ChIP-seq of H3.3K4A/K36A mutant ESCs and neurons
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PROVIDER: PRJEB26774 | ENA |
REPOSITORIES: ENA
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Project description:Study to investigate the role of histone residues H3K4 and H3K36 for gene expression, histone localization and neuronal lineage specification by mutation of K4 and K36 in H3.3 to alanine. Histone variant H3.3 differs from the canonical H3.1/H3.2 by only 4 to 5 amino acids, which are necessary for nucleosome assembly independent of DNA replication, and is encoded by two gene copies. Complete loss of the two H3.3 genes (H3f3a and H3f3b) leads to embryonic lethality while single gene knockout yields viable mice. We used CRISPR-Cas9 to delete H3f3a and introduce homozygous point-mutations into H3f3b, thus ensuring that the entire pool of H3.3 protein carries the mutation of interest. We differentiated H3.3ctrl (H3f3a knock-out; H3f3b wild type), H3.3K4A mutant (H3f3a knock-out; H3f3b K4A) and H3.3K36A mutant (H3f3a knock-out; H3f3b K36A) ESCs into glutamatergic neurons. Genomic localization of H3.3 protein was determined by ChIP-Sequencing in ESCs (D0). Histone modifications patterns of H3K4me1, H3K4me3 and H3K27ac were measured by ChIP-Sequencing in ESCs (D0) to assess the impact of the H3.3K4A mutation on the epigenetic landscape. Levels of H3K36me3 were measured by ChIP-Sequencing in WT and H3.3K36A mutant ESCs (D0), NPCs (D8) and neurons (D12) to assess the impact of the H3.3K36A mutation on H3K36me3 levels in development.
2020-01-28 | E-MTAB-6822 | biostudies-arrayexpress
Project description:ChIP-seq of H3.3K4A/K36A mutant ESCs 2
| PRJEB35808 | ENA
Project description:RNA-seq of H3.3K4A/K36A mutant ESCs and neurons
| PRJEB27315 | ENA
Project description:Study to investigate the role of histone residues H3K4 and H3K36 for gene expression, histone localization and neuronal lineage specification by mutation of K4 and K36 in H3.3 to alanine. Histone variant H3.3 differs from the canonical H3.1/H3.2 by only 4 to 5 amino acids, which are necessary for nucleosome assembly independent of DNA replication, and is encoded by two gene copies. Complete loss of the two H3.3 genes (H3f3a and H3f3b) leads to embryonic lethality while single gene knockout yields viable mice. We used CRISPR-Cas9 to delete H3f3a and introduce homozygous point-mutations into H3f3b, thus ensuring that the entire pool of H3.3 protein carries the mutation of interest. We differentiated H3.3ctrl (H3f3a knock-out; H3f3b wild type), H3.3K4A mutant (H3f3a knock-out; H3f3b K4A) and H3.3K36A mutant (H3f3a knock-out; H3f3b K36A) ESCs into glutamatergic neurons. Gene expression profiles were measured by mRNA-Sequencing in undifferentiated ESCs (D0), neurodevelopment (D8) and differentiated neurons (D12) to assess the impact of the mutation on gene expression and development.
