CpG islands and GC content dictate nucleosome depletion in a transcription independent manner at mammalian promoters
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Romain Fenouil
PROVIDER: E-GEOD-38577 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:One clear hallmark of mammalian promoters is the presence of CpG islands (CGIs) at more than two thirds of genes whereas TATA boxes are only present at a minority of promoters. Using genome-wide approaches, we show that GC content and CGIs are major promoter elements in mammalian cells, able to govern open chromatin conformation and support paused transcription. First, we define three classes of promoters with distinct transcriptional directionality and pausing properties which correlate with their GC content. We further analyze the direct influence of GC content on nucleosome positioning and depletion, and show that CGIs correlate with nucleosome depletion both in vivo and in vitro. We also show that transcription is not essential for nucleosome exclusion but influences both a weak +1 and a well-positioned nucleosome at CGI borders. Altogether our data support the idea that CGIs have become an essential feature of promoter structure defining novel regulatory properties in mammals. Nucleosome density and positioning were studied by high-throughput sequencing of DNA previously treated with Mnase. In parallel, chIPseq for PolII and H3K27ac were performed in mouse and human with different conditions to assess a potential effect of transcription on nucleosomes properties.
2012-06-12 | E-GEOD-38563 | biostudies-arrayexpress
Project description:One clear hallmark of mammalian promoters is the presence of CpG islands (CGIs) at more than two thirds of genes whereas TATA boxes are only present at a minority of promoters. Using genome-wide approaches, we show that GC content and CGIs are major promoter elements in mammalian cells, able to govern open chromatin conformation and support paused transcription. First, we define three classes of promoters with distinct transcriptional directionality and pausing properties which correlate with their GC content. We further analyze the direct influence of GC content on nucleosome positioning and depletion, and show that CGIs correlate with nucleosome depletion both in vivo and in vitro. We also show that transcription is not essential for nucleosome exclusion but influences both a weak +1 and a well-positioned nucleosome at CGI borders. Altogether our data support the idea that CGIs have become an essential feature of promoter structure defining novel regulatory properties in mammals. Nucleosome density and positioning were studied by high-throughput sequencing of DNA previously treated with Mnase. In parallel, chIPseq for PolII and H3K27ac were performed in mouse and human with different conditions to assess a potential effect of transcription on nucleosomes properties.
2012-06-12 | E-GEOD-38561 | biostudies-arrayexpress
Project description:One clear hallmark of mammalian promoters is the presence of CpG islands (CGIs) at more than two thirds of genes whereas TATA boxes are only present at a minority of promoters. Using genome-wide approaches, we show that GC content and CGIs are major promoter elements in mammalian cells, able to govern open chromatin conformation and support paused transcription. First, we define three classes of promoters with distinct transcriptional directionality and pausing properties which correlate with their GC content. We further analyze the direct influence of GC content on nucleosome positioning and depletion, and show that CGIs correlate with nucleosome depletion both in vivo and in vitro. We also show that transcription is not essential for nucleosome exclusion but influences both a weak +1 and a well-positioned nucleosome at CGI borders. Altogether our data support the idea that CGIs have become an essential feature of promoter structure defining novel regulatory properties in mammals. Nucleosome density and positioning were studied by high-throughput sequencing of DNA previously treated with Mnase. In parallel, ChIP-seq for PolII and H3K27ac were performed in mouse and human with different conditions to assess a potential effect of transcription on nucleosomes properties. To exclude a possible MNase digestion bias or sequencing artefact within CGIs, we performed additional controls such as nucleosome mapping using an alternative method based on DNA digestion by a chemical agent (phenanthroline) in human Raji B-cells.
2012-06-12 | E-GEOD-38564 | biostudies-arrayexpress
Project description:One clear hallmark of mammalian promoters is the presence of CpG islands (CGIs) at more than two thirds of genes whereas TATA boxes are only present at a minority of promoters. Using genome-wide approaches, we show that GC content and CGIs are major promoter elements in mammalian cells, able to govern open chromatin conformation and support paused transcription. First, we define three classes of promoters with distinct transcriptional directionality and pausing properties which correlate with their GC content. We further analyze the direct influence of GC content on nucleosome positioning and depletion, and show that CGIs correlate with nucleosome depletion both in vivo and in vitro. We also show that transcription is not essential for nucleosome exclusion but influences both a weak +1 and a well-positioned nucleosome at CGI borders. Altogether our data support the idea that CGIs have become an essential feature of promoter structure defining novel regulatory properties in mammals. Nucleosome density and positioning were studied by high-throughput sequencing of DNA previously treated with Mnase. In parallel, chIPseq for PolII and H3K27ac were performed in mouse and human with different conditions to assess a potential effect of transcription on nucleosomes properties. Sonicated genomic DNA has been sequenced to quantify and exclude sequencing bias in the largest CGIs regions studied in this article.
