Elucidation of the molecular mechanism for accurately and efficiently retrograde cargo transport from the Golgi to the ER

• Project/Area Number: 20K06650
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 44010:Cell biology-related
• Research Institution: Institute of Physical and Chemical Research
• Principal Investigator: Kurokawa Kazuo 国立研究開発法人理化学研究所, 光量子工学研究センター, 専任研究員 (40333504)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Project Status: Completed (Fiscal Year 2022)
• Budget Amount: ¥4,420,000 (Direct Cost: ¥3,400,000、Indirect Cost: ¥1,020,000); Fiscal Year 2022: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2021: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000); Fiscal Year 2020: ¥2,340,000 (Direct Cost: ¥1,800,000、Indirect Cost: ¥540,000)
• Keywords: ゴルジ体 / 4Dライブイメージング / 小胞体 / 積荷受容体 / 逆行輸送 / COPI / ゴルジ体シス槽 / 多色4次元イメージング / 積荷タンパク質

Outline of Research at the Start

真核細胞には、微小な膜構造により積荷となるタンパク質を目的地へと輸送する機構（膜交通）がある。そこには、新たに合成された積荷を目的地へと順行輸送する機構とともに、誤って運ばれた積荷や、ゴルジ体で働く酵素を絶えず本来働くべき場所へと戻す逆行輸送の機構も存在する。ではどのように多様なタンパク質を見分けて、順行および逆行輸送するのであろうか。本研究では、小胞体ゴルジ体間で順行・逆行輸送を制御している積荷受容体に焦点を当て、多色同時高時空間分解能4Dイメージングと出芽酵母の分子機能変異株の利用を組み合わせたアプローチで、タンパク質を正確に逆行輸送する制御機構の解明を目指している。

Outline of Final Research Achievements

To elucidate the mechanism of retrograde transport from the Golgi to the ER, I performed simultaneous multicolor 4D live imaging analysis of the cargo receptor Emp24, cis-Golgi, and ERES. Emp24 showed quick anterograde transport to cis-Golgi during the contact between cis-Golgi and the ERES. During this contact, Emp24 was then retrogradely transported to the ERES. It was shown that both anterograde and retrograde transport of the cargo receptor is achieved during the contact of cis-Golgi with the ERES (hug-and-kiss action).

Academic Significance and Societal Importance of the Research Achievements

本研究課題では、生きた細胞内の複数の分子の4D動態を捉える多色同時高時空間分解能4Dイメージングと出芽酵母の自由度の高い遺伝学とを組み合わせた研究手法を進めることで、膜交通の新たなモデルを提案する結果を得ることができた。

