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Elucidation of the function of the pre-autophagosomal structure through in vitro reconstitution

Research Project

Project/Area Number 18H03989
Research Category

Grant-in-Aid for Scientific Research (A)

Allocation TypeSingle-year Grants
Section一般
Review Section Medium-sized Section 43:Biology at molecular to cellular levels, and related fields
Research InstitutionMicrobial Chemistry Research Foundation

Principal Investigator

Noda Nobuo  公益財団法人微生物化学研究会, 微生物化学研究所, 部長 (40396297)

Project Period (FY) 2018-04-01 – 2021-03-31
Project Status Completed (Fiscal Year 2020)
Budget Amount *help
¥44,330,000 (Direct Cost: ¥34,100,000、Indirect Cost: ¥10,230,000)
Fiscal Year 2020: ¥14,560,000 (Direct Cost: ¥11,200,000、Indirect Cost: ¥3,360,000)
Fiscal Year 2019: ¥14,560,000 (Direct Cost: ¥11,200,000、Indirect Cost: ¥3,360,000)
Fiscal Year 2018: ¥15,210,000 (Direct Cost: ¥11,700,000、Indirect Cost: ¥3,510,000)
Keywords液-液相分離 / オートファジー / Atg1複合体 / 天然変性タンパク質 / 高速AFM / リン酸化 / 液滴 / 再構成 / オートファゴソーム形成場 / 液-液相分離
Outline of Final Research Achievements

Despite the importance in autophagy, the entity of the PAS has been a long-standing mystery. We showed that the PAS behaves as a liquid droplet with high fluidity in budding yeast. In vitro experiments using purified proteins revealed that the Atg1 complex can undergo liquid-liquid phase separation to form a liquid droplet. Mutations inhibiting phase separation of the Atg1 complex impaired PAS formation in vivo, suggesting that the entity of the PAS is a liquid droplet formed by phase separation of the Atg1 complex. Further in vitro analyses revealed that phospho-regulation of Atg13 by TORC1 and Ptc2 regulate the phase separation of the Atg1 complex, thereby regulating the PAS formation. Finally, using synthetic liposomes and protein droplets, we reconstituted the PAS attached to the vacuole in vitro. These data suggest that the PAS is a liquid droplet formed on the vacuole, which will function as a place for autophagosome formation by concentrating various Atg proteins and membranes.

Academic Significance and Societal Importance of the Research Achievements

細胞内の液-液相分離は様々な生命現象への関与が示唆されている新しい概念であるが、それがオートファジーのマシーナリーの制御に働くという今回の発見は、オートファジーの基本的なメカニズム解明に多大な貢献をするばかりでなく、液-液相分離の分野においてもその関与の対象がこれまでの想定以上に大きいことを強く示唆する画期的な成果である。オートファジーは生体の恒常性維持の機構として重要な役割を担い、その異常は神経変性疾患などの重篤な疾病につながることが知られている。本研究成果は、相分離の制御を通した全く新しいオートファジー制御剤の開発の可能性を提唱するものであり、今後の応用研究が期待される。

Report

(4 results)
  • 2020 Annual Research Report   Final Research Report ( PDF )
  • 2019 Annual Research Report
  • 2018 Annual Research Report
  • Research Products

    (35 results)

All 2021 2020 2019 2018 Other

All Int'l Joint Research (2 results) Journal Article (18 results) (of which Int'l Joint Research: 3 results,  Peer Reviewed: 15 results,  Open Access: 7 results) Presentation (15 results) (of which Int'l Joint Research: 3 results,  Invited: 11 results)

  • [Int'l Joint Research] Max Planck Institute(ドイツ)

    • Related Report
      2019 Annual Research Report
  • [Int'l Joint Research] Max Planck Institute(ドイツ)

    • Related Report
      2018 Annual Research Report
  • [Journal Article] Biomolecular condensates in autophagy regulation.2021

    • Author(s)
      Fujioka Y, Noda NN
    • Journal Title

      Current Opinion in Cell Biology

      Volume: 69 Pages: 23-29

    • DOI

      10.1016/j.ceb.2020.12.011

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Structural catalog of core Atg proteins opens new era of autophagy research2021

    • Author(s)
      Matoba Kazuaki、Noda Nobuo N
    • Journal Title

      The Journal of Biochemistry

      Volume: - Issue: 5 Pages: 1-9

    • DOI

      10.1093/jb/mvab017

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Atg2 and Atg9: Intermembrane and interleaflet lipid transporters driving autophagy2021

    • Author(s)
      Noda Nobuo N.
    • Journal Title

      Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids

      Volume: 1866 Issue: 8 Pages: 158956-158956

    • DOI

      10.1016/j.bbalip.2021.158956

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Liquid-liquid phase separation in autophagy2020

