Specialized cytonemes induce self-organization of stem cells

Spatial cellular organization is fundamental for embryogenesis. Remarkably, coculturing embryonic stem cells (ESCs) and trophoblast stem cells (TSCs) recapitulates this process, forming embryo-like structures. However, mechanisms driving ESC–TSC interaction remain elusive. We describe specialized ESC-generated cytonemes that react to TSC-secreted Wnts. Cytoneme formation and length are controlled by actin, intracellular calcium stores, and components of the Wnt pathway. ESC cytonemes select self-renewal–promoting Wnts via crosstalk between Wnt receptors, activation of ionotropic glutamate receptors (iGluRs), and localized calcium transients. This crosstalk orchestrates Wnt signaling, ESC polarization, ESC–TSC pairing, and consequently synthetic embryogenesis. Our results uncover ESC–TSC contact–mediated signaling, reminiscent of the glutamatergic neuronal synapse, inducing spatial self-organization and embryonic cell specification.

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Super Enhancer Profiles Identify Key Cell Identity Genes During Differentiation From Embryonic Stem Cells to Trophoblast Stem Cells Super Enhencers in Trophoblast Differentiation

Frontiers in Genetics ◽

10.3389/fgene.2021.762529 ◽

2021 ◽

Vol 12 ◽

Author(s):

Rongpu Jia ◽

Yu Gao ◽

Song Guo ◽

Si Li ◽

Liangji Zhou ◽

...

Keyword(s):

Stem Cells ◽

Embryo Implantation ◽

Control Cell ◽

Embryonic Stem ◽

Growth Factor Receptor ◽

Chromatin Accessibility ◽

Integrated Analysis ◽

Cell Identity ◽

Trophoblast Stem Cells ◽

Trophoblast Stem

Trophoblast stem cells (TSCs) are derived from blastocysts and the extra-embryonic ectoderm (ExE) of post-implantation embryos and play a significant role in fetal development, but the roles that TSCs play in the earlier status of fetal diseases need further exploration. Super enhancers (SEs) are dense clusters of stitched enhancers that control cell identity determination and disease development and may participate in TSC differentiation. We identified key cell identity genes regulated by TSC-SEs via integrated analysis of H3K27ac and H3K4me1 chromatin immunoprecipitation sequencing (ChIP-seq), RNA-sequencing (RNA-seq) and ATAC-sequencing (ATAC-seq) data. The identified key TSC identity genes regulated by SEs, such as epidermal growth factor receptor (EGFR), integrin β5 (ITGB5) and Paxillin (Pxn), were significantly upregulated during TSC differentiation, and the transcription network mediated by TSC-SEs enriched in terms like focal adhesion and actin cytoskeleton regulation related to differentiation of TSCs. Additionally, the increased chromatin accessibility of the key cell identity genes verified by ATAC-seq further demonstrated the regulatory effect of TSC-SEs on TSC lineage commitment. Our results illustrated the significant roles of the TSC-SE-regulated network in TSC differentiation, and identified key TSC identity genes EGFR, ITGB5 and Pxn, providing novel insight into TSC differentiation and lays the foundation for future studies on embryo implantation and related diseases.

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Epigenetic regulation of Nanog gene in embryonic stem and trophoblast stem cells

Genes to Cells ◽

10.1111/j.1365-2443.2007.01058.x ◽

2007 ◽

Vol 12 (3) ◽

pp. 387-396 ◽

Cited By ~ 130

Author(s):

Naoko Hattori ◽

Yuko Imao ◽

Koichiro Nishino ◽

Naka Hattori ◽

Jun Ohgane ◽

...

Keyword(s):

Stem Cells ◽

Epigenetic Regulation ◽

Embryonic Stem ◽

Trophoblast Stem Cells ◽

Trophoblast Stem ◽

Nanog Gene

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Human Naïve Epiblast Cells Possess Unrestricted Lineage Potential

10.1101/2020.02.04.933812 ◽

2020 ◽

Cited By ~ 3

Author(s):

Ge Guo ◽

Giuliano Giuseppe Stirparo ◽

Stanley Strawbridge ◽

Daniel Spindlow ◽

Jian Yang ◽

...

