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Development of tagged protein imaging during translation and folding steps by using genome editing

Research Project

Project/Area Number 16K01931
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeMulti-year Fund
Section一般
Research Field Chemical biology
Research InstitutionTokyo Medical and Dental University

Principal Investigator

Nomura Wataru  東京医科歯科大学, 生体材料工学研究所, 准教授 (80463909)

Project Period (FY) 2016-04-01 – 2019-03-31
Project Status Completed (Fiscal Year 2018)
Budget Amount *help
¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000)
Fiscal Year 2018: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2017: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2016: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
Keywordsゲノム編集 / 蛍光イメージング / ペプチド / タンパク質 / タグ-プローブ / タグープローブ / 分子イメージング / ゲノム編集技術
Outline of Final Research Achievements

In this project, a method to observe processes of protein translation, folding, and translocation was developed using our tag-probe system. Membrane receptor and histone protein were chosen as targets and their fusion with fluorescent protein were expressed in cells. Expression and localization of target proteins were observed by fluorescent microscopy. To introduce tag gene after starting codon, guide RNA was designed and genome editing by CRISPR-Cas9 was performed. We have established a method to select cells with drug-resistant marker to increase editing efficiency. The method can widely applied to introduce various gene to genome.

Academic Significance and Societal Importance of the Research Achievements

細胞内に存在するタンパク質はその機能自体の解明も重要であるが,細胞の中にどれくらいの量が存在し,どこに存在するのか、存在場所は機能とともに変化するか,などを顕微鏡で観察することが必要になる。それを解決する手段として、細胞のゲノムに存在するタンパク質を定義している遺伝子に目印となるタグを付けることで細胞内のタンパク質の絶対量や動きを正確に観察することができる。本研究では独自のタグをゲノムに組み入れる方法を確立することができた。

Report

(4 results)
  • 2018 Annual Research Report   Final Research Report ( PDF )
  • 2017 Research-status Report
  • 2016 Research-status Report
  • Research Products

    (49 results)

All 2019 2018 2017 2016 Other

All Journal Article (19 results) (of which Int'l Joint Research: 6 results,  Peer Reviewed: 14 results,  Open Access: 7 results,  Acknowledgement Compliant: 1 results) Presentation (29 results) (of which Int'l Joint Research: 13 results,  Invited: 7 results) Remarks (1 results)

  • [Journal Article] Delivery of a Proapoptotic Peptide to EGFR-Positive Cancer Cells by a Cyclic Peptide Mimicking the Dimerization Arm Structure of EGFR.2019

    • Author(s)
      Kei Toyama, Wataru Nomura, Takuya Kobayakawa, Hirokazu Tamamura
    • Journal Title

      Bioconjugate Chemistry

      Volume: 29 Issue: 6 Pages: 2050-2057

    • DOI

      10.1021/acs.bioconjchem.8b00250

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Benzolactam-related compounds promote apoptosis of HIV-infected human cells via protein kinase C-induced HIV latency reversal.2019

    • Author(s)
      Kouki Matsuda, Takuya Kobayakawa, Kiyoto Tsuchiya, Shin-ichiro Hattori, Wataru Nomura, Hiroyuki Gatanaga, Kazuhisa Yoshimura, Shinichi Oka, Yasuyuki Endo, Hirokazu Tamamura, Hiroaki Mitsuya, Kenji Maeda
    • Journal Title

      Journal of Biological Chemistry

      Volume: 294 Issue: 1 Pages: 116-129

    • DOI

      10.1074/jbc.ra118.005798

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Design, synthesis and biological evaluation of low molecular weight CXCR4 ligands2019

    • Author(s)
      3.Sakyiamah MM., Kobayakawa T., Fujino M., KonnoM., Narumi T., Tanaka T., Nomura W., Yamamoto N., Murakami T., and Tamamura H.
    • Journal Title

      Bioorg. Med. Chem

      Volume: 2 Issue: 6 Pages: 1130-1138

    • DOI

      10.1016/j.bmc.2019.02.013

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Inhibition of EGFR Activation by Bivalent Ligands Based on a Cyclic Peptide Mimicking the Dimerization Arm Structure of EGFR2018

    • Author(s)
      Kei Toyama, Takuya Kobayakawa, Wataru Nomura, Hirokazu Tamamura
    • Journal Title

      Chemical and Pharmaceutical Bulletin

      Volume: 66 Issue: 11 Pages: 1083-1089

    • DOI

      10.1248/cpb.c18-00539

    • NAID

      130007502876

    • ISSN
      0009-2363, 1347-5223
    • Year and Date
      2018-11-01
    • Related Report
      2018 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Efficient and Orthogonal Transcription Regulation by Chemically Inducible Artificial Transcription Factors.2018