    • Author(s)
      Noda Nobuo N.、Wang Zheng、Zhang Hong
    • Journal Title

      Journal of Cell Biology

      Volume: 219 Issue: 8

    • DOI

      10.1083/jcb.202004062

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] In vitro reconstitution of autophagic processes2020

    • Author(s)
      Alam Jahangir Md.、Noda Nobuo N.
    • Journal Title

      Biochemical Society Transactions

      Volume: 48 Issue: 5 Pages: 2003-2014

    • DOI

      10.1042/bst20200130

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Atg9 is a lipid scramblase that mediates autophagosomal membrane expansion2020

    • Author(s)
      Matoba Kazuaki、Kotani Tetsuya、Tsutsumi Akihisa、Tsuji Takuma、Mori Takaharu、Noshiro Daisuke、Sugita Yuji、Nomura Norimichi、Iwata So、Ohsumi Yoshinori、Fujimoto Toyoshi、Nakatogawa Hitoshi、Kikkawa Masahide、Noda Nobuo N.
    • Journal Title

      Nature Structural & Molecular Biology

      Volume: 27 Issue: 12 Pages: 1185-1193

    • DOI

      10.1038/s41594-020-00518-w

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Secret of Atg9: lipid scramblase activity drives de novo autophagosome biogenesis2020

    • Author(s)
      Matoba Kazuaki、Noda Nobuo N.
    • Journal Title

      Cell Death & Differentiation

      Volume: 27 Issue: 12 Pages: 3386-3388

    • DOI

      10.1038/s41418-020-00663-1

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Formation of Autophagy Initiation Complex Mediated by an Intrinsically Disordered Protein2020

    • Author(s)
      FUJIOKA Yuko、NODA Nobuo N.
    • Journal Title

      Seibutsu Butsuri

      Volume: 60 Issue: 3 Pages: 171-173

    • DOI

      10.2142/biophys.60.171

    • NAID

      130007847786

    • ISSN
      0582-4052, 1347-4219
    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] 液-液相分離によるオートファゴソームの形成部位の構築2020

    • Author(s)
      藤岡優子,野田展生
    • Journal Title

      実験医学

      Volume: 38 Pages: 1354-1357

    • Related Report
      2020 Annual Research Report
  • [Journal Article] Phase separation organizes the site of autophagosome formation2020

    • Author(s)
      Fujioka Yuko、Alam Jahangir Md.、Noshiro Daisuke、Mouri Kazunari、Ando Toshio、Okada Yasushi、May Alexander I.、Knorr Roland L.、Suzuki Kuninori、Ohsumi Yoshinori、Noda Nobuo N.
    • Journal Title

      Nature

      Volume: 578 Issue: 7794 Pages: 301-305

    • DOI

      10.1038/s41586-020-1977-6

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Liquidity Is a Critical Determinant for Selective Autophagy of Protein Condensates2020

    • Author(s)
      Yamasaki Akinori、Alam Jahangir Md.、Noshiro Daisuke、Hirata Eri、Fujioka Yuko、Suzuki Kuninori、Ohsumi Yoshinori、Noda Nobuo N.
    • Journal Title

      Molecular Cell

      Volume: 77 Issue: 6 Pages: 1163-1175

    • DOI

      10.1016/j.molcel.2019.12.026

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed
  • [Journal Article] 柔らかい構造の可視化-LLPSと膜動態を例に2020

    • Author(s)
      能代大輔、野田展生
    • Journal Title

      実験医学

      Volume: 38 Pages: 84-89

    • Related Report
      2019 Annual Research Report
  • [Journal Article] 選択的オートファジーの構造生物学的基盤2020

    • Author(s)
      野田展生
    • Journal Title

      医学のあゆみ

      Volume: 272 Pages: 769-775

    • Related Report
      2019 Annual Research Report
  • [Journal Article] Human ATG2B possesses a lipid transfer activity which is accelerated by negatively charged lipids and WIPI42019

    • Author(s)
      Osawa Takuo、Ishii Yuki、Noda Nobuo N.
    • Journal Title

      Genes to Cells

      Volume: 25 Issue: 1 Pages: 65-70

    • DOI

      10.1111/gtc.12733

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Atg2: A novel phospholipid transfer protein that mediates de novo autophagosome biogenesis2019

    • Author(s)
      Osawa T, Noda NN
    • Journal Title

      Protein Science

      Volume: 28 Issue: 6 Pages: 1005-1012

    • DOI

      10.1002/pro.3623

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Atg2 mediates direct lipid transfer between membranes for autophagosome formation.2019

    • Author(s)
      Takuo Osawa, Tetsuya Kotani, Tatsuya Kawaoka, Eri Hirata, Kuninori Suzuki, Hitoshi Nakatogawa, Yoshinori Ohsumi, Nobuo N. Noda
    • Journal Title