2020-01-28 | E-MTAB-6821 | biostudies-arrayexpress
Project description:Study to investigate the role of histone residues H3K4 and H3K36 for gene expression, histone localization and neuronal lineage specification by mutation of K4 and K36 in H3.3 to alanine. Histone variant H3.3 differs from the canonical H3.1/H3.2 by only 4 to 5 amino acids, which are necessary for nucleosome assembly independent of DNA replication, and is encoded by two gene copies. Complete loss of the two H3.3 genes (H3f3a and H3f3b) leads to embryonic lethality while single gene knockout yields viable mice. We used CRISPR-Cas9 to delete H3f3a and introduce homozygous point-mutations into H3f3b, thus ensuring that the entire pool of H3.3 protein carries the mutation of interest. We differentiated H3.3ctrl (H3f3a knock-out; H3f3b wild type), H3.3K4A mutant (H3f3a knock-out; H3f3b K4A) and H3.3K36A mutant (H3f3a knock-out; H3f3b K36A) ESCs into glutamatergic neurons. Genomic localization of H3.3 protein was determined by ChIP-Sequencing in ESCs (D0). Distribution patterns of RNA Polymerase II Phosphorylated on Serine 5 (RNA Pol II Ser5P), of histone modification H3K27me3 and chromatin remodeler components Brg1/Smarca4 (Swi/Snf) and Chd4 (NuRD) were measured by ChIP-Sequencing in ESCs (D0) to assess the impact of the H3.3K4A mutation on the epigenetic landscape. Distribution patterns of H3.3 were assessed by ChIP-Sequencing in HEK293T cells after depletion of Brg1/Smarca4 (Swi/Snf) and Chd4 (NuRD).
2020-01-28 | E-MTAB-8614 | biostudies-arrayexpress
Project description:Mouse WT129 ESCs and differentiated from them within 12 days glutamatergic neurons were used for the ChiP-seq experiment with an antibody against Sox2 protein. ES stands for mESCs, NP for neurons, inputs are provided for both cell types.
2020-05-21 | E-MTAB-8196 | biostudies-arrayexpress
Project description:Study to investigate the role of histone residues H3K4 and H3K36 for gene expression, histone localization and neuronal lineage specification by mutation of K4 and K36 in H3.3 to alanine. Histone variant H3.3 differs from the canonical H3.1/H3.2 by only 4 to 5 amino acids, which are necessary for nucleosome assembly independent of DNA replication, and is encoded by two gene copies. Complete loss of the two H3.3 genes (H3f3a and H3f3b) leads to embryonic lethality while single gene knockout yields viable mice. We used CRISPR-Cas9 to delete H3f3a and introduce homozygous point-mutations into H3f3b, thus ensuring that the entire pool of H3.3 protein carries the mutation of interest. We differentiated H3.3ctrl (H3f3a knock-out; H3f3b wild type), H3.3K4A mutant (H3f3a knock-out; H3f3b K4A) and H3.3K36A mutant (H3f3a knock-out; H3f3b K36A) ESCs into glutamatergic neurons. To assess the effect of the K4A mutation on Pol II activity, nascent RNA levels were mesaured by PRO-seq.
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Project description:We analyzed the genome-wide binding of Sox2 and POU factor partner factors, Oct4 in ESCs (using published datasets PMID:18692474 and GSM307137, GSM307154, GSM307155) and Brn2 in NPCs. We found that Sox2 and Oct4 co-occupied a large subset of promoters and enhancers in ESCs, but that Sox2 and Brn2 co-occupy predominantly enhancers. Further, we overexpressed Brn2 in differentiating ESCs and showed that ectopic Brn2 recruited Sox2 to NPC-specific targets, resulting in skewed differentiation towards the neural lineage. Examination of transcription factor binding in ESCs, NPCs, and differentiating ESCs by ChIP-Seq.
2013-02-20 | E-GEOD-35496 | biostudies-arrayexpress
Project description:Suz12(Bgal/Bgal) ESCs express a truncated form of Suz12 fused to Beta-galactosidase. These cells maintain a reduced level of H3K27me3 despite this mutation to a core component of PRC2, unlike Eed-/- ESCs whose H3K27me3 is ablated. Two ESC lines mutant in genes of core components of Polycomb Repressive Complex 2 were assessed for H3K27me3 by ChIP-seq, as compared to a wild type ESC line.
2013-07-22 | E-GEOD-47483 | biostudies-arrayexpress
Project description:We performed microarray analyses on human ESCs-derived NPs and neurons carrying loss-of-function mutation in the MeCP2 gene. Neural precursors and differentiating neurons at 2 and 4-week were used for RNA extraction and affymetrix microarrays. We added ERCC RNA spike-in controls to normalize to cell number input.
2013-10-01 | E-GEOD-50584 | biostudies-arrayexpress