Single-cell RNA-sequencing of mouse embryonic fibroblasts, ESCs, and control and Hic2 overexpressing intermediate reprogramming cells
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ABSTRACT:
PROVIDER: PRJEB62854 | ENA |
REPOSITORIES: ENA
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Project description:We performed bulk mRNA sequencing of either control or Zfp266 KO mouse embryonic fibroblasts (MEFs), intermediate reprogramming populations, iPSCs and ESCs. We reveal enhanced and early upregulation of pluripotent and ESC associated genes in Zfp266 KO reprogramming populations.
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Project description:Mouse embryonic fibroblasts (MEFs) with doxycycline (Dox)-inducible reprogramming cassette MKOS-ires-mOrange and a Nanog-GFP reporter were transduced with lentiviral Dox-inducible Ty1-BFP (blue fluoresce protein with Ty1 tag in the N-terminus) or Hic2-Ty1 (Hic2 with Ty1 tag in the C-terminus) expression vector with MOI 5. One day later reprogramming was initiated by the administration of Dox. 48 hours later, the cells for Hic2 ChIP-seq with anti-Ty1 antibody were crosslinked with 2mM of Disuccinimidyl Glutarate (DSG) for 45 min at room temperature (RT) under constant agitation, before being cross-linked with 1% formaldehyde solution for 10 min at room temperature. For KLF4 ChIP-seq, cells were only cross-linked with 1% formaldehyde solution for 10 min at room temperature. ChIP-seq library was generated with the NEBNext Ultra-II DNA Library Prep kit. Each library was uniquely barcoded using NEBNext Multiplex Oligos for Illumina. Samples were sequenced with the NextSeq High 40PE.
2024-04-26 | E-MTAB-13170 | biostudies-arrayexpress
Project description:mRNA Sequencing of control and Zfp266 KO mouse embryonic fibroblasts, intermediate reprogramming cells, iPSCs and ESCs
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Project description:HIC2 is required for normal development of the heart. Loss of function mutants exhibit early embryonic lethality while heterozygous mutants have a partially penetrant ventricular septal defect. The aim of this study was to understand the function of HIC2 in early heart development by identifying potential transcriptional targets. We harvested whole hearts (which contain circulating erythrocytes) at embryonic day E9.5 and compared expression in a conditional loss of function mutant (Mesp1 Cre/+; Hic2 Fl/Fl) to controls (Hic2 Fl/Fl).
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Project description:The fetal hemoglobin (HBG) regulator BCL11A is repressed by HIC2 to regulate hemoglobin switching. HIC2 itself is under developmental control, highly expressed in fetal tissues and decreased in adult cells, but the mechanism of this developmental regulation is unclear. Here, we demonstrate that HIC2 is developmentally controlled by miRNAs, and loss of global miRNA biogenesis through DICER1 depletion leads to upregulation of HIC2 and HBG. We subsequently identify the adult-expressed let-7 miRNA family as a repressor of HIC2 expression. Ectopic expression of let-7 in fetal-type cells repressed HIC2 levels, while let-7 inhibition in adult erythroblasts upregulated HIC2 levels. Let-7 regulates BCL11A and HBG through induction of HIC2 and reduction of GATA1 binding at the BCL11A erythroid enhancer. Finally, HIC2 depletion rescues BCL11A-mediated repression of fetal hemoglobin in let-7 inhibited cells. Together these data demonstrate a key miRNA-mediated mechanism for developmental regulation of BCL11A expression and fetal hemoglobin repression in adult erythroid cells.
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Project description:The fetal hemoglobin (HBG) regulator BCL11A is repressed by HIC2 to regulate hemoglobin switching. HIC2 itself is under developmental control, highly expressed in fetal tissues and decreased in adult cells, but the mechanism of this developmental regulation is unclear. Here, we demonstrate that HIC2 is developmentally controlled by miRNAs, and loss of global miRNA biogenesis through DICER1 depletion leads to upregulation of HIC2 and HBG. We subsequently identify the adult-expressed let-7 miRNA family as a repressor of HIC2 expression. Ectopic expression of let-7 in fetal-type cells repressed HIC2 levels, while let-7 inhibition in adult erythroblasts upregulated HIC2 levels. Let-7 regulates BCL11A and HBG through induction of HIC2 and reduction of GATA1 binding at the BCL11A erythroid enhancer. Finally, HIC2 depletion rescues BCL11A-mediated repression of fetal hemoglobin in let-7 inhibited cells. Together these data demonstrate a key miRNA-mediated mechanism for developmental regulation of BCL11A expression and fetal hemoglobin repression in adult erythroid cells.