Research Products

• [Int'l Joint Research] University of Seville/Institute de Biomedica de Seville(スペイン)
• [Int'l Joint Research] Institute of Protein Biochemistry(イタリア)
• [Int'l Joint Research] University of Geneva/University of Fribourg(スイス)
• [Int'l Joint Research] University of Seville/Instituto de Biomedicina de Sevilla(スペイン)
• [Journal Article] Dissociation of ERMES clusters plays a key role in attenuating the endoplasmic reticulum stress (2022)
  • Author(s): Kakimoto-Takeda Yuriko、Kojima Rieko、Shiino Hiroya、Shinmyo Manatsu、Kurokawa Kazuo、Nakano Akihiko、Endo Toshiya、Tamura Yasushi
  • Journal Title: iScience Volume: 25 Issue: 11 Pages: 105362-105362
  • DOI: 10.1016/j.isci.2022.105362
  • Peer Reviewed / Open Access
• [Journal Article] Quality-controlled ceramide-based GPI-anchored protein sorting into selective ER exit sites (2022)
  • Author(s): Rodriguez-Gallardo Sofia、Sabido-Bozo Susana、Ikeda Atsuko、Araki Misako、Okazaki Kouta、Nakano Miyako、Aguilera-Romero Auxiliadora、Cortes-Gomez Alejandro、Lopez Sergio、Waga Miho、Nakano Akihiko、Kurokawa Kazuo、Muniz Manuel、Funato Kouichi
  • Journal Title: Cell Reports Volume: 39 Issue: 5 Pages: 110768-110768
  • DOI: 10.1016/j.celrep.2022.110768
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Assay for dual cargo sorting into endoplasmic reticulum exit sites imaged by 3D Super-resolution Confocal Live Imaging Microscopy (SCLIM) (2021)
  • Author(s): Rodriguez-Gallardo Sofia、Kurokawa Kazuo、Sabido-Bozo Susana、Cortes-Gomez Alejandro、Perez-Linero Ana Maria、Aguilera-Romero Auxiliadora、Lopez Sergio、Waga Miho、Nakano Akihiko、Muniz Manuel
  • Journal Title: PLOS ONE Volume: 16 Issue: 10 Pages: e0258111-e0258111
  • DOI: 10.1371/journal.pone.0258111
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Golgi maturation-dependent glycoenzyme recycling controls glycosphingolipid biosynthesis and cell growth via GOLPH3. (2021)
  • Author(s): Riccardo Rizzo, Domenico Russo, Kazuo Kurokawa, Alfredo Budillon, Franck Perez, Seetharaman Parashuraman, Yusuf A Hannun, Akihiko Nakano, Daniela Corda, Giovanni D’Angelo and Alberto Luini et al.,.
  • Journal Title: The EMBO Journal Volume: 40 Issue: 8
  • DOI: 10.15252/embj.2020107238
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Cargo sorting zones in the trans-Golgi network visualized by super-resolution confocal live imaging microscopy in plants (2021)
  • Author(s): Shimizu Yutaro、Takagi Junpei、Ito Emi、Ito Yoko、Ebine Kazuo、Komatsu Yamato、Goto Yumi、Sato Mayuko、Toyooka Kiminori、Ueda Takashi、Kurokawa Kazuo、Uemura Tomohiro、Nakano Akihiko
  • Journal Title: Nature Communications Volume: 12 Issue: 1 Pages: 1901-1901
  • DOI: 10.1038/s41467-021-22267-0
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] 超解像ライブイメージング顕微鏡SCLIMの開発とSCLIMによるゴルジ体タンパク質輸送機構の解明 (2021)
  • Author(s): 黒川量雄, 宮代大輔, 中野明彦
  • Journal Title: レーザー研究 Volume: 49
  • Peer Reviewed
• [Journal Article] Tricalbins are required for nonvesicular ceramide transport at ER-Golgi contacts and modulate lipid droplet biogenesis. (2020)
  • Author(s): Atsuko Ikeda, Philipp Schlarmann, Kazuo Kurokawa, Akihiko Nakano, Howard Riezman, and Kouichi Funato.
  • Journal Title: iScience Volume: 23 Issue: 10 Pages: 101603-101603
  • DOI: 10.1016/j.isci.2020.101603
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Ceramide chain length-dependent protein sorting into selective endoplasmic reticulum exit sites. (2020)
  • Author(s): Sofia Rodriguez-Gallardo, Kazuo Kurokawa, Susana Sabido-Bozo, Alejandro Cortes-Gomez, Atsuko Ikeda, Valeria Zoni, Auxiliadora Aguilera-Romero, Ana Maria Perez-Linero, Sergio Lopez, Miho Waga, Misako Araki, Miyako Nakano, Howard Riezman, Kouichi Funato, Stefano Vanni, Akihiko Nakano, Manuel Muñiz.
  • Journal Title: Science Advances Volume: 6 Issue: 50
  • DOI: 10.1126/sciadv.aba8237
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Recycling endosomes attach to the trans-side of Golgi stacks in Drosophila and mammalian cells (2020)
  • Author(s): Fujii Syara、Kurokawa Kazuo、Inaba Ryota、Hiramatsu Naoki、Tago Tatsuya、Nakamura Yuri、Nakano Akihiko、Satoh Takunori、Satoh Akiko K.
  • Journal Title: Journal of Cell Science Volume: 133 Issue: 4 Pages: 236935-236935
  • DOI: 10.1242/jcs.236935
  • Peer Reviewed
• [Journal Article] Sec71 separates Golgi stacks in Drosophila S2 cells. (2020)
  • Author(s): Syara Fujii, Kazuo Kurokawa, Tatsuya Tago, Ryota Inaba, Arata Takiguchi, Akihiko Nakano, *Takunori Satoh, and *Akiko K Satoh.
  • Journal Title: Journal of Cell Science Volume: 133 Issue: 24
  • DOI: 10.1242/jcs.245571
  • Peer Reviewed
• [Journal Article] Live-cell Imaging by Super-resolution Confocal Live Imaging Microscopy (SCLIM): Simultaneous Three-color and Four-dimensional Live Cell Imaging with High Space and Time Resolution. (2020)
  • Author(s): Kazuo Kurokawa and Akihiko Nakano
  • Journal Title: Bio-Protocol Volume: 10 Issue: 17
  • DOI: 10.21769/bioprotoc.3732
  • Peer Reviewed
• [Presentation] 4Dライブイメージングによる小胞体，ゴルジ体の積荷タンパク質輸送機構の解明 (2020)
  • Author(s): 黒川量雄
  • Organizer: 第93回日本生化学会大会
  • Invited
• [Presentation] ミトコンドリア- 小胞体間結合因子ERMES 複合体クラスターの解離がER ストレス軽減に寄与する (2020)
  • Author(s): 柿元百合子，小島理恵子，新名真夏，黒川量雄，中野明彦，遠藤斗志也，田村康
  • Organizer: 第72回日本細胞生物学会大会
• [Presentation] 小胞体における積荷タンパク質選別輸送機構 (2020)
  • Author(s): 黒川量雄，Sofia Gallardo，Manuel Muniz，中野明彦
  • Organizer: 第72回日本細胞生物学会大会
• [Presentation] 膜交通関連オルガネラの減数分裂に伴う再構成 (2020)
  • Author(s): 須田恭之，舘川宏之，黒川量雄，中野明彦，入江賢児
  • Organizer: 第72回日本細胞生物学会大会
• [Remarks] 理化学研究所・光量子工学研究センター・生細胞超解像イメージング研究チーム
  • URL: http://www.riken.jp/research/labs/rap/live_cell_superresolution_img/