    • Author(s)
      Wataru Nomura, Daisuke Matsumoto, Taisuke Sugii, Takuya Kobayakawa, Hirokazu Tamamura
    • Journal Title

      Biochemistry

      Volume: 57 Issue: 45 Pages: 6452-6459

    • DOI

      10.1021/acs.biochem.8b00741

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Development of Toolboxes for Precision Genome/Epigenome Editing and Imaging of Epigenetics.2018

    • Author(s)
      Wataru Nomura
    • Journal Title

      The Chemical Record

      Volume: 18 Issue: 12 Pages: 1717-1726

    • DOI

      10.1002/tcr.201800036

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed
  • [Journal Article] ゲノム編集がもたらす蛍光バイオイメージングの革新2018

    • Author(s)
      野村渉
    • Journal Title

      Precision Medicine

      Volume: 1 Pages: 80-85

    • Related Report
      2018 Annual Research Report
  • [Journal Article] Synthesis of a Chloroalkene Dipeptide Isostere-Containing Peptidomimetic and Its Biological Application2018

    • Author(s)
      Takuya Kobayakawa, Yudai Matsuzaki, Kentaro Hozumi, Wataru Nomura, Motoyoshi Nomizu, Hirokazu Tamamura
    • Journal Title

      ACS Med. Chem. Lett.

      Volume: 9 Issue: 1 Pages: 6-10

    • DOI

      10.1021/acsmedchemlett.7b00234

    • Related Report
      2017 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Creation of Functional Molecules Based on Biomolecular Interactions; Development toward Chemical Biology2017

    • Author(s)
      野村 渉
    • Journal Title

      YAKUGAKU ZASSHI

      Volume: 137 Issue: 10 Pages: 1223-1231

    • DOI

      10.1248/yakushi.17-00125

    • NAID

      130006109870

    • ISSN
      0031-6903, 1347-5231
    • Year and Date
      2017-10-01
    • Related Report
      2017 Research-status Report
    • Open Access
  • [Journal Article] DNAメチル化制御から精密エピゲノム編集へ2017

    • Author(s)
      野村 渉
    • Journal Title

      MEDCHEM NEWS

      Volume: 27 Pages: 200-207

    • NAID

      130007755934

    • Related Report
      2017 Research-status Report
  • [Journal Article] Synthesis and Evaluation of Dimeric Derivatives of Diacylglycerol-lactones as PKC Ligands2017

    • Author(s)
      Nami Ohashi, Ryosuke Kobayashi, Wataru Nomura, Takuya Kobayakawa, Agnes Czikora, Brienna Herold, Nancy E. Lewin, Peter M. Blumberg, Hirokazu Tamamura
    • Journal Title

      Bioconjugate Chem.

      Volume: 28 Issue: 8 Pages: 2135-2144

    • DOI

      10.1021/acs.bioconjchem.7b00299

    • Related Report
      2017 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Development of Anti-HIV Peptides Based on a Viral Capsid Protein2017

    • Author(s)
      Takaaki Mizuguchi, Nami Ohashi, Daichi Matsumoto, Chie Hashimoto, Wataru Nomura, Naoki Yamamoto, Tsutomu Murakami, Hirokazu Tamamura
    • Journal Title

      Biopolymers: Peptide Science

      Volume: 印刷中 Issue: 1

    • DOI

      10.1002/bip.22920

    • Related Report
      2016 Research-status Report
    • Peer Reviewed
  • [Journal Article] Bivalent 14-mer Peptide Ligands of CXCR4 with Polyproline Linkers with Anti-Chemotactic Activity against Jurkat Cells2017

    • Author(s)
      Tomohiro Tanaka, Toru Aoki, Wataru Nomura, Hirokazu Tamamura
    • Journal Title

      J. Pept. Sci.

      Volume: 印刷中 Issue: 7-8 Pages: 574-580

    • DOI

      10.1002/psc.2946

    • Related Report
      2016 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Microwave-Assisted Synthesis of Azacoumarin Fluorophores and the Fluorescence Characterization2017

    • Author(s)
      Hikaru Takano, Tetsuo Narumi, Wataru Nomura, Hirokazu Tamamura
    • Journal Title

      J. Org. Chem.

      Volume: 82 Issue: 5 Pages: 2739-2744

    • DOI

      10.1021/acs.joc.6b02656

    • Related Report
      2016 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] 多価型CXCR4リガンドの創製とがん細胞イメージング・検出2016

