In vitro expansion of mouse primordial germ cell‐like cells recapitulates an epigenetic blank slate |
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| Authors: | Hiroshi Ohta Kazuki Kurimoto Ikuhiro Okamoto Tomonori Nakamura Yukihiro Yabuta Hidetaka Miyauchi Takuya Yamamoto Yukiko Okuno Masatoshi Hagiwara Kenjiro Shirane Hiroyuki Sasaki Mitinori Saitou |
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| Institution: | 1. Department of Anatomy and Cell Biology, Graduate School of Medicine, Kyoto University, Yoshida‐Konoe‐cho, Sakyo‐ku, Kyoto, Japan;2. JST, ERATO, Yoshida‐Konoe‐cho, Sakyo‐ku, Kyoto, Japan;3. Center for iPS Cell Research and Application, Kyoto University, Shogoin, Sakyo‐ku, Kyoto, Japan;4. Institute for Integrated Cell‐Material Sciences, Kyoto University, Yoshida‐Ushinomiya‐cho, Sakyo‐ku, Kyoto, Japan;5. AMED‐CREST, AMED, Chiyoda‐ku, Tokyo, Japan;6. Medical Research Support Center, Graduate School of Medicine, Kyoto University, Yoshida‐Konoe‐cho, Sakyo‐ku, Kyoto, Japan;7. Department of Anatomy and Developmental Biology, Graduate School of Medicine, Kyoto University, Yoshida‐Konoe‐cho, Kyoto, Japan;8. Division of Epigenomics and Development, Medical Institute of Bioregulation, Epigenome Network Research Center, Kyushu University, Higashi‐ku, Fukuoka, Japan;9. Graduate School of Medical Sciences, Kyushu University, Higashi‐ku, Fukuoka, Japan |
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| Abstract: | The expansion of primordial germ cells (PGCs), the precursors for the oocytes and spermatozoa, is a key challenge in reproductive biology/medicine. Using a chemical screening exploiting PGC‐like cells (PGCLCs) induced from mouse embryonic stem cells (ESCs), we here identify key signaling pathways critical for PGCLC proliferation. We show that the combinatorial application of Forskolin and Rolipram, which stimulate cAMP signaling via different mechanisms, expands PGCLCs up to ~50‐fold in culture. The expanded PGCLCs maintain robust capacity for spermatogenesis, rescuing the fertility of infertile mice. Strikingly, during expansion, PGCLCs comprehensively erase their DNA methylome, including parental imprints, in a manner that precisely recapitulates genome‐wide DNA demethylation in gonadal germ cells, while essentially maintaining their identity as sexually uncommitted PGCs, apparently through appropriate histone modifications. By establishing a paradigm for PGCLC expansion, our system reconstitutes the epigenetic “blank slate” of the germ line, an immediate precursory state for sexually dimorphic differentiation. |
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| Keywords: | cAMP signaling epigenetic reprogramming in vitro expansion PGC‐like cells primordial germ cells |
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