Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Principles of early human development and germ cell program from conserved model systems

Abstract

Human primordial germ cells (hPGCs), the precursors of sperm and eggs, originate during weeks 2–3 of early post-implantation development1. Using in vitro models of hPGC induction2,3,4, recent studies have suggested that there are marked mechanistic differences in the specification of human and mouse PGCs5. This may be due in part to the divergence in their pluripotency networks and early post-implantation development6,7,8. As early human embryos are not accessible for direct study, we considered alternatives including porcine embryos that, as in humans, develop as bilaminar embryonic discs. Here we show that porcine PGCs originate from the posterior pre-primitive-streak competent epiblast by sequential upregulation of SOX17 and BLIMP1 in response to WNT and BMP signalling. We use this model together with human and monkey in vitro models simulating peri-gastrulation development to show the conserved principles of epiblast development for competency for primordial germ cell fate. This process is followed by initiation of the epigenetic program9,10,11 and regulated by a balanced SOX17BLIMP1 gene dosage. Our combinatorial approach using human, porcine and monkey in vivo and in vitro models provides synthetic insights into early human development.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Specification of PGCs in gastrulating porcine embryos.
Figure 2: Competence for pPGC specification.
Figure 3: Simulation of human peri-gastrulation and hPGC competency.
Figure 4: Induction of hPGCs by SOX17–BLIMP1 and combined representation of hPGC and pPGC specification.

Accession codes

Primary accessions

Gene Expression Omnibus

References

  1. 1

    Tang, W. W., Kobayashi, T., Irie, N., Dietmann, S. & Surani, M. A. Specification and epigenetic programming of the human germ line. Nat. Rev. Genet. 17, 585–600 (2016)

    CAS  Article  Google Scholar 

  2. 2

    Sugawa, F. et al. Human primordial germ cell commitment in vitro associates with a unique PRDM14 expression profile. EMBO J. 34, 1009–1024 (2015)

    CAS  Article  Google Scholar 

  3. 3

    Sasaki, K. et al. Robust in vitro induction of human germ cell fate from pluripotent stem cells. Cell Stem Cell 17, 178–194 (2015)

    CAS  Article  Google Scholar 

  4. 4

    Irie, N. et al. SOX17 is a critical specifier of human primordial germ cell fate. Cell 160, 253–268 (2015)

    CAS  Article  Google Scholar 

  5. 5

    Hayashi, K., Ohta, H., Kurimoto, K., Aramaki, S. & Saitou, M. Reconstitution of the mouse germ cell specification pathway in culture by pluripotent stem cells. Cell 146, 519–532 (2011)

    CAS  Article  Google Scholar 

  6. 6

    Wu, J. & Izpisua Belmonte, J. C. Stem cells: a renaissance in human biology research. Cell 165, 1572–1585 (2016)

    CAS  Article  Google Scholar 

  7. 7

    Rossant, J. Mouse and human blastocyst-derived stem cells: vive les differences. Development 142, 9–12 (2015)

    CAS  Article  Google Scholar 

  8. 8

    Davidson, K. C., Mason, E. A. & Pera, M. F. The pluripotent state in mouse and human. Development 142, 3090–3099 (2015)

    CAS  Article  Google Scholar 

  9. 9

    Tang, W. W. et al. A unique gene regulatory network resets the human germline epigenome for development. Cell 161, 1453–1467 (2015)

    CAS  Article  Google Scholar 

  10. 10

    Guo, F. et al. The transcriptome and DNA methylome landscapes of human primordial germ cells. Cell 161, 1437–1452 (2015)

    CAS  Article  Google Scholar 

  11. 11

    Gkountela, S. et al. DNA demethylation dynamics in the human prenatal germline. Cell 161, 1425–1436 (2015)

    CAS  Article  Google Scholar 

  12. 12

    Klisch, K. et al. The Sda/GM2-glycan is a carbohydrate marker of porcine primordial germ cells and of a subpopulation of spermatogonia in cattle, pigs, horses and llama. Reproduction 142, 667–674 (2011)

    CAS  Article  Google Scholar 

  13. 13

    Seki, Y. et al. Cellular dynamics associated with the genome-wide epigenetic reprogramming in migrating primordial germ cells in mice. Development 134, 2627–2638 (2007)

    CAS  Article  Google Scholar 

  14. 14

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

    CAS  Article  Google Scholar 

  15. 15

    Valdez Magaña, G., Rodríguez, A., Zhang, H., Webb, R. & Alberio, R. Paracrine effects of embryo-derived FGF4 and BMP4 during pig trophoblast elongation. Dev. Biol. 387, 15–27 (2014)

