• Search Research Projects
  • Search Researchers
  • How to Use
  1. Back to previous page

Molecular Biological Analysis for Mechanism of Thombosis Regulation and Its APlication for Clinical Desease.

Research Project

Project/Area Number 07457231
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Hematology
Research InstitutionNagoya University

Principal Investigator

SAITO Hidehiko  Nagoya U Sch Med, 1st Dep Int Med, Proffeser, 医学部, 教授 (20153819)

Co-Investigator(Kenkyū-buntansha) KOJIMA Tetsuhito  Nagoya U Sch Med, 1st Dep Int Med, Assistant Proffeser, 医学部, 助手 (40161913)
TANIMOTO Mitsune  Nagoya U Sch Med, 1st Dep Int Med, Assiatant Proffeser, 医学部, 助手 (10240805)
Project Period (FY) 1995 – 1996
Project Status Completed (Fiscal Year 1996)
Budget Amount *help
¥7,200,000 (Direct Cost: ¥7,200,000)
Fiscal Year 1996: ¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 1995: ¥4,400,000 (Direct Cost: ¥4,400,000)
KeywordsThrombophilia / protein C deficiency / protein S deficiency / impaired secretion / endothelium / heparan sulfate proteoglycan / ryudocan gene / promoter activity / 生理的凝固阻止因子 / プロテインC / プロテインS / Ryudocan
Research Abstract

To determine the subcellular localization of the protein C (PC) Nagoya, an elongated variant of the human PC,the recombinant PC bearing this mutation was expressed in Chinese hamster ovary cells. Immunoelectron microscopy indicated that PC Nagoya was relained in the ER,whereas wild type PC was observed in both the ER and the Golgi apparatus. Metabolic radiolabeling with [35S] methionine in combination with chemical cross-linking revealed that the PC Nagoya existed in the ER as an complex with GRP78 and GRP94. Because both GRP78 and GRP94 associate far lesser degree with wild type PC than with PC Nagoya, our data suggest that both stress proteins function as molecular chaperones, and work in concert with the folding and assembly of PC.
DNA sequence analysis in the proband with a hereditary type I protein S (PS) deficiency showed a novel missense mutation substituting Cys (TGT) for Arg474 (CGT). Stable expression and pulse-chase experiments demonstrated an intracellular degradation and an impaired secretion of the recombinant Cys-mutant PS.Furthermore, the substitution of Arg 474 by Ala or Clu, but not Lys. markedly reduced the secretion of the recombinant PS mutant, suggesting that a positively charged basic amino acid might be needed at residue 474 and might play a key role in the protein structure and conformation of the sex hormone binding globulin-homology domain of the PS molecule.
It was found that basic fibroblast growth factor (bFGF), midkine (MK), and tissue factor pathway inhibitor (TFPI) exhibited significant ryudocan binding through its heparan sulfate chains. Immuno-histochemical analysis revealed that ryudocan was expressed in peripheral nerve tissues, fibrous connective tissues, and placental trophoblasts. These observations suggest that ryudocan may possess multiple biologic functions, such as bFGF modulation, neurite growth promotion, and anticoagulation, via heparan sulfate binding effectors present in the cellular microenvironment.

Report

(3 results)
  • 1996 Annual Research Report   Final Research Report Summary
  • 1995 Annual Research Report
  • Research Products

    (30 results)

All Other

All Publications (30 results)

