トランスポゾンを用いた方法論に基づく脊椎動物フェノーム解析の基盤形成

2014 Fiscal Year Annual Research Report

• Project/Area Number: 23241063
• Research Institution: National Institute of Genetics
• Principal Investigator: 川上 浩一 国立遺伝学研究所, 個体遺伝研究系, 教授 (70195048)
• Project Period (FY): 2011-04-01 – 2015-03-31
• Keywords: ゲノム / 遺伝学 / 遺伝子 / 神経科学 / 発生・分化

Outline of Annual Research Achievements

（１）ゼブラフィッシュにおいて、Gal4を組み込んだ遺伝子トラップトランスポゾンコンストラクトをゲノムにランダムに組み込ませたトランスジェニックフィッシュから、細胞・組織・器官特異的にGal4を発現するトランスジェニックフィッシュを選別する大規模スクリーニングを行い、平成26年度、新規100系統を作製した。これらをかけあわせ単一挿入をもつ系統を樹立し、インバースPCR等分子生物学的解析を行うことにより、トランスポゾン挿入部位を決定した。これら発現パターン、ゲノム情報をデータベースzTrapに登録・整備し、閲覧・検索可能にした。
（２）本研究により作製してきたトランスジェニックフィッシュを用いて、国内外の研究者と初期発生、器官形成、神経科学に関する共同研究を行った。脊髄神経細胞の増殖・分化におけるNotchシグナルの役割（千葉大伊藤博士との共同研究）、幹細胞の増殖や器官の再生におけるレチノイン酸受容体の役割（英国アストン大Johnson博士との共同研究）、側線鱗（管器）の骨リモデリング過程による形成（国立遺伝研和田博士との共同研究）、小脳神経細胞特異的なトランスジェニックゼブラフィッシュ系統（名古屋大日比博士との共同研究）、神経細胞におけるナトリウムチャンネルの蛋白質レベルでの調節機構（国立遺伝研平田博士との共同研究）を明らかにし、英文論文５報、総説１報を発表した。また本研究に関連した遺伝学的方法論に基づき、脊椎動物の性決定、哺乳動物細胞への遺伝子導入などの英文論文３報、及び方法の論文１報を発表した。
（３）脳の様々な領域でGal4を特異的に発現するトランスジェニックフィッシュ系統を脳切片の作製により明らかにした。これらをGal4発現神経細胞の機能を神経毒素遺伝子発現によ阻害し行動解析実験を行うことにより、学習・記憶に必要な脳神経回路の同定に成功した。