    Article  Google Scholar 

  16. 16

    Yoshida, M. et al. Conserved and divergent expression patterns of markers of axial development in eutherian mammals. Dev. Dyn. 245, 67–86 (2016)

    CAS  Article  Google Scholar 

  17. 17

    Aramaki, S. et al. A mesodermal factor, T, specifies mouse germ cell fate by directly activating germline determinants. Dev. Cell 27, 516–529 (2013)

    CAS  Article  Google Scholar 

  18. 18

    Loh, K. M. et al. Efficient endoderm induction from human pluripotent stem cells by logically directing signals controlling lineage bifurcations. Cell Stem Cell 14, 237–252 (2014)

    CAS  Article  Google Scholar 

  19. 19

    D’Amour, K. A. et al. Efficient differentiation of human embryonic stem cells to definitive endoderm. Nat. Biotechnol. 23, 1534–1541 (2005)

    Article  Google Scholar 

  20. 20

    Murry, C. E. & Keller, G. Differentiation of embryonic stem cells to clinically relevant populations: lessons from embryonic development. Cell 132, 661–680 (2008)

    CAS  Article  Google Scholar 

  21. 21

    Nakamura, T. et al. A developmental coordinate of pluripotency among mice, monkeys and humans. Nature 537, 57–62 (2016)

    ADS  CAS  Article  Google Scholar 

  22. 22

    Lim, J. & Thiery, J. P. Epithelial–mesenchymal transitions: insights from development. Development 139, 3471–3486 (2012)

    CAS  Article  Google Scholar 

  23. 23

    Sasaki, K. et al. The germ cell fate of cynomolgus monkeys is specified in the nascent amnion. Dev. Cell 39, 169–185 (2016)

    CAS  Article  Google Scholar 

  24. 24

    Kuckenberg, P., Kubaczka, C. & Schorle, H. The role of transcription factor Tcfap2c/TFAP2C in trophectoderm development. Reprod. Biomed. Online 25, 12–20 (2012)

    CAS  Article  Google Scholar 

  25. 25

    Robertson, E. J. et al. Blimp1 regulates development of the posterior forelimb, caudal pharyngeal arches, heart and sensory vibrissae in mice. Development 134, 4335–4345 (2007)

    CAS  Article  Google Scholar 

  26. 26

    Carter, A. M. & Enders, A. C. Placentation in mammals: definitive placenta, yolk sac, and paraplacenta. Theriogenology 86, 278–287 (2016)

    CAS  Article  Google Scholar 

  27. 27

    Viotti, M., Nowotschin, S. & Hadjantonakis, A. K. SOX17 links gut endoderm morphogenesis and germ layer segregation. Nat. Cell Biol. 16, 1146–1156 (2014)

    CAS  Article  Google Scholar 

  28. 28

    Murakami, K. et al. NANOG alone induces germ cells in primed epiblast in vitro by activation of enhancers. Nature 529, 403–407 (2016)

    ADS  CAS  Article  Google Scholar 

  29. 29

    Yamaji, M. et al. Critical function of Prdm14 for the establishment of the germ cell lineage in mice. Nat. Genet. 40, 1016–1022 (2008)

    CAS  Article  Google Scholar 

  30. 30

    Lin, I. Y. et al. Suppression of the SOX2 neural effector gene by PRDM1 promotes human germ cell fate in embryonic stem cells. Stem Cell Reports 2, 189–204 (2014)

    CAS  Article  Google Scholar 

  31. 31

    Alberio, R., Croxall, N. & Allegrucci, C. Pig epiblast stem cells depend on activin/nodal signaling for pluripotency and self-renewal. Stem Cells Dev. 19, 1627–1636 (2010)

    CAS  Article  Google Scholar 

  32. 32

    Chen, G. et al. Chemically defined conditions for human iPSC derivation and culture. Nat. Methods 8, 424–429 (2011)

    CAS  Article  Google Scholar 

  33. 33

    Gafni, O. et al. Derivation of novel human ground state naive pluripotent stem cells. Nature 504, 282–286 (2013)

    ADS  CAS  Article  Google Scholar 

  34. 34

    Wang, H., Luo, X., Yao, L., Lehman, D. M. & Wang, P. Improvement of cell survival during human pluripotent stem cell definitive endoderm differentiation. Stem Cells Dev. 24, 2536–2546 (2015)

