Ｇｅｍｉｎｉｎ発現制御による白血病幹細胞の活性制御機構の解析

• Project Area: Development of Novel Treatment Strategies Targeting Cancer Stem Cells
• Project/Area Number: 23130511
• Research Category: Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
• Allocation Type: Single-year Grants
• Review Section: Biological Sciences
• Research Institution: Hiroshima University
• Principal Investigator: 安永 晋一郎 広島大学, 原爆放射線医科学研究所, 准教授 (50336111)
• Project Period (FY): 2011-04-01 – 2013-03-31
• Project Status: Completed (Fiscal Year 2012)
• Budget Amount: ¥8,580,000 (Direct Cost: ¥6,600,000、Indirect Cost: ¥1,980,000); Fiscal Year 2012: ¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000); Fiscal Year 2011: ¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000)
• Keywords: 白血病幹細胞 / Geminin / ユビキチン化 / Hox / ポリコーム / 細胞周期 / DNA複製 / クロマチンリモデリング

Outline of Annual Research Achievements

本研究において、レトロウイルスベクターを用いたGemininの過剰発現系及びGemininのshRNAノックダウン系を確立した。また、spinoculationを用いた極めて効率的に未分化造血細胞にレトロウイルスを感染させる方法をも確立した。これらの実験系を用いて、MLL遺伝子の転座を有する白血病細胞において発現が著しく亢進しており、その予後を規定する因子であるHoxa9が、従来知られてきた転写制御因子としての機能だけではなく、Roc1-Cul4a-Ddb1コアE3ユビキチンリガーゼ複合体と結合し、DNA複製ライセンス化制御因子であり幹細胞の未分化性維持因子でもあるGemininに対するE3ユビキチンリガーゼを構成し、Gemininタンパク質を分解に導くことを生化学的に証明し、Hoxa9の過剰発現による造血幹細胞や造血前駆細胞の活性化においてHoxa9によるGemininのユビキチン化を介したタンパク質発現減少が、重要な役割を果たしていることを明らかにした。次いで、本研究ではGemininの発現動態をin vivoで可視化するためにGeminin-EYFPノックインマウスを作成し、これを用いた血液学的実験系を確立した。このノックインマウスの骨髄細胞にMLL-AF9、Hoxa9+Meis1a及びBCR-ABLを導入し形質転換した細胞を用いてコロニー形成能を調べる実験系を確立し、さらに上記のGeminin発現制御系を用いて、コロニー形成能に与えるGemininの発現量の影響を検討した。そして、Geminin高発現下でこれら形質転換細胞を誘導するに当たりHoxa9の高発現の維持が必須であることを見出した。

