• Search Research Projects
  • Search Researchers
  • How to Use
  1. Back to previous page

Screening of autophagosomal outer membrane proteins required for membrane fusion

Research Project

Project/Area Number 16K14720
Research Category

Grant-in-Aid for Challenging Exploratory Research

Allocation TypeMulti-year Fund
Research Field Cell biology
Research InstitutionThe University of Tokyo

Principal Investigator

Yamamoto Hayashi  東京大学, 大学院医学系研究科(医学部), 講師 (80551283)

Co-Investigator(Renkei-kenkyūsha) MIZUSHIMA Noboru  東京大学, 大学院医学系研究科, 教授 (10353434)
Research Collaborator UEMATSU Masaaki  東京大学, 大学院医学系研究科, 大学院生
Project Period (FY) 2016-04-01 – 2018-03-31
Project Status Completed (Fiscal Year 2017)
Budget Amount *help
¥3,770,000 (Direct Cost: ¥2,900,000、Indirect Cost: ¥870,000)
Fiscal Year 2017: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2016: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Keywordsオートファジー / 膜融合 / SNARE / プロテオーム / 膜動態
Outline of Final Research Achievements

Autophagy is a fundamental degradation system conserved in eukaryotes. Upon induction of autophagy, a double-membrane structure, called an autophagosome, is generated and fuses with lysosomes to degrade its contents. Although many ATG proteins have been identified, it remains unclear how autophagosome-lysosome fusion is regulated. In this study, we tried to develop a biochemical method to purify autophagosomes and to identify autophagosomal outer membrane proteins involved in the membrane fusion. For this purpose, we prepared GFP-STX17DN cells to accumulate autophagosomes, harvested an autophagosome-enriched fraction by OptiPrep flotation, and purified autophagosomes using 3xFLAG-LC3. Finally, outer membrane proteins were labeled by a membrane-impermeable biotinylation reagent. By mass spectrometry of the biotinylated proteins, we obtained several candidates of outer membrane proteins. We prepared KO cells (CRISPR) or KD cells (siRNA), however, significant phenotypes were not observed.

Report

(3 results)
  • 2017 Annual Research Report   Final Research Report ( PDF )
  • 2016 Research-status Report
  • Research Products

    (6 results)

All 2017 2016

All Journal Article (4 results) (of which Int'l Joint Research: 1 results,  Peer Reviewed: 4 results,  Open Access: 2 results,  Acknowledgement Compliant: 1 results) Presentation (2 results) (of which Invited: 1 results)

  • [Journal Article] Differential requirement for ATG2A domains for localization to autophagic membranes and lipid droplets2017

    • Author(s)
      Tamura Norito、Nishimura Taki、Sakamaki Yuriko、Koyama-Honda Ikuko、Yamamoto Hayashi、Mizushima Noboru
    • Journal Title

      FEBS Letters

      Volume: 591 Issue: 23 Pages: 3819-3830

    • DOI

      10.1002/1873-3468.12901

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Accumulation of undegraded autophagosomes by expression of dominant-negative STX17 (syntaxin 17) mutants2017

    • Author(s)
      Uematsu Masaaki、Nishimura Taki、Sakamaki Yuriko、Yamamoto Hayashi、Mizushima Noboru
    • Journal Title

      Autophagy

      Volume: 13 Issue: 8 Pages: 1452-1464

    • DOI

      10.1080/15548627.2017.1327940

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Autophagosome formation is initiated at phosphatidylinositol synthase-enriched ER subdomains2017

    • Author(s)
      Nishimura, T., Tamura, N., Kono, N., Shimanaka, Y., Arai, H., Yamamoto, H., Mizushima, N.
    • Journal Title

      The EMBO Journal

      Volume: 36 Issue: 12 Pages: 1719

    • DOI

      10.15252/embj.201695189

    • Related Report
      2017 Annual Research Report 2016 Research-status Report
    • Peer Reviewed
  • [Journal Article] The Intrinsically Disordered Protein Atg13 Mediates Supramolecular Assembly of Autophagy Initiation Complexes.2016

    • Author(s)
      Yamamoto H, Fujioka Y, Suzuki SW, Noshiro D, Suzuki H, Kondo-Kakuta C, Kimura Y, Hirano H, Ando T, Noda NN, Ohsumi Y
    • Journal Title

      Developmental Cell

      Volume: 38 Issue: 1 Pages: 86-99

    • DOI

      10.1016/j.devcel.2016.06.015

    • Related Report
      2016 Research-status Report
    • Peer Reviewed / Open Access / Acknowledgement Compliant
  • [Presentation] オートファジータンパク質群の動的相互作用と分子集合形態の解析2016

    • Author(s)
      山本 林
    • Organizer
      第54回 日本生物物理学会年会
    • Place of Presentation
      つくば国際会議場(筑波)
    • Year and Date
      2016-11-25
    • Related Report
      2016 Research-status Report
    • Invited
  • [Presentation] 天然変性タンパク質Atg13によるオートファジー始動複合体の高次集積機構の解明2016

    • Author(s)
      山本 林,藤岡 優子,鈴木 翔,野田 展生,大隅 良典
    • Organizer
      第89回 日本生化学会大会
    • Place of Presentation
      仙台国際センター(仙台)
    • Year and Date
      2016-09-25
    • Related Report
      2016 Research-status Report

URL: 

Published: 2016-04-21   Modified: 2019-03-29  

Information User Guide FAQ News Terms of Use Attribution of KAKENHI

Powered by NII kakenhi