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シナプスにおける時・空間近接分子群の同定手法の開発

Research Project

Project/Area Number 19K22439
Research Category

Grant-in-Aid for Challenging Research (Exploratory)

Allocation TypeMulti-year Fund
Review Section Medium-sized Section 44:Biology at cellular to organismal levels, and related fields
Research InstitutionNational Institute for Physiological Sciences

Principal Investigator

深田 優子  生理学研究所, 分子細胞生理研究領域, 准教授 (40416186)

Project Period (FY) 2019-06-28 – 2022-03-31
Project Status Granted (Fiscal Year 2020)
Budget Amount *help
¥6,500,000 (Direct Cost: ¥5,000,000、Indirect Cost: ¥1,500,000)
Fiscal Year 2020: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
Fiscal Year 2019: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
Keywordsプロテオミクス
Outline of Research at the Start

生体蛋白質は時々刻々と多様な分子群と相互作用することにより、成熟、輸送を経て適所に局在化することで、生理機能を発揮する。これらいずれの過程が障害されても蛋白質はその機能を果たせず、時には疾患の要因となる。本研究では、超解像プロテオミクス法の開発に挑戦する。これまで見過されてきた蛋白質相互作用を明らかにして、シナプス制御機構やシナプス病態機構の理解につながる成果を得ることを目指す。

Outline of Annual Research Achievements

本研究では、生体タンパク質、とりわけ疾患関連タンパク質の相互作用分子を高い時・空間的解像度で同定するために、(1)不溶性が高い細胞膜に起こるタンパク質相互作用、(2)酵素と基質間結合のような一過的なタンパク質相互作用や(3)細胞種特異的なタンパク質相互作用の同定・解析手法を確立することを目指す。具体的には、てんかん関連リガンド-受容体や脂質修飾関連酵素をモデルタンパク質として、超解像プロテオミクス法の開発に挑戦する。2020年度は、2019年度に構築した実験条件とリソースを活用し、シナプス不溶性画分からタンパク質複合体の精製と構成分子の同定を進めた。具体的には、ADAM22を含むシナプスタンパク質複合体を網羅的、特異的、定量的に同定した。タグを挿入したノックインマウスを作製し、野生型マウス脳をネガティブコントロールとすることで特異性を担保した。シナプス画分のADAM22複合体構成タンパク質には、NMDA受容体、AMPA受容体等の受容体、イオンチャネルやLRRTM4-Neurexinといった接着分子などが含まれていた。また、これら膜タンパク質との結合はPSD-95を代表とするMAGUK足場タンパク質を介することを明らかにした(Fukata et al, PNAS 2021)。また、酵素と基質間結合についても、ある脂質修飾関連酵素に関して、実験リソースの整備と精製を終え、新たな酵素-基質ペアの候補を得た。さらに、シナプスタンパク質複合体の同定技術を初代培養神経細胞に応用し、細胞種特異的な相互作用分子の同定のための実験を開始した。

Current Status of Research Progress
Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

(1) シナプス不溶性画分からADAM22のシナプスタンパク質複合体を精製、同定し、論文発表を行った(Fukata et al, PNAS 2021)。
(2) 脂質修飾関連酵素の相互作用分子を探索し、候補分子を得た。
(3) 神経細胞種特異的なシナプスタンパク質複合体の同定に向けた実験を推進した。

Strategy for Future Research Activity

(1) 脂質修飾関連酵素の相互作用分子の機能解析を進める。
(2) 神経細胞種特異的なシナプスタンパク質複合体を同定する。

Report

(2 results)
  • 2020 Research-status Report
  • 2019 Research-status Report

Research Products

(31 results)

All 2021 2020 2019 Other

All Int'l Joint Research (11 results) Journal Article (13 results) (of which Int'l Joint Research: 9 results,  Peer Reviewed: 11 results,  Open Access: 9 results) Presentation (4 results) (of which Int'l Joint Research: 2 results) Book (1 results) Remarks (2 results)

  • [Int'l Joint Research] UCSF/UCSD(米国)

    • Related Report
      2020 Research-status Report
  • [Int'l Joint Research] DZNE(ドイツ)

    • Related Report
      2020 Research-status Report
  • [Int'l Joint Research] IST(オーストリア)

    • Related Report
      2020 Research-status Report
  • [Int'l Joint Research] UCL/The University of Edinburgh(英国)

    • Related Report
      2020 Research-status Report
  • [Int'l Joint Research] University of Helsinki(フィンランド)

    • Related Report
      2020 Research-status Report
  • [Int'l Joint Research] UCSF(米国)

    • Related Report
      2019 Research-status Report
  • [Int'l Joint Research] Institut Curie(フランス)

    • Related Report
      2019 Research-status Report
  • [Int'l Joint Research] DZNE(ドイツ)

    • Related Report
      2019 Research-status Report
  • [Int'l Joint Research] UCL Institute of Neurology(英国)

