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内在化αシヌクレインにおけるESCRTを中心とした小胞輸送に関する研究

Research Project

Project/Area Number 25461264
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeMulti-year Fund
Section一般
Research Field Neurology
Research InstitutionTohoku University

Principal Investigator

菅野 直人  東北大学, 大学病院, 助教 (30509550)

Co-Investigator(Kenkyū-buntansha) 武田 篤  東北大学, 医学(系)研究科(研究院), 大学院非常勤講師 (70261534)
長谷川 隆文  東北大学, 大学病院, 講師 (70361079)
Project Period (FY) 2013-04-01 – 2014-03-31
Project Status Discontinued (Fiscal Year 2013)
Budget Amount *help
¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000)
Fiscal Year 2015: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2014: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2013: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Keywordsalpha-synuclein / ESCRT / Nedd4 / 小胞輸送
Research Abstract

神経変性疾患における異常凝集タンパクの細胞間伝播が注目されているが、その機構に関しては未だ不明な点が多い。近年E3ユビキチンライゲースNedd4がαシヌクレイン(aS)に標的タンパクの細胞内移動に関わるK63ユビキチンを付加する事が報告された。Nedd4は膜結合ドメインを持つ事から、内在化し膜近傍に存在するaSの輸送制御への関連に着目し研究を行った。まず、aSをリコンビナントタンパクとして大腸菌で合成し、サイズクロマトグラフィーで<100kDaに精製。細胞内局在の顕微鏡下での可視化のために一部は蛍光色素標識した。実験細胞SH-SY5Yの培地にaSを5μMで添加すると、時間・濃度依存性に後期エンドソームへの蓄積がみられた。in vivoでNedd4はaSをUbcH5, H6存在下でK63ポリユビキチン化した。Nedd4過剰発現条件下ではaSのエンドソーム蓄積が促進し、一方、ユビキチン・リガーゼ欠損変異型(C867A)および基質結合部位欠損変異型(ΔWW)はエンドソーム蓄積に影響を与えなかった。siRNAによるCHMP2Bのノックダウンでエンドソームに蓄積するaSが抑制された事から、aSはESCRTによってエンドソームへターゲッティングされているものと考えられた。基質のプロリンを多く含む領域にNedd4が結合するため、C末端に存在すると予想される部位(PR)を欠損させたaSをリコンビナントタンパクとして精製。これらの変異型aSはNedd4によるユビキチン化が抑制され、またエンドソームへの内在化も抑制された。さらに、PR配列に含まれる2つのプロリンをアラニンに変換した変異体を作製し、うちP120A変異型はユビキチン化・エンドソーム輸送ともに抑制され、P120がaS内在化において重要な役割を持つものと考えられた。

Report

(1 results)
  • 2013 Annual Research Report
  • Research Products

    (16 results)

All 2014 2013

All Journal Article (10 results) (of which Peer Reviewed: 10 results,  Open Access: 1 results) Presentation (6 results)

  • [Journal Article] K63-linked ubiquitination by E3 ubiquitin ligase Nedd4-1 facilitates endosomal sequestration of internalized alpha-synuclein2014

    • Author(s)
      Sugeno N, Hasegawa T, Tanaka N, Fukuda M, Wakabayashi K, Oshima R, Konno M, Miura E, Kikuchi A, Baba T, Anan T, Nakao M, Geisler S, Aoki M, Takeda A.
    • Journal Title

      Journal of Biological Chemistry

      Volume: in press

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Retrospective analysis of Guillain-Barre syndrome and Fisher syndrome after the Great East Japan Earthquake2014

    • Author(s)
      Hirofumi Tsuboi, Naoto Sugeno, Maki Tateyama, Ichiro Nakashima, Takafumi Hasegawa, Hiroshi Kuroda, Kimihiko Kaneko, Michiko Kobayashi, Aya Ishigaki, Juichi Fujimori and Masashi Aoki
    • Journal Title

      Brain and Behavior

      Volume: in press

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Isolated posterior column lesion in a case with mixed connective tissue disease2014

    • Author(s)
      Tomomi Shijo, Naoto Sugeno, Hiroshi Kuroda, Takafumi Hasegawa, Atsushi Takeda and Masashi Aoki
    • Journal Title

      Neurology and Clinical Neuroscience

      Volume: in press Issue: 3 Pages: 95-95

    • DOI

      10.1111/ncn3.88

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Improvement of freezing of gait in patients with Parkinson’s disease by imagining bicycling.2014

    • Author(s)
      Kikuchi A, Baba T, Hasegawa T, Sugeno N, Konno M, Miura E, Oshima R, Aoki M, Takeda A.
    • Journal Title

      Case Rep Neurol

      Volume: 6 Issue: 1 Pages: 92-95

    • DOI

      10.1159/000362119

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Olfactory Dysfunction and Dementia in Parkinson's Disease.2014

    • Author(s)
      Takeda A, Baba T, Kikuchi A, Hasegawa T, Sugeno N, Konno M, Miura E, Mori E.
    • Journal Title

      J Parkinsons Dis.

      Volume: in press

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Postural leg tremor in X-linked spinal and bulbar muscular atrophy.2014

    • Author(s)
      Nishiyama A, Sugeno N, Tateyama M, Nishiyama S, Kato M, Aoki M.
    • Journal Title

      J Clin Neurosci.