      Nature Structural and Molecular Biology

      Volume: 26 Issue: 4 Pages: 281-288

    • DOI

      10.1038/s41594-019-0203-4

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Membrane-binding domains in autophagy2019

    • Author(s)
      Osawa T, Alam JM, Noda NN
    • Journal Title

      Chemistry and Physics of Lipids

      Volume: 218 Pages: 1-9

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Lipidation-independent vacuolar functions of Atg8 rely on its noncanonical interaction with a vacuole membrane protein2018

    • Author(s)
      Liu Xiao-Man、Yamasaki Akinori、Du Xiao-Min、Coffman Valerie C、Ohsumi Yoshinori、Nakatogawa Hitoshi、Wu Jian-Qiu、Noda Nobuo N、Du Li-Lin
    • Journal Title

      eLife

      Volume: 7

    • DOI

      10.7554/elife.41237

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Presentation] 多様な構造生物学を駆使したオートファジー研究2021

    • Author(s)
      野田展生
    • Organizer
      基礎から学ぶ最新NMR 解析法 第2回ワークショップ
    • Related Report
      2020 Annual Research Report
    • Invited
  • [Presentation] Autophagy regulation by liquid-liquid phase separation.2020

    • Author(s)
      Nobuo N. Noda
    • Organizer
      4th NanoLSI Symposium
    • Related Report
      2020 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] 液-液相分離によるオートファジーの始動制御2020

    • Author(s)
      野田展生
    • Organizer
      第72回 日本細胞生物学会大会
    • Related Report
      2020 Annual Research Report
    • Invited
  • [Presentation] 相分離したタンパク質の選択的オートファジーの試験管内再構成2019

    • Author(s)
      野田展生
    • Organizer
      第19回蛋白質科学会年会・第71回日本細胞生物学会大会 合同年次大会
    • Related Report
      2019 Annual Research Report
    • Invited
  • [Presentation] 液-液相分離とオートファジー2019

    • Author(s)
      野田展生
    • Organizer
      第3回LLPS研究会/大阪大学蛋白質研究所セミナー
    • Related Report
      2019 Annual Research Report
    • Invited
  • [Presentation] 液-液相分離によるオートファジーの始動制御2019

    • Author(s)
      野田展生
    • Organizer
      第92回 日本生化学会大会
    • Related Report
      2019 Annual Research Report
    • Invited
  • [Presentation] Mechanisms of autophagosomal membrane expansion and shaping2019

    • Author(s)
      Nobuo N. Noda
    • Organizer
      9th International Symposium on Autophagy
    • Related Report
      2019 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] オートファジー関連液滴と脂質膜の相互作用基盤2019

    • Author(s)
      野田展生
    • Organizer
      第42回 日本分子生物学会年会
    • Related Report
      2019 Annual Research Report
    • Invited
  • [Presentation] Molecular mechanisms of initial steps of autophagy2018

    • Author(s)
      Nobuo N. Noda
    • Organizer
      The 24th IUBMB Congress&15th FAOBMB Congress
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] 再構成生物学から迫るオートファジーの分子機構2018

    • Author(s)
      野田 展生
    • Organizer
      第29回新薬創製談話会
    • Related Report
      2018 Annual Research Report
    • Invited
  • [Presentation] 液-液相分離したタンパク質の選択的オートファジーの分子機構2018

    • Author(s)
      野田 展生
    • Organizer
      第91回日本生化学会大会
    • Related Report
      2018 Annual Research Report
    • Invited
  • [Presentation] オートファジーの足場タンパク質Atg17-Atg29-Atg31複合体の高速AFM観察2018

    • Author(s)
      能代大輔、藤岡優子、野田展生、安藤敏夫
    • Organizer
      第91回日本生化学会大会
    • Related Report
      2018 Annual Research Report
  • [Presentation] PASは液-液相分離で形成される非膜型オルガネラの一種である2018

    • Author(s)
      野田 展生
    • Organizer
      第11回オートファジー研究会
    • Related Report
      2018 Annual Research Report
  • [Presentation] 高速AFMによるAtg17-Atg29-Atg31のS字構造とAtg13-Atg17-Atg29-Atg31複合体により形成される液滴の観察2018

    • Author(s)
      能代大輔、藤岡優子、安藤敏夫、野田展生
    • Organizer
      第11回オートファジー研究会
    • Related Report
      2018 Annual Research Report
  • [Presentation] オートファゴソーム形成場 PASの液-液相分離を介した構築原理2018

    • Author(s)
      藤岡 優子,Jahangir Md. Alam,野田 展生
    • Organizer
      第41回日本分子生物学会年会
    • Related Report
      2018 Annual Research Report

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Published: 2018-04-23   Modified: 2022-01-27  

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