Time-series measurement of totipotent state and establishment of totipotent stem cells using a novel long-term single-cell tracking method

• Project/Area Number: 18K19275
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Review Section: Medium-sized Section 42:Veterinary medical science, animal science, and related fields
• Research Institution: Nara Medical University
• Principal Investigator: Horie Kyoji 奈良県立医科大学, 医学部, 教授 (30333446)
• Project Period (FY): 2018-06-29 – 2021-03-31
• Project Status: Completed (Fiscal Year 2020)
• Budget Amount: ¥6,370,000 (Direct Cost: ¥4,900,000、Indirect Cost: ¥1,470,000); Fiscal Year 2019: ¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000); Fiscal Year 2018: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
• Keywords: 全能性 / 幹細胞 / ES細胞 / iPS細胞 / 再生医学

Outline of Final Research Achievements

In this study, we tried to visualize the totipotent state in mice and humans using fluorescent proteins and observed it over time to elucidate the factors that regulate the totipotent state and to establish totipotent stem cells. To visualize the totipotent state, we used markers that have been reported to be upregulated in the totipotent state. We also evaluated the relationship between the totipotent state and a newly identified gene whose expression changes in the undifferentiated state. In particular, for the latter gene, we identified a group of transcription factors that regulate the expression of this gene by gene disruption experiments using CRISPR/Cas9.

Academic Significance and Societal Importance of the Research Achievements

「全能性幹細胞」とは、全ての細胞に分化誘導が可能で、かつ、安定的に培養できる細胞を意味する。これまで、ES細胞、生殖幹細胞、神経幹細胞など、様々な幹細胞が樹立され、幹細胞状態の理解や再生医療に役立ってきたが、「全能性幹細胞」の樹立は極めて遅れている。本研究では、マウスとヒトの両方において、全能性状態の検出系の開発と全能性状態の誘導を試みた。マウスを用いた実験は、初期胚実験を自在に行える点で利点があり、一方、ヒト細胞を用いた実験は、再生医療を指向した研究を行う上で必須である。今後、マウスとヒトの各々の実験系の利点を生かしながら、哺乳動物の全能性の共通原理の解明と再生医療への応用を目指したい。

Research Products

• [Presentation] Identifying the heterogeneity of ground state pluripotency in mouse embryonic stem cells and elucidating its regulatory mechanism (2019)
  • Author(s): Kyoji Horie, Hitomi Watanabe, Yosuke Nishimura, Hikaru Watanabe, Masahide Seki, Akio Seita, Kagayaki Kato, Yuichi Wakayama, Jun Sese, Yutaka Suzuki, Takuji Yamada, Gen Kondoh, Junko Yoshida
  • Organizer: 第42回日本分子生物学会年会 2019.12.4 福岡
  • Invited
• [Presentation] Identifying heterogeneity of ground state pluripotency in mouse embryonic stem cells (2018)
  • Author(s): Kyoji Horie, Junko Yoshida
  • Organizer: International Society for Stem Cell Research Annual Meeting
  • Int'l Joint Research
• [Presentation] Ground stateにおけるマウスES細胞の不均一性の同定 (2018)
  • Author(s): 堀江恭二, 渡邊仁美, 西村陽介, 渡邊日佳流, 関真秀, 清田晃央, 加藤輝, 若本祐一, 鈴木穣, 山田拓司, 近藤玄, 吉田純子
  • Organizer: 第41回日本分子生物学会年会