3D-CLEM analysis of the membrane structure in autophagy
| Project/Area Number |
17K15550
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| Research Category |
Grant-in-Aid for Young Scientists (B)
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| Allocation Type | Multi-year Fund |
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| Research Field |
General anatomy (including histology/embryology)
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| Research Institution | Juntendo University |
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Principal Investigator |
Kakuta Soichiro 順天堂大学, 医学(系)研究科(研究院), 准教授 (80551209)
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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 |
¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2019: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Fiscal Year 2018: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Fiscal Year 2017: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
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| Keywords | オートファジー / 電子顕微鏡 / 形態学 / 共焦点レーザー顕微鏡 / CLEM / 光-電子相関顕微鏡法 / 三次元電子顕微鏡解析 / メンブレントラフィック / Atg9 / 解剖学 |
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| Outline of Final Research Achievements |
To better understand the morphology of autophagosome membrane formation, we performed a three-dimensional electron microscopy analysis. The primary objective of this research is to establish a method for selective observation of target structures and sites using light-electron correlative microscopy (CLEM). In addition to the labeling of the target of autophagy formation site, the mitochondria were fluorescently labeled. We developed the method for accurate positional correction by overlaying the fluorescence image with the electron microscope image using image analysis software. Normal cells showed autophagosomes and autolysosomes, and ATG7 KO cells showed more isolation membranes. Some isolation membrane-like structures were observed in ATG9A KO cells, where membrane formation is not supposed to occur.
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| Academic Significance and Societal Importance of the Research Achievements |
オートファジーは様々な疾病への関与が盛んに報告されている細胞内機構である。顕微鏡を用いた形態学的な研究は初期から行われていたが、蛍光顕微鏡では微細形態までは捉えることが出来ず、一方で電子顕微鏡では立体的な情報を得ることが難しく、また形態だけでは目的の構造かどうか判断できないことも多い。本研究では蛍光顕微鏡像と「FIB/SEMによる3次元電子顕微鏡像」を組み合わせた解析を行うことでそれぞれの欠点を補うことが出来た。これによりオートファジー関連構造体の変化についての新しい知見が得られ、またこれを進めることによってオートファジー現象全体の理解へと繋がることが期待出来る。
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Report
(4 results)
Research Products
(16 results)
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[Journal Article] Three-dimensional electron microscopy analysis reveals endopolygeny-like nuclear architecture segregation in Plasmodium oocyst development.2020
Author(s)
Araki T, Kawai S, Kakuta S, Kobayashi H, Umeki Y, Saito-Nakano Y, Sasaki T, Nagamune K, Yasutomi Y, Nozaki T, Franke-Fayard B, Khan SM, Hisaeda H, Annoura T
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Journal Title
Related Report
Peer Reviewed
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[Journal Article] Atg9a deficiency causes axon-specific lesions including neuronal circuit dysgenesis.2017
Author(s)
Yamaguchi J, Suzuki C, Nanao T, Kakuta S, Ozawa K, Tanida I, Saitoh T, Sunabori T, Komatsu M, Tanaka K, Aoki S, Sakimura K, Uchiyama Y.
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Journal Title
Autophagy
Volume: 17
Issue: 5
Pages: 1-14
DOI
NAID
Related Report
Peer Reviewed / Open Access / Int'l Joint Research
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