Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Genome wide profiling of the RNAPII, Î³H2AX, H3K4Me1, H3K4M2 and H3K4Me3 in MLL2 F/F and FC/FC MEF cells

ABSTRACT: We investigated the RNAPII and Î³H2AX occupancy genome wide by ChIP-Seq in MLL2 F/F and FC/FC80 MEF cells. We found that a week after MLL2 excision (FC/FC cells), a group of genes present higher levels of Î³H2AX and RNAPII near the TSS, as compared to the control (F/F cells). H3K4Me1, H3K4M2 and H3K4Me3 levels near the TSS were also studied. There is a total of 52 samples. 3 independent replicates for each experiment were performed. H3, H2AX and IgG ChIPs were used for normalisation or as controls.The experiments were performed using immortalised mouse embryonic fibroblasts (MEF) in which both MLL2 alleles were targeted by the loxp system (F/F cells). Tamoxifen treatment of the F/F cells for 24 hours results in the excision of both MLL2 alleles (FC/FC cells).

ORGANISM(S): Mus musculus

SUBMITTER: Theo Kantidakis

PROVIDER: E-GEOD-73124 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Mutation of cancer driver MLL2 results in transcription stress and genome instability.

Kantidakis Theodoros T Kantidakis Theodoros T Saponaro Marco M Mitter Richard R Horswell Stuart S Kranz Andrea A Boeing Stefan S Aygün Ozan O Kelly Gavin P GP Matthews Nik N Stewart Aengus A Stewart A Francis AF Svejstrup Jesper Q JQ

Genes & development 20160201 4

Genome instability is a recurring feature of tumorigenesis. Mutation in MLL2, encoding a histone methyltransferase, is a driver in numerous different cancer types, but the mechanism is unclear. Here, we present evidence that MLL2 mutation results in genome instability. Mouse cells in which MLL2 gene deletion can be induced display elevated levels of sister chromatid exchange, gross chromosomal aberrations, 53BP1 foci, and micronuclei. Human MLL2 knockout cells are characterized by genome instabi ...[more]

PMID: 26883360

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Project description:We used human or mouse MLL2-impaired cells (including a human DSET MLL2 cell line,) as well as their parental controls, and measured mutations in histones and a few other genes that were affected in our gH2AX ChIP-Seq experiments. We found that cells lacking or having impaired MLL2 were presenting statistically signifficant, higher levels of mutations at the tested genes as compared to their parental controls. The experiments were performed using human and mouse cells. For the human cells, the parental HCT116 cells were used as control. Two different MLL2 KO clones (KO19 and KO34) and a DSET MLL2 cell line were employed. For the mouse cells, the F/F cells were used, in which both MLL2 alleles are targeted by loxp. Tamoxifen treatment for 24 hours results in the excision of both MLL2 alleles (FC/FC cells). F/F cells were treated with tamoxifen (FC/FC) or vehicle (DMSO, F/F) for 24 hours. Half of the cells were then harvested and used as controls (T0), while the other half were cultured for another 30 days (T30). Mutations after 30 days (T30) of tamoxifen/vehicle treatment were measured compared to the T0 controls. In all cases, two different pools of primers were employed. All histone genes available to target with high coverage were studied, in either human or mouse cells. For the human cells, 32 histone (HIST1H1A HIST1H1B HIST1H1C HIST1H1D HIST1H1E HIST1H1T HIST1H2AC HIST1H2AD HIST1H2AG HIST1H2AH HIST1H2AI HIST1H2AJ HIST1H2AK HIST1H2BF HIST1H2BG HIST1H2BH HIST1H2BI HIST1H2BJ HIST1H2BK HIST1H2BN HIST1H3B HIST1H3C HIST1H3D HIST1H3E HIST1H3F HIST1H3G HIST1H3H HIST1H3I HIST1H4B HIST1H4F HIST1H4J HIST2H2AC) and 6 other genes (BTG2 SGK1 MDM2 CDKN1A EGR1 RPLP0) were targeted. For the mouse cells, 21 histone (HIST1H1A HIST1H1B HIST1H1C HIST1H1D HIST1H1E HIST1H1T HIST1H2AK HIST1H2BQ HIST1H3B HIST1H3C HIST1H3F HIST1H4F HIST1H4J HIST1H4N HIST1H4N HIST2H2AA1 HIST2H2AA1 HIST2H2AA2 HIST2H2AA2 HIST2H2AC HIST2H2BB) and 6 other genes (BTG2 SGK1 MDM2 CDKN1A EGR1 RPLP0) were targeted. Processed data to be submitted to appropriate variation archive.

Project description:Heat shock response (HSR) is critical for survival of organisms undergoing proteotoxic stress. Heat shock factor 1 (HSF1) is widely believed to be the master regulator of this response. Here, we examined the kinetics of the transcriptional response and HSF1 binding changes genome-wide after heat shock (HS) with high sensitivity and high spatial and temporal resolution using PRO-seq and ChIP-seq assays respectively in wild type and in hsf1-deletion mouse embryonic fibroblasts. The transcriptional response is rapid, dynamic, and extensive with activation of several hundred genes and repression of several thousands of genes. Although HSF1 is critically required for classical inducible Heat Shock Protein (HSP) genes, it is not required for the majority of gene activation. HSF1 acts mechanistically to promote RNA polymerase II (Pol II) release from the promoter-proximal pause, while recruitment and initiation of Pol II are predominantly HSF1-independent. Surprisingly, a major class of cytoskeleton genes is immediately (within 2.5 minutes) and transiently activated in an HSF1-independent manner. A second wave of regulation is characterized by robust activation or repression of new sets of genes. The broad repression of thousands of genes, a quarter of which are repressed immediately upon HS and the rest are repressed in the second wave of regulation, is mediated at the level of decreasing Pol II pause release and not at prior steps of Pol II recruitment or initiation. Notably, heat shock does not induce transcription of some previously defined HSP genes, and HSF2 â a homolog of HSF1 â does not compensate for the lack of HSF1. Together, these findings indicate that mammalian cells cope with stress by rapidly inducing pervasive and dramatic changes in transcription that are largely modulated at the step of pause release, and only a fraction of this regulation is HSF1-dependent. Examination of transcriptional regulation before and after heat shock in WT, HSF1-/-, and HSF1&2-/- MEFs

Dataset Information

Genome wide profiling of the RNAPII, Î³H2AX, H3K4Me1, H3K4M2 and H3K4Me3 in MLL2 F/F and FC/FC MEF cells

Publications

Mutation of cancer driver MLL2 results in transcription stress and genome instability.

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