A molecular roadmap for the emergence of early-embryonic-like cells in culture

Rodriguez-Terrones, Diego; Gaume, Xavier; Ishiuchi, Takashi; Weiss, Amélie; Kopp, Arnaud; Kruse, Kai; Penning, Audrey; Vaquerizas, Juan M.; Brino, Laurent; Torres-Padilla, Maria-Elena

doi:10.1038/s41588-017-0016-5

Cited by 159 publications

(242 citation statements)

References 74 publications

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“…Instead, to directly assess whether earlyembryonic-like cells exhibit an overall distinct pattern of metabolic activity than ESCs, we first measured oxygen consumption in 2-celllike cells. We observed no differences in OCR between these two cell types after rotenone addition, suggesting similar levels of extra-mitochondrial oxygen consumption rates (Fig EV2A and B) [16]. We optimized conditions for low cell numbers, which we set at 50,000 cells per well.…”

Section: Resultsmentioning

confidence: 94%

“…To discern between the above possibilities, we next determined glycolytic activity in 2-cell-like cells by measuring the extracellular acidification rate and the glucose uptake rate of ESCs, Zscan4 + and 2cell-like cells. Because Zscan4 + and 2-cell-like cells derive primarily from naïve ESCs [16], we addressed whether the observed differences in glucose uptake between ESCs and 2-cell-like cells are related to their preferential origin from the naïve ESC state. Under standard ESC culture medium conditions, Zscan4 + and 2cell-like cells exhibited a lower extracellular acidification rate than ESCs, suggesting lower glycolytic output ( Fig 2B).…”

Section: Resultsmentioning

confidence: 99%

“…The induction of 2-cell-like cells by acetate is in line with the known increase in levels of histone acetylation in 2-cell-like cells [11,15] and suggests that at least part of the effect observed upon acetate supplementation might be linked to increased levels of histone acetylation. We also addressed whether sodium acetate and L-lactate increase the 2-cell-like cell population by promoting either maintenance or induction using timelapse microscopy with a Zscan4 reporter [16] after removal of Zscan4 + and 2-cell-like cells. We also addressed whether sodium acetate and L-lactate increase the 2-cell-like cell population by promoting either maintenance or induction using timelapse microscopy with a Zscan4 reporter [16] after removal of Zscan4 + and 2-cell-like cells.…”

Section: Resultsmentioning

confidence: 99%

“…A 2C::turboGFP reporter cell line with constitutive tdTomato expression driven by the CAG promoter was used for the electron microscopy experiments and is described in further detail in [16]. This cell line incorporates a Zscan4 reporter construct (kindly provided by M. Ko) and is described elsewhere [16]. This cell line incorporates a Zscan4 reporter construct (kindly provided by M. Ko) and is described elsewhere [16].…”

Section: Reporter Cell Linesmentioning

confidence: 99%

“…All measurements on the Zscan4 + and 2-cell-like populations were performed on a triple reporter cell line carrying 2C::tbGFP, Zscan4c::mCherry and constitutive H2B-iRFP constructs. The Rex1 reporter cell line used for the glucose uptake measurements on naïve and primed pluripotent ES cells was kindly provided by A. Smith [4], and the generation of the Rex1 reporter line with the Zscan4 reporter has been described previously [16]. The Rex1 reporter cell line used for the glucose uptake measurements on naïve and primed pluripotent ES cells was kindly provided by A. Smith [4], and the generation of the Rex1 reporter line with the Zscan4 reporter has been described previously [16].…”

Section: Reporter Cell Linesmentioning

confidence: 99%

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A distinct metabolic state arises during the emergence of 2‐cell‐like cells

et al. 2019

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Pluripotent stem cells are thought of as a surrogate of early developmental stages that sustain the capacity to generate all cell types in the body, thereby constituting an invaluable tool to address the mechanisms underlying cellular plasticity. In the mouse, cells resembling totipotent 2-cell-stage embryos (2-cell-like cells) arise at a very low frequency in embryonic stem cell (ESC) cultures. However, the extent to which these early-embryonic-like cells recapitulate the molecular features of the early embryo is unclear. Here, we have undertaken a characterization of some of the metabolic features of early-embryonic-like cells in culture. Our data indicate that early-embryonic-like cells exhibit decreased glycolytic and respiratory activity, lower levels of reactive oxygen species and increased glucose uptake, suggesting a shift of the metabolic programme during 2-cell-like cell reprogramming. Accordingly, we find that 2-cell-like cells can be induced by defined metabolites. Thus, in addition to their transcriptional and chromatin features, 2-cell-like cells recapitulate some of the metabolic features of their in vivo counterpart. Altogether, our work underscores a distinct metabolic state of early-embryonic-like cells and identifies compounds that can induce their emergence in vitro.

