title: Membrane translocation of t-SNARE protein syntaxin-4 abrogates ground-state pluripotency in mouse embryonic stem cells.
abstract: Embryonic stem (ES) and induced pluripotent stem (iPS) cells are attractive tools for regenerative medicine therapies. However, aberrant cell populations that display flattened morphology and lose ground-state pluripotency often appear spontaneously, unless glycogen synthase kinase 3β (GSK3β) and mitogen-activated protein kinase kinase (MEK1/2) are inactivated. Here, we show that membrane translocation of the t-SNARE protein syntaxin-4 possibly is involved in this phenomenon. We found that mouse ES cells cultured without GSK3β/MEK1/2 inhibitors (2i) spontaneously extrude syntaxin-4 at the cell surface and that artificial expression of cell surface syntaxin-4 induces appreciable morphological changes and mesodermal differentiation through dephosphorylation of Akt. Transcriptome analyses revealed several candidate elements responsible for this, specifically, an E-to P-cadherin switch and a marked downregulation of Zscan4 proteins, which are DNA-binding proteins essential for ES cell pluripotency. Embryonic carcinoma cell lines F9 and P19CL6, which maintain undifferentiated states independently of Zscan4 proteins, exhibited similar cellular behaviors upon stimulation with cell surface syntaxin-4. The functional ablation of E-cadherin and overexpression of P-cadherin reproduced syntaxin-4-induced cell morphology, demonstrating that the E- to P-cadherin switch executes morphological signals from cell surface syntaxin-4. Thus, spontaneous membrane translocation of syntaxin-4 emerged as a critical element for maintenance of the stem-cell niche.
authors.affiliation: Department of Biomedical Chemistry, Graduate School of Science and Technology, Kwansei Gakuin University, Sanda, Japan.
authors.forename: Natsumi
authors.initials: N
authors.lastname: Hagiwara-Chatani
authors.affiliation: Department of Biomedical Chemistry, Graduate School of Science and Technology, Kwansei Gakuin University, Sanda, Japan.
authors.forename: Kota
authors.initials: K
authors.lastname: Shirai
authors.affiliation: Department of Biomedical Chemistry, Graduate School of Science and Technology, Kwansei Gakuin University, Sanda, Japan.
authors.forename: Takumi
authors.initials: T
authors.lastname: Kido
authors.affiliation: Department of Biomedical Chemistry, Graduate School of Science and Technology, Kwansei Gakuin University, Sanda, Japan.
authors.forename: Tomoatsu
authors.initials: T
authors.lastname: Horigome
authors.affiliation: Department of Orthodontics and Dentofacial Orthopedics, Institute of Health Biosciences, The University of Tokushima, Tokushima, Japan.
authors.forename: Akihiro
authors.initials: A
authors.lastname: Yasue
authors.affiliation: Department of Biomedical Chemistry, Graduate School of Science and Technology, Kwansei Gakuin University, Sanda, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Adachi
authors.affiliation: Department of Biomedical Chemistry, Graduate School of Science and Technology, Kwansei Gakuin University, Sanda, Japan.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Hirai
mesh_terms: D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism; D048369:MAP Kinase Kinase 1 / Q000378:metabolism; D048370:MAP Kinase Kinase 2 / Q000378:metabolism; D051379:Mice; D047428:Protein Kinase Inhibitors / Q000032:analysis; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D050765:Qa-SNARE Proteins / Q000378:metabolism*; D055153:Stem Cell Niche; D059467:Transcriptome
chemical_list: D015820:Cadherins; D003470:Culture Media; D047428:Protein Kinase Inhibitors; D050765:Qa-SNARE Proteins; D000071679:Glycogen Synthase Kinase 3 beta; D051057:Proto-Oncogene Proteins c-akt; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; C482129:Map2k2 protein, mouse