2012-06-11 | E-GEOD-38562 | biostudies-arrayexpress
Project description:In mammals, the carboxy-terminal domain (CTD) of RNA polymerase (Pol) II consists of 52 conserved heptapeptide repeats containing the consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. Post-translational modifications of the CTD coordinate the transcription cycle and various steps of mRNA maturation. Here we describe Tyr1 phosphorylation (Tyr1P) as a hallmark of initiating Pol II in mammalian cells, in contrast to what was described in yeast. Tyr1P is predominantly found in antisense orientation at promoters but is also specifically enriched at active enhancers. Mutation of Tyr1 to phenylalanine (Y1F) prevents the formation of the hyper-phosphorylated Pol IIO form, induces degradation of Pol II to the Pol IIB form and results in a lethal phenotype. Our results suggest that Tyr1P has evolved specialized and essential functions in higher eukaryotes associated with antisense promoter and enhancer transcription, and Pol II stability. This study was performed in a human Raji cell line. It contains ChIP-seq data for H3K36me3 (two replicates), H3K4me1 (two replicates), H3K4me3 (two replicates), Pol II (three replicates), Ser2P (two replicates), Ser5P (two replicates), Ser7P (two replicates), Tyr1P 3D12 (two replicates) and Tyr1P 8G5 (one replicate). MNase-experiment for nucleosomes was performed in paired-end sequencing on one replicate, 4 replicates for the input genomic DNA was used and one replicate was generated for the short strand specific RNA experiment.
2014-06-01 | E-GEOD-52914 | biostudies-arrayexpress
Project description:Recent works have shown that RNA Polymerase (Pol) II can be recruited to, and transcribe, distal regulatory regions. Here, we analyzed transcription initiation and elongation through genome-wide localization of Pol II, General Transcription Factors (GTFs) and active chromatin in developing T-cells. We show that Pol II and GTFs are recruited to known, highly T-cell-specific enhancers. We extend this observation for the first time to many new putative enhancers, a majority of which are transcribed with or without polyadenylation. Importantly, we also identify genomic features characterized by large Pol II/GTFs recruitment and Transcriptional Initiation Platforms (TIPs) at promoters, intergenic and intragenic regions. TIPs are characterized by variable widths (0.4 to 10 kb), and correlate with high CpG content and tissue-specificity at promoters. Finally, we also report differential recruitment of TFIID and other GTFs at promoters and enhancers. Overall, we propose that TIPs represent important novel regulatory hallmarks of the genome. This study is based on the analysis of 20 samples in mouse double-positive thymocytes, 15 of which cover a range of general and specific transcription factors, chromatin modifying enzymes as well as histone modifications. 2 samples were dedicated for strand-specific RNA sequencing either from the total RNA population or after a selection of Poly(A) RNA. A Formaldehyde-Assisted Isolation of Regulatory Elements (FAIRE) sample has been sequenced in order to isolate open chromatin regions. Finally, input and IG control samples were designed and sequenced for data processing purposes. Biological and technical replicates were processed as described in the supplementary methods.
2011-05-20 | E-GEOD-29362 | biostudies-arrayexpress
Project description:Eukaryotic RNA polymerase II (Pol II) has evolved an array of heptad repeats with the consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7 at the carboxy-terminal domain (CTD) of the large subunit (Rpb1). Differential phosphorylation of Ser2, Ser5, and Ser7 in the 5M-bM-^@M-^Y and 3M-bM-^@M-^Y regions of genes coordinates the binding of transcription and RNA processing factors to the initiating and elongating polymerase complexes. Here, we report phosphorylation of Thr4 by Polo-like-kinase-3 in mammalian cells. ChIPseq analyses indicate an increase of Thr4-P levels in the 3M-bM-^@M-^Y region of genes occurring subsequently to an increase of Ser2-P levels. A Thr4/Ala mutant of Pol II displays a lethal phenotype. This mutant reveals a global defect in RNA elongation, while initiation is largely unaffected. Since Thr4 replacement mutants are viable in yeast we conclude that this amino acid has evolved an essential function(s) in the CTD of Pol II for gene transcription in mammalian cells. In this study, we investigated the function and ChIPseq genome-wide profiling of Thr4P residue (using the 6D7 antibody) of the Pol II CTD in Raji human B cells in comparison with either total Pol II profiling (N20 antibody, santa-cruz sc-899x), Ser5P CTD (3E8) or Ser2P (3E10) profiling in WT Raji cells. In another set of experiments, we also analysed total Pol II profiling (using an HA tag at the N-terminus of RPB1 and HA antibody Abcam ab9110) when endogenous enzyme is shut down by alpha-amanitin and replaced by either a recominant Pol II with 48 consensus repeats of the CTD (con48) or a mutated version where Thr4 residues were replaced by Ala (Thr4-Ala).In total 6 experimental sets (Pol IIt, Ser5P, Ser2P, Thr4P, con48, Thr4-Ala) were generated for our analysis and for each a biological replicate was performed. Biological replicates were merged when the data showed comparable signal noise ratio. Otherwise a unique replicate, showing the best noise ratio, was chosen for further analysis although the second replicate (for Ser2P and Thr4-Ala experiments). An input control (genomic DNA extracted after reverse crosslinking of the nuclear chip extracts) was performred and used for substraction to the ChIP experiments. One specific input material was used for wt cells, one for con48 and one for Thr4-Ala. Our data were processed to generate final wig files using our in house analysis pipeline essentially as described in Koch et al, (2011) NSMB 18 (8) p956.In brief, after alignment, sequence tags are: (i) artefact removed, (ii) elongated to an in silico optimized actual size of the initial fragments , (iii) input substracted, (iv) merged if applicable, (v) scaled for all experiments to correct for variation of tag number in between experiments. Several of the raw data files were no longer available.
