Functional analysis of newly identified DNA demethylation factors in mouse ES cells and its application to somatic cell reprogramming

• Project/Area Number: 15K18545
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Developmental biology
• Research Institution: Institute of Physical and Chemical Research
• Principal Investigator: Hatanaka Yuki 国立研究開発法人理化学研究所, バイオリソースセンター, 特別研究員 (70719450)
• Project Period (FY): 2015-04-01 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥4,030,000 (Direct Cost: ¥3,100,000、Indirect Cost: ¥930,000); Fiscal Year 2016: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000); Fiscal Year 2015: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
• Keywords: 受精卵 / ES細胞 / DNA脱メチル化 / エピジェネティックリプログラミング / エピゲノム / ヒストン修飾 / 5mCの酸化

Outline of Final Research Achievements

To explore the application to improvements of somatic nuclear reprogramming, we focused on GSE and H3R17me2a enzyme Mettl23 which are newly identified DNA demethylation factors in mouse zygotes. To this end, we first produced their KO mice and investigated the effect of GSE and Mettl23 deficient on the DNA demethylation in ESCs. The levels of 5mC and its oxidative products were not significantly different between wild type (WT) ESCs and these KO. Interestingly, the level of 5fC was significantly increased in their KOESCs on the promoter regions that undergo DNA demethylation in zygote. The loss of GSE and Mettl23 resulted in the impairment of recruitment of TDG, which excises 5fC and 5CaC. The treatment of TBBD (ellagic acid), which specifically inhibits methylation at H3R17, resulted in the reduction of recruitment of GSE and Histone H3.3 on the target regions. These results suggested that GSE and Mettl23 are responsible for DNA demethylation on defined regions in ESCs.

Research Products

• [Journal Article] The rodent-specific microRNA cluster within the Sfmbt2 gene is imprinted and essential for placental development. (2017)
  • Author(s): Inoue K, Hirose M, Inoue H, Hatanaka Y, Honda A, Hasegawa A, Mochida K, Ogura A.
  • Journal Title: Cell Reports Volume: 印刷中
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] CRISPR/Cas9-mediated genome editing in wild-derived mice: generation of tamed wild-derived strains by mutation of the a (nonagouti) gene (2017)
  • Author(s): Michiko Hirose, Ayumi Hasegawa, Keiji Mochida, Shogo Matoba, Yuki Hatanaka, Kimiko Inoue, Tatsuhiko Goto, Hideki Kaneda, Ikuko Yamada, Tamio Furuse, Kuniya Abe, Yoshihisa Uenoyama, Hiroko Tsukamura, Shigeharu Wakana, Arata Honda, and Atsuo Ogura
  • Journal Title: Scientific Reports Volume: 7 Issue: 1 Pages: 42476-42476
  • DOI: 10.1038/srep42476
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Mouse D1Pas1, a DEAD-box RNA helicase, is required for the completion of first meiotic prophase in male germ cells (2016)
  • Author(s): Inoue H, Ogonuki N, Hirose M, Hatanaka Y, Matoba S, Chuma S, Kobayashi K, Wakana S, Noguchi J, Inoue K, Tanemura K, *Ogura A
  • Journal Title: Biochem Biophys Res Commun Volume: 478 Issue: 2 Pages: 592-8
  • DOI: 10.1016/j.bbrc.2016.07.109
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Histone chaperone CAF-1 mediates repressive histone modifications to protect preimplantation mouse embryos from endogenous retrotransposon. (2015)
  • Author(s): Hatanaka Y, Inoue K, Oikawa M, Kamimura S, Ogonuki N, Kodam E, Ohkawa Y, Tsukada Y, Ogura A.
  • Journal Title: Proc Natl Acad Sci USA Volume: 112 Issue: 47 Pages: 14641-14646
  • DOI: 10.1073/pnas.1512775112
  • Peer Reviewed / Open Access
• [Presentation] マウス受精卵における能動的DNA脱メチル化制御因子の役割 (2016)
  • Author(s): 畑中勇輝, 越後貫成美, 廣瀬美智子, 本多新, 井上貴美子, 松本和也, 小倉淳郎
  • Organizer: 第109回日本繁殖生物学会
  • Place of Presentation: 麻布大学獣医学部 (神奈川県相模原市)
  • Year and Date: 2016-09-11
• [Presentation] Histone H3R17me2a mark recruits Tet3 to initiate active DNA demethylation in mouse zygotes (2016)
  • Author(s): Hatanaka Y, Shimizu N, Morita K, Satoh M, Honda A, Hirose M, Kamimura S, Ogonuki N, Nakamura N, Inoue K, Hosoi Y, Nakano T, Matsumoto K, Ogura A.
  • Organizer: Mouse Genetics 2016
  • Place of Presentation: Florida, USA
  • Year and Date: 2016-07-13
  • Int'l Joint Research
• [Presentation] 着床前胚においてヒストンシャペロンCAF-1 は抑制性ヒストン修飾を調節することでレトロトランスポゾン抑制に機能する。 (2015)
  • Author(s): 畑中勇輝，井上貴美子，及川真実，上村悟氏，越後貫成美，児玉栄一，大川恭行，束田裕一，小倉淳郎
  • Organizer: 第38回日本分子生物学会大会
  • Place of Presentation: 神戸ポートアイランド (神戸)
  • Year and Date: 2015-12-01
• [Presentation] Histone H3R17me2a Mark Recruits Tet3 to Initiate Active DNA Demethylation in Mouse Zygotes (2015)
  • Author(s): Hatanaka Y, Shimizu N, Morita K, Satoh M, Honda A, Hirose M, Kamimura S, Ogonuki N, Nakamura T, Inoue K, Hosoi Y, Nakano T, Matsumoto K, Ogura A
  • Organizer: THE 40th NAITO CONFERENCE “Epigenetics―From Histone Code to Therapeutic Strategy”
  • Place of Presentation: シャトレーゼ ガトーキングダム サッポロ（北海道）
  • Year and Date: 2015-09-15
  • Int'l Joint Research / Invited
• [Presentation] 受精卵における能動的DNA脱メチル化機構 (2015)
  • Author(s): 畑中勇輝
  • Organizer: 第60回日本生殖医学会学術講演会
  • Place of Presentation: パシフィコ横浜 (横浜)
  • Year and Date: 2015-04-26
  • Invited