3D ultrastructural analysis of plant endomembrane with high pressure freezing and correlative array tomography
Project/Area Number |
17K07477
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Research Category |
Grant-in-Aid for Scientific Research (C)
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Allocation Type | Multi-year Fund |
Section | 一般 |
Research Field |
Morphology/Structure
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Research Institution | Institute of Physical and Chemical Research |
Principal Investigator |
Toyooka Kiminori 国立研究開発法人理化学研究所, 環境資源科学研究センター, 上級技師 (10360596)
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Project Period (FY) |
2017-04-01 – 2020-03-31
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Project Status |
Completed (Fiscal Year 2019)
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Budget Amount *help |
¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000)
Fiscal Year 2019: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2018: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2017: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
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Keywords | 高圧凍結技法 / エンドソーム / アレイトモグラフィー / 光電子相関顕微鏡 / CLEM / 電子顕微鏡 / 植物細胞 / オルガネラ / 3次元超微形態 / 細胞内輸送 / 膜交通 / 3次元超微形態 / 光電子相関顕微鏡法 / 相関アレイトモグラフィー法 / 細胞内膜系 |
Outline of Final Research Achievements |
The purpose of this research is to develop an ultrastructural observation for fluorescent proteins (FP) -labeled organelles by combining high-pressure freeze technique (HPF) and correlative light & electron microscopy (CLEM), and to identify the ultrastructure of FP-labeled intracellular membrane systems by array tomography (AT). As obtained results, we developed the CLEM method, which recovers the fluorescence of FP-labeled organelles in chemically fixed resin-embedded sections using antifade reagents, and established the HFP method suitable for plants by using the latest HPF equipment. The CLEM method, which combines a high-sensitivity confocal laser scanning microscope and an AT-equipped scanning electron microscope using ultra-thin sections, has been studied and the possibility of capturing the microstructure of intracellular membrane organelles has been obtained by using a fixed-resin-resistant FPs.
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Academic Significance and Societal Importance of the Research Achievements |
CLEM法及び凍結技法は組織・細胞内の蛍光標識した超微細構造を限りなく生きた状態で可視化する技術として、これからの研究に重要である。本研究により、これら技術を発展させた。電顕メーカーとともにCLEMシステムを改良するとともに、蛍光顕微鏡や共焦点顕微鏡で試料中の蛍光を効率よく感度良く検出し、電顕で観察するために微細構造を保持する試料調製を開発した。また、最新の高圧凍結装置を用いて生物の超微形態を保持する試料調製法を確立した。これらの技術は、植物だけでなく動物や微生物など幅広い基礎研究及び応用研究に活用できると期待される。
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Report
(4 results)
Research Products
(49 results)
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[Journal Article] A conserved regulatory mechanism mediates the convergent evolution of plant shoot lateral organs.2019
Author(s)
Naramoto S, Jones VAS, Nicola, N, Sato M,Toyooka K, Shimamura M, Ishida S, Nishitani K,Ishizaki K, Nishihama R, Kohchi T, Dolan L, Kyozuka J.
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Journal Title
PLoS Biology
Volume: 17
Issue: 12
Pages: 12-12
DOI
NAID
Related Report
Peer Reviewed / Open Access / Int'l Joint Research
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[Journal Article] The RopGEF KARAPPO Is Essential for the Initiation of Vegetative Reproduction in Marchantia polymorpha2019
Author(s)
Hiwatashi T., Goh H., Yasui Y., Koh L.Q., Takami H., Kajikawa M., Kirita H., Kanazawa T., Minamino N., Togawa T., Sato M., Wakazaki M., Yamaguchi K., Shigenobu S., Fukaki H., Mimura T., Toyooka K., Sawa S., Yamato K.T.,Ueda T., Urano D., Kohchi T., Ishizaki K.
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Journal Title
Current Biology
Volume: 29
Issue: 20
Pages: 3525-3531
DOI
Related Report
Peer Reviewed / Open Access / Int'l Joint Research
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[Journal Article] Plastid gene expression is essential for lateral root stem-cell patterning, symplastic connectivity, and root plastid distribution in Arabidopsis thaliana2018
Author(s)
Nakata MT, Sato M, Wakazaki M, Sato N, Kojima K Sekine A, Nakamura S, Shikanai T, Toyooka K, Tsukaya H, Horiguchi G
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Journal Title
Biology Open
Volume: 7
Pages: 028175-028175
DOI
Related Report
Peer Reviewed / Open Access
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[Journal Article] Ectopic colonization of oral bacteria in the intestine drives TH1 cell induction and inflammation" Ectopic colonization of oral bacteria in the intestine drives TH1 cell induction and inflammationx2017
Author(s)
Atarashi K, Suda W, Luo C, Kawaguchi T, Motoo I, Narushima S, Kiguchi Y, Yasuma K, Watanabe E, Tanoue T, Thaiss CA, Sato M, Toyooka K, Said HS, Yamagami H, Rice SA, Gevers D, Johnson RC, Segre JA, Chen K, Kolls JK, Elinav E, Morita H, Xavier RJ, Hattori M, Honda K.
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Journal Title
Science
Volume: 358
Issue: 6361
Pages: 359-365
DOI
Related Report
Peer Reviewed / Int'l Joint Research
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[Journal Article] Protonema of the moss Funaria hygrometrica can function as a lead (Pb) adsorbent2017
Author(s)
Itouga M, Hayatsu M, Sato M, Tsuboi Y, Kato Y, Toyooka K, Suzuki S, Nakatsuka S, Kawakami S, Kikuchi J, Sakakibara H
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Journal Title
PLOS ONE
Volume: 12
Issue: 12
Pages: e0189726-e0189726
DOI
Related Report
Peer Reviewed / Open Access
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[Journal Article] The karrikin receptor KAI2 promotes drought resistance in Arabidopsis thaliana2017
Author(s)
Li W, Nguyen KH, Chu HD, Ha CV, Watanabe Y, Osakabe Y, Leyva-Gonzalez MA, Sato M, Toyooka K, Voges L, Tanaka M, Mostofa MG, Seki M, Seo M, Yamaguchi S, Nelson DC, Herrera-Estrella L, Tran LSran
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Journal Title
PLoS Genetics
Volume: -
Issue: 11
Pages: e1007076-e1007076
DOI
Related Report
Peer Reviewed / Open Access / Int'l Joint Research
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