Elevated O-GlcNAc levels activate epigenetically repressed genes and delay mouse ES cell differentiation without affecting naive to primed cell transition

Christopher M. Speakman, Tanja C. E. Domke, Wikrom Wongpaiboonwattana, Kelly Sanders, Manikhandan Mudaliar, Daan M. F. van Aalten, Geoffrey J. Barton, Marios P. Stavridis (Lead / Corresponding author)

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    Abstract

    The differentiation of mouse embryonic stem (ES) cells is controlled by the interaction of multiple signaling pathways, typically mediated by post-translational protein modifications. The addition of O-linked N-acetylglucosamine (O-GlcNAc) to serine and threonine residues of nuclear and cytoplasmic proteins is one such modification (O-GlcNAcylation), whose function in ES cells is only now beginning to be elucidated. Here we demonstrate that the specific inhibition of O-GlcNAc hydrolase (Oga) causes increased levels of protein O-GlcNAcylation and impairs differentiation of mouse ES cells both in serum-free monolayer and in embryoid bodies (EBs). Use of reporter cell lines demonstrates that Oga inhibition leads to a reduction in the number of Sox1-expressing neural progenitors generated following induction of neural differentiation, as well as maintained expression of the ES cell marker Oct4 (Pou5f1). In EBs expression of mesodermal and endodermal markers is also delayed. However, the transition of naive cells to primed pluripotency indicated by Rex1 (Zfp42), Nanog, Esrrb and Dppa3 downregulation and Fgf5 upregulation remains unchanged. Finally, we demonstrate that increased O-GlcNAcylation results in upregulation of genes normally epigenetically silenced in ES cells, supporting the emerging role for this protein modification in the regulation of histone modifications and DNA methylation.
    Original languageEnglish
    Pages (from-to)2605-2615
    JournalStem Cells
    Volume32
    Issue number10
    Early online date6 Jun 2014
    DOIs
    Publication statusPublished - Oct 2014

    Fingerprint

    Embryonic Stem Cells
    Embryoid Bodies
    Cell Differentiation
    Hydrolases
    Up-Regulation
    Histone Code
    Genes
    Acetylglucosamine
    DNA Methylation
    Threonine
    Post Translational Protein Processing
    Nuclear Proteins
    Serine
    Proteins
    Down-Regulation
    Cell Line
    Serum
    Mouse Embryonic Stem Cells

    Keywords

    • Embryonic stem cells
    • Cell differentiation
    • O-GlcNAc
    • Post-translational modification
    • Signal transduction
    • OLIGONUCLEOTIDE MICROARRAYS

    Cite this

    Speakman, Christopher M. ; Domke, Tanja C. E. ; Wongpaiboonwattana, Wikrom ; Sanders, Kelly ; Mudaliar, Manikhandan ; van Aalten, Daan M. F. ; Barton, Geoffrey J. ; Stavridis, Marios P. . / Elevated O-GlcNAc levels activate epigenetically repressed genes and delay mouse ES cell differentiation without affecting naive to primed cell transition. In: Stem Cells. 2014 ; Vol. 32, No. 10. pp. 2605-2615.
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    title = "Elevated O-GlcNAc levels activate epigenetically repressed genes and delay mouse ES cell differentiation without affecting naive to primed cell transition",
    abstract = "The differentiation of mouse embryonic stem (ES) cells is controlled by the interaction of multiple signaling pathways, typically mediated by post-translational protein modifications. The addition of O-linked N-acetylglucosamine (O-GlcNAc) to serine and threonine residues of nuclear and cytoplasmic proteins is one such modification (O-GlcNAcylation), whose function in ES cells is only now beginning to be elucidated. Here we demonstrate that the specific inhibition of O-GlcNAc hydrolase (Oga) causes increased levels of protein O-GlcNAcylation and impairs differentiation of mouse ES cells both in serum-free monolayer and in embryoid bodies (EBs). Use of reporter cell lines demonstrates that Oga inhibition leads to a reduction in the number of Sox1-expressing neural progenitors generated following induction of neural differentiation, as well as maintained expression of the ES cell marker Oct4 (Pou5f1). In EBs expression of mesodermal and endodermal markers is also delayed. However, the transition of naive cells to primed pluripotency indicated by Rex1 (Zfp42), Nanog, Esrrb and Dppa3 downregulation and Fgf5 upregulation remains unchanged. Finally, we demonstrate that increased O-GlcNAcylation results in upregulation of genes normally epigenetically silenced in ES cells, supporting the emerging role for this protein modification in the regulation of histone modifications and DNA methylation.",
    keywords = "Embryonic stem cells, Cell differentiation, O-GlcNAc, Post-translational modification, Signal transduction, OLIGONUCLEOTIDE MICROARRAYS",
    author = "Speakman, {Christopher M.} and Domke, {Tanja C. E.} and Wikrom Wongpaiboonwattana and Kelly Sanders and Manikhandan Mudaliar and {van Aalten}, {Daan M. F.} and Barton, {Geoffrey J.} and Stavridis, {Marios P.}",
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    Elevated O-GlcNAc levels activate epigenetically repressed genes and delay mouse ES cell differentiation without affecting naive to primed cell transition. / Speakman, Christopher M.; Domke, Tanja C. E.; Wongpaiboonwattana, Wikrom; Sanders, Kelly; Mudaliar, Manikhandan; van Aalten, Daan M. F. ; Barton, Geoffrey J.; Stavridis, Marios P. (Lead / Corresponding author).