Keyword(s):

Stem Cells ◽

Human Embryo ◽

Embryonic Stem ◽

Trophoblast Stem Cells ◽

Assisted Reproduction Technology ◽

Trophoblast Stem ◽

Genetic Perturbations ◽

Cell Propagation ◽

Post Implantation ◽

Regulative Capacity

SUMMARYClassical mouse embryology has established a paradigm of early development driven by sequential lineage bifurcations. Accordingly, mouse embryonic stem cells derived from early epiblast have lost the potency to produce extraembryonic trophectoderm. We show in contrast that human naïve epiblast cells readily make trophectoderm. Inhibition of ERK signalling, instrumental in naïve stem cell propagation, unexpectedly potentiates trophectoderm formation, an effect enhanced by Nodal inhibition. Transcriptome analyses authenticate conversion into trophectoderm with subsequent production of syncitiotrophoblast, cytotrophoblast and trophoblast stem cells. Genetic perturbations indicate that NANOG suppresses and TFAP2C enables trophectoderm induction. Consistent with post-implantation progression, trophectoderm potential is extinguished in conventional human pluripotent stem cells, which instead make amnion. Finally, human embryo epiblasts from late blastocysts efficiently generate trophectoderm and differentiated trophoblast. Thus, pluripotent cells in the human embryo retain extraembryonic lineage plasticity and regenerative potential until implantation. Harnessing this unanticipated regulative capacity may be beneficial for assisted reproduction technology.

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Pluripotency-Independent Induction of Human Trophoblast Stem Cells from Fibroblasts

10.1101/2021.11.10.468044 ◽

2021 ◽

Author(s):

Yosef Buganim ◽

Moriyah Naama ◽

Ahmed Radwan ◽

Valery Zayat ◽

Shulamit Sebban ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem ◽

Gene Signature ◽

Chromatin Accessibility ◽

Double Knockout ◽

Human Trophoblast ◽

Trophoblast Stem Cells ◽

Pluripotent State ◽

Trophoblast Stem ◽

Specific Factors

Recent studies demonstrated that human trophoblast stem-like cells (hTS-like cells) can be derived from naive embryonic stem cells or be induced from somatic cells by the pluripotency factors, OSKM. This raises two main questions; (i) whether human induced TSCs (hiTSCs) can be generated independently to pluripotent state or factors and (ii) what are the mechanisms by which hTSC state is established during reprogramming. Here, we identify GATA3, OCT4, KLF4 and MYC (GOKM) as a pluripotency-independent combination of factors that can generate stable and functional hiTSCs, from both male and female fibroblasts. By using single and double knockout (KO) fibroblasts for major pluripotency genes (i.e. SOX2 or NANOG/PRDM14) we show that GOKM not only is capable of generating hiTSCs from the KO cells, but rather that the efficiency of the process is increased. Through H3K4me2 and chromatin accessibility profiling we demonstrate that GOKM target different loci and genes than OSKM, and that a significant fraction of them is related to placenta and trophoblast function. Moreover, we show that GOKM exert a greater pioneer activity compared to OSKM. While GOKM target many specific hTSC loci, OSKM mainly target hTSC loci that are shared with hESCs. Finally, we reveal a gene signature of trophoblast-related genes, consisting of 172 genes which are highly expressed in blastocyst-derived TSCs and GOKM-hiTSCs but absent or mildly expressed in OSKM-hiTSCs. Taken together, these results imply that not only is the pluripotent state, and SOX2 specifically, not required to produce functional hiTSCs, but that pluripotency-specific factors actually interfere with the acquisition of the hTSC state during reprogramming.

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Conserved mechanism of nucleoporin regulation of the Kcnq1ot1 imprinted domain with divergence in embryonic and trophoblast stem cells

10.1101/430694 ◽

2018 ◽

Author(s):

Saqib S. Sachani ◽

William A. MacDonald ◽

Ashley M. Foulkrod ◽

Carlee R. White ◽

Liyue Zhang ◽

...

