The germ-cell lineage ensures the continuity of life through the generation of male and female gametes, which unite to form a totipotent zygote. We have previously demonstrated that, by using cytokines, embryonic stem cells and induced pluripotent stem cells can be induced into epiblast-like cells (EpiLCs) and then into primordial germ cell (PGC)-like cells with the capacity for both spermatogenesis and oogenesis1,2, creating an opportunity for understanding and regulating mammalian germ-cell development in both sexes in vitro. Here we show that, without cytokines, simultaneous overexpression of three transcription factors, Blimp1 (also known as Prdm1), Prdm14 and Tfap2c (also known as AP2γ), directs EpiLCs, but not embryonic stem cells, swiftly and efficiently into a PGC state. Notably, Prdm14 alone, but not Blimp1 or Tfap2c, suffices for the induction of the PGC state in EpiLCs. The transcription-factor-induced PGC state, irrespective of the transcription factors used, reconstitutes key transcriptome and epigenetic reprogramming in PGCs, but bypasses a mesodermal program that accompanies PGC or PGC-like-cell specification by cytokines including bone morphogenetic protein 4. Notably, the transcription-factor-induced PGC-like cells contribute to spermatogenesis and fertile offspring. Our findings provide a new insight into the transcriptional logic for PGC specification, and create a foundation for the transcription-factor-based reconstitution and regulation of mammalian gametogenesis.
Your institute does not have access to this article
Open Access articles citing this article.
Nature Communications Open Access 19 May 2022
Stem Cell Research & Therapy Open Access 21 February 2022
Nature Communications Open Access 25 November 2021
Subscribe to Journal
Get full journal access for 1 year
only $3.90 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Get time limited or full article access on ReadCube.
All prices are NET prices.
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)
Hayashi, K. et al. Offspring from oocytes derived from in vitro primordial germ cell-like cells in mice. Science 338, 971–975 (2012)
Saitou, M. & Yamaji, M. Primordial germ cells in mice. Cold Spring Harb. Perspect. Biol. 4, 223–241 (2012)
Vincent, S. D. et al. The zinc finger transcriptional repressor Blimp1/Prdm1 is dispensable for early axis formation but is required for specification of primordial germ cells in the mouse. Development 132, 1315–1325 (2005)
Ohinata, Y. et al. Blimp1 is a critical determinant of the germ cell lineage in mice. Nature 436, 207–213 (2005)
Yamaji, M. et al. Critical function of Prdm14 for the establishment of the germ cell lineage in mice. Nature Genet. 40, 1016–1022 (2008)
Weber, S. et al. Critical function of AP-2γ/TCFAP2C in mouse embryonic germ cell maintenance. Biol. Reprod. 82, 214–223 (2010)
Saitou, M., Barton, S. C. & Surani, M. A. A molecular programme for the specification of germ cell fate in mice. Nature 418, 293–300 (2002)
Sato, M. et al. Identification of PGC7, a new gene expressed specifically in preimplantation embryos and germ cells. Mech. Dev. 113, 91–94 (2002)
Ohinata, Y., Sano, M., Shigeta, M., Yamanaka, K. & Saitou, M. A comprehensive, non-invasive visualization of primordial germ cell development in mice by the Prdm1-mVenus and Dppa3-ECFP double transgenic reporter. Reproduction 136, 503–514 (2008)
Hochedlinger, K., Yamada, Y., Beard, C. & Jaenisch, R. Ectopic expression of Oct-4 blocks progenitor-cell differentiation and causes dysplasia in epithelial tissues. Cell 121, 465–477 (2005)
Yabuta, Y., Kurimoto, K., Ohinata, Y., Seki, Y. & Saitou, M. Gene expression dynamics during germline specification in mice identified by quantitative single-cell gene expression profiling. Biol. Reprod. 75, 705–716 (2006)
Kurimoto, K. et al. Complex genome-wide transcription dynamics orchestrated by Blimp1 for the specification of the germ cell lineage in mice. Genes Dev. 22, 1617–1635 (2008)
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)
Subramanian, A. et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc. Natl Acad. Sci. USA 102, 15545–15550 (2005)
Yamaji, M. et al. PRDM14 ensures naive pluripotency through dual regulation of signaling and epigenetic pathways in mouse embryonic stem cells. Cell Stem Cell 12, 368–382 (2013)
