title: NANOG alone induces germ cells in primed epiblast in vitro by activation of enhancers.
abstract: Nanog, a core pluripotency factor in the inner cell mass of blastocysts, is also expressed in unipotent primordial germ cells (PGCs) in mice, where its precise role is yet unclear. We investigated this in an in vitro model, in which naive pluripotent embryonic stem (ES) cells cultured in basic fibroblast growth factor (bFGF) and activin A develop as epiblast-like cells (EpiLCs) and gain competence for a PGC-like fate. Consequently, bone morphogenetic protein 4 (BMP4), or ectopic expression of key germline transcription factors Prdm1, Prdm14 and Tfap2c, directly induce PGC-like cells (PGCLCs) in EpiLCs, but not in ES cells. Here we report an unexpected discovery that Nanog alone can induce PGCLCs in EpiLCs, independently of BMP4. We propose that after the dissolution of the naive ES-cell pluripotency network during establishment of EpiLCs, the epigenome is reset for cell fate determination. Indeed, we found genome-wide changes in NANOG-binding patterns between ES cells and EpiLCs, indicating epigenetic resetting of regulatory elements. Accordingly, we show that NANOG can bind and activate enhancers of Prdm1 and Prdm14 in EpiLCs in vitro; BLIMP1 (encoded by Prdm1) then directly induces Tfap2c. Furthermore, while SOX2 and NANOG promote the pluripotent state in ES cells, they show contrasting roles in EpiLCs, as Sox2 specifically represses PGCLC induction by Nanog. This study demonstrates a broadly applicable mechanistic principle for how cells acquire competence for cell fate determination, resulting in the context-dependent roles of key transcription factors during development.
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.|Laboratory for Pluripotent Cell Studies, Center for Developmental Biology, RIKEN, 2-2-3 Minatojima-minamimachi, Chuo-ku, Kobe, Hyogo 650-0047, Japan.|Laboratory for Molecular and Cellular Biology, Faculty of Advanced Life Science, Hokkaido University, Kita21 Nishi11, Kita-ku, Sapporo, Hokkaido 001-0021, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Murakami
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: Ufuk
authors.initials: U
authors.lastname: Günesdogan
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: Jan J
authors.initials: JJ
authors.lastname: Zylicz
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: Walfred W C
authors.initials: WWC
authors.lastname: Tang
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: Roopsha
authors.initials: R
authors.lastname: Sengupta
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Kobayashi
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: Shinseog
authors.initials: S
authors.lastname: Kim
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.
authors.forename: Richard
authors.initials: R
authors.lastname: Butler
authors.affiliation: Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: Sabine
authors.initials: S
authors.lastname: Dietmann
authors.affiliation: Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY, UK.|Wellcome Trust Medical Research Council Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.
authors.forename: M Azim
authors.initials: MA
authors.lastname: Surani
mesh_terms: D028341:Activins / Q000494:pharmacology; D000818:Animals; D055415:Bone Morphogenetic Protein 4 / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D044127:Epigenesis, Genetic; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D018507:Gene Expression Regulation, Developmental; D016678:Genome / Q000235:genetics; D005854:Germ Cells / Q000166:cytology* / Q000378:metabolism*; D005855:Germ Layers / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D018398:Homeodomain Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008297:Male; D051379:Mice; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects; D000071317:Nanog Homeobox Protein; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects; D000074462:Positive Regulatory Domain I-Binding Factor 1; D011485:Protein Binding; D016601:RNA-Binding Proteins; D055748:SOXB1 Transcription Factors / Q000378:metabolism; D050656:Transcription Factor AP-2 / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
chemical_list: C525810:Bmp4 protein, mouse; D055415:Bone Morphogenetic Protein 4; D002843:Chromatin; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D000071317:Nanog Homeobox Protein; C475540:Nanog protein, mouse; C513258:Prdm1 protein, mouse; C531066:Prdm14 protein, mouse; D016601:RNA-Binding Proteins; D055748:SOXB1 Transcription Factors; C527783:Sox2 protein, mouse; C583890:Tfap2c protein, mouse; D050656:Transcription Factor AP-2; D014157:Transcription Factors; C422405:activin A; D016222:Fibroblast Growth Factor 2; D028341:Activins; D000074462:Positive Regulatory Domain I-Binding Factor 1