    • Related Report
      2019 Research-status Report
  • [Int'l Joint Research] University of Helsinki(フィンランド)

    • Related Report
      2019 Research-status Report
  • [Int'l Joint Research]

    • Related Report
      2019 Research-status Report
  • [Journal Article] LGI1-ADAM22-MAGUK configures transsynaptic nanoalignment for synaptic transmission and epilepsy prevention2021

    • Author(s)
      Fukata Yuko、Chen Xiumin、Chiken Satomi、Hirano Yoko、Yamagata Atsushi、Inahashi Hiroki、Sanbo Makoto、Sano Hiromi、Goto Teppei、Hirabayashi Masumi、Kornau Hans-Christian、Pruss Harald、Nambu Atsushi、Fukai Shuya、Nicoll Roger A.、Fukata Masaki
    • Journal Title

      Proceedings of the National Academy of Sciences

      Volume: 118

    • DOI

      10.1073/pnas.2022580118

    • NAID

      120006976050

    • Related Report
      2020 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Canonical versus non-canonical transsynaptic signaling of neuroligin 3 tunes development of sociality in mice2021

    • Author(s)
      Yoshida T, Yamagata A, Imai A, Kim J, Izumi H, Maenaka K, Shimada T, Fukata M, Nishijo H, Takao K, Okabe S, Tabuchi K, Mishina M, Mori H, Fukai S.
    • Journal Title

      Nature Communications

      Volume: 12(1) Issue: 1 Pages: 1848-1848

    • DOI

      10.1038/s41467-021-22059-6

    • NAID

      120006996978

    • Related Report
      2020 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Encephalitis patient derived monoclonal GABAA receptor antibodies cause catatonia and epileptic seizures2021

    • Author(s)
      Kreye Jakob、Wright Sukhvir K.、van Casteren Adriana、Machule Marie-Luise、Reincke S. Momsen、Nikolaus Marc、Stoffler Laura、van Hoof Scott、Sanchez-Sendin Elisa、Kornau Hans-Christian、Kaindl Angela M.、Wilson Max A.、Greenhill Stuart、他5名、Geis Christian、Fukata Yuko、Fukata Masaki、Pruss Harald
    • Journal Title

      bioRxiv

      Volume: -

    • DOI

      10.1101/2021.01.28.428602

    • Related Report
      2020 Research-status Report
    • Open Access / Int'l Joint Research
  • [Journal Article] APEGS法によるタンパク質パルミトイル化修飾の定量2021

    • Author(s)
      深田正紀、横井紀彦、深田優子
    • Journal Title

      電気泳動

      Volume: 65

    • NAID

      130008072916

    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] Forelimb movements evoked by optogenetic stimulation of the macaque motor cortex2020

    • Author(s)
      Watanabe H, Sano H, Chiken S, Kobayashi K, Fukata Y, Fukata M, Mushiake H, Nambu A
    • Journal Title

      Nature Commun

      Volume: 11 Pages: 3253-3253

    • DOI

      10.1038/s41467-020-16883-5

    • Related Report
      2020 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] A genetically encoded red fluorescence dopamine biosensor enables dual imaging of dopamine and norepinephrine2020

    • Author(s)
      Nakamoto Chihiro、Goto Yuhei、Tomizawa Yoko、Fukata Yuko、Fukata Masaki、Harpsoe Kasper、Gloriam David E.、Aoki Kazuhiro、Takeuchi Tomonori
    • Journal Title

      bioRxiv

      Volume: -

    • DOI

      10.1101/2020.05.25.115162

    • Related Report
      2020 Research-status Report
    • Open Access / Int'l Joint Research
  • [Journal Article] The extracellular domain of angulin-1 and palmitoylation of its cytoplasmic region are required for angulin-1 assembly at tricellular contacts2020

    • Author(s)
      Yukako Oda, Taichi Sugawara, Yuko Fukata, Yasushi Izumi, Tetsuhisa Otani, Tomohito Higashi, Masaki Fukata, Mikio Furuse
    • Journal Title

      Journal of Biological Chemistry

      Volume: 295 Issue: 13 Pages: 4289-4302

    • DOI

      10.1074/jbc.ra119.010491

    • Related Report
      2019 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Human Cerebrospinal Fluid Monoclonal LGI1 Autoantibodies Increase Neuronal Excitability2020

    • Author(s)
      Kornau Hans‐Christian、Kreye Jakob、Stumpf Alexander、Fukata Yuko、Parthier Daniel、Sammons Rosanna P.、Imbrosci Barbara、Kurpjuweit Sarah、Kowski Alexander B.、Fukata Masaki、Pruss Harald、Schmitz Dietmar
    • Journal Title

      Annals of Neurology

      Volume: 87 Pages: 405-418

    • DOI

      10.1002/ana.25666

    • Related Report
      2019 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Dynamic palmitoylation controls the microdomain localization of the DKK1 receptors CKAP4 and LRP62019

