Project description:We report the derivation of two distinct stem cells of the trophectoderm lineage from human pluripotent stem cells. The first is a CDX2- stem cell equivalent to primary hTSCs – they both exhibit identical expression of key markers, are maintained in culture and differentiate under similar conditions, and share high transcriptome similarity. The second is a CDX2+ putative human trophectoderm stem cell (hTESC) with distinct cell culture requirements and differences in gene expression and differentiation relative to hTSCs.
Project description:The first lineage choice made in human embryo development separates trophectoderm from the inner cell mass, which proceeds to form the pluripotent epiblast and primitive endoderm. Trophectoderm on the other hand gives rise to the placenta. Naïve pluripotent stem cells are derived from the pluripotent epiblast of the blastocyst and offer possibilities to explore how lineage integrity is maintained. Here, we discover that Polycomb repressive complex 2 (PRC2) restricts an intrinsic capacity of naïve pluripotent stem cells to give rise to trophectoderm. Through quantitative epigenome profiling, we find that broad histone H3 lysine 27 trimethylation (H3K27me3) hypermethylation is a common feature of naïve pluripotency across species. We define a previously unappreciated, naïve-specific set of bivalent promoters, featuring PRC2-mediated H3K27me3 concomitant with H3K4me3. Naïve bivalency maintains key trophectoderm transcription factors in a transcriptionally poised state that is resolved to an active state upon depletion of H3K27me3 via inhibition of the enzymatic subunits of PRC2, EZH1/2. Conversely, primed human embryonic stem cells cannot be driven towards trophectoderm development via PRC2 inhibition. While naïve and primed hESCs share the majority of bivalent promoters, PRC2 contributes to the repression of largely non-overlapping subsets of these promoters in each state, hence H3K27me3-mediated repression provides a highly adaptive mechanism to restrict lineage potential during early human development.
Project description:The first lineage choice made in human embryo development separates trophectoderm from the inner cell mass, which proceeds to form the pluripotent epiblast and primitive endoderm. Trophectoderm on the other hand gives rise to the placenta. Naïve pluripotent stem cells are derived from the pluripotent epiblast of the blastocyst and offer possibilities to explore how lineage integrity is maintained. Here, we discover that Polycomb repressive complex 2 (PRC2) restricts an intrinsic capacity of naïve pluripotent stem cells to give rise to trophectoderm. Through quantitative epigenome profiling, we find that broad histone H3 lysine 27 trimethylation (H3K27me3) hypermethylation is a common feature of naïve pluripotency across species. We define a previously unappreciated, naïve-specific set of bivalent promoters, featuring PRC2-mediated H3K27me3 concomitant with H3K4me3. Naïve bivalency maintains key trophectoderm transcription factors in a transcriptionally poised state that is resolved to an active state upon depletion of H3K27me3 via inhibition of the enzymatic subunits of PRC2, EZH1/2. Conversely, primed human embryonic stem cells cannot be driven towards trophectoderm development via PRC2 inhibition. While naïve and primed hESCs share the majority of bivalent promoters, PRC2 contributes to the repression of largely non-overlapping subsets of these promoters in each state, hence H3K27me3-mediated repression provides a highly adaptive mechanism to restrict lineage potential during early human development.
Project description:The trophoblast cell lineage is specified as early as the blastocyst stage, leading to the individualization of trophectoderm from pluripotent cells of the inner cell mass. We used a double in vitro transcription mRNA amplification technique and compared trophectoderm with pluripotent stem cells.
Project description:Single-cell RNA-seq reveals that Polycomb repressive complex 2 (PRC2) maintains naïve pluripotency and restricts an intrinsic capacity of pre-implantation pluripotent stem cells to give rise to trophectoderm and mesoderm lineages. Inhibition of PRC2 forces naïve hESC into an ‘activated’ state through which differentiation into either trophectoderm or mesoderm lineages is triggered. This trajectory is distinct from embryonic lineage specification out of the post-implantation pluripotent state, hence PRC2-mediated repression provides a highly adaptive mechanism to restrict lineage potential during early human development.
Project description:To elucidate the molecular basis of BMP4-induced differentiation of human pluripotent stem cells (PSCs) toward progeny with trophectoderm characteristics, we produced transcriptome, epigenome H3K4me3, H3K27me3, and CpG methylation maps of trophoblast progenitors, purified using the surface marker APA. We combined them with the temporally resolved transcriptome of the preprogenitor phase and of single APA+ cells. This revealed a circuit of bivalent TFAP2A, TFAP2C, GATA2, and GATA3 transcription factors, coined collectively the “trophectoderm four” (TEtra), which are also present in human trophectoderm in vivo. At the onset of differentiation, the TEtra factors occupy multiple sites in epigenetically inactive placental genes and in OCT4. Functional manipulation of GATA3 and TFAP2A indicated that they directly couple trophoblast-specific gene induction with suppression of pluripotency. In accordance, knocking down GATA3 in primate embryos resulted in a failure to form trophectoderm. The discovery of the TEtra circuit indicates how trophectoderm commitment is regulated in human embryogenesis.
Project description:The implantation process begins with attachment of the trophectoderm (TE) of the blastocyst to the maternal endometrial epithelium. Herein we have investigated the transcriptome of mural TE cells from 13 human blastocysts and compared these with those of human embryonic stem cell (hESC)-derived-TE (hESCtroph). The transcriptomes of hESFtroph at days 8, 10, and 12 had the greatest consistency with TE. Among genes coding for secreted proteins of the TE of human blastocysts and of hESCtroph are several molecules known to be involved in the implantation process as well as novel ones, such as CXCL12, HBEGF, inhibin A, DKK3, Wnt 5A, follistatin. The similarities between the two lineages underscore some of the known mechanisms and offer discovery of new mechanisms and players in the process of the very early stages of human implantation. We propose that the hESCtroph is a viable functional model of human trophoblasts to study trophoblast-endometrial interactions. Furthermore, the data derived herein offer the promise of novel diagnostics and therapeutics aimed at practical challenges in human infertility and pregnancy disorders associated with abnormal embryonic implantation. Transcriptome analyses suggest human embryonic stem cell-derived-trophoblast as a viable functional model of human trophoblast to study trophoblast-endometrial interactions. Five pools of trophectoderm cells were subjected to RNA isolation and microarray. The resulting data were compared to the transcriptomes of H7 human embryonic stem cells differentiated to the trophoblast lineage after 0, 2, 4, 6, 8, 10 and 12 days of induction with BMP-4. This submission represents the trophectoderm cells from blastocysts.