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Project description:The fetal hemoglobin (HBG) regulator BCL11A is repressed by HIC2 to regulate hemoglobin switching. HIC2 itself is under developmental control, highly expressed in fetal tissues and decreased in adult cells, but the mechanism of this developmental regulation is unclear. Here, we demonstrate that HIC2 is developmentally controlled by miRNAs, and loss of global miRNA biogenesis through DICER1 depletion leads to upregulation of HIC2 and HBG. We subsequently identify the adult-expressed let-7 miRNA family as a repressor of HIC2 expression. Ectopic expression of let-7 in fetal-type cells repressed HIC2 levels, while let-7 inhibition in adult erythroblasts upregulated HIC2 levels. Let-7 regulates BCL11A and HBG through induction of HIC2 and reduction of GATA1 binding at the BCL11A erythroid enhancer. Finally, HIC2 depletion rescues BCL11A-mediated repression of fetal hemoglobin in let-7 inhibited cells. Together these data demonstrate a key miRNA-mediated mechanism for developmental regulation of BCL11A expression and fetal hemoglobin repression in adult erythroid cells.
2024-02-05 | GSE216194 | GEO
Project description:The fetal hemoglobin (HBG) regulator BCL11A is repressed by HIC2 to regulate hemoglobin switching. HIC2 itself is under developmental control, highly expressed in fetal tissues and decreased in adult cells, but the mechanism of this developmental regulation is unclear. Here, we demonstrate that HIC2 is developmentally controlled by miRNAs, and loss of global miRNA biogenesis through DICER1 depletion leads to upregulation of HIC2 and HBG. We subsequently identify the adult-expressed let-7 miRNA family as a repressor of HIC2 expression. Ectopic expression of let-7 in fetal-type cells repressed HIC2 levels, while let-7 inhibition in adult erythroblasts upregulated HIC2 levels. Let-7 regulates BCL11A and HBG through induction of HIC2 and reduction of GATA1 binding at the BCL11A erythroid enhancer. Finally, HIC2 depletion rescues BCL11A-mediated repression of fetal hemoglobin in let-7 inhibited cells. Together these data demonstrate a key miRNA-mediated mechanism for developmental regulation of BCL11A expression and fetal hemoglobin repression in adult erythroid cells.
2024-02-05 | GSE216190 | GEO
Project description:The switch from fetal to adult hemoglobin production has been studied in great depth in part because of its relevance to the treatment of hemolobinopathies. Transcription factor BCL11A, which is essential for repression of the fetal beta-type globin (γ-globin) genes after birth, is largely controlled at the level of transcription but the mechanism of BCL11A developmental control is unknown. Here, using a CRISPR-Cas9 screen in human erythroblasts, we identify transcription factor HIC2 as a repressor of BCL11A transcription. HIC2 and BCL11A expression are anti-correlated in fetal and adult erythroblasts. Forced expression of HIC2 in adult erythroblasts silences BCL11A transcription and markedly induces γ-globin expression. HIC2 binds selectively to constituent erythroid developmental BCL11A enhancer to reduce chromatin accessibility and impair access by transcription factor GATA1, resulting in loss of enhancer activity and enhancer-promoter contacts. Conversely, loss of HIC2 in fetal erythroblasts increases enhancer accessibility, enables GATA1 binding and induces BCL11A transcription. HIC2 is unveiled as a critical evolutionarily conserved regulator of globin gene switching by imposing developmental control on the BCL11A gene.
2022-05-26 | GSE173584 | GEO
Project description:The switch from fetal to adult hemoglobin production has been studied in great depth in part because of its relevance to the treatment of hemolobinopathies. Transcription factor BCL11A, which is essential for repression of the fetal beta-type globin (γ-globin) genes after birth, is largely controlled at the level of transcription but the mechanism of BCL11A developmental control is unknown. Here, using a CRISPR-Cas9 screen in human erythroblasts, we identify transcription factor HIC2 as a repressor of BCL11A transcription. HIC2 and BCL11A expression are anti-correlated in fetal and adult erythroblasts. Forced expression of HIC2 in adult erythroblasts silences BCL11A transcription and markedly induces γ-globin expression. HIC2 binds selectively to constituent erythroid developmental BCL11A enhancer to reduce chromatin accessibility and impair access by transcription factor GATA1, resulting in loss of enhancer activity and enhancer-promoter contacts. Conversely, loss of HIC2 in fetal erythroblasts increases enhancer accessibility, enables GATA1 binding and induces BCL11A transcription. HIC2 is unveiled as a critical evolutionarily conserved regulator of globin gene switching by imposing developmental control on the BCL11A gene.
2022-05-26 | GSE173583 | GEO