Keyword(s):

Stem Cells ◽

Control Region ◽

Noncoding Rna ◽

Nuclear Periphery ◽

Embryonic Stem ◽

Trophoblast Stem Cells ◽

Parental Allele ◽

Trophoblast Stem ◽

Summary Statement ◽

Imprinting Control Region

AbstractGenomic imprinting is an epigenetic phenomenon, whereby dual chromatin states lead to expression of one, and silencing of the other parental allele. Recently, we identified a nucleoporin-mediated mechanism of Kcnq1ot1 imprinted domain regulation in extraembryonic endoderm stem cells by nucleoporins NUP107, NUP62 and NUP153. Here, we investigate their role in Kcnq1ot1 imprinted domain regulation in embryonic and trophoblast stem cells. Nucleoporin depletion in both lineages reduced Kcnq1ot1 noncoding RNA expression and volume, reduced Kcnq1ot1 paternal domain positioning at the nuclear periphery, and altered histone modifications along with histone modifier enrichment at the imprinting control region. However, while CTCF and cohesin were enriched at nucleoporin binding sites in the imprinting control region in embryonic stem cells, with reduction upon nucleoporin depletion, neither CTCF or cohesin occupied these sites in trophoblast stem cells. Finally, different subsets of silent paternal alleles were reactivated via altered histone modification upon nucleoporin depletion in embryonic and trophoblast stem cells. These results demonstrate a conserved mechanism with divergent regulation of the Kcnq1ot1 imprinted domain by NUP107, NUP62 and NUP153 in embryonic and extraembryonic lineages.Summary StatementInvestigation of nucleoporins, NUP107, NUP62, and NUP153, revealed a conserved nucleoporin-dependent mechanism that mediates Kcnq1ot1 imprinted domain regulation in ES and TS cells, although notable lineage-specific divergence was also observed.

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Esrrb plays important roles in maintaining self-renewal of trophoblast stem cells (TSCs) and reprogramming somatic cells to induced TSCs

Journal of Molecular Cell Biology ◽

10.1093/jmcb/mjy054 ◽

2018 ◽

Vol 11 (6) ◽

pp. 463-473 ◽

Cited By ~ 1

Author(s):

Haibo Gao ◽

Rui Gao ◽

Linfeng Zhang ◽

Wenchao Xiu ◽

Ruge Zang ◽

...

Keyword(s):

Stem Cells ◽

Molecular Mechanisms ◽

Target Genes ◽

Cell Model ◽

Embryonic Stem ◽

Placental Development ◽

Self Renewal ◽

Trophoblast Stem Cells ◽

Trophoblast Stem

Abstract Trophoblast stem cells (TSCs), which can be derived from the trophoectoderm of a blastocyst, have the ability to sustain self-renewal and differentiate into various placental trophoblast cell types. Meanwhile, essential insights into the molecular mechanisms controlling the placental development can be gained by using TSCs as the cell model. Esrrb is a transcription factor that has been shown to play pivotal roles in both embryonic stem cell (ESC) and TSC, but the precise mechanism whereby Esrrb regulates TSC-specific transcriptome during differentiation and reprogramming is still largely unknown. In the present study, we elucidate the function of Esrrb in self-renewal and differentiation of TSCs, as well as during the induced TSC (iTSC) reprogramming. We demonstrate that the precise level of Esrrb is critical for stem state maintenance and further trophoblast differentiation of TSCs, as ectopically expressed Esrrb can partially block the rapid differentiation of TSCs in the absence of fibroblast growth factor 4. However, Esrrb depletion results in downregulation of certain key TSC-specific transcription factors, consequently causing a rapid differentiation of TSCs and these Esrrb-deficient TSCs lose the ability of hemorrhagic lesion formation in vivo. This function of Esrrb is exerted by directly binding and activating a core set of TSC-specific target genes including Cdx2, Eomes, Sox2, Fgfr4, and Bmp4. Furthermore, we show that Esrrb overexpression can facilitate the MEF-to-iTSC conversion. Moreover, Esrrb can substitute for Eomes to generate GEsTM-iTSCs. Thus, our findings provide a better understanding of the molecular mechanism of Esrrb in maintaining TSC self-renewal and during iTSC reprogramming.

