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2004 Fiscal Year Final Research Report Summary

Analysis of nuclear pore translocation mechanism : single molecule analysis and biochemical approach

Research Project

Project/Area Number 15370090
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Cell biology
Research InstitutionRIKEN

Principal Investigator

IMAMOTO Naoko  RIKEN, Cellular Dynamics Laboratory, Chief Scientist, 今本細胞核機能研究室, 主任研究員 (20202145)

Co-Investigator(Kenkyū-buntansha) KOSE Shingo  RIKEN, Cellular Dynamics Laboratory, Research Scientist, 今本細胞核機能研究室, 研究員 (90333278)
KOIKE Makiko  RIKEN, Cellular Dynamics Laboratory, Contract Technical Scientist, 今本細胞核機能研究室, 協力研究員 (30391949)
Project Period (FY) 2003 – 2004
Keywordsnuclear transport / single molecule imaging / nuclear pore comlex / importin β / semi-intact cells / 70kDa heat-shock cognate protein
Research Abstract

Using novel microscopy based on total internal reflection fluorescent microscopy (TIRF), we were able to clearly visualize single fluorescent molecules of green fluorescent protein (GFP)-tagged importin β, a carrier protein, and GFP-tagged cargo protein during transport on the nuclear envelope in both permeabilized cells and in living cells. Image analysis of single nuclear pores showed that two point resolution of 70 nm was achieved. Kinetic parameters of the interactions between translocating molecules and NPCs were obtained through quantitative analysis. Two types of binding were found ; weaker binding sites, which gathers up to 〜100 molecules/NPC, concentrating substrates locally ; and stronger binding sites, where up to 〜8 molecules/NPC are bound. Retention times by single molecule analysis and translocation rates by single nuclear analysis showed a significant correlation with a coefficient of 〜8 molecules/NPC, which exhibits the stoichiometry of transport. We termed weaker binding sites as a "multiplex sites", which is likely to consist from FG-repeat containing nucleoporins. Based on correlation coefficient, we proposed presence of 8 "transit site" in nuclear pore complex. Whether this transit site is equivalent to strong binding sites observed remains to be elucidated in the further study. Whether the findings account for different pathways, which show a wide range of transport rates, also remains to be elucidated. In the parallel experiments, we have purified, and identified 70kDa heat-shock cognate protein (hsc70) as a molecule that facilitates recycling of importin β. The effect of hsc70 was observed, not only in the case of the nuclear export of importin β but also for that of another import receptors, transportin and importin α. These results suggest that hsc70 broadly modulates nucleocytoplasmic transport systems by regulating nuclear export of receptor proteins (Kose et al., J Cell Biol. 2005).

  • Research Products

    (30 results)

All 2006 2005 2004 2003

All Journal Article (28 results) Book (2 results)

  • [Journal Article] he nuclear import of RNA helicase A is mediated by m portin-α3.2006

    • Author(s)
      Satoko Aratani
    • Journal Title

      BBRC 340

      Pages: 125-133

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] 核膜孔複合体 : 核-細胞質間分子流通のメディエーターとしての機能と構造2006

    • Author(s)
      今本尚子
    • Journal Title

      遺伝子医学Mook 5号

      Pages: 186-194

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] The nuclear import of RNA helicase A is mediated by importin-α32006

    • Author(s)
      Satoko Aratani, Takayuki Oishi, Hidetoshi Fujita, Minako Nkazawa, Ryouji Fujii, Noko Imamoto, Yoshihiro Yoneda, Akiyoshi Fukamizu, Toshihiro Nakajima
    • Journal Title

      BBRC 340

      Pages: 125-133

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Structure and Function of Nuclear Pore Complex2006

    • Author(s)
      Naoko Imamoto
    • Journal Title

      Gene&Medicine Mook 5

      Pages: 186-194

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] The 70-kD heat shock cognate protein (hsc70) facilitates the nuclear export of the import receptors.2005

    • Author(s)
      Shingo Kose
    • Journal Title

      J. Cell Biol. 171

      Pages: 19-25

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] Dissociation of heterochromatin protein 1 from lamin B receptor induced by human polyomavirus agnoprotein : role uclear egress of viral particles2005

    • Author(s)
      Yuki Okada
    • Journal Title

      EMBO Rep. 6

      Pages: 452-457

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] 1枚の写真館「核の表面を観て思う」2005

    • Author(s)
      今本尚子
    • Journal Title

      細胞工学 Vol.24

      Pages: 783

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] 生細胞への複数種cDNAの共導入と発現量の制御2005

    • Author(s)
      今本文男
    • Journal Title

      蛋白質核酸酵素 Vol.50

      Pages: 1637-1648

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] The 70-kD heat shock cognate protein(hsc70) facilitates the nuclear export of the import receptors2005

    • Author(s)
      Shingo Kose, Maiko Furuta, Makiko Koike, Yoshihiro Yoneda, Naoko Imamoto
    • Journal Title

      J.Cell Biol. 171

      Pages: 19-25

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Dissociation of heterochromatin protein 1 from lamin B receptor induced by human polyomavirus agnoprotein : role nuclear egress of viral particles2005

    • Author(s)
      Yuki Okada, Tadaki Suzuki, Yuji Sunden, Yasuko Orba, Shingo Kose, Noko Imamoto, Hidehiro Takahashi, Shinya Tanaka, William W.Hall, Kazuo Nagashima, Hirofumi Sawa
    • Journal Title

      EMBO Rep. 6

      Pages: 452-457

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Ichimai no Shashinkan : Kaku no Hyomen wo mite omou2005

    • Author(s)
      Naoko Imamoto
    • Journal Title

      CELL TECHNOLOGY Vol.24

      Pages: 783

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Multi-gene expression clone design for conditional expression at near physiological levels of multiple heterologous genes in living cells2005