      Volume: 21 Issue: 5 Pages: 799-802

    • DOI

      10.1016/j.jocn.2013.07.026

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Corticospinal tract signal changes in a patient with spinal and bulbar muscular atrophy2013

    • Author(s)
      Naoto Sugeno, Ayumu Sunaguchi, Ayumi Nishiyama, Masaaki Kato and Masashi Aoki
    • Journal Title

      Neurology and Clinical Neuroscience

      Volume: 1 Issue: 3 Pages: 125-125

    • DOI

      10.1111/j.2049-4173.2013.00025.x

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Choreoathetosis in a patient with neuromyelitis optica spectrum disorder2013

    • Author(s)
      Naoto Sugeno, Rumiko Izumi, Kazuhiro Kato, Ichiro Nakashima, Atsushi Takeda and Masashi Aoki
    • Journal Title

      Neurology and Clinical Neuroscience

      Volume: 1 Issue: 4 Pages: 154-156

    • DOI

      10.1111/ncn3.36

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Hereditary neuropathy with liability to pressure palsy emerging after hypothyroidism2013

    • Author(s)
      Kimihiko Kaneko, Naoto Sugeno, Maki Tateyama, Tatsuro Misu, Naoki Suzuki and Masashi Aoki
    • Journal Title

      Neurology and Clinical Neuroscience

      Volume: 1 Issue: 4 Pages: 160-161

    • DOI

      10.1111/ncn3.41

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Hypometabolism in the supplementary and anterior cingulate cortices is related to dysphagia in Parkinson's disease: a cross-sectional and 3-year longitudinal cohort study2013

    • Author(s)
      Akio Kikuchi, Toru Baba, Takafumi Hasegawa, Michiko Kobayashi, Naoto Sugeno, Masatoshi Konno, Emiko Miura, Yoshiyuki Hosokai, Toshiyuki Ishioka, Yoshiyuki Nishio, Kazumi Hirayama, Kyoko Suzuki, Masashi Aoki, ShokiTakahashi, Hiroshi Fukuda, Yasuto Itoyama, Etsuro Mori, Atsushi Takeda
    • Journal Title

      BMJ Open

      Volume: 3 Issue: 3 Pages: e002249-e002249

    • DOI

      10.1136/bmjopen-2012-002249

    • Related Report
      2013 Annual Research Report
    • Peer Reviewed
  • [Presentation] K63-linked ubiquitination by E3 ubiquitin ligase Nedd4-1 facilitates endosomal sequestration of internalized alpha-synuclein2014

    • Author(s)
      Sugeno N, Hasegawa T, Tanaka N, Fukuda M, Wakabayashi K, Oshima R, Konno M, Miura E, Kikuchi A, Baba T, Anan T, Nakao M, Geisler S, Aoki M, Takeda A.
    • Organizer
      18th International Congress of Parkinson's Disease and Movement Disorders
    • Place of Presentation
      スウェーデン、ストックホルム
    • Related Report
      2013 Annual Research Report
  • [Presentation] E3 ubiquitin ligase Nedd4はαシヌクレインのエンドソームへのターゲッティングに重要である2013

    • Author(s)
      菅野直人、長谷川隆文、今野昌俊、三浦永美子、菊池昭夫、青木正志、武田篤
    • Organizer
      第54回日本神経学会学術大会
    • Place of Presentation
      東京、東京国際フォーラム
    • Related Report
      2013 Annual Research Report
  • [Presentation] E3 ubiquitin ligase Nedd4 is a key regulator of the endosomal targeting of a-synuclein2013

    • Author(s)
      Naoto Sugeno, Takafumi Hasegawa, Masatoshi Konno, Emiko Miura, Akio Kikuchi, Masashi Aoki, Atsushi Takeda
    • Organizer
      17th International Congress of Parkinson's Disease and Movement Disorders
    • Place of Presentation
      オーストラリア、シドニー
    • Related Report
      2013 Annual Research Report
  • [Presentation] E3ユビキチンライゲースNedd4は細胞外αシヌクレインのエンドソームへのターゲッティングを制御する2013

    • Author(s)
      菅野直人、長谷川隆文、今野昌俊、三浦永美子、大嶋龍司、菊池昭夫、青木正志、武田篤
    • Organizer
      第7回パーキンソン病・運動障害疾患コングレス
    • Place of Presentation
      東京、京王プラザホテル
    • Related Report
      2013 Annual Research Report
  • [Presentation] Nedd4 E3 ubiquitin ligase facilitates the endosomal targeting of extracellular alpha-synuclein2013

    • Author(s)
      Naoto Sugeno, Takafumi Hasegawa, Masatoshi Konno, Emiko Miura, Akio Kikuchi, Masashi Aoki, Atsushi Takeda
    • Organizer
      第86回日本生化学会大会
    • Place of Presentation
      横浜、パシフィコ横浜
    • Related Report
      2013 Annual Research Report
  • [Presentation] K63-linked ubiquitination by Nedd4 facilitates endosomal sequestration of internalized α-synuclein2013

    • Author(s)
      Naoto Sugeno, Takafumi Hasegawa, Ryuji Oshima, Nobuyuki Tanaka, Masashi Konno, Emiko Miura, Akio Kikuchi, Toru Baba, Mitsunori Fukuda, Tadashi Anan, Mitsuyoshi Nakao, Sven Geisler, Masashi Aoki and Atsushi Takeda
    • Organizer
      Neuroscience 2013
    • Place of Presentation
      アメリカ合衆国、サン・ディエゴ
    • Related Report
      2013 Annual Research Report

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Published: 2014-07-25   Modified: 2019-07-29  

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