Chuma, S. et al. Spermatogenesis from epiblast and primordial germ cells following transplantation into postnatal mouse testis. Development 132, 117–122 (2005)
Kimura, Y. & Yanagimachi, R. Intracytoplasmic sperm injection in the mouse. Biol. Reprod. 52, 709–720 (1995)
Ying, Q. L. et al. The ground state of embryonic stem cell self-renewal. Nature 453, 519–523 (2008)
Woltjen, K. et al. piggyBac transposition reprograms fibroblasts to induced pluripotent stem cells. Nature 458, 766–770 (2009)
Kurimoto, K. et al. An improved single-cell cDNA amplification method for efficient high-density oligonucleotide microarray analysis. Nucleic Acids Res. 34, e42 (2006)
Niwa, H., Masui, S., Chambers, I., Smith, A. G. & Miyazaki, J. Phenotypic complementation establishes requirements for specific POU domain and generic transactivation function of Oct-3/4 in embryonic stem cells. Mol. Cell. Biol. 22, 1526–1536 (2002)
Guo, G. & Smith, A. A genome-wide screen in EpiSCs identifies Nr5a nuclear receptors as potent inducers of ground state pluripotency. Development 137, 3185–3192 (2010)
Niwa, H., Yamamura, K. & Miyazaki, J. Efficient selection for high-expression transfectants with a novel eukaryotic vector. Gene 108, 193–199 (1991)
Lucifero, D., Mertineit, C., Clarke, H. J., Bestor, T. H. & Trasler, J. M. Methylation dynamics of imprinted genes in mouse germ cells. Genomics 79, 530–538 (2002)
Kumaki, Y., Oda, M. & Okano, M. QUMA: quantification tool for methylation analysis. Nucleic Acids Res. 36, W170–W175 (2008)
Li, C. & Wong, W. H. Model-based analysis of oligonucleotide arrays: expression index computation and outlier detection. Proc. Natl Acad. Sci. USA 98, 31–36 (2001)
R. Development Core Team. R: A Language and Environment for Statistical Computinghttp://www.R-project.org (R Foundation for Statistical Computing, 2012)
Storey, J. D. & Tibshirani, R. Statistical significance for genomewide studies. Proc. Natl Acad. Sci. USA 100, 9440–9445 (2003)
Kanatsu-Shinohara, M. et al. Allogeneic offspring produced by male germ line stem cell transplantation into infertile mouse testis. Biol. Reprod. 68, 167–173 (2003)
We thank A. Bradley, A. Smith, G. Guo, H. Niwa and G. Nagamatsu for providing plasmids. We are grateful to the Center for Anatomical Studies (Graduate School of Medicine, Kyoto University) for performing the histological analyses. We thank M. Yamaji for advice and T. Mori for encouragement. F.N. is a Japan Society for the Promotion of Science (JSPS) Research Fellow. This study was supported in part by a Grant-in-Aid from the Ministry of Education, Culture, Sports, Science, and Technology of Japan; by JST-CREST/ERATO; by the Takeda Science Foundation; and by the Academia for Repro-regenerative Medicine.
The authors declare no competing financial interests.
This file contains Supplementary Figures 1-10. (PDF 5684 kb)
This file contains the top 100 genes contributing to the PC1 and PC2 of Figure 2a. (XLS 51 kb)
This file contains core PGC genes and somatic mesodermal genes (related to Supplementary Figure 6b). (XLS 78 kb)
This file contains the top 100 genes up-/down-regulated in TF(BP14A)-PGCLCs compared to EpiLCs and in SC-ESCs by BP14A compared to ESCs (related to Supplementary Figure 6c). (XLS 74 kb)
This file contains the top 100 genes contributing to the PC1 and PC2 of Figure 3a. (XLS 50 kb)
This file contains lists of genes exhibiting differential expression between P14 and B cells (related to Supplementary Figure 9). (XLS 102 kb)
This file contains spermatogenesis by TF- or Ck-PGCLCs. (XLS 22 kb)
This file contains development of embryos derived from TF- and Ck-PGCLC-derived spermatozoa. (XLS 20 kb)
This file contains tagged sequences for exogenous transcription factors. (XLS 14 kb)
This file contains primer sequences used in this study. (XLS 24 kb)
About this article
Cite this article
Nakaki, F., Hayashi, K., Ohta, H. et al. Induction of mouse germ-cell fate by transcription factors in vitro . Nature 501, 222–226 (2013). https://doi.org/10.1038/nature12417
Stem Cell Research & Therapy (2022)
Nature Communications (2022)
Nature Cell Biology (2022)
Bend family proteins mark chromatin boundaries and synergistically promote early germ cell differentiation
Protein & Cell (2022)
Nature Communications (2021)