    • Author(s)
      Fumio Imamoto, Takeshi Sone, Kazuhide Yahata, Yukari Sasaki, Masatoshi Takagi, Kazuhiro Maeshima, Naoko Imamoto
    • Journal Title

      PROTEIN, NUCLEIC ACID AND ENZYME Vol.50

      Pages: 1637-1648

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Molecular Mechanism of Nucleocytoplasmic Transport2005

    • Author(s)
      Naoko Imamoto
    • Journal Title

      Molecular Biology of Cell Nucleus -Chromatin・Chromosomes・Nuclear Structure -

      Pages: 155-175

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Specific monoclonal antibody against nuclear import factor, importin α1/Rch1.2004

    • Author(s)
      Yuji Kamikubo
    • Journal Title

      HYBRIDOMA AND HYBRIDOMICS 23

      Pages: 301-304

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] β-Catenin shows an overlapping sequence requirement but distinct molecular interaction for its hi-directional passage through nuclear pores.2004

    • Author(s)
      Makiko Koike
    • Journal Title

      J. Biol. Chem. 279

      Pages: 34038-34047

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] Heat-shock induced nuclear retention and recycling inhibition of importin α2004

    • Author(s)
      Maiko Furuta
    • Journal Title

      Genes Cells 9

      Pages: 429-441

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] 核膜孔複合体と核-細胞質問輸送2004

    • Author(s)
      小瀬真吾
    • Journal Title

      蛋白質核酸酵素 Vo1.49

      Pages: 907-909

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] Specific monoclonal antibody against nuclear import factor, importin α1/Rch12004

    • Author(s)
      Yuji Kamikubo, Naoko Sakaguchi, Keiji Shikata, Maiko Furuta, Yoichi Miyamoto, Naoko Imamoto, Yoshihiro Yoneda, Kenji Ogino, Taro Tachibana
    • Journal Title

      HYBRIDOMA AND HYBRIDOMICS 23

      Pages: 301-304

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] β-Catenin shows an overlapping sequence requirement but distinct molecular interaction for its bi-directional passage through nuclear pores2004

    • Author(s)
      Makiko Koike, Shingo Kose, Maiko Furuta, Naoko Taniguchi, Fumihiko Yokota, Yoshihiro Yoneda, Naoko Imamoto
    • Journal Title

      J.Biol.Chem. 279

      Pages: 34038-34047

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Heat-shock induced nuclear retention and recycling inhibition of importin α2004

    • Author(s)
      Maiko Furuta, Shingo Kose, Makiko Koike, Takeshi Shimi, Yasushi Hiraoka, Yoshihiro Yoneda, Tokuko Haraguchi, Naoko Imamoto
    • Journal Title

      Genes Cells 9

      Pages: 429-441

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Nuclear pore complex and nucleocytoplasmic transport2004

    • Author(s)
      Shingo Kose, Naoko Imamoto
    • Journal Title

      PROTEIN, NUCLEIC ACID AND ENZYME Vol.49

      Pages: 907-909

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Architecture and Dynamics of Nuclear Pore Complex2004

    • Author(s)
      Naoko Imamoto, Maiko Furuta, Makiko Koike, Shingo Kose
    • Journal Title

      Dynamics of Cell Nucleus

      Pages: 195-201

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] The structure of importin-β bound to SREBP-2 : Nuclear import of a transcription factor.2003

    • Author(s)
      Soo Jae Lee
    • Journal Title

      Science 302

      Pages: 1571-1575

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] 核 : 細胞質問輸送の分子機構と輸送経路の多様性2003

    • Author(s)
      今本尚子
    • Journal Title

      実験医学 Vol.21

      Pages: 1842-1849

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] 細胞内輸送研究 : 細胞と個体形成の秩序を知る2003

    • Author(s)
      今本尚子
    • Journal Title

      実験医学 Vol. 21

      Pages: 1836-1841

    • Description
      「研究成果報告書概要(和文)」より
  • [Journal Article] The structure of importin-β bound to SREBP-2 : Nuclear import of a transcription factor2003

    • Author(s)
      Soo Jae Lee, Toshihiro Sekimoto, Eiki Yamashita, Emi Nagoshi, Atsushi Nakagawa, Naoko Imamoto, Masato Yoshimura, Hiroaki Sakai, Khoon Tee Chong, Tomitake Ysukihara, Yoshihiro Yoneda
    • Journal Title

      Science 302

      Pages: 1571-1575

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Nucleocytoplasmic transport : mechanism and pathways2003

    • Author(s)
      Naoko Imamoto, Makiko Koike, Maiko Furuta
    • Journal Title

      Experimental Medicine Vol.21

      Pages: 1842-1849

    • Description
      「研究成果報告書概要(欧文)」より
  • [Journal Article] Intracellular trafficking of molecules and biogenesis of eukaryotic organelles2003

    • Author(s)
      Naoko Imamoto
    • Journal Title

      Experimental Medicine Vol.21

      Pages: 1836-1841

    • Description
      「研究成果報告書概要(欧文)」より
  • [Book] 細胞核の分子生物学-クロマチン・染色体・核構造- 第6章「核と細胞質問の分子流通メカニズム」2005

    • Author(s)
      今本尚子
    • Total Pages
      21
    • Publisher
      朝倉書店
    • Description
      「研究成果報告書概要(和文)」より
  • [Book] 細胞核ダイナミクス「核膜孔複合体の分子機構とダイナミクス」2004

    • Author(s)
      今本尚子
    • Total Pages
      7
    • Publisher
      フェアラーク東京株式会社
    • Description
      「研究成果報告書概要(和文)」より

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Published: 2007-12-13  

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