Project description:We performed nascent RNA sequencing of inducible FOXD3 KO cells with and without synchronization to determine primary response genes.
Project description:We performed streptavidin pulldown sequencing of endogenously and exogenously Avi-tagged FOXD3 to determine genome-wide binding pattern of this transcription factor in asynchronous or cell cycle synchronized naive mouse ESCs.
Project description:Transcription factor/enhancer interactions determine cell specific gene expression. Here, we followed enhancers during differentiations of embryonic stem (ESCs) to epiblast like cells (EpiLCs). There were highly dynamic changes in histone lysine 27 acetylation at enhancer sites throughout the genome. These sites were enriched for a Foxd3 binding motif, a forkhead transcription factor essential in early embryonic development. Surprisingly, Foxd3 occupied largely mutually exclusive sites in the ESCs versus EpiLCs. Foxd3 bound to nucleosome occupied regions, simultaneously evicting the histones while inhibiting full gene expression through the recruitment of histone deacetylases. Knockout of Foxd3 resulted in hyperacetylation and transcriptional upregulation of neighboring genes, many of which were further upregulated at later stages of differentiation. These data show that Foxd3 primes enhancer sites during pregastrulation by removing nucleosomes, yet suppresses neighboring histone hyperacetylation. Such a mechanism may be common to many transcription factors that prepare enhancers for later gene activation during development. ChIP-seq of H3K4me1, H3K27ac, H3K27me3, p300, H3K4me3, RNA Pol2 and Oct4 in four pluripotent states: embryonic stem cells (ESCs) day 1 ESC differentiation, Epi-like stem cells (EpiLCs), and epiblast stem cells (EpiSCs); ChIP-seq of 3XFlag tagged Foxd3 in ESCs and EpiLCs; ChIP-seq of H3K4me1, H3K27ac, H3K27me3, p300 and H3K4me3 in Foxd3 conditional knockout cells (tamoxifen-inducible) -/+ 36h Tamoxifen treatemnt. ChIP seq of Flag-Foxd3 (third replicate), ChIP-seq of HDAC1 and Brg1 in WT and Foxd3 KO cells and MNase-ChIP-seq of H3K4me1
Project description:RNA-seq were performed with six samples, WT KH2 ESCs treated with or without PD0325901 for 48 hours (KH2_PD and KH2, respectively), iErk1; Erk KO ESCs cultured in the presence Dox (P0), 48 and 96 hours after Dox withdrawal (P1 and P2, respectively), and iErk1; Erk KO ESCs cultured without Dox for 96 hours, and treated with PD0325901 in the last 48 hours (P2_PD). iErk1; Erk KO ESCs cultured without Dox does not express Erk, thus mimicking Erk KO.
Project description:To gain more insights about the role of Hbxip in mESC, we performed RNA-Seq for WT and Hbxip-KO E14 ESCs in pluripotent condition and during differentiation. The goals of this study are to compare the transcriptome changes upon Hbxip knockout in pluripotent condition and during differentiation.
Project description:We performed CUT&Tag of inducible FOXD3 KO cells with or without cell cycle phase sorting to determine impact of FOXD3 loss of active chromatin marks.
Project description:We performed ATAC seq of inducible FOXD3 KO cells after cell cycle phase sorting to determine impact of FOXD3 loss on chromatin accessibility.
Project description:Transcription factor/enhancer interactions determine cell specific gene expression. Here, we followed enhancers during differentiations of embryonic stem (ESCs) to epiblast like cells (EpiLCs). There were highly dynamic changes in histone lysine 27 acetylation at enhancer sites throughout the genome. These sites were enriched for a Foxd3 binding motif, a forkhead transcription factor essential in early embryonic development. Surprisingly, Foxd3 occupied largely mutually exclusive sites in the ESCs versus EpiLCs. Foxd3 bound to nucleosome occupied regions, simultaneously evicting the histones while inhibiting full gene expression through the recruitment of histone deacetylases. Knockout of Foxd3 resulted in hyperacetylation and transcriptional upregulation of neighboring genes, many of which were further upregulated at later stages of differentiation. These data show that Foxd3 primes enhancer sites during pregastrulation by removing nucleosomes, yet suppresses neighboring histone hyperacetylation. Such a mechanism may be common to many transcription factors that prepare enhancers for later gene activation during development. Total RNA obtained Foxd3 knockout embryonic stem cells (ESCs) and epiblast-like cells (EpiLCs) (treated with 1uM tamoxifen for 36h to induce knockout) compared to wild-type controls