Research Progress Status

26年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

26年度が最終年度であるため、記入しない。

Research Products

• [Journal Article] Establishment of Gal4 transgenic zebrafish lines for analysis of development of cerebellar neural circuitry (2015)
  • Author(s): Takeuchi, M., Matsuda, K., Yamaguchi, S., Asakawa, K., Miyasaka, N., Lal, P., Yoshihara, Y., Koga, A., Kawakami, K., Shimizu, T., and Hibi, M.
  • Journal Title: Developmental Biology Volume: 397 Pages: 1-17
  • DOI: 10.1016/j.ydbio.2014.09.030
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] The Tol2 transposon system mediates the genetic engineering of T-cells with CD19-specific chimeric antigen receptors for B-cell malignancies (2015)
  • Author(s): Tsukahara, T., Iwase, N., Kawakami, K., Iwasaki, M., Yamamoto, C., Ohmine, K., Uchibori, R., Teruya, T., Ido, H., Saga, Y., Urabe, M., Mizukami, H., Kume, A., Nakamura, M., Brentjens, R., and Ozawa, K.
  • Journal Title: Gene Therapy Volume: 22 Pages: 209-215
  • DOI: 10.1038/gt.2014.104
  • Peer Reviewed
• [Journal Article] RING finger protein 121 facilitates the degradation and membrane localization of voltage-gated sodium channels (2015)
  • Author(s): Ogino, K., Low, S.E., Yamada, K., Saint-Amant, L., Zhou, W., Muto, A., Asakawa, K., Nakai, J., Kawakami, K., Kuwada, J.Y., and Hirata, H.
  • Journal Title: Proc. Natl. Acad. Sci. USA Volume: 112 Pages: 2859-2864
  • DOI: 10.1073/pnas.1414002112
  • Peer Reviewed
• [Journal Article] Size control during organogenesis: Development of the lateral line organs in zebrafish (2015)
  • Author(s): Wada, H. and Kawakami, K.
  • Journal Title: Development, Growth and Differentiation Volume: 57 Pages: 169-178
  • DOI: 10.1111/dgd.12196
  • Peer Reviewed
• [Journal Article] Tol2-mediated transgenesis and its application to electroporation (2015)
  • Author(s): Kawakami, K.
  • Journal Title: Springer Protocols -Electroporation Methods in Neuroscience Volume: 102 Pages: 179-185
• [Journal Article] Different combinations of Notch ligands and receptors regulate V2 interneuron progenitor proliferation and V2a/V2b cell fate determination (2014)
  • Author(s): Okigawa, S., Mizoguchi, T., Okano, M., Tanaka, H., Isoda, M., Jiang, Y.J., Suster, M., Higashijima, S.-I., Kawakami, K., and Itoh, M.
  • Journal Title: Developmental Biology Volume: 391 Pages: 196-206
  • DOI: 10.1016/j.ydbio.2014.04.011
  • Peer Reviewed
• [Journal Article] Development of the lateral line canal system through a bone remodeling process in zebrafish (2014)
  • Author(s): Wada, H., Iwasaki, M., and Kawakami, K.
  • Journal Title: Developmental Biology Volume: 392 Pages: 1-14
  • DOI: doi: 10.1016/j.ydbio.2014.05.004
  • Peer Reviewed
• [Journal Article] Involvement of androgen receptor in sex determination in an amphibian species (2014)
  • Author(s): Fujii, J., Kodama, M., Oike, A., Matsuo, Y., Min, M.S., Hasebe, T., Ishizuya-Oka, A., Kawakami, K., and Nakamura, M.
  • Journal Title: PLoS One Volume: 9 Pages: e93655
  • DOI: 10.1371/journal.pone.0093655
  • Peer Reviewed
• [Journal Article] Co-option of Sox3 as the male-determining factor on the Y chromosome in the fish Oryzias dancena (2014)
  • Author(s): Takehana, Y., Matsuda, M., Myosho, T., Suster, M.L., Kawakami, K., Shin-I, T., Kohara, Y., Kuroki, Y., Toyoda, A., Fujiyama, A., Hamaguchi, S., Sakaizumi, M., and Naruse, K.
  • Journal Title: Nature Communications Volume: 5 Pages: 4157
  • DOI: 10.1038/ncomms5157
  • Peer Reviewed