2012-06-04 | E-GEOD-37519 | biostudies-arrayexpress
Project description:In higher eukaryotes, an important mechanism to tune translation in different tissues and conditions is mTORC1-dependent regulation of tRNAs transcription by RNA polymerase III (Pol III), as the relative amount of tRNAs tightly coordinates with the translational needs of the cell. mTORC1 contributes to regulate protein synthesis through its direct substrate MAF1, which functions as a negative regulator of Pol III transcription in response to stimuli such as serum starvation or rapamycin treatment. Here, we applied ChIP-seq to examine the Pol III occupancy profile in human fibroblasts and report evidence of a genome wide, MAF1-dependent coordinated response to favorable or stress growth conditions. Strikingly, while a set of genes is extremely responsive in terms of Pol III binding, other genes are mostly unperturbed, yet associated with transcriptionally engaged polymerases as revealed by nascent EU-labeled RNA-seq (neuRNA-seq). As shown by DamIP-seq, the responsiveness of a subset of genes is tightly connected to the rapid and transient interaction of MAF1 with DNA-bound Pol III. We performed duplicate ChIP-seq experiments for the Rpc4 (POLR3D) subunit of RNA polymerase III in IMR90hTert cells grown in the presence of fetal bovine serum (FBS), serum starved (SS), serum starved and treated with insulin (SS+I), serum starved and treated with insulin and rapamycin (SS+R+I). Additional ChIP-seq profiles were generated in cells treated with MAF1 siRNAs and serum starved. MAF1 binding was addressed by DamIP-seq, using two replicates per clone of IMR90hTert cells expressing HA-tagged MAF1-DamK9A (2 different clones) or EGFP-DamK9A (2 different clones). To monitor dynamic transcription profiles we did neusRNA-seq in IMR90hTert cells EU-labeled or mock (DMSO)-labeled. For both DamIP-seq and neusRNA-seq, cells were either unperturbed or serum starved.
2016-02-15 | E-GEOD-73929 | biostudies-arrayexpress
Project description:Ssk1-type response regulator proteins are the core elements of histidine-to-aspartate systems that mediate fungal stress tolerance, a determinant to the biocontrol potential of fungal entomopathogens. We characterized for the first time the functions of Beauveria bassiana Ssk1 (Bbssk1) by analyzing multi-phenotypic changes in DBbssk1 and differentially expressed genes (DEGs) in the digital gene expression (DGE) libraries of DBbssk1 and wild-type constructed under osmotic stress. The results revealed 1003 DEGs, of which many associated with conidiation, xenotics transport, cell wall integrity, and protein and carbohydrate metabolism were greatly down-regulated. Total RNA obtained from Bbssk1 disruption mutant subjected to 0.5 M NaCl for 30 minutes compared to the wild type strain under the same stress treatment.
2012-08-21 | E-GEOD-40229 | biostudies-arrayexpress
Project description:Transcription factor Msn2 played crucial roles in mediating fungal stress tolerance, a determinant to the biocontrol potential of fungal entomopathogens. We characterized for the first time the functions of Beauveria bassiana Msn2 (BbMsn2) and Metarhizium robertsii (MrMsn2) by analyzing multi-phenotypic changes in Msn2-deletion and investigating transcription patterns of WT versus M-NM-^TMsn2 of B. bassiana and M. robertsii under thermal and oxidative stresses by using high throughput sequencing (RNA-Seq). Our transcriptional profiles revealed that numerous differentially expressed genes (DEGs), of which involved in transportation, detoxification, signal transduction, and energy metabolism, were significantly repressed in expression level. Total RNA obtained from Bbmsn2 and MrMsn2 disruption mutant subjected to 2 mM menadione and 40 M-BM-0C for 3-h response compared to the wild type strain under the same stress treatment.
2015-02-01 | E-GEOD-40765 | biostudies-arrayexpress