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Section: Resultsmentioning

confidence: 94%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Reporter Cell Linesmentioning

confidence: 99%

Section: Reporter Cell Linesmentioning

confidence: 99%

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A distinct metabolic state arises during the emergence of 2‐cell‐like cells

et al. 2019

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Chromatin Modifier EP400 Regulates Oocyte Quality and Zygotic Genome Activation in Mice

Tian,

Yin,

Tian

et al. 2024

Advanced Science

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Epigenetic modifiers that accumulate in oocytes, play a crucial role in steering the developmental program of cleavage embryos and initiating life. However, the identification of key maternal epigenetic regulators remains elusive. In the findings, the essential role of maternal Ep400, a chaperone for H3.3, in oocyte quality and early embryo development in mice is highlighted. Depletion of Ep400 in oocytes resulted in a decline in oocyte quality and abnormalities in fertilization. Preimplantation embryos lacking maternal Ep400 exhibited reduced major zygotic genome activation (ZGA) and experienced developmental arrest at the 2‐to‐4‐cell stage. The study shows that EP400 forms protein complex with NFYA, occupies promoters of major ZGA genes, modulates H3.3 distribution between euchromatin and heterochromatin, promotes transcription elongation, activates the expression of genes regulating mitochondrial functions, and facilitates the expression of rate‐limiting enzymes of the TCA cycle. This intricate process driven by Ep400 ensures the proper execution of the developmental program, emphasizing its critical role in maternal‐to‐embryonic transition.

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A Single Cell Transcriptomic Fingerprint of Stressed Premature, Imbalanced Differentiation of Embryonic Stem Cells

L Ruden,

Singh,

Marben

et al. 2024

Birth Defects Research

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BackgroundMiscarriages cause a greater loss‐of‐life than cardiovascular diseases, but knowledge about environmentally induced miscarriages is limited. Cultured naïve pluripotent embryonic stem cells (ESC) differentiate into extra‐embryonic endoderm/extraembryonic endoderm (XEN) or formative pluripotent ESC, during the period emulating maximal miscarriage of peri‐implantation development. In previous reports using small marker sets, hyperosmotic sorbitol, or retinoic acid (RA) decreased naïve pluripotency and increased XEN by FACS quantitation.MethodsBulk and single cell (sc)RNAseq analyses of two cultured ESC lines was done, corroborated by qPCR. Transcriptomic responses were analyzed of cultured ESC stressed by Sorbitol, with Leukemia inhibitory factor (LIF + ; stemness growth factor), RA without LIF to control for XEN induction, and compared with normal differentiation (LIF − , ND).ResultsSorbitol and RA increase subpopulations of 2‐cell embryo‐like (2CEL) and XEN sub‐lineages; primitive, parietal, and visceral endoderm (VE) cells and suppress formative pluripotency, imbalancing alternate lineage choices of initial naïve pluripotent cultured ESC compared with ND. Although bulk RNAseq and gene ontology (GO) group analyses suggest that stress induces anterior VE‐head organizer and placental markers, scRNAseq reveals relatively few cells. But VE and placental markers/cells were in adjacent stressed cell clusters in the UMAP, like recent, normal UMAP of conceptuses. UMAPs show that dose‐dependent stress overrides stemness to force premature lineage imbalance.ConclusionsHyperosmotic stress, and other toxicological stresses, like drugs with active ingredient RA, may cause premature, lineage imbalance, resulting in miscarriages or birth defects.

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A molecular roadmap for the emergence of early-embryonic-like cells in culture

Cited by 159 publications

References 74 publications

A distinct metabolic state arises during the emergence of 2‐cell‐like cells

A distinct metabolic state arises during the emergence of 2‐cell‐like cells

Chromatin Modifier EP400 Regulates Oocyte Quality and Zygotic Genome Activation in Mice

A Single Cell Transcriptomic Fingerprint of Stressed Premature, Imbalanced Differentiation of Embryonic Stem Cells

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