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Reprogramming of Extraembryonic Trophoblast Stem Cells into Embryonic Pluripotent State by Fusion with Embryonic Stem Cells

Stem Cells and Development ◽

10.1089/scd.2018.0034 ◽

2018 ◽

Vol 27 (19) ◽

pp. 1350-1359

Author(s):

Yean Ju Hong ◽

Kwonho Hong ◽

Seki Byun ◽

Hyun Woo Choi ◽

Jeong Tae Do

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Embryonic Stem ◽

Trophoblast Stem Cells ◽

Pluripotent State ◽

Trophoblast Stem

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Generation of Trophoblast Stem Cells from Rabbit Embryonic Stem Cells with BMP4

PLoS ONE ◽

10.1371/journal.pone.0017124 ◽

2011 ◽

Vol 6 (2) ◽

pp. e17124 ◽

Cited By ~ 15

Author(s):

Tao Tan ◽

Xianghui Tang ◽

Jing Zhang ◽

Yuyu Niu ◽

Hongwei Chen ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Embryonic Stem ◽

Trophoblast Stem Cells ◽

Trophoblast Stem

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Direct reprogramming of human embryonic to trophoblast stem cells

10.1101/2021.08.18.456785 ◽

2021 ◽

Author(s):

Norah M.E. Fogarty ◽

Ahmed Abdelbaki ◽

Afshan McCarthy ◽

Liani Devito ◽

Alice E. Chen ◽

...

Keyword(s):

Stem Cells ◽

Transcription Factors ◽

Embryonic Stem Cells ◽

Human Embryonic Stem Cells ◽

Embryonic Stem ◽

Transcriptional Analysis ◽

Differentiation Potential ◽

Trophoblast Stem Cells ◽

Cellular Models ◽

Trophoblast Stem

AbstractDuring the first week of development, human embryos form a blastocyst comprised of an inner cell mass and trophectoderm (TE) cells, the latter of which are progenitors of placental trophoblast. Here we investigated the expression of transcripts in the human TE from early to late blastocyst stages. We identified enrichment of transcription factors GATA2, GATA3, TFAP2C and KLF5 and characterised their protein expression dynamics across TE development. By inducible overexpression and mRNA transfection we determined that these factors, together with MYC, are sufficient to establish induced trophoblast stem cells (iTSCs) from human embryonic stem cells. These iTSCs self-renew and recapitulate morphological characteristics, gene expression profiles and directed differentiation potential similar to existing human TSCs. Systematic omission of each or combinations of factors, revealed the critical importance of GATA2 and GATA3 for successful iTSC reprogramming. Altogether, these findings provide insights into the transcription factor network that may be operational in the human TE and broaden the methods for establishing cellular models of early human placental progenitor cells, which may be useful in the future to model placental-associated diseases.Summary statementTranscriptional analysis of human blastocysts reveals transcription factors sufficient to derive induced trophoblast stem cells from primed human embryonic stem cells.

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Functional evaluation of transposable elements as transcriptional enhancers in mouse embryonic and trophoblast stem cells

10.1101/500322 ◽

2018 ◽

Author(s):

Christopher D. Todd ◽

Özgen Deniz ◽

Miguel R. Branco

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Gene Regulation ◽

Transposable Elements ◽

Regulation Of Gene Expression ◽

Embryonic Stem ◽

Specific Gene ◽

Functional Evaluation ◽

Trophoblast Stem Cells ◽

Trophoblast Stem

AbstractThe recurrent invasion and expansion of transposable elements (TEs) throughout evolution brought with it a vast array of coding and non-coding sequences that can serve as substrates for natural selection. Namely, TEs are thought to have contributed to the establishment of gene regulatory networks via their cis-acting elements. Both the embryonic and extraembryonic lineages of the early mouse embryo are thought to have benefited from the co-option of TEs as distal enhancer elements. However, there is little to no evidence that these particular TEs play significant roles in the regulation of gene expression. Here we tested for roles of TEs as enhancers in mouse embryonic and trophoblast stem cells by combining bioinformatic analyses with genetic and epigenetic editing experiments. Epigenomic and transcriptomic data from wildtype cells suggested that a large number of TEs played a role in the establishment of highly tissue-specific gene expression programmes. Through genetic editing of individual TEs we confirmed a subset of these regulatory relationships. However, a wider survey via CRISPR interference of RLTR13D6 elements in embryonic stem cells revealed that only a minority play significant roles in gene regulation. Our results suggest that a small proportion of TEs contribute to the mouse pluripotency regulatory network, and highlight the importance of functional experiments when evaluating the role of TEs in gene regulation.

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