal domain of Pol II and should therefore be relatively specific for active, elongating Pol II, and it worked well for the first two goals above. In order to investigate accumulation of Pol II at promoters, we used the S2 antibody and complemeted it with antibody 4H8 (Abcam ab5408), which according to the manafacturer recognizes phosphorylated and non-phosphorylated Pol II, and antibody 8WG16 (Abcam ab817), which binds primarily to non-phosphorylated Pol II but also relatively weakly to phosphorylated Pol II. We were somewhat surprised to find that the results obtained with each of these antibodies were very similar, though upon deeper analysis we discovered relatively subtle differences consistent with the relative specificities of each antibody and existing models regarding phosphorylation state of Pol II and its activity and location. In summary, we find that while Pol II continues transcribing starvation genes, it is M-bM-^@M-^XpausedM-bM-^@M-^Y accumulates on the promoters of growth and development genes during L1 arrest. Consistent with it poising arrested larvae for recovery, pausingpromoter accumulation decreases in response to feeding, while elongation and mRNA levels increase. These results demonstrate that Pol II pausing is widespread in C. elegans and that it is nutritionally controlled during development.These results demonstrate that accumulation of Pol II at promoters of growth and development genes is common in C. elegans and that promoter accumulation anticipates nutritionally controlled gene expression during development. RNA Pol II binding was examined with three different antibodies (S2, 4H8, and 8WG16) during either L1 arrest (starvation) or after 1 hr recovery by feeding. For the S2 antibody two different time points during L1 arrest were examined (6 and 12 hr). 14 total samples are included: 4 independent control samples (Input), a pair biological replicates with the S2 antibody at 6 hr L1 arrest, a pair of biological replicates with the 4H8 antibody at 12 hr L1 arrest and at 1 hr recovery, and singletons for the S2 and 8WG16 antibodies at 12 hr L1 arrest and at 1 hr recovery. "GSE13973_PolII_ChIP-Seq.xls.gz" contains 5 worksheets with the following contents: "geneDensity" includes the number of reads per million mapping to each gene model normalized to gene length. Units are reads per million per kilobase. "TSSdensity" includes the number of reads per million mapping to a 200 bp window spanning the most upstream transcription start site for each gene and normalized by length (200 bp). Units are reads per million per kilobase. "geneEnrichment" includes the fold-enrichment of read density over each gene model relative to input. Where input values were below the median of all input values then the median was used. Units are fold-enrichment. "TSSenrichment" includes the fold-enrichment of read density over the 200 bp TSS window relative to input. Where input values were below the median of all input values then the median was used. Units are fold-enrichment. "5' bias" includes 5' bias calculation for each gene and each antibody in each condition. "GSE13973_CelegansWS190_GeneCoordinates.xls.gz" contains the gene coordinates based on version WS190 of the C. elegans genome.
2009-02-27 | E-GEOD-13973 | biostudies-arrayexpress
Project description:Broadly expressed transcriptions factors (TFs) control tissue-specific programs of gene expression through interactions with local TF networks. Prime examples are the circadian clock TFs CLOCK (CLK) and CYCLE (CYC or BMAL1): while they control a core transcriptional circuit throughout animal bodies, downstream clock target genes and circadian physiology are tissue-specific. Here, we use ChIP-seq to determine the regulatory targets of Drosophila CLK and CYC, which we epitope-tagged by homologous recombination. Both TFs have distinct binding sites in heads versus bodies, suggesting that they directly control tissue-specific downstream target genes. Analysis of these context-specific binding sites revealed distinct sequence motifs for putative clock partner factors, including a motif for the GATA factor SERPENT (SRP). SRP indeed synergistically enhances CLK/CYC-mediated activity of a cis-regulatory region bound by CLK/CYC specifically in bodies. These results reveal how universal clock circuits can generate tissue-specific outputs and demonstrate an approach to dissect regulatory interactions more generally. We sequenced ChIP and input samples, as well as M-bM-^@M-^\mockM-bM-^@M-^] samples for which we performed ChIP with the V5 antibody from wildtype w- flies (not carrying the V5 tag) for two independent biological replicates each, summing to 24 libraries in total.
2013-06-01 | E-GEOD-40467 | biostudies-arrayexpress
Project description:To investigate the chromatin transcription cycle, we determined genome-wide occupancy profiles for RNA polymerase (Pol) II, its phosphorylated forms, and transcription factors in growing yeast. ChIP-chip was performed to identify the genomic binding locations for Rpb3, TFIIB, Tfg1, Kin28, Cet1, Spt4, Spt5, Spt6, Elf1, Spn1, Bur1, Ctk1, Paf1, Spt16, Pcf11, and Rpb1 phosphorylated at serine 2, 5, and 7 residues of the CTD, respectively.
2010-08-23 | E-TABM-1033 | biostudies-arrayexpress
Project description:Spt6 is a highly conserved histone chaperone that interacts directly with both RNA polymerase II and histones to regulate gene expression. To gain a comprehensive understanding of the requirements for this critical factor, we have performed genome-wide analyses of transcription, chromatin structure, and histone modifications in an S. pombe spt6 mutant. Our results demonstrate several dramatic changes to transcription and chromatin structure in the spt6 mutant, including an elevation of antisense transcripts at over 70 percent of all genes and general loss of the +1 nucleosome. Furthermore, Spt6 is required for the trimethylation of histone H3 on lysines 4 and 36, marks associated with active transcription. Taken together, our results indicate that Spt6 is critical for the accuracy of transcription and the integrity of chromatin, likely via its direct interactions with RNA polymerase II and histones. ChIP-seq experiments were performed on wild type and spt6-1 strains on the following proteins: RNA polymerase II (Rpb1), Paf1 Complex (Ctr9), COMPASS (Swd1), Set2, Spt6, histones H2B and H3, histone modifications H3K4me3 and H3K36me3. Experiments were performed in replicates and matching inputs were also sequenced.
2013-10-09 | E-GEOD-49574 | biostudies-arrayexpress
Project description:ChIP-chip was performed to identify the genomic binding locations for the termination factors Nrd1, and Rtt103, and for RNA polymerase (Pol) II phosphorylated at the tyrosine 1 and threonine 4 position of its C-terminal domain (CTD). In different phases of the transcription cycle, Pol II recruits different factors via its CTD, which consists of heptapeptide repeats with the sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. Here we show that the CTD of transcribing yeast Pol II is phosphorylated at Tyr1, and that this impairs recruitment of termination factors. Tyr1 phosphorylation levels rise downstream of the transcription start site (TSS), and decrease before the polyadenylation (pA) site. Tyr1-phosphorylated gene bodies are depleted of CTD-binding termination factors Nrd1, Pcf11, and Rtt103. Tyr1 phosphorylation blocks CTD binding by these termination factors, but stimulates binding of elongation factor Spt6. These results show that CTD modifications can not only stimulate but also block factor recruitment, and lead to an extended CTD code for transcription cycle coordination.
2012-06-29 | E-MTAB-1060 | biostudies-arrayexpress
Project description:The aim of the experiment is to define p-AMPK binding sites across the genome upon mice starvation and treatment with Dexamethasone (GR ligand) and GW7647 (PPAR alpha ligand).
2020-10-01 | E-MTAB-7301 | biostudies-arrayexpress
Project description:29-32 days old male mice where either treated with Phenobarbital or untreated Replicated control/treated study with IP and Input per group and antibody
2012-11-10 | E-GEOD-41399 | biostudies-arrayexpress