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Plasmalemmal caveolae and intracellular calcium regulation

Research Project

Project/Area Number 08457001
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field General anatomy (including Histology/Embryology)
Research InstitutionGunma University

Principal Investigator

FUJIMOTO Toyoshi  Gunma University School of Medicine, Department of Anatomy and Cell Biology, Professor, 医学部, 教授 (50115929)

Co-Investigator(Kenkyū-buntansha) KOGO Hiroshi  ibid, 医学部, 助手 (20282387)
AOKI Takeo  ibid, 医学部, 助手 (70150919)
HAGIWARA Haruo  Gunma University School of Medicine, Department of Anatomy and Cell Biology, Pro, 医学部, 講師 (80189464)
Project Period (FY) 1996 – 1997
Project Status Completed (Fiscal Year 1997)
Budget Amount *help
¥7,700,000 (Direct Cost: ¥7,700,000)
Fiscal Year 1997: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1996: ¥5,800,000 (Direct Cost: ¥5,800,000)
KeywordsCaveola / Plasma membrane / Calcium / Gi2a / Cholesterol / Freeze fracture / Immunoelectron microscopy
Research Abstract

1) A simple quick freezing method to observe the plasma membrane from monolayr cultured cells was invented. The metal sandwich method is simple, does not require any expensive equipment, and provides a large fracture plane of the plasma membrane for subsequent histochemical manipulation.
2) rheta-Toxin (perfringolysin O), a cholesterol-binding toxin, was partially proteolysed and biotinylated (BCrheta) and used as a cytochemical probe. Because the plasma membrane keeps integrity even after the binding of BCrheta, the probe can be used not only for cytochemical labeling of fixed cells, but for pursuing the behavior of cross-linked cholesterol molecules in living cells. By using this new probe, it was revealed that cross-linked cholesterol in the plasma membrane are sequestered to caveolae.
3) By immunoelectron microscopy, whether Gi2a is constitutively concentrated in caveolae was critically assessed. First, behavior of caveolin and Gi2a in density-equilibrium ultracentrifugation was reexamined. By collecting fractions efficiently, caveolin and Gi2a were found to distribute differently. Secondly, by novel immunocytochemical methods it was found that the labeling density of Gi2a was 2.29 times more in caveolae than in the non-caveolar plasma membrane. The results indicate that concentration of Gi2a in caveolae is less than deduced from most biochemical studies.

Report

(3 results)
  • 1997 Annual Research Report   Final Research Report Summary
  • 1996 Annual Research Report
  • Research Products

    (22 results)

All Other

All Publications (22 results)

  • [Publications] Fujimot, T.: "GPI-anchored protcins,glycasphingolipids,and sphingomgelin are disffisely in untrested cells and Sefuestered to caveolae only after cross-linking" J.Histochem.Cytochem. 44. 929-941 (1966)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto, K.M.Umeda, and T.Fujimoto: "Transmenbrone phospholipid distribution revealed by freceze-frafture replica labeling" J.Call Sci. 109. 2453-2460 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto, K.M.Umeda, and T.Fujimoto: "Cross-linked plasmaleramal chalestrel is suquestered to cuveolae:analysio with a new cytochemicl prelke" J.Histochem.Cytochem. 45. 1197-1205 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto, K.M.Umeda, and T.Fujimoto: "Metal sandwich method to quicls-freeze monolayer cultured cells for feeze fracture" J.Histochem.Cytochem. 45. 595-598 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Nomura, R.C.Inoue, F.Takahashi, T.Asamo, and T.Fujimoto: "Two-dimensionaidistribution fo Gi2a in the plasma membraue:a critical evaluation by immunovytothemisxry" FEBS Lett. 415. 139-144 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Amino.K, Y.Honda, C.Inue and T.Fujimoto: "Distribution of plasmalemmal Ca^<2+>-pumpand caveclin in the corneal epithelium during wonud healing process" Carr.Eye Res. 16. 1088-1095 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto, T.: "GPI-anchored proteins, glycosphingolipids, and sphingomyelin are distributed diffusely in untreated cells and sequestered to caveolae only after cross-linking." HisJ.tochem.Cytochem. 44. 929-941 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto, K., M.Umeda, and T.Fujimoto: "Transmembrane phospholipid distribution revealed by freeze-fracture replica labeling." J.Cell Sci.109. 2453-2460 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto, T., M.Hayashi, M.Iwamoto and Y.Ohno-Iwashita: "Cross-linked plasmalemmal cholesterol is sequestered to caveolae : analysis with a new cytochemical probe." J.Histochem.Cytochem.45. 1197-1205 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto, T.and K.Fujimoto: "Metal sandwich method to quick-freeze monolayr cultured cells for freeze fracture." J.Histochem.Cytochem.45. 595-598 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Nomura, R., C.Inuo, Y.Takahashi, T.Asano, and T.Fujimoto: "Two-dimensional distribution of Gi_<2alpha> in the plasma membrane : a critical evaluation by immunocytochemistry." FEBS Lett.415. 139-144 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Amino, K., Y.Honda, C.Ide, and T.Fujimoto: "Distribution of plasmalemmal Ca^<2+>-pump and caveolin in the corneal epithelium during wound healing process." Curr.Eye Res.16. 1088-1095 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Fujimoto,T. and K.Fujimoto: "Metal sandwich method to quich-fraeze monolayer cultwed cells for freeze fracture" J.Histochem.Cytochem.45. 595-598 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Fujimoto,T., M.Hayashi, M.Iwamoto and Y.Oho.Iwashita: "Crass-lonhed plasmalemmal cholerterol is sequestened to caveolal : analysis with a new cytchemical probe" J.Histochem.Cytochem.45. 1197-1205 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Amino.K., Y.Honda, C Ide, and T.Fujimoto: "Distribution of plasmalemmal Ca^<2+>-pump and caveolin in the corneal epithlium during wound healing process" Curr.Eye Res.16. 1088-1095 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Nomura,R., C.Inuo, Y.Takahashi, A.Asano and T.Fujimoto: "Two-dimensional distribution of Gi2a in the plasma momhrane : a critical evaluation by immunocyto chemistry" FEBS Lett. 415. 139-144 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Imhiki,M., J.Ando, R.Korenaga, H.Kogo T.Fujimoto, T.Fujita and A.Kamiya: "End・thelial Ca^<2+> waves preferentially ariginate at specific eoct in caveolin-rich cell ectges.," Proc.Natl.Acad.Sci.USA.in press.

    • Related Report
      1997 Annual Research Report
  • [Publications] Fujimoto,T.: "GPI-anchored proteins,glycosphingolipids,and sphingomyelin are distributed diffusely in untreated cells and sequestered to caveolae only after cross-linking." J. Histochem. Cytochem.44. 929-941 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Fujimoto,K.,M.Umeda,and T.Fujimoto: "Transmembrane phospholipid distribution revealed by freeze-fracture replica labeling." J. Cell Sci.109. 2453-2460 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] 藤本豊士: "細胞膜カベオラの機能" 医学のあゆみ. 176(6). 390-391 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] 藤本豊士: "細胞膜カベオラの新しい概念" MEBIO. 13(1). 157-159 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] 藤本豊士: "カベオラ-古くて新しいオルガネラ-" 解剖学雑誌. 71. 611-614 (1996)

    • Related Report
      1996 Annual Research Report

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Published: 1996-04-01   Modified: 2016-04-21  

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