| Project/Area Number |
17K19533
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| Research Category |
Grant-in-Aid for Challenging Research (Exploratory)
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| Allocation Type | Multi-year Fund |
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| Research Field |
Biomedical structure and function and related fields
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| Research Institution | Toho University |
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Principal Investigator |
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| Co-Investigator(Kenkyū-buntansha) |
村井 晋 東邦大学, 医学部, 助教 (90287540)
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| Research Collaborator |
SHINDO Ryodai
NAKABAYASHI Osamu
YAMAZAKI Soh
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| Project Period (FY) |
2017-06-30 – 2019-03-31
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| Project Status |
Completed (Fiscal Year 2018)
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| Budget Amount *help |
¥6,500,000 (Direct Cost: ¥5,000,000、Indirect Cost: ¥1,500,000)
Fiscal Year 2018: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
Fiscal Year 2017: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
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| Keywords | ネクロプトーシス / FRET / イメージング / DAMPs / RIPK3 / MLKL / 全反射顕微鏡 / トランスジェニックマウス / SMART / ２光子顕微鏡 / HMGB1 / Histone H3 |
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| Outline of Final Research Achievements |
Necroptosis is a regulated form of necrosis that depends on receptor-interacting protein kinase (RIPK)3 and mixed lineage kinase domain-like (MLKL). While danger-associated molecular pattern (DAMP)s are involved in various pathological conditions and released from dead cells, the underlying mechanisms are not fully understood. Here we develop a fluorescence resonance energy transfer (FRET) biosensor, termed SMART (a sensor for MLKL activation by RIPK3 based on FRET). SMART is composed of a fragment of MLKL and monitors necroptosis, but not apoptosis or necrosis. SMART monitors plasma membrane translocation of oligomerized MLKL, which is induced by RIPK3. SMART in combination with imaging of the release of nuclear DAMPs and Live-Cell Imaging for Secretion activity (LCI-S) reveals two different modes of the release of High Mobility Group Box 1 from necroptotic cells. We also generated mice stably expressing SMART that will be useful to investigate necroptotic cells in vivo.
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| Academic Significance and Societal Importance of the Research Achievements |
本研究は、蛍光イメージング法によるネクロプトーシスの可視化を実現した世界で初めての研究であり、死細胞から放出され周囲の細胞に影響を与え、様々な病態に深く関与しているDAMPsの放出パターンが2種類があることを1細胞レベルで初めて明らかにした研究である。今後、ネクロプトーシスやDAMPsが関与する病態の解明や、治療技術の開発に貢献することが期待される。また新規に開発したセンサータンパク質を発現する遺伝子改変マウスなどを作成することで、ネクロプトーシスが体の中でどのような状況で起こっているかの新たな解析手法を提供できる可能性がある。
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