    CAS  Article  Google Scholar 

  35. 35

    Loh, K. M. et al. Mapping the pairwise choices leading from pluripotency to human bone, heart, and other mesoderm cell types. Cell 166, 451–467 (2016)

    CAS  Article  Google Scholar 

  36. 36

    Ng, E. S., Davis, R., Stanley, E. G. & Elefanty, A. G. A protocol describing the use of a recombinant protein-based, animal product-free medium (APEL) for human embryonic stem cell differentiation as spin embryoid bodies. Nat. Protocols 3, 768–776 (2008)

    CAS  Article  Google Scholar 

  37. 37

    Kobayashi, T ., Alberio, R . & Surani, M. A. Simulating gastrulation development and germ cell fate in vitro using human and monkey pluripotent stem cells. Protoc. Exch. (2017)

  38. 38

    Magnúsdóttir, E. et al. A tripartite transcription factor network regulates primordial germ cell specification in mice. Nat. Cell Biol. 15, 905–915 (2013)

    Article  Google Scholar 

  39. 39

    Grabole, N. et al. Prdm14 promotes germline fate and naive pluripotency by repressing FGF signalling and DNA methylation. EMBO Rep. 14, 629–637 (2013)

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank R. Campbell and C. Lee for help with animals and hPSCs, A. Riddell for FACS and T. Otani for advice on cmPSCs. T.K. was supported by JSPS, the Uehara and Kanae Foundations; H.Z. by CSC; D.A.C. by CONACYT. The work was funded by BBSRC grant to R.A., C.A. and M.A.S (BB/M001466/1). M.A.S is a Wellcome Investigator.

Author information

Affiliations

Authors

Contributions

T.K. designed the experiments and performed cell culture, plasmid construction, immunofluorescence, qPCR, RNA-seq, western blots, data analysis and wrote the paper. W.W.C.T. designed experiments and analysed RNA-seq. N.I. performed preliminary work and designed experiments, and S.D. performed bioinformatics. A.S. helped with a hPSC reporter. H.Z., S.W., D.K. and C.A. designed and performed immunofluorescence and culture of pig embryos and epiblasts. D.A.C. and R.W. designed and performed in situ hybridization experiments and immunofluorescence. W.W.C.T., N.I. and S.W. made equal contributions. R.A. supervised the project, designed experiments, performed dissections and wrote the paper. M.A.S. supervised the project, designed experiments, and wrote the paper. All authors contributed to the manuscript.

Corresponding authors

Correspondence to Ramiro Alberio or M. Azim Surani.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Additional information

Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data figures and tables

Extended Data Figure 1 Expression of key germ cell genes in early pPGCs.

a, Representation of mouse, pig and human embryos before gastrulation. b, Sections of early-PS and primitive-streak-stage embryo showing SOX17, BLIMP1, NANOG and OCT4. Yellow dashed insets show cells at high magnification and white dashed lines mark SOX17+ and/or BLIMP1 cells. Scale bar, 20 μm. c, Primitive-streak-stage embryo (~E12) with a cluster of pPGCs (arrow) with multiple combinations of PGC gene expression (SOX17, BLIMP1, NANOG, TFAP2C, OCT-4, Sda/GM2 and mesoderm gene, T). Arrowheads at the anterior streak point to primitive endoderm (SOX17+BLIMP1+ and NANOG cells. Scale bar, 20 μm. d, Late-PS (~E12.5–E13.5) embryo with a pPGC cluster (arrow) showing NANOG, SOX17 (split colour image of e), BLIMP1, and T expression. Arrowheads mark early migratory PGCs in the primitive endoderm. Scale bar, 25 μm. e, E14 embryo stained for SOX17, BLIMP1 and TFAP2C. Yellow dashed insets show cells at high magnification and white dashed lines mark SOX17+BLIMP1+ and TFAP2C cells. Scale bar, 20 μm. f, Immunostaining for PRDM14 co-stained with Sda/GM2 and SOX17 in E14 (pPGC cluster) embryos and E26 gonads. Arrows point to pPGCs in the gonad. Scale bar, 20 μm.