• [Journal Article] The development and growth of tissues derived from cranial neural crest and primitive mesoderm is dependent on the ligation status of retinoic acid receptor (RAR) γ: evidence that RARγ functions to maintain stem/progenitor cells in the absence of retinoic acid (2014)
  • Author(s): Wai, H.A., Wada, H., Kawakami, K., Müller, F., Vernallis, A.B., Brown, G., and Johnson, W.E.
  • Journal Title: Stem Cells and Development Volume: 24 Pages: 507-519
  • DOI: 10.1089/scd.2014.0235
  • Peer Reviewed
• [Presentation] Genetic dissection of the zebrafish brain by the Gal4-UAS approach (2015)
  • Author(s): Koichi Kawakami
  • Organizer: 6th Strategic Conference of Zebrafish Investigators
  • Place of Presentation: Pacific Grove, California, USA
  • Year and Date: 2015-01-17 – 2015-01-21
• [Presentation] Genetic identification of prey detector neurons in zebrafish (2015)
  • Author(s): Akira Muto, Koichi Kawakami
  • Organizer: 6th Strategic Conference of Zebrafish Investigators
  • Place of Presentation: Pacific Grove, California, USA
  • Year and Date: 2015-01-17 – 2015-01-21
• [Presentation] Genetic dissection of the zebrafish brain by the Gal4-UAS approach (2014)
  • Author(s): Koichi Kawakami
  • Organizer: 7th Model Organism Symposium
  • Place of Presentation: Daejeon, Korea
  • Year and Date: 2014-12-12 – 2014-12-12
  • Invited
• [Presentation] Genetic dissection of the zebrafish brain by the Gal4-UAS approach (2014)
  • Author(s): Koichi Kawakami
  • Organizer: The 3rd Imaging Structure & Function in the Zebrafish Brain Symposium
  • Place of Presentation: Paris, France
  • Year and Date: 2014-12-04 – 2014-12-07
  • Invited
• [Presentation] An activity-based somatosensory map in the zebrafish hindbrain as revealed by in vivo two photon calcium imaging (2014)
  • Author(s): Kazuhide Asakawa, Gembu Abe, Koichi Kawakami
  • Organizer: The 37th Annual Meeting of the Molecular Biology Society of Japan
  • Place of Presentation: 横浜
  • Year and Date: 2014-11-25 – 2014-11-27
• [Presentation] Genetic dissection of the zebrafish brain by the Gal4-UAS approach (2014)
  • Author(s): Koichi Kawakami
  • Organizer: Cold Spring Harbor Asia Meeting: Genetics, Genomics & Phenomics of Fish
  • Place of Presentation: Suzhou, China
  • Year and Date: 2014-10-20 – 2014-10-24
  • Invited
• [Presentation] The Tol2 transposon technology and its applications in zebrafish (2014)
  • Author(s): Koichi Kawakami
  • Organizer: 12th Transgenic Technology Meeting: Zebrafish workshop 2014
  • Place of Presentation: Edinburgh, UK
  • Year and Date: 2014-10-09 – 2014-10-09
  • Invited
• [Presentation] The Tol2 transposon technology in vertebrates (2014)
  • Author(s): Koichi Kawakami
  • Organizer: 12th Transgenic Technology Meeting by International Society for Transgenic Technologies
  • Place of Presentation: Edinburgh, UK
  • Year and Date: 2014-10-06 – 2014-10-08
  • Invited
• [Presentation] Genetic dissection of neural circuits mediating fear –learning in zebrafish (2014)
  • Author(s): Lal, P., Suster, M.L., Tanabe, H., Iwasaki, M., Hiratani, M. and Kawakami, K.
  • Organizer: The 20th Japanese Medaka and Zebrafish Meeting
  • Place of Presentation: 東京
  • Year and Date: 2014-09-20 – 2014-09-21
• [Presentation] Visual perception of a prey in zebrafish (2014)
  • Author(s): Muto, A., and Kawakami, K.
  • Organizer: The 20th Japanese Medaka and Zebrafish Meeting
  • Place of Presentation: 東京
  • Year and Date: 2014-09-20 – 2014-09-21
• [Presentation] Precise assembly of the basement membrane is important for the axogenesis of cerebellar granule cells (2014)
  • Author(s): Takeuchi, M., Yamaguchi, S., Yonemura, S., Asakawa, K., Kawakami, K., Takada, S., Shimizu, T., and Hibi, M.
  • Organizer: The 20th Japanese Medaka and Zebrafish Meeting
  • Place of Presentation: 東京
  • Year and Date: 2014-09-20 – 2014-09-21
• [Presentation] An activity-based somatosensory map in the zebrafish hindbrain as revealed by calcium imaging (2014)
  • Author(s): Asakawa, K., Abe, G., and Kawakami, K.
  • Organizer: The 20th Japanese Medaka and Zebrafish Meeting
  • Place of Presentation: 東京
  • Year and Date: 2014-09-20 – 2014-09-21
• [Presentation] Neuronal dysfunction caused by TDP-43 overexpression in a transgenic ALS model (2014)
  • Author(s): Kazuhide Asakawa, Koichi Kawakami
  • Organizer: Neuroscience 2014: The 37th Annual Meeting of the Japan Neuroscience Society
  • Place of Presentation: 横浜
  • Year and Date: 2014-09-11 – 2014-09-13
• [Presentation] Optogenetic analysis of the functional role of Mauthner cell on the sound/vibration-evoked fast escapes in larval zebrafish (2014)
  • Author(s): Masashi Tanimoto, Atsuko Sugimoto, Sena Yokomichi, Kazuhide Asakawa, Koichi Kawakami, Yoichi Oda
  • Organizer: Neuroscience 2014: The 37th Annual Meeting of the Japan Neuroscience Society
  • Place of Presentation: 横浜
  • Year and Date: 2014-09-11 – 2014-09-13
• [Presentation] Imaging of the neural circuit activities during retrieval of behavioral program for active avoidance in zebrafish (2014)
  • Author(s): Hisaya Kakinuma, Ryo Aoki, Tazu Aoki, Masako Yamazaki, Toshiyuki Shiraki, Mikako Takahoko, Kaori Eizumi, Tetsuya Koide, Yoshihiro Yoshihara, Junichi Nakai, Koichi Kawakami, Hitoshi Okamoto
  • Organizer: Neuroscience 2014: The 37th Annual Meeting of the Japan Neuroscience Society
  • Place of Presentation: 横浜
  • Year and Date: 2014-09-11 – 2014-09-13
• [Presentation] The Gal4 driver transgenic resource for developmental biology and neuroscience (2014)
  • Author(s): Koichi Kawakami
  • Organizer: ZDM7: Zebrafish Disease Models Conference
  • Place of Presentation: Madison, Wisconsin, USA
  • Year and Date: 2014-06-28 – 2014-07-01
  • Invited
• [Presentation] Genetic dissection of the adult zebrafish brain by the Gal4-UAS approach (2014)
  • Author(s): Koichi Kawakami
  • Organizer: 11th International Conference on Zebrafish Development and Genetics
  • Place of Presentation: Madison, Wisconsin, USA
  • Year and Date: 2014-06-24 – 2014-06-28
  • Invited
• [Presentation] An endoplasmic reticulum-resident ubiquitin ligase regulates the transport of voltage-gated sodium channels in zebrafish sensory neurons (2014)
  • Author(s): Ogino, K., Low, S., Yamada, K., Saint-Amant, L., Zhou, W., Cui, W., Muto, A., Asakawa, K., Nakai, J., Kawakami, K., Kuwada, J., Hirata, H.
  • Organizer: 11th International Conference on Zebrafish Development and Genetics
  • Place of Presentation: Madison, Wisconsin, USA
  • Year and Date: 2014-06-24 – 2014-06-28
• [Presentation] Control of axon guidance of cerebellar granule cells by the basement membrane (2014)
  • Author(s): Takeuchi, M., Hayashi, T., Yamaguchi, S., Yonemura, S., Asakawa, K., Kawakami, K., Takada, S., Kuraku, S., Shimizu, T., and Hibi, M.
  • Organizer: 11th International Conference on Zebrafish Development and Genetics
  • Place of Presentation: Madison, Wisconsin, USA
  • Year and Date: 2014-06-24 – 2014-06-28
• [Presentation] Behavioral recovery from an aversive stimulus corresponds with left habenula activity (2014)
  • Author(s): Erik R. Dubue, Akira Muto, Koichi Kawakami, Marnie Halpern
  • Organizer: 11th International Conference on Zebrafish Development and Genetics
  • Place of Presentation: Madison, Wisconsin, USA
  • Year and Date: 2014-06-24 – 2014-06-28
• [Presentation] The study of the hippocampal function in zebrafish (2014)
  • Author(s): Hideyuki Tanabe, Pradeep Lal, Akira Muto, Koichi Kawakami
  • Organizer: 11th International Conference on Zebrafish Development and Genetics
  • Place of Presentation: Madison, Wisconsin, USA
  • Year and Date: 2014-06-24 – 2014-06-28
• [Presentation] A Gal4 driver transgenic fish resource for developmental biology and neuroscience (2014)
  • Author(s): Koichi Kawakami
  • Organizer: The 66th Annual Meeting of the Japanese Society for Cell Biology
  • Place of Presentation: 奈良
  • Year and Date: 2014-06-11 – 2014-06-13
  • Invited
• [Presentation] Exploring mechanisms of neurogenesis in the developing brain with live zebrafish embryos (2014)
  • Author(s): Tomomi Sato, Fuminori Sato, Aosa Kamezaki, Koichi Kawakami, Atsuko Sehara-Fujisawa
  • Organizer: The 66th Annual Meeting of the Japanese Society for Cell Biology
  • Place of Presentation: 奈良
  • Year and Date: 2014-06-11 – 2014-06-13
• [Presentation] Directional control of the escape locomotion in zebrafish (2014)
  • Author(s): Kazuhide Asakawa, Gembu Abe, Koichi Kawakami
  • Organizer: 47th Annual Meeting of the Japanese Society of Developmental Biologists
  • Place of Presentation: 名古屋
  • Year and Date: 2014-05-27 – 2014-05-30
• [Presentation] Development of the lateral line canal system through a bone remodeling process in zebrafish (2014)
  • Author(s): Hironori Wada, Koichi Kawakami
  • Organizer: 47th Annual Meeting of the Japanese Society of Developmental Biologists Oral presentation
  • Place of Presentation: 名古屋
  • Year and Date: 2014-05-27 – 2014-05-30
• [Presentation] Dissection of cerebellar neural circuitry with zebrafish gene/enhancer trap lines (2014)
  • Author(s): Koji Matsuda, Miki Takeuchi, Kazuhide Asakawa, Masamichi Ohkura, Nobuhiko Miyasaka, Yoshihiro Yoshihara, Tetsutaro Hayashi, Shigehiro Kuraku, Koichi Kawakami, Masahiko Hibi, Takashi Shimizu
  • Organizer: 47th Annual Meeting of the Japanese Society of Developmental Biologists Oral presentation
  • Place of Presentation: 名古屋
  • Year and Date: 2014-05-27 – 2014-05-30
• [Presentation] Precise axogenesis of the cerebellar granule cells requires interaction between their axons and the basement membrane (2014)
  • Author(s): Miki Takeuchi, Shigenobu Yonemura, Kazuhide Asakawa, Koichi Kawakami, Shinji Takada, Takashi Shimizu, Masahiko Hibi
  • Organizer: 47th Annual Meeting of the Japanese Society of Developmental Biologists
  • Place of Presentation: 名古屋
  • Year and Date: 2014-05-27 – 2014-05-30
• [Presentation] Genetic dissection of the zebrafish brain by the Gal4-UAS method (2014)
  • Author(s): Koichi Kawakami, Lal Pradeep, Mari Itoh
  • Organizer: 16th Annual Meeting of the International Behavioural and Neural Genetics Society
  • Place of Presentation: Chicago, USA
  • Year and Date: 2014-05-10 – 2014-05-13
• [Patent(Industrial Property Rights)] ボツリヌス毒素遺伝子トランスジェニックフィッシュ (2011)
  • Inventor(s): 川上浩一
  • Industrial Property Rights Holder: 大学共同利用機関法人 情報・システム研究機構
  • Industrial Property Rights Type: 特許
  • Industrial Property Number: 5527684号
  • Filing Date: 2011-04-01
  • Acquisition Date: 2014-04-25