    • Author(s)
      野村 渉, 玉村啓和
    • Journal Title

      医学のあゆみ 第5土曜特集

      Volume: 5 Pages: 430-437

    • Related Report
      2016 Research-status Report
  • [Journal Article] 細胞内タンパク質を可視化するタグ-蛍光プローブシステム2016

    • Author(s)
      野村渉, 玉村啓和
    • Journal Title

      「生化学」みにれびゅう

      Volume: 89 Pages: 115-120

    • NAID

      40021119837

    • Related Report
      2016 Research-status Report
    • Acknowledgement Compliant
  • [Journal Article] A minimally cytotoxic CD4 mimic as an HIV entry inhibitor2016

    • Author(s)
      Takaaki Mizuguchi, Shigeyoshi Harada, Tomoyuki Miura, Nami Ohashi, Tetsuo Narumi, Hiromi Mori, Yu Irahara, Yuko Yamada, Wataru Nomura, Shuzo Matsushita, Kazuhisa Yoshimura, Hirokazu Tamamura
    • Journal Title

      Bioorganic & Medicinal Chemistry Letters

      Volume: 26 Issue: 2 Pages: 397-400

    • DOI

      10.1016/j.bmcl.2015.11.103

    • Related Report
      2016 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Small-Molecule CD4 Mimics Containing Mono-cyclohexyl Moieties as HIV Entry Inhibitors.2016

    • Author(s)
      Nami Ohashi1, Shigeyoshi Harada, Takaaki Mizuguchi1, Yu Irahara1, Yuko Yamada, Misato Kotani, Wataru Nomura, Shuzo Matsushita, Kazuhisa Yoshimura, Hirokazu Tamamura.
    • Journal Title

      Chem Med Chem

      Volume: 19 Issue: 8 Pages: 940-946

    • DOI

      10.1002/cmdc.201500590

    • Related Report
      2016 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Evaluation of Fluorescein-Labeled Derivatives of a Peptide Inhibitor of the EGF Receptor Dimerization2016

    • Author(s)
      Kei Toyama, Takaaki Mizuguchi, Wataru Nomura, Hirokazu Tamamura
    • Journal Title

      Bioorg. Med. Chem.

      Volume: 24 Issue: 16 Pages: 3406-3412

    • DOI

      10.1016/j.bmc.2016.05.026

    • Related Report
      2016 Research-status Report
    • Peer Reviewed
  • [Presentation] Chemically-controlled orthogonal regulation of multiple endogenous genes2018

    • Author(s)
      Wataru Nomura, Daisuke Matsumoto, Taisuke Sugii, Takuya Kobayakawa, Hirokazu Tamamura
    • Organizer
      ASBMB Annual meeting: Experimental Biology 2018
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Differential endogenous gene activation by orthogonally-controlled chemically inducible systems2018

    • Author(s)
      Wataru Nomura, Daisuke Matsumoto, Taisuke Sugii, Takuya Kobayakawa, Hirokazu Tamamura
    • Organizer
      FASEB Summer Research Conference
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research
  • [Presentation] 化合物を用いた直交的な内因性遺伝子制御系の構築2018

    • Author(s)
      野村渉、松本大亮、杉井太亮、小早川拓也、玉村啓和
    • Organizer
      第12回バイオ関連化学シンポジウム
    • Related Report
      2018 Annual Research Report
  • [Presentation] Fluorogenic and genetically encodable tag-probe system for in-cell imaging of protein synthesis2018

    • Author(s)
      Wataru Nomura, Takumi Kamimura, Daisuke Matsumoto, Hirokazu Tamamura
    • Organizer
      10th International Peptide Symposium
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Distinct chemically inducible system for orthogonal differential regulation of endogenous genes2018

    • Author(s)
      Wataru Nomura, Daisuke Matsumoto, Takuya Kobayakawa, Taisuke Sugii, Hirokazu Tamamura
    • Organizer
      Keystone Symposia Conference A2: Precision Genome Engineering
    • Related Report
      2017 Research-status Report
    • Int'l Joint Research
  • [Presentation] Genome editing with a FokI-based chemically inducible nuclease system2018

    • Author(s)
      Daisuke Matsumoto, Wataru Nomura, Hirokazu Tamamura
    • Organizer
      Keystone Symposia Conference A2: Precision Genome Engineering
    • Related Report
      2017 Research-status Report
    • Int'l Joint Research
  • [Presentation] Orthogonal differential regulation of endogenous genes by distinct chemically inducible systems2018

    • Author(s)
      Wataru Nomura, Daisuke Matsumoto, Takuya Kobayakawa, Taisuke Sugii, Hirokazu Tamamura
    • Organizer
      武田薬科学シンポジウム
    • Related Report
      2017 Research-status Report
  • [Presentation] Development of tools for the modification, regulation, and molecular imaging of gene and epigenetic functions2018

    • Author(s)
      Wataru Nomura, Hirokazu Tamamura
    • Organizer
      NCU Global Young Investigator Forum 2018
    • Related Report
      2017 Research-status Report
    • Int'l Joint Research / Invited
  • [Presentation] 標的タンパク質の動きをみるペプチド性蛍光イメージングツール2018

    • Author(s)
      野村 渉, 神村拓美, 森あつみ, 小早川拓也, 玉村啓和
    • Organizer
      日本薬学会第138年会シンポジウム
    • Related Report
      2017 Research-status Report
    • Invited
  • [Presentation] 分割型部位特異的ヌクレアーゼを利用した化合物誘導型ゲノム編集技術の開発2017

    • Author(s)
      松本大亮, 野村 渉, 玉村啓和
    • Organizer
      日本化学会第97春季年会
    • Place of Presentation
      慶應義塾大学日吉キャンパス(神奈川県横浜市)
    • Year and Date
      2017-03-19
    • Related Report
      2016 Research-status Report
  • [Presentation] Controllable genome editing by chemically inducible split site-specific nucleases2017

    • Author(s)
      Daisuke Matsumoto, Wataru Nomura, HIrokazu Tamamura
    • Organizer
      2017 Keystone Symposia Conference A2: Precision Genome Engineering
    • Place of Presentation
      Breckenridge, USA
    • Year and Date
      2017-01-10
    • Related Report
      2016 Research-status Report
    • Int'l Joint Research
  • [Presentation] Chemical-inducible artificial transcription factors based on sequence-specificity of TALE and dCas92017

    • Author(s)
      Wataru Nomura, Taisuke Sugii, HIrokazu Tamamura
    • Organizer
      2017 Keystone Symposia Conference A2: Precision Genome Engineering
    • Place of Presentation
      Breckenridge, USA
    • Year and Date
      2017-01-10
    • Related Report
      2016 Research-status Report
    • Int'l Joint Research
  • [Presentation] FokIをDNA切断ドメインとして用いた化合物誘導型配列特異的ヌクレアーゼの開発2017

    • Author(s)
      松本大亮, 野村 渉, 杉井太亮, 玉村啓和
    • Organizer
      日本ケミカルバイオロジー学会第12回年会
    • Related Report
      2017 Research-status Report
  • [Presentation] Development of chemical-inducible artificial transcription factors based on sequence-specific DNA binders2017

    • Author(s)
      Wataru Nomura, Daisuke Matsumoto, Tsukasa Hashimoto, Taisuke Sugii, Hirokazu Tamamura
    • Organizer
      The 254th ACS National Meeting and Exposition
    • Related Report
      2017 Research-status Report
    • Int'l Joint Research
  • [Presentation] 生体分子間相互作用を制御する機能性ペプチド/タンパク質の設計と応用2017

    • Author(s)
      野村 渉
    • Organizer
      熊本大学先端科学研究部セミナー
    • Related Report
      2017 Research-status Report
    • Invited
  • [Presentation] Development of genome editing and gene regulation systems utilizing specific DNA binding domains2017

    • Author(s)
      Wataru Nomura
    • Organizer
      熊本大学 HIGO program seminar
    • Related Report
      2017 Research-status Report
    • Invited
  • [Presentation] Differential regulation of endogenous genes in an orthogonal manner by distinct chemically inducible systems2017

    • Author(s)
      Wataru Nomura, Daisuke Matsumoto, Takuya Kobayakawa, Taisuke Sugii, Hirokazu Tamamura
    • Organizer
      International Conference on Epigenetics and Bioengineering
    • Related Report
      2017 Research-status Report
    • Int'l Joint Research
  • [Presentation] 化合物による誘導が可能な分割型人工ヌクレアーゼの構築とその評価2016

    • Author(s)
      松本大亮, 野村 渉, 玉村啓和
    • Organizer
      第39回日本分子生物学会年会
    • Place of Presentation
      パシフィコ横浜(神奈川県横浜市)
    • Year and Date
      2016-12-01
    • Related Report
      2016 Research-status Report
  • [Presentation] 化学誘導二量体形成法を応用したゲノム編集ツールの開発2016

    • Author(s)
      松本大亮, 野村 渉, 玉村啓和
    • Organizer
      「細胞を創る」研究会 9.0
    • Place of Presentation
      早稲田大学井深大記念ホール(東京都新宿区)
    • Year and Date
      2016-11-21
    • Related Report
      2016 Research-status Report
  • [Presentation] 生体分子間相互作用を基盤とする機能分子の創製とケミカルバイオロジーへの展開2016