Extended Data Figure 2 Proliferation and development of early pPGCs.

a, OCT4 RNA in situ hybridization identifies the pPGC cluster (arrow) in the posterior end of E13.5–E15.5 embryos. Insets show whole embryos. b, Whole-mount OCT4 immunohistochemistry of a porcine embryo. Dashed square marks the area shown at higher magnification on the top right. Arrow points to the pPGC cluster. Bottom right: cross section of the embryo (line in the wholemount image) shows migratory pPGCs (red cells). c, Number of pPGCs at different stages as indicated. d, Immunostaining of EdU-labelled embryos at the indicated stages with different antibody combinations identifying the PGCs. The pPGC cluster is highlighted with dashed white line. Arrows show SOX17+EdU+ and SOX17+BLIMP1+EdU+ cells. Scale bar, 20 μm. e, Immunostained migratory pPGCs (arrows); inset show cells at higher magnification. f, Immunostained gonadal pPGCs. Inset shows SOX2+ neural tissue.

Source data

Extended Data Figure 3 Epigenetic reprogramming in pre- and early migratory pPGCs, and key germ cell markers in migratory pPGC and cultured porcine epiblast.

a, A cluster of pPGCs (dashed line) at E13 stained for TET1 and OCT4. Scale bar, 20 μm. b, Serial sections of E14 embryos immunostained for different epigenetic markers combined with BLIMP1, NANOG and SOX17. Dashed lines highlight pPGC clusters. Scale bar, 20 μm. c, Quantification of 5-methylcytosine and H3K9me2 in embryos of different stages. Numbers of cells analysed are indicated (boxes, mean and interquartile ranges; whiskers, maximum and minimum; *P < 0.01; Mann–Whitney U-test). d, Serial sections of E16 embryos showing migratory pPGCs (arrows) immunostained for the indicated epigenetic marks. Scale bar, 20 μm. e, Triple immunostaining of epiblasts cultured under different conditions. Scale bar, 10 μm.

Source data

Extended Data Figure 4 Characterization of NANOS3–tdTomato reporter hPSC.

a, Targeting strategy for making NANOS3–tdTomato (NT) reporter. b, Representative genotyping of targeted clones using genomic DNA. c, Conventional (conv) and PGC-competent (comp) hPSCs states are reversible; the latter is equivalent to pre-ME (12 h at mesendoderm induction). Conv-hPSCs are cultured in Essential 8 medium on vitronectin coated dishes (see Methods). Comp-hPSCs are cultured in the hPSC medium containing inhibitors (i) (GSK3i, ERKi, p38i, JNKi) on MEF (see Methods). d, NANOS3–tdTomato reporter conv-hPSCs and comp-hPSCs, and day 1–4 embryoids induced with or without cytokines. e, FACS pattern and percentage of NANOS3–tdTomato+AP+ cells shown in Extended Data Fig. 4d. f, FACS pattern and percentage of NANOS3–tdTomato+AP+ cells in multiple clones derived from Wis2 or H9 hESC lines. g, Immunostaining of day 4 embryoids induced from pre-ME (12 h) or comp-hPSCs by BMP containing cytokines. Scale bar, 50 μm. h, Comparison of sensitivity of two NANOS3 reporter cell lines. FACS patterns of day 4 embryoids induced from comp-hPSCs (harbouring NANOS3–mCherry reporter or NANOS3–tdTomato reporter) with or without cytokines. i, Comparison of hPGC induction efficiency derived from pre-ME (12 h) or comp-hPSCs. Representative images and FACS patterns are shown. j, Scatter plot (mean ± s.d.) shows percentage of NANOS3–tdTomato+AP+ cells in indicated condition (n = 6). Paired t-test was used to test for statistical significance (*P < 0.05).

Source data

Extended Data Figure 5 Characterization of pre-ME and mesendoderm induced from Conv-hPSC.

a, Immunostaining of conv-hPSCs during 12–24 h mesendoderm induction. Scale bar, 50 μm. b, Gene expression (RT–qPCR) change during mesendoderm induction. c, FACS patterns of day 4 embryoids induced from pre-ME (12 h) with cytokines. Pre-ME was induced with or without GSK3i or activin A. d, FACS patterns of day 4 embryoids induced from pre-ME (12 h) with cytokines. Pre-ME was induced with or without BMP2 or the inhibitor. e, Schematics of definitive endoderm or lateral mesoderm (LM) differentiation from mesendoderm. f, FACS patterns of day 2 definitive endoderm (percentage of CXCR4+) and day 1 lateral mesoderm (percentage of PDGFRa+) induced from 24 h of mesendoderm induction. g, Relative induction efficiency of definitive endoderm or lateral mesoderm from mesendoderm induced with or without BMP2 (definitive endoderm, n = 5; lateral mesoderm, n = 6). Paired t-test was used to test for statistical significance (*P < 0.05). h, Immunostaining of definitive endoderm and lateral mesoderm in Extended Data Fig. 5f. Scale bar, 50 μm. i, Schematic of spatial-temporal progression from conv-hPSCs to mesendoderm and the signalling.

