Nanog safeguards pluripotency and mediates germline development


Nanog is a divergent homeodomain protein found in mammalian pluripotent cells and developing germ cells1,2. Deletion of Nanog causes early embryonic lethality2, whereas constitutive expression enables autonomous self-renewal of embryonic stem cells1. Nanog is accordingly considered a core element of the pluripotent transcriptional network3,4,5,6,7. However, here we report that Nanog fluctuates in mouse embryonic stem cells. Transient downregulation of Nanog appears to predispose cells towards differentiation but does not mark commitment. By genetic deletion we show that, although they are prone to differentiate, embryonic stem cells can self-renew indefinitely in the permanent absence of Nanog. Expanded Nanog null cells colonize embryonic germ layers and exhibit multilineage differentiation both in fetal and adult chimaeras. Although they are also recruited to the germ line, primordial germ cells lacking Nanog fail to mature on reaching the genital ridge. This defect is rescued by repair of the mutant allele. Thus Nanog is dispensible for expression of somatic pluripotency but is specifically required for formation of germ cells. Nanog therefore acts primarily in construction of inner cell mass and germ cell states rather than in the housekeeping machinery of pluripotency. We surmise that Nanog stabilizes embryonic stem cells in culture by resisting or reversing alternative gene expression states.

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Figure 1: Nanog expression within the undifferentiated embryonic stem-cell population is reversible.
Figure 2: Nanog -/- embryonic stem cells retain expression of pluripotency markers and the capacity for in vitro self renewal.
Figure 3: Nanog -/- cells retain the potential for embryo colonization and somatic contribution to chimaeras.
Figure 4: Nanog is required for cell state transitions during germ cell development and for cell state reversions in embryonic stem-cell cultures.


  1. 1

    Chambers, I. et al. Functional expression cloning of Nanog, a pluripotency sustaining factor in embryonic stem cells. Cell 113, 643–655 (2003)

    CAS  Article  Google Scholar 

  2. 2

    Mitsui, K. et al. The homeoprotein Nanog is required for maintenance of pluripotency in mouse epiblast and ES cells. Cell 113, 631–642 (2003)

    CAS  Article  Google Scholar 

  3. 3

    Boyer, L. A. et al. Core transcriptional regulatory circuitry in human embryonic stem cells. Cell 122, 947–956 (2005)

    CAS  Article  Google Scholar 

  4. 4

    Loh, Y. H. et al. The Oct4 and Nanog transcription network regulates pluripotency in mouse embryonic stem cells. Nature Genet. 38, 431–440 (2006)

    CAS  Article  Google Scholar 

  5. 5

    Wang, J. et al. A protein interaction network for pluripotency of embryonic stem cells. Nature 444, 364–368 (2006)

    CAS  ADS  Article  Google Scholar 

  6. 6

    Chickarmane, V., Troein, C., Nuber, U. A., Sauro, H. M. & Peterson, C. Transcriptional dynamics of the embryonic stem cell switch. PLoS Comput. Biol. 2, e123 (2006)

    ADS  Article  Google Scholar 

  7. 7

    Ivanova, N. et al. Dissecting self-renewal in stem cells with RNA interference. Nature 442, 533–538 (2006)

    CAS  ADS  Article  Google Scholar 

  8. 8

    Ying, Q. L., Nichols, J., Chambers, I. & Smith, A. BMP induction of Id proteins suppresses differentiation and sustains embryonic stem cell self-renewal in collaboration with STAT3. Cell 115, 281–292 (2003)

    CAS  Article  Google Scholar 

  9. 9

    Hatano, S. Y. et al. Pluripotential competence of cells associated with Nanog activity. Mech. Dev. 122, 67–79 (2005)

    CAS  Article  Google Scholar 

  10. 10

    Vallier, L. et al. An efficient system for conditional gene expression in embryonic stem cells and in their in vitro and in vivo differentiated derivatives. Proc. Natl Acad. Sci. USA 98, 2467–2472 (2001)

    CAS  ADS  Article  Google Scholar 

  11. 11

    Robertson, M. et al. Nanog retrotransposed genes with functionally conserved open reading frames. Mamm. Genome 17, 732–743 (2006)

    CAS  Article  Google Scholar 

  12. 12

    Brons, I. G. et al. Derivation of pluripotent epiblast stem cells from mammalian embryos. Nature 448, 191–195 (2007)

    CAS  ADS  Article  Google Scholar 

  13. 13

    Tesar, P. J. et al. New cell lines from mouse epiblast share defining features with human embryonic stem cells. Nature 448, 196–199 (2007)

    CAS  ADS  Article  Google Scholar 

  14. 14

    Niwa, H. How is pluripotency determined and maintained? Development 134, 635–646 (2007)

    CAS  Article  Google Scholar 

  15. 15

    Chambers, I. & Smith, A. Self-renewal of teratocarcinoma and embryonic stem cells. Oncogene 23, 7150–7160 (2004)

    CAS  Article  Google Scholar 

  16. 16

    Lowell, S., Benchoua, A., Heavey, B. & Smith, A. G. Notch promotes neural lineage entry by pluripotent embryonic stem cells. PLoS Biol. 4, e121 (2006)

    Article  Google Scholar 

  17. 17

    Ying, Q. L., Stavridis, M., Griffiths, D., Li, M. & Smith, A. Conversion of embryonic stem cells into neuroectodermal precursors in adherent monoculture. Nature Biotechnol. 21, 183–186 (2003)