    • Author(s)
      野村 渉
    • Organizer
      第60回日本薬学会関東支部大会
    • Place of Presentation
      東京大学山上会館 他(東京都文京区)
    • Year and Date
      2016-09-17
    • Related Report
      2016 Research-status Report
    • Invited
  • [Presentation] 末端架橋型ZIPタグプローブペアの創製2016

    • Author(s)
      斉藤 歩, 大橋南美, 野村 渉, 玉村啓和
    • Organizer
      第60回日本薬学会関東支部大会
    • Place of Presentation
      東京大学山上会館 他(東京都文京区)
    • Year and Date
      2016-09-17
    • Related Report
      2016 Research-status Report
  • [Presentation] TALEおよびdCas9を利用した化合物による標的遺伝子特異的な転写活性制御技術2016

    • Author(s)
      野村 渉, 杉井太亮, 玉村啓和
    • Organizer
      第10回 バイオ関連化学シンポジウム
    • Place of Presentation
      石川県立音楽堂 他(石川県金沢市)
    • Year and Date
      2016-09-07
    • Related Report
      2016 Research-status Report
  • [Presentation] 化合物による制御可能なゲノム編集を目指した分割型人工ヌクレアーゼの構築2016

    • Author(s)
      松本大亮, 野村 渉, 玉村啓和
    • Organizer
      第10回 バイオ関連化学シンポジウム
    • Place of Presentation
      石川県立音楽堂 他(石川県金沢市)
    • Year and Date
      2016-09-07
    • Related Report
      2016 Research-status Report
  • [Presentation] 分割型人工ヌクレーゼを用いた化合物誘導型ゲノム編集系の構築2016

    • Author(s)
      松本大亮, 野村 渉, 玉村啓和
    • Organizer
      第48回若手ペプチド夏の勉強会
    • Place of Presentation
      大学セミナーハウス(東京都八王子市)
    • Year and Date
      2016-08-01
    • Related Report
      2016 Research-status Report
  • [Presentation] Protein dynamics imaging using tag-probe systems2016

    • Author(s)
      Wataru Nomura, Nami Ohashi, HIrokazu Tamamura
    • Organizer
      The 14th Chinese International Peptide Symposium & the 5th Asia-Pacific International Peptide Symposium
    • Place of Presentation
      Nanjing, China
    • Year and Date
      2016-07-06
    • Related Report
      2016 Research-status Report
    • Int'l Joint Research / Invited
  • [Presentation] ケミカルツールを利用した遺伝子発現調節機構の構築と応用2016

    • Author(s)
      松本大亮, 野村 渉, 杉井太亮, 玉村啓和
    • Organizer
      創薬懇話会 2016 in 蓼科
    • Place of Presentation
      エクシブ蓼科(長野県茅野市)
    • Year and Date
      2016-06-30
    • Related Report
      2016 Research-status Report
  • [Presentation] Construction of gene regulation system utilizing chemical tools2016

    • Author(s)
      野村 渉, 杉井太亮, 玉村啓和
    • Organizer
      日本ケミカルバイオロジー学会第11回年会
    • Place of Presentation
      京都テルサ(京都府京都市)
    • Year and Date
      2016-06-15
    • Related Report
      2016 Research-status Report
  • [Presentation] Chemically-inducible gene regulation system utilizing sequence-specificity of TALE and dCas92016

    • Author(s)
      Wataru Nomura, Taisuke Sugii, HIrokazu Tamamura
    • Organizer
      FASEB SRC: Genome Engineering: Cutting-Edge Research and Applications
    • Place of Presentation
      Lisbon, Portugal
    • Year and Date
      2016-06-08
    • Related Report
      2016 Research-status Report
    • Int'l Joint Research
  • [Presentation] Tag-probe systems for protein dynamics imaging2016

    • Author(s)
      Wataru Nomura, Nami Ohashi, HIrokazu Tamamura
    • Organizer
      The 16th Akabori Conference: Japanese-German Symposium on Peptide Science
    • Place of Presentation
      六甲山ホテル(兵庫県神戸市)
    • Year and Date
      2016-05-24
    • Related Report
      2016 Research-status Report
    • Int'l Joint Research / Invited
  • [Remarks] 東京医科歯科大学生体材料工学研究所メディシナルケミストリー分野ホームページ

    • URL

      http://tamamura-tmd.sakura.ne.jp/members/teachers/nomura/

    • Related Report
      2017 Research-status Report 2016 Research-status Report

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Published: 2016-04-21   Modified: 2020-03-30  

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