    • Author(s)
      Sada Ryota、Kimura Hirokazu、Fukata Yuko、Fukata Masaki、Yamamoto Hideki、Kikuchi Akira
    • Journal Title

      Science Signaling

      Volume: 12 Pages: 9519-9519

    • DOI

      10.1126/scisignal.aat9519

    • Related Report
      2019 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] ABHD10 is an S-depalmitoylase affecting redox homeostasis through peroxiredoxin-52019

    • Author(s)
      Cao Yang、Qiu Tian、Kathayat Rahul S.、Azizi Saara-Anne、Thorne Anneke K.、Ahn Daniel、Fukata Yuko、Fukata Masaki、Rice Phoebe A.、Dickinson Bryan C.
    • Journal Title

      Nature Chemical Biology

      Volume: 15 Pages: 1232-1240

    • DOI

      10.1038/s41589-019-0399-y

    • Related Report
      2019 Research-status Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Targeting CCR5 trafficking to inhibit HIV-1 infection2019

    • Author(s)
      Boncompain Gaelle、Herit Floriane、Tessier Sarah、Lescure Aurianne、Del Nery Elaine、Gestraud Pierre、Staropoli Isabelle、Fukata Yuko、Fukata Masaki、Brelot Anne、Niedergang Florence、Perez Franck
    • Journal Title

      Science Advances

      Volume: 5

    • DOI

      10.1126/sciadv.aax0821

    • Related Report
      2019 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Systematic Screening of Depalmitoylating Enzymes and Evaluation of Their Activities by the Acyl-PEGyl Exchange Gel-Shift (APEGS) Assay2019

    • Author(s)
      Kanadome T, Yokoi N, Fukata Y, Fukata M.
    • Journal Title

      Methods in Molecular Biology

      Volume: 印刷中 Pages: 83-98

    • DOI

      10.1007/978-1-4939-9532-5_7

    • Related Report
      2019 Research-status Report
    • Peer Reviewed
  • [Journal Article] Limbic Encephalitis in a Cat with Hippocampal Necrosis, putatively associated with DCC (deleted in colorectal cancer; Netrin-1 Receptor) Antibodies2019

    • Author(s)
      Hasegawa D, Ohnishi Y, Koyama E, Matsunaga S, Ohtani S, Nakanishi A, Shiga T, Chambers JK, Uchida K, Yokoi N, Fukata Y, Fukata M.
    • Journal Title

      Journal of Veterinary Internal Medicine

      Volume: 印刷中 Pages: 1440-1445

    • DOI

      10.1111/jvim.15492

    • Related Report
      2019 Research-status Report
    • Peer Reviewed
  • [Presentation] Acyl-PEGyl exchange gel-shift (APEGS) assay for palmitoylation quantification2020

    • Author(s)
      Fukata Masaki、Yokoi Norihiko、Fukata Yuko
    • Organizer
      第71回 日本電気泳動学会総会
    • Related Report
      2020 Research-status Report
  • [Presentation] Synaptic function regulated by palmitate cycling on PSD-95 and trans-synaptic LGI1 and ADAM222020

    • Author(s)
      Fukata Masaki、Yuri Miyazaki、Yokoi Norihiko、Fukata Yuko
    • Organizer
      第43回日本分子生物学会年会
    • Related Report
      2020 Research-status Report
  • [Presentation] Physiological roles of trans-synaptic LGI1-ADAM22-MAGUK complex2019

    • Author(s)
      Fukata Yuko、Hirano Yoko、Inahashi Hiroki、Miyazaki Yuri、Yokoi Norihiko、Sanbo Makoto、Goto Teppei、Hirabayashi Masumi、Fukata Masaki
    • Organizer
      Neuroscience Meeting SfN 2019
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research
  • [Presentation] Functions of trans-synaptic protein complex of epilepsy-related ligand-receptor, LGI1 and ADAM222019

    • Author(s)
      Fukata Yuko
    • Organizer
      McGill-NIPS Joint Workshop
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research
  • [Book] 膜タンパク質工学ハンドブック2020

    • Author(s)
      平野瑶子、宮﨑裕理、横井紀彦、平田哲也、深田正紀、深田優子
    • Total Pages
      17
    • Publisher
      エヌ・ティー・エス
    • ISBN
      9784860435370
    • Related Report
      2020 Research-status Report
  • [Remarks] 研究室ホームページ

    • URL

      https://www.nips.ac.jp/fukata/

    • Related Report
      2020 Research-status Report
  • [Remarks] プレスリリース:神経シナプス間の軸調整を担う分子群を発見―新たなてんかん病態を解明―

    • URL

      https://www.nips.ac.jp/release/2021/01/post_426.html

    • Related Report
      2020 Research-status Report

URL: 

Published: 2019-07-04   Modified: 2021-12-27  

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