    In: Stem Cells, Vol. 32, No. 10, 10.2014, p. 2605-2615.

    Research output: Contribution to journalArticle

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    T1 - Elevated O-GlcNAc levels activate epigenetically repressed genes and delay mouse ES cell differentiation without affecting naive to primed cell transition

    AU - Speakman, Christopher M.

    AU - Domke, Tanja C. E.

    AU - Wongpaiboonwattana, Wikrom

    AU - Sanders, Kelly

    AU - Mudaliar, Manikhandan

    AU - van Aalten, Daan M. F.

    AU - Barton, Geoffrey J.

    AU - Stavridis, Marios P.

    PY - 2014/10

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    N2 - The differentiation of mouse embryonic stem (ES) cells is controlled by the interaction of multiple signaling pathways, typically mediated by post-translational protein modifications. The addition of O-linked N-acetylglucosamine (O-GlcNAc) to serine and threonine residues of nuclear and cytoplasmic proteins is one such modification (O-GlcNAcylation), whose function in ES cells is only now beginning to be elucidated. Here we demonstrate that the specific inhibition of O-GlcNAc hydrolase (Oga) causes increased levels of protein O-GlcNAcylation and impairs differentiation of mouse ES cells both in serum-free monolayer and in embryoid bodies (EBs). Use of reporter cell lines demonstrates that Oga inhibition leads to a reduction in the number of Sox1-expressing neural progenitors generated following induction of neural differentiation, as well as maintained expression of the ES cell marker Oct4 (Pou5f1). In EBs expression of mesodermal and endodermal markers is also delayed. However, the transition of naive cells to primed pluripotency indicated by Rex1 (Zfp42), Nanog, Esrrb and Dppa3 downregulation and Fgf5 upregulation remains unchanged. Finally, we demonstrate that increased O-GlcNAcylation results in upregulation of genes normally epigenetically silenced in ES cells, supporting the emerging role for this protein modification in the regulation of histone modifications and DNA methylation.

    AB - The differentiation of mouse embryonic stem (ES) cells is controlled by the interaction of multiple signaling pathways, typically mediated by post-translational protein modifications. The addition of O-linked N-acetylglucosamine (O-GlcNAc) to serine and threonine residues of nuclear and cytoplasmic proteins is one such modification (O-GlcNAcylation), whose function in ES cells is only now beginning to be elucidated. Here we demonstrate that the specific inhibition of O-GlcNAc hydrolase (Oga) causes increased levels of protein O-GlcNAcylation and impairs differentiation of mouse ES cells both in serum-free monolayer and in embryoid bodies (EBs). Use of reporter cell lines demonstrates that Oga inhibition leads to a reduction in the number of Sox1-expressing neural progenitors generated following induction of neural differentiation, as well as maintained expression of the ES cell marker Oct4 (Pou5f1). In EBs expression of mesodermal and endodermal markers is also delayed. However, the transition of naive cells to primed pluripotency indicated by Rex1 (Zfp42), Nanog, Esrrb and Dppa3 downregulation and Fgf5 upregulation remains unchanged. Finally, we demonstrate that increased O-GlcNAcylation results in upregulation of genes normally epigenetically silenced in ES cells, supporting the emerging role for this protein modification in the regulation of histone modifications and DNA methylation.

    KW - Embryonic stem cells

    KW - Cell differentiation

    KW - O-GlcNAc

    KW - Post-translational modification

    KW - Signal transduction

    KW - OLIGONUCLEOTIDE MICROARRAYS

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    SN - 1066-5099

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