Transcriptional state during endodermal differentiation
Ontology highlight
ABSTRACT: ChIP-seq was performed to determine the transcriptional activity of genes during endodermal differentiation RNA pol II occupancy and histone marks of elongation and gene repression were analyzed during early endodermal differentiation
Project description:Many long non-coding RNA (lncRNA) species have been identified in mammalian cells, but the genomic origin, regulation and function of these molecules in individual cell types is poorly understood. We have generated comprehensive catalogs of lncRNA species expressed in human and murine embryonic stem cells (ESCs) and mapped their genomic origin. A surprisingly large fraction of these transcripts (>60%) originate from divergent transcription at promoters of active protein-coding genes. The divergently transcribed lncRNA/mRNA gene pairs exhibit coordinated changes in transcription when ESCs are differentiated into endoderm. A significant number of the divergently transcribed lncRNAs/mRNA pairs are conserved between human and mouse ESCs. Disruption of promoter-associated lncRNA orthologs in a zebrafish model of early development causes gross developmental defects. Our results reveal that transcription of most lncRNA genes is coordinated with transcription of protein-coding genes and that these lncRNAs have roles in early development. Analysis of genome-wide lncRNA transcription in human embryonic stem cells and early differentiation using RNA-seq, GRO-seq and ChIP-seq
Project description:Cohesin interacts with Mediator to link the enhancers and core promoters of actively transcribed genes bound by RNA polymerase II. Activin signaling directs human embryonic stem cells to differentiate into endoderm. ChIP-seq was performed to determine the gene targets of these factors in human embryonic stem cells. Human ES cells were grown without feeders using mTESR1. ChIP-seq was performed against RNA Pol2, Cohesin subunits (Smc1 and Smc3) and IgG for cells grown in mTESR1. Cells were differentiated into endoderm after resting for 24 hours in RMPI-B27 (0hr), and then treating with 50 ng/ml Activin A for two hours (2hr) and 48 hours (48hr). DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing.
Project description:Unraveling complex signaling programs animating developmental lineage-decisions is pivotal to differentiate human pluripotent stem cells (hPSC) into pure populations of desired lineages for regenerative medicine. Developmental signals are strikingly temporally dynamic: BMP and Wnt initially specify primitive streak (progenitor to endoderm) yet 24 hours later suppress endoderm and induce mesoderm. At lineage bifurcations we show mutually-exclusive embryonic lineages are segregated through cross-repressive signals: TGFM-NM-2 and BMP/MAPK duel to respectively specify pancreas versus liver from endoderm. Unilateral endodermal differentiation requires blockade of alternative fates at every stage, revealing a universal developmental strategy for efficient differentiation and anterior-posterior patterning of diverse hPSC lines into highly-pure endodermal populations. This culminated in hPSC-derived hepatic progenitors that, for the first time, engraft long-term in genetically-unconditioned mouse livers and secrete human albumin. Finally, thirty transcriptional and chromatin state maps capturing endoderm commitment revealed endodermal enhancers reside in an unanticipated diversity of "pre-enhancer" chromatin states before activation. Endoderm RNA-seq and ChIP-seq data sets
Project description:Oct4, Sox2 and Nanog cooperate to form the transcriptional regulatory circuitry of embyronic stem cells. To gain insight into the genes regulated by the core circuitry of transcription factors, chromatin immunoprecipitation coupled with massive parallel sequencing (ChIP-seq) was performed to determine the genome-wide binding targets of Oct4. DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. ChIP was performed using an antibody against IgG as a background control. ChIP was performed using an antibody against Oct4 in two biological replicates. The first replicate was sequenced twice (technical replicate 1 and technical replicate 2).
Project description:To understand in detail the molecular phases that occur during the differentiation of human embryonic stem cells to hepatocyte-like cells we performed an RNA-seq time course of the stages of differentiation across the 21 day differentiation time course. We used a chemically defined (serum-free) protocol, modified from Si-Tayeb et al., 2010 and Song et al., 2009. We describe acquisition of definitive endoderm characteristics at day 3, followed by hepatoblast charcter at day 7-13 and then maturation to hepatocyte-like cells at day 21. Time course of differentiation of human embryoic stem cells to hepatocyte-like cells over 21 days, using an adapted chemically defined protocol of Song et al., 2009; Cell Research
Project description:Here we show that by simple modulation of extrinsic signaling pathways, a new class of pluripotent stem cells, referred to as region selective epiblast stem cells (rsEpiSCs), could be efficiently derived from different stages of the early embryo. rsEpiSCs share features of primed pluripotency yet are distinct from EpiSCs in their molecular characteristics and ability to colonize post-implantation embryos. We performed ChIP-seqencing experiments using antibodies against H3K4me3 and H3K27me3, comparing conventional EpiSCs and rsEpiSCs, as well as comparing conventional human H1 ESCs and human H1 ESCs cultured in mouse rsEpiSCs based culture conditions (H1 rsESCs). Examination of 2 different histone modifications (H3K4me3, H3K27me3) in 2 pluripotent stem cell types in mouse and human.
Project description:Here we show that by simple modulation of extrinsic signaling pathways, a new class of pluripotent stem cells, referred to as region selective epiblast stem cells (rsEpiSCs), could be efficiently derived from different stages of the early embryo. rsEpiSCs share features of primed pluripotency yet are distinct from EpiSCs in their molecular characteristics and ability to colonize post-implantation embryos. We performed RNA-sequencing experiments and examined the global gene expression profiles of EpiSCs, rsEpiSCs, in vivo isolated four regions of E6.5 mouse epiblasts: AP (anterior-proximal), AD (anterior-distal), PP (posterior-proximal) and PD (posterior-distal), human H1 ESCs, H1 rsESCs, H9 ESCs, H9 rsESCs, rhesus monkey ORMES23 rsESCs, and chimpanzee rsiPSCs. Examination of global gene expression profiles in 2 pluripotent stem cell types across multiple species.
Project description:In order to investigate the genome-wide binding profile of the forkhead transcription factor FOXK2 in human embryonic stem cells (ESCs) and downstream cell types, we generated the ChIP-seq data of FOXK2 in H1 ESC cells and two differentiated types of cells, mesendoderm cells and NPC cells.
Project description:The conversion of mouse embryonic fibroblasts (MEFs) to induced pluripotent stem cells (iPS) by forced expression of Oct4, Sox2 and Klf4 is among the earliest demonstrations of reprogramming to a pluripotent state by forced expression of transcription factors. To gain insights into the chromatin state of genes required for reprogramming, we profiled H3K4me3, H3K27me3 and H3K9me3. DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. ChIP was performed using an antibody against H3K4me3, H3K27me3 and H3K9me3.