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24 h differentiation in N2B27 of wild-type and Med12-mutant mESCs [bulkRNA-seq]

ABSTRACT: Cell differentiation results from coordinated changes in gene transcription in response to combinations of signals. FGF, Wnt, and mTOR signals regulate the differentiation of pluripotent mammalian cells towards embryonic and extraembryonic lineages, but how these signals cooperate with general transcriptional regulators is not fully resolved. Here, we report a genome-wide CRISPR screen that reveals both signaling components and general transcriptional regulators for differentiation-associated gene expression in mESCs. Focusing on the Mediator subunit Med12 as one of the strongest hits in the screen, we show that it regulates gene expression in parallel to FGF and mTOR signals. Loss of Med12 is compatible with differentiation along both the embryonic epiblast and the extraembryonic primitive endoderm lineage, but pluripotency transitions are slowed down, and the transcriptional separation between epiblast and primitive endoderm identities is enhanced in Med12-mutant cells. These cellular phenotypes correlate with reduced biological noise upon loss of Med12. These findings suggest that Med12 regulates cellular plasticity through the priming of transcriptional changes during differentiation, thereby modulating the effects of a broad range of signals.

ORGANISM(S): Mus musculus

PROVIDER: GSE253607 | GEO | 2024/01/25

REPOSITORIES: GEO

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6 h FGF4 stimulation of Fgf4-mutant mESCs [bulkRNA-seq]

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2024-01-25 | GSE253606 | GEO

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RNA-seq of H9 cell line during differentiation from naïve extraembryonic endoderm to extraembryonic mesoderm.

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2012-06-04 | E-GEOD-38477 | biostudies-arrayexpress

Tissue-intrinsic beta-catenin signals antagonize Nodal-driven anterior visceral endoderm differentiation

Project description:The anterior-posterior axis of the mammalian embryo is laid down by the anterior visceral endoderm (AVE), an extraembryonic signaling center that is specified within the visceral endoderm. Current models posit that AVE differentiation is promoted globally by epiblast-derived Nodal signals, and spatially restricted by a BMP gradient established by the extraembryonic ectoderm. Here, we report spatially restricted AVE differentiation in bilayered embryo-like aggregates made from mouse embryonic stem cells that lack an extraembryonic ectoderm. Notably, clusters of AVE cells also form in pure visceral endoderm cultures upon activation of Nodal signaling, indicating that tissue-intrinsic factors can restrict AVE differentiation. We identify β-catenin activity as a tissue-intrinsic factor that antagonizes AVE-inducing Nodal signals. Together, our results show how an AVE-like population can arise through interactions between epiblast and visceral endoderm alone. This mechanism may be a flexible solution for axis patterning in a wide range of embryo geometries, and provide robustness to axis patterning when coupled with signal gradients.

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BMP4 signalling directs primitive endoderm-derived XEN cells to an extraembryonic visceral endoderm identity

Project description:The visceral endoderm (VE) is an epithelial tissue in the early postimplantation mouse embryo that encapsulates the pluripotent epiblast distally and the extraembryonic ectoderm proximally. In addition to facilitating nutrient exchange before the establishment of a circulation, the VE is critical for patterning the epiblast. Since VE is derived from the primitive endoderm (PrE) of the blastocyst, and PrE-derived eXtraembryonic ENdoderm (XEN) cells can be propagated in vitro, XEN cells should provide an important tool for identifying factors that direct VE differentiation. In this study, we demonstrated that BMP4 signalling induces the formation of a polarized epithelium in XEN cells. This morphological transition was reversible, and was associated with the acquisition of a molecular signature comparable to extraembryonic (ex) VE. Resembling exVE which will form the endoderm of the visceral yolk sac, BMP4-treated XEN cells regulated hematopoiesis by stimulating the expansion of primitive erythroid progenitors. We also observed that LIF exerted an antagonistic effect on BMP4-induced XEN cell differentiation, thereby impacting the extrinsic conditions used for the isolation and maintenance of XEN cells in an undifferentiated state. Taken together, our data suggest that XEN cells can be differentiated towards an exVE identity upon BMP4 stimulation, and therefore represent a valuable tool for investigating PrE lineage differentiation. Total RNA isolated in triplicate from XEN stem cell cultures that were untreated (samples 1-3) or treated with BMP4 growth factor (samples 4-6). Total RNA isolated in triplicate from XEN stem cells that were treated with BMP4 and were flow sorted as Afp::GFP-positive (samples 7-9) or Afp::GFP-negative (samples 10-12).

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BMP4 signalling directs primitive endoderm-derived XEN cells to an extraembryonic visceral endoderm identity

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FGF inhibition directs BMP4-mediated differentiation of Human Embryonic Stem Cells to syncytiotrophoblast

Project description:Bone morphogenetic protein (BMP) signaling is known to support differentiation of human embryonic stem cells (hESCs) into mesoderm and extraembryonic lineages, whereas other signaling pathways can largely influence this lineage specification. Here, we set out to reinvestigate the influence of ACTIVIN/NODAL and fibroblast growth factor (FGF) pathways on the lineage choices made by hESCs during BMP4-driven differentiation. We show that BMP activation, coupled with inhibition of both ACTIVIN/NODAL and FGF signaling, induces differentiation of hESCs, specifically to M-NM-2hCG hormone-secreting multinucleated syncytiotrophoblast and does not support induction of embryonic and extraembryonic lineages, extravillous trophoblast, and primitive endoderm. It has been previously reported that FGF2 can switch BMP4-induced hESC differentiation outcome to mesendoderm. Here, we show that FGF inhibition alone, or in combination with either ACTIVIN/NODAL inhibition or BMP activation, supports hESC differentiation to hCG-secreting syncytiotrophoblast. We show that the inhibition of the FGF pathway acts as a key in directing BMP4-mediated hESC differentiation to syncytiotrophoblast. Human embryonic Stem Cells (hESCs) were treated under defined conditions (N2B27) with BMP4 (B), SB431542 (SB) (ACTIVIN/NODAL inhibitor), SU5402 (SU) (FGFR1 inhibitor), FGF2 (F) either alone or in various combinations as mentioned, followed by isolation of total RNA.

2012-10-17 | E-GEOD-30125 | biostudies-arrayexpress

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