    CAS  Article  Google Scholar 

  18. 18

    Yamaguchi, S., Kimura, H., Tada, M., Nakatsuji, N. & Tada, T. Nanog expression in mouse germ cell development. Gene Expr. Patterns 5, 639–646 (2005)

    CAS  Article  Google Scholar 

  19. 19

    Scholer, H. R., Dressler, G. R., Balling, R., Rohdewohld, H. & Gruss, P. Oct-4: a germline-specific transcription factor mapping to the mouse t-complex. EMBO J. 9, 2185–2195 (1990)

    CAS  Article  Google Scholar 

  20. 20

    Toyooka, Y. et al. Expression and intracellular localization of mouse Vasa-homologue protein during germ cell development. Mech. Dev. 93, 139–149 (2000)

    CAS  Article  Google Scholar 

  21. 21

    Szabo, P. E., Hubner, K., Scholer, H. & Mann, J. R. Allele-specific expression of imprinted genes in mouse migratory primordial germ cells. Mech. Dev. 115, 157–160 (2002)

    CAS  Article  Google Scholar 

  22. 22

    Hajkova, P. et al. Epigenetic reprogramming in mouse primordial germ cells. Mech. Dev. 117, 15–23 (2002)

    CAS  Article  Google Scholar 

  23. 23

    Monk, M. & McLaren, A. X-chromosome activity in foetal germ cells of the mouse. J. Embryol. Exp. Morphol. 63, 75–84 (1981)

    CAS  Google Scholar 

  24. 24

    Shi, W. et al. Regulation of the pluripotency marker Rex-1 by Nanog and Sox2. J. Biol. Chem. 281, 23319–23325 (2006)

    CAS  Article  Google Scholar 

  25. 25

    Suzuki, A. et al. Maintenance of embryonic stem cell pluripotency by Nanog-mediated reversal of mesoderm specification. Nature Clin. Pract. Cardiovasc. Med. 3 (suppl. 1). S114–S122 (2006)

    CAS  Article  Google Scholar 

  26. 26

    Hart, A. H., Hartley, L., Ibrahim, M. & Robb, L. Identification, cloning and expression analysis of the pluripotency promoting Nanog genes in mouse and human. Dev. Dyn. 230, 187–198 (2004)

    CAS  Article  Google Scholar 

  27. 27

    Silva, J., Chambers, I., Pollard, S. & Smith, A. Nanog promotes transfer of pluripotency after cell fusion. Nature 441, 997–1001 (2006)

    CAS  ADS  Article  Google Scholar 

  28. 28

    Smith, A. G. Culture and differentiation of embryonic stem cells. J. Tissue Cult. Methods 13, 89–94 (1991)

    Article  Google Scholar 

  29. 29

    Ashfield, R. et al. MAZ-dependent termination between closely spaced human complement genes. EMBO J. 13, 5656–5667 (1994)

    CAS  Article  Google Scholar 

  30. 30

    Schwartzberg, P. L., Goff, S. P. & Robertson, E. J. Germ-line transmission of a c-abl mutation produced by targeted gene disruption in ES cells. Science 246, 799–803 (1989)

    CAS  ADS  Article  Google Scholar 

  31. 31

    Nagy, A., Gertsenstein, M., Vintersten, K. & Behringer, R. Manipulating the Mouse Embryo: A Laboratory Manual (Cold Spring Harbor Press, New York, 2003)

    Google Scholar 

  32. 32

    Chambers, I. Mechanisms and factors in embryonic stem cell self-renewal. Rend. Fis. Acc. Lincei s.9, v.16. 83–97 (2004)

  33. 33

    Lowell, S., Benchoua, A., Heavey, B. & Smith, A. G. Notch promotes neural lineage entry by pluripotent embryonic stem cells. PLoS Biol. 4, e121 (2006)

    Article  Google Scholar 

  34. 34

    Laemmli, U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685 (1970)

    CAS  ADS  Article  Google Scholar 

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We are grateful to V. Karwacki, A. Waterhouse, R. Wilkie, R. MacLay and J. Ure for technical assistance, to C. Manson, J. Verth and colleagues for animal husbandry, and to V. Wilson for comments on the manuscript. This research was supported by the Wellcome Trust, the Juvenile Diabetes Research Foundation, the Medical Research Council and the Biotechnological and Biological Sciences Research Council of the United Kingdom, and a Human Frontier Science Program Fellowship (to L.G.).

Author Contributions I.C. designed the experimental strategy and analysed the data; J.S., J.N. and A.S. contributed to the experimental design. J.S., J.N. and K.J. performed the chimaera study, and J.S. the confocal analyses. D.C. conducted gene targeting and cell biological analysis, and together with J.V. ran the FACS experiments. M.R. and B.N. performed molecular biological analyses. L.G. produced and characterized the ROSA26-Cre-ERT2 cells. I.C. and A.S. conceived the study and wrote the paper.

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Correspondence to Ian Chambers.

Supplementary information

Supplementary Information

The file contains Supplementary Tables 1-4 and Supplementary Figures 1-12 (PDF 4603 kb)

Supplementary Video 1

This file contains Supplementary Video 1 showing bright field of GFP- TNG cells from 24-44h post-plating (MOV 32601 kb)

Supplementary Video 2

This file contains Supplementary Video 2 showing fluorescent field of GFP TNG cells from 24-44h post-plating (MOV 42173 kb)

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Chambers, I., Silva, J., Colby, D. et al. Nanog safeguards pluripotency and mediates germline development. Nature 450, 1230–1234 (2007).

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