Source data

Extended Data Figure 6 Robust induction of cmPGCs from cells during mesendoderm differentiation.

a, Schematic of in vitro differentiation of cmPSCs. The same system was adopted as shown for conv-hPSCs differentiation in Fig. 3. b, Bright field image of undifferentiated cmPSCs. c, Immunostaining of day 2 embryoids induced with cytokines from cells at 0 h (cmPSCs), 12 h and 24 h during mesendoderm differentiation. Dashed lines highlight SOX17+BLIMP1+TFAP2C+ cmPGCs. Scale bar, 50 μm. d, Immunostaining of day 2 pre-ME-derived embryoids (12 h) in Extended Data Fig. 6c for pluripotency markers. Notably, cmPGCs express SOX17, but not SOX2. By contrast, cmPSC colonies express SOX2 but not SOX17. e, Immunostaining of day 2 monkey definitive endoderm induced from pre-ME (12 h) and mesendoderm (24 h). Scale bar, 50 μm. f, Immunostaining of day 1 monkey lateral mesoderm induced from mesendoderm (24 h). Mesendoderm were induced with or without BMP. Notably, adding BMP during mesendoderm differentiation increased the efficiency for FOXF1+HAND1+ lateral mesoderm cells, as shown in conv-hPSCs (Extended Data Fig. 5e–i). Scale bar, 50 μm.

Extended Data Figure 7 Chronology of transcription factors expression during hPGC induction.

a, Schematic of hPGC induction from pre-ME (12 h). b, Images of day 2 and 4 embryoids in response to BMP2 alone or BMP2 with LIF, SCF and EGF (cytokines). Notably, BMP2 alone can induce hPGC at almost the same efficiency as the full cytokines, but do not survive during extended culture, as shown previously. c, Immunostaining of embryoids induced with BMP2 alone or BMP2 with LIF, SCF and EGF showing expression of SOX17, BLIMP1 and TFAP2C. Scale bar, 50 μm. d, Proportion of SOX17+ cells indicated in Extended Data Fig. 7c. e, Immunostaining of embryoids induced with BMP2 alone or BMP2 with LIF, SCF and EGF showing expression of SOX17, BLIMP1 and NANOG. Scale bar, 50 μm. f, Proportion of SOX17+ cells in Extended Data Fig. 7e.

Source data

Extended Data Figure 8 Effect of NANOG on hPGC induction, characterization of NANOS3–tdTomato reporter hPSCs containing inducible SOX17, BLIMP1 with or without TFAP2C, and similarity between cytokine- and SOX17–BLIMP1-induced hPGCs.

a, Represents overexpression of dex-inducible NANOG transgenes in NANOS3–tdTomato reporter comp-hPSC. b, Day 4 embryoids following induction of NANOG (by dex), with or without cytokines as indicated. c, FACS patterns after induction of hPGCs; NANOS3–tdTomato+AP+ cells (%) shown in Extended Data Fig. 8b. d, Represents overexpression of dex-inducible SOX17, dox-inducible BLIMP1, Shield1(S1)-inducible TFAP2C transgenes in NANOS3–tdTomato reporter comp-hPSCs. e, Immunostaining of NANOS3–tdTomato reporter comp-hPSCs + inducible SOX17/BLIMP1/TFAP2C (iSBT) 1 day after induction of SOX17, BLIMP1 and TFAP2C by addition of dex, dox or S1. Scale bar, 50 μm. f, Immunostaining of NANOS3–tdTomato reporter comp-hPSCs + iSB 1 day after induction of SOX17 or BLIMP1 by addition of dex or dox. Scale bar, 50 μm. g, Immunostaining of day 2 embyroid induced with or without dex to induce nuclear localization of SOX17. Notably, accumulation of SOX17 signal is observed in +dex condition. h, Changes in gene expression (RT-qPCR) during hPGC induction: comp-hPSCs control (AP+ cells); NANOS3–tdTomato+ hPGCs induced by SOX17–BLIMP1 or cytokines; NANOS3–tdTomato cells in cells exposed to cytokines (error bars, mean ± s.d.). i, Unsupervised hierarchical clustering (UHC) of gene expression. j, Gene set enrichment analysis (GSEA) of 123 hPGC-specific genes (Supplementary Table 1) on the transcriptome of cytokine- and SOX17–BLIMP1-induced hPGCs. k, Heat map showing expression of epigenetic modifiers related to global DNA demethylation. Same datasets as shown in Fig. 4e were used for analysis.