Research Progress Status

24年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

24年度が最終年度であるため、記入しない。

Research Products

• [Journal Article] Scmh1 has E3 ubiquitin ligase activity for Geminin and histone H2A and regulates Geminin stability directly or indirectly via transcription repression of Hoxa9 and Hoxb4 (2013)
  • Author(s): Yasunaga, S., Ohtsubo, M., Ohno, Y., Saeki, K., Kurogi. T., Tanaka-Okamoto, M., Ishizaki, H., Shirai, M., Mihara, K., Brock, H. W., Miyoshi, J. & Takihara Y
  • Journal Title: Mol. Cell. Biol. Volume: 33(4) Issue: 4 Pages: 644-660
  • DOI: 10.1128/mcb.00974-12
  • Peer Reviewed
• [Journal Article] Hoxa9 transduction induces mouse hematopoietic stem and progenitor cell activity through direct down-regulation of Geminin. (2013)
  • Author(s): Ohno, Y., Yasunaga, S., Janmohamed, S., Ohtubo, M., Mihara, K., Iscove, N., Takihara, Y.
  • Journal Title: PLoS ONE Volume: 8(1) Issue: 1 Pages: e53161-e53161
  • DOI: 10.1371/journal.pone.0053161
  • Peer Reviewed
• [Journal Article] oly-ADP ribosylation of Miki by tankyrase-1 promotes centrosome maturation (2012)
  • Author(s): Ozaki, Yuko
  • Journal Title: Molecular Cell Volume: 47 Issue: 5 Pages: 694-706
  • DOI: 10.1016/j.molcel.2012.06.033
  • Peer Reviewed
• [Journal Article] Dominant-negative STAT1 SH2 domain mutations in unrelated patients with Mendelian susceptibility to mycobacterial disease. (2012)
  • Author(s): Tsumura M.
  • Journal Title: Human Mutation Human Mutation Volume: 33 Issue: 9 Pages: 1377-1387
  • DOI: 10.1002/humu.22113
  • Peer Reviewed
• [Journal Article] Overexpression of BMI-1 correlates with drug resistance in B-cell lymphoma cells through the stabilization of survivin expression (2012)
  • Author(s): Bhattacharyya, J., Mihara, K., Ohtsubo, M., Yasunaga, S., Takei, Y., Yanagihara, Y., Sakai, A., Hoshi, M., Takihara, Y., Kimura, A.
  • Journal Title: Cancer Sci. Volume: 103(1) Issue: 1 Pages: 34-41
  • DOI: 10.1111/j.1349-7006.2011.02121.x
  • Peer Reviewed
• [Journal Article] Decreased expression in nuclear factor-kB essential modulator due to a novel splice-site mutation causes X-linked ectodermal dysplasia with immunedeficiency (2011)
  • Author(s): Karakawa, S., Okada, S., Tsumura, M., Mizoguchi, Y., Hara, K., Ohno, N., Yasunaga, S., Ohtsubo M., Kawai, T., Nishikimori, R., Sakaguchi, T., Hata, I., Sakura, N., Takihara, Y., Kobayashi, M.
  • Journal Title: Clin.Immunol. Volume: 31(5) Pages: 762-772
  • Peer Reviewed
• [Presentation] cmh1 is required for maintaining hematopoietic homeostasis through the molecular network governing Geminin expression (2012)
  • Author(s): Ohno Y.
  • Organizer: he 35th Annual Meeting of the Molecular Biology of Japan
  • Place of Presentation: Fukuoka, Japan
  • Year and Date: 2012-12-11
• [Presentation] Molecular role for Geminin in E2F-mediated transcriptional regulation (2012)
  • Author(s): Saeki K.
  • Organizer: The 35th Annual Meeting of the Molecular Biology of Japan
  • Place of Presentation: Fukuoka, Japan
  • Year and Date: 2012-12-11
• [Presentation] Scmh1 regulates Geminin through the ubiquitin-proteasome systems to govern hematopoietic homeostasis (2012)
  • Author(s): Yasunaga S.
  • Organizer: The 74th Annual Meeting of the Japanese Society of Hematology
  • Place of Presentation: Kyoto, Japan
  • Year and Date: 2012-10-19
• [Presentation] A new approach for activating mouse hematopoietic stem and progenitor cells by manipulatin (2012)
  • Author(s): Yasunaga S.
  • Organizer: The 10th Annual Meeting of International Society for Stem Cell Research (ISSCR)
  • Place of Presentation: Yokohama, Japan
  • Year and Date: 2012-06-13
• [Presentation] A novel molecular function for Hoxa9 in hematopoiesis and leukemogenesis (2012)
  • Author(s): Yasunaga S.
  • Organizer: The 10th Stem Cell Research Symposium
  • Place of Presentation: Awaji, Japan
  • Year and Date: 2012-05-31
• [Presentation] A new strategy for manipulating HSC activity through the modulation of Geminin expression (2011)
  • Author(s): Yasunaga S., Ohno Y., Ohtsubo M., Furutani T., Kawashima A., Tetsuguchi H., Saeki K., Nakashima Y., Takihara Y.
  • Organizer: The 73^<rd> Annual Meeting of the Japanese Society of Hematology
  • Place of Presentation: Nagoya, Japan
• [Presentation] HOXA9 provides hematopoietic cells with proliferation potential through the degradation of Geminin (2011)
  • Author(s): Ohno Y., Yasunga S., Ohtsubo M., Tetsuguchi H., Furutani T., Kawashima A., Saeki K., Nakashima Y., Takihara Y.
  • Organizer: The 73^<rd> Annual Meeting of the Japanese Society of Hematology
  • Place of Presentation: Nagoya, Japan
• [Presentation] Visualization and modulation of Geminin expression for inducing the hematopoietic stem cell activity (2011)
  • Author(s): Furutani T., Yasunaga S., Ohno Y., Kawashima A., Tetsuguchi H., Saeki K., Nakashima Y., Ohtsubo M., Takihara Y.
  • Organizer: The 34^<th> Annual Meeting of the Molecular Biology of Japan
  • Place of Presentation: Yokohama, Japan
• [Presentation] HOXA9 directly regulates Geminin by composing the E3 ubiquitin ligase that plays an important role in HOXA9-mediated induction of hematopoiesis and leukemogenesis (2011)
  • Author(s): Tetsuguchi H., Ohno Y., Yasunaga S., Furutani T., Kawashima A., Saeki K., Nakashima Y., Ohtsubo M., Takihara Y.
  • Organizer: The 34^<th> Annual Meeting of the Molecular Biology of Japan
  • Place of Presentation: Yokohama, Japan
• [Presentation] Molecular roles for Scmh1, substoichiometric component of Polycomb-group complex 1 (2011)
  • Author(s): Kawashima A., Yasunaga S., Ohno Y., Furutani T., Tetsuguchi H., Saeki K., Nakashima Y., Ohtsubo M., Takihara Y.
  • Organizer: The 34^<th> Annual Meeting of the Molecular Biology of Japan
  • Place of Presentation: Yokohama, Japan
• [Remarks] 広島大学 原爆放射線医科学研究所 幹細胞機能学研究分野
  • URL: http://home.hiroshima-u.ac.jp/dscb
• [Remarks]
  • URL: http://home.hiroshima-u.ac.jp/dscb