ChIP-seq experiments to investigate the role of HOXB1 overexpression in cardiac progenitors
Ontology highlight
ABSTRACT: ChIP-seq experiments for HOXB1-FLAG in human embryonic stem cell line HOXB1_H11 on day 8 of cardiac differentiation. HOXB1-FLAG was overexpressed from day4 of cardiac differentiation using doxycycline. The accompanying input DNA samples were used as controls in downstream ChIP-seq analyses.
Project description:ChIP-seq experiments for MEIS1, GATA6, histone mark H3K27ac, KMT2D in human embryonic stem cells NKX2-5 eGFP/w (https://doi.org/10.1038/nmeth.1740) and its derivative MEIS1/MEIS2 KO on day 5 of cardiac differentiation.
Project description:Open chromatin profiling (ATAC-seq) of human embryonic stem cells NKX2-5eGFP/w (https://doi.org/10.1038/nmeth.1740) and its derivative MEIS1/MEIS2 KO on day 5 and day 12 of cardiac differentiation.
Project description:Identification of Zic3 cistrome at ESC, d1EpiLC and d2EpiLC. ChIPmentaion experiments were performed using ZIC3 (Abcam, ab222124) antibody in the Rex1GFPd2 mouse embryonic stem cell line (ESC; parental line; E14Tg2a) and d1- and d2-EpiLC differentiated states.
Project description:T cells were extracted from spleens of CBLA mice and activated in vitro with CD3/28 on day 0. They were spinoculated with retroviruses overexpressing a 3xFLAG-gene of interest on day 1. On day 4, BFP-positive (infected) cells were FACS sorted and measured by ChIPmentation. For input ChIP controls, see E-MTAB-6276
Project description:Scl/Tal1 confers hemogenic competence and prevents cardiomyogenesis in embryonic endothelium. Here we show that Scl both directly activates a broad gene regulatory network required for hematopoietic stem/progenitor cell (HS/PC) development, and represses transcriptional regulators required for cardiogenesis. Cardiac repression occurs during a short developmental window through Scl binding to distant cardiac enhancers that harbor H3K4me1 at this stage. Scl binding to hematopoietic regulators extends throughout HS/PC and erythroid development and spreads from distant enhancers to promoters. Surprisingly, Scl complex partners Gata 1 and 2 are dispensable for hematopoietic versus cardiac specification and Scl binding to the majority of its target genes. Nevertheless, Gata factors co-operate with Scl to activate selected transcription factors to facilitate HS/PC emergence from hemogenic endothelium. These results uncover a dual function for Scl in dictating hematopoietic versus cardiac fate choice and suggest a mechanism by which lineage-specific bHLH factors direct the divergence of competing fates. ChIP-seq with Scl, Hand1, Lsd1, Ezh2, H3K4me1 and H3K27ac in different cell types with mesodermal origin. Scl ChIP-seq in WT, SclKO, SclKO-iScl and Gata12KO mES cell derived day4 EB (embryoid body) Flk1+ mesodermal cells, SclKO-iScl ES cells and MEL cells; Hand1 ChIP-seq in WT mES cell derived day4 EB Flk1+ mesodermal cells; Lsd1 and Ezh2 ChIP-seq in WT and SclKO mES cell derived day4 EB Flk1+ mesodermal cells. ChIP-seq of histone modifications H3K4me1 and H3K27ac in WT, SclKO and Gata12KO mES cell derived day4 EB Flk1+ mesodermal cells, HPC7 hematopoietic progenitor cells and HL1 cardiomyogenic cells
Project description:We have developed a protocol to generate cardiopharyngeal mesoderm (CPM) in vitro by Mesp1 induction in ES cells. The goal of this study is to compare the transcriptome of CPM-derived cardiac and skeletal myogenic progenitors to identify novel lineage-specific markers. mRNA profiles of CPM-derived D6 (early) and D12 (late), cardiac (BMP) and skeletal myogenic (control) progenitors were generated
Project description:Genome wide mapping of RNA polymearase III binding sites in Saccharomyces cerevisiae under normal growth and nutrient starved condition using ChIP-seq. Chromatin Immuno-precipitation (ChIP) was performed for FLAG tagged version of pol III subunit RPC128 after crosslinking the log-phase cells with formaldehyde. MOCK and IP DNA was sequenced and coverage of pol III was calculated at each base of the genome. RPC128-FLAG ChIP-seq single end seqquencing on Illumina GAII. 2 replicates of IP samples and 1 MOCK sample. Done in under normal growth and nutrient deprivation (4 hours).
Project description:Rationale: Cardiogenesis is regulated by a complex interplay between transcription factors and chromatin-modifying enzymes. However, little is known about how these interactions regulate the transition from mesodermal precursors to cardiac progenitor cells (CPCs). Objective: To identify novel regulators of mesodermal cardiac lineage commitment. Methods and Results: We performed a bioinformatic-based transcription factor-binding site analysis on upstream promoter regions of genes that are enriched in ES cell-derived CPCs. From 32 candidate transcription factors screened, we found that YY1, a repressor of sarcomeric gene expression, is present in CPCs in vivo. Interestingly, we uncovered the ability of YY1 to transcriptionally activate Nkx2.5, a key marker of early cardiogenic commitment. YY1 regulates Nkx2.5 expression via a 2.1 kb cardiac-specific enhancer as demonstrated by in vitro luciferase-based assays and in vivo chromatin immunoprecipitation (ChIP) and genome-wide sequencing analysis. Furthermore, the ability of YY1 to activate Nkx2.5 expression depends on its cooperative interaction with GATA4 at a nearby chromatin. Cardiac mesoderm-specific loss-of-function of YY1 resulted in early embryonic lethality. This was corroborated in vitro by ES cell-based assays where we show that the over-expression of YY1 enhanced the cardiogenic differentiation ES cells into CPCs in a cell autonomous manner. Conclusion: These results demonstrate an essential and unexpected role for YY1 to promote cardiogenesis as a transcriptional activator of Nkx2.5 and other CPC-enriched genes. We report the identification of putative YY1 target genes in cardiac progenitor cells (CPCs). Two samples of independently FACS-purified eGFP+ CPCs were examined against the input.
Project description:We report the derivation of 2 different methods of generating cardiac myocytes from human ESCs. The traditional route is via cardiac progenitor cells and the second, new approach is through re-directing hemogenic endothelium into the cardiac lineage using inhibition of Wnt/b-catenin signaling Examination of 2 different cardiac populations using RNA-seq
Project description:In this study, time-course transcriptome profiling of caidiomyocyte differentiation derived from human hESCs and hiPSCs was investigated. Two hiPSC lines (C15 and C20) and two hESC lines (H1 and H9) were differentiated to caidiomyocytes. The cells were collected for RNA-seq analysis at day0(undifferentiated cells) day2 (mesoderm), day4 (cardiac mesoderm) and day30 (cardiomyocytes) using Illumina HiSeq 2000 sequencer.