Source data

Extended Data Figure 9 Response of SOX17-knockout comp-hPSC to SOX17.

a, Overexpression of dex-inducible SOX17 (iS) in SOX17-knockout comp-hPSCs. b, Gene expression (RT–qPCR) on day 4 of FACS-sorted NANOS3–mCherry (NC)+alkaline phosphatase (AP)+ hPGCs by RT–qPCR. c, FACS analysis of day 2 embryoids induced from wild-type comp-hPSCs, SOX17-knockout comp-hPSCs and SOX17-knockout comp-hPSCs rescued with SOX17GR transgene (iS) (percentage of CXCR4+AP+ cells). d, FACS pattern of day 2 embyoid induced from NANOS3–tdTomato reporter comp-hPSCs showing AP+CXCR4+ cells expressing NANOS3–tdTomato. e, Represents SOX17-inducible system (iSdd). Expression of SOX17 fused with destabilized domain (DD) can be induced by doxycycline (dox); addition of Shield1 (S1) can stabilize SOX17–DD protein. f, Western blots showing SOX17 expression level in day 5 embryoids from SOX17-knockout + inducible SOX17–DD (iSdd) comp-hPSCs. Embryoids were induced with cytokines. To induce SOX17, different concentration of dox and S1 were added. As controls, NANOS3–mCherry+AP+ hPGCs and NANOS3–mCherryAP cells from wild-type comp-hPSC-derived embryoids induced with cytokines were used. Histone H3 (H3) was used for internal control. g, Immunostaining of day 4 embryoids from SOX17-knockout + iSdd comp-hPSCs. Embryoids from SOX17-knockout and wild-type comp-hPSC-induced with cytokines were used as controls. Scale bar, 50 μm. h, Quantification of immunostaining data in Extended Data Fig. 9g. The numbers of OCT4+BLIMP1+ hPGCs, FOXA2+ endodermal cells and OCT4+BLIMP1+ hPGCs expressing FOXA2 were counted from 3 different embryoids. The proportions of the 3 populations are shown. i, Expression of SOX17 and BLIMP1 (RT–qPCR) in day 4 embyoids in response to different SOX17 dosage.

Source data

Extended Data Figure 10 Response of SOX17-knockout comp-hPSCs to SOX17 and BLIMP1, and changes in epigenetic modifier expression after overexpression of SOX17 and BLIMP1.

a, Overexpression of dex-inducible SOX17 (iS) and dox-inducible BLIMP1 (iB) in SOX17-knockout comp-hPSCs. b, Immunostaining of comp-hPSC 1 day after induction of SOX17 (dex) or BLIMP1 (dox), or both, and day 2 embryoids following dex-induced SOX17 with endogenous or dox-induced BLIMP1. Scale bar, 50 μm. c, Gene expression (RT–qPCR) in day 4 embryoids following induction by dex (+SOX17), dox (+BLIMP1), or dex + dox (+SOX17 and BLIMP1). Bulk cells of embryoids induced from SOX17-knockout and wild-type comp-hPSCs with cytokines were used as controls. d, Upon specification, hPGCs become refractory to activin or Wnt signalling. Left schematic shows the experimental design. The embryoids were transferred to the medium with or without GSK3i (3 μM) or activin A (100 ng ml−1) at day 0, 1 or 2 to see the effect on PGC induction. FACS patterns (right) show the induction efficiency of hPGCs (percentage of NANOS3–tdTomato+AP+) at day 4. e, Day 4 embryoids induced by cytokines with or without activin A or GSK3i.

Source data

Supplementary information

Supplementary Information

This file contains the uncropped gels and Supplementary Tables 2-3. (PDF 5375 kb)

Supplementary Table 1

This file contains a list of hPGC specific genes used for GSEA analysis. (XLSX 36 kb)

PowerPoint slides

Source data

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Kobayashi, T., Zhang, H., Tang, W. et al. Principles of early human development and germ cell program from conserved model systems. Nature 546, 416–420 (2017). https://doi.org/10.1038/nature22812

Download citation

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing