Unknown,Transcriptomics,Genomics,Proteomics

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ChIP-seq to profile LPS-induced RNA polymerase II transcription in bone marrow-derived macrophages from WT and Hes1 KO mice

ABSTRACT: We characterized the RNA polymerase II occupancy on gene loci in WT and Hes1 KO BMDMs under untreated and LPS-stimulated conditions WT and Hes1 KO BMDMs were left untreated or stimulated with LPS for 1 hour. Pol II ChIP was performed and the DNA products were subject to ChIPseq

ORGANISM(S): Mus musculus

SUBMITTER: Yingli Shang

PROVIDER: E-GEOD-73568 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:BMDMs or iMACs were generated via differentiation of bone marrow cells or immortalized precursors, respectively, in culture media containing m-CSF for 7 day. Upon differentiation, BMDMs or iMacs were left untreated or stimulated as described below. 1) BMDM WT were stimulated with IFNa, LPS, PGE2, IL-4, IL-10, LPS+PGE2, LPS+IL-4, LPS+IL-10, IFNa+PGE2, IFNa+IL-10: each condition in duplicate. This experiment was performed to investigate transcriptional cross-antagonism in the concomitant presence of pro-inflammatory (LPS or IFNa) and anti-inflammatory stimuli (IL-4, IL-10, or PGE2). 2) BMDM WT were stimulated with LPS, in the presence or absence of PFI-1 (Brd2/4 inhibitor) or SGC-CBP30 (CBP/p300 inhibitor): each condition in triplicate. This experiment was performed to define pro-inflammatory genes dependent or not on chromatin remodeling for their induction. 3) BMDM Mef2C/D proficient (Vav-Cre) or BMDM Mef2C/D KO (obtained by crossing Mef2d-/- Mef2cfl/fl with Vav-Cre mice) were stimulated with LPS, PGE2, or LPS+PGE2: each condition in triplicate. This experiment was performed to investigate the role of MEF2C-D on LPS transcritptional response. 4) BMDM WT were pretreated for 40 minutes with an anti-IL10R antibody or the corresponding isotope control, and then left untreated or stimulated with LPS, PGE2 or LPS+PGE2: each condition in duplicate. This experiment was performed to investigate the crosstalk between IL-10 and PGE2. 5) BMDM WT were left untreated or stimulated with PGE2, LPS, LPS+PGE2, or LPS+PGE2+IFNb. In the latter condition, IFNb was added 1 hour after the beginning of the costimulation: each condition in triplicate. This experiment was performed to asses the impact of PGE2 into the LPS-mediated IFNb induction. 5) MEF2A-deficient iMac clones (D7, A7, A8, C7) and MEF2A-proficient (referred to as wild-type) iMac clones (NE, B3 and D10) were generated via CRISPR/Cas9 and stimulated or not with LPS: each condition in single. This experiment was performed to investigate the role of MEF2A on LPS transcritptional response.

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ChIP-seq to profile LPS-induced RNA polymerase II transcription in bone marrow-derived macrophages from WT and Hes1 KO mice

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