  • [Publications] T. Yamazaki, H. Saito, etal.: "A. Quantitative Protein S Deficiency Associated with a Novel Nonsense Mutation and Markedly Reduced Levels of Mutated mRNA." Thromb. Haemost.74(2). 590-595 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H. Toyozumi, H. Saito, et al.: "Diagnosis of Hemophilia B Carriers Using Two Novel Dinucleotide Polymorphisms and Hha I RFLP of the Factor IX Gene in Japanese Subjects." Thromb. Haemost.74(4). 1009-1014 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T. Kojima, H. Saito, et al.: "Human Ryudocan from Endothelium-like Cells Binds Basic Fibroblast Growth Factor, Midkine, and Tissue Factor Pathway Inhibitor." J. Biol. Chem.271(10). 5914-5920 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] A. Takagi, H. Saito, et al.: "Structural Organization and Promoter Activity of the Human Ryudocan Gene.21GC04:J. Biochem. (Tokyo)" 119(5). 979-984 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] A. Katsumi, H. Saito, et al.: "Protein C Nagoya, an Elongated Mutant of Protein C, Is Retained Within the Endoplasmic Reticulum and Is Associated With GRP78 and GRP94." Blood. 87(10). 4164-4175 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T. Yamazaki, H. Saito, et al.: "Molecular Basis of a Hereditary Type I Protein S Deficiency Caused By a Substitution of Cys for Arg474." Blood. 87(11). 4643-4650 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H. Saito, and T. Kojima: "Factor XII, prekallikrein, and high-molecular-weight kininogen. K.A. High, and H. R. Roberts, eds ; Molecular Basis of Thrombosis and Hemostasis." Marcel Dekker, Inc., New York・Basel・Hong kong, 269-285 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H. Saito, J. Takamatsu: "Disorders of prothrombin conversion. Gross S. and Roath S edits ; Hematology, A problem-oriented approach." Williams & Wilkins, Baltimore, 569-578 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T.Yamazaki, H.Saito, et al.: "A Quantitative Protein S Deficiency Associated with a Novel Nonsense Mutation and Markedly Reduced Levels of Mutated mRNA." Thromb.Haemost.74(2). 590-595 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H.Toyozumi, H.Saito, et al.: "Diagnosis of Hemophilia B Carriers Using Two Novel Dinucleotide Polymorphisms and Hha I RFLP of the Factor IX Gene in Japanese Subjects." Thromb.Haemost.74(4). 1009-1014 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T.Kojima, H.Saito, et al.: "Human Ryudocan from Endothelium-like Cells Binds Basic Fibroblast Growth Factor, Midkine, and Tissue Factor Pathway Inhibitor." J.Biol.Chem.271(10). 5914-5920 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] A.Takagi, H.Saito, et al.: "Structural Organization and Promoter Activity of the Human Ryudocan Gene." J.Biochem., (Tokyo). 119(5). 979-984 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] A.Katsumi, H.Saito, et al.: "Protein C Nagoya, an Elongated Mutant of Protein C,Is Retained Within the Endoplasmic Reticulum and Is Associated With GRP78 and GRP94." Blood. 87(10). 4164-4175 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T.Yamazaki, H.Saito, et al.: "Molecular Basis of a Hereditary Type I Protein S Deficiency Caused By a Substitution of Cys for Arg474." Blood. 87(11). 4643-4650 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H.Saito, and T.Kojima: Factor XII,prekallikrein, and high-molecular-weight kininogen.Molecular Basis of Thrombosis and Hemostasis. : K.A.High, and H.R.Roberts, eds, Marcel Dekker, Inc., New York・Basel・Hong Kong, 269-285 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] H.Saito, J.Takamatsu: Disorders of prothrombin conversion.Hematology, A problem-oriented approach. : Gross S.and Roath S edits, Williams & Wilkins, Baltimore, 569-578 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] T.Kojima,H.Saito,et al.: "Human Ryudocan from Endothelium-like Cells Binds Basic Fibroblast Growth Factor,Midkine,and Tissue Factor Pathway Inhibitor." J.Biol.Chem.271(10). 5914-5920 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] A.Takagi,H,Saito,et al.: "Structural Organization and Promoter Activity of the Human Ryudocan Gene." J.Biochem.(Tokyo). 119(5). 979-984 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] T.Yamazaki,H.Saito,et al.: "Analysis for anthrombin gene polymorphisms in Japanese subjects and cosegregation studies families with hereditary antithrombin deficiency." Thromb.Res.82(3). 275-280 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] A.Katsumi,H.Saito,et al.: "Protein C Nagoya,an Elongated Mutant of Protein C,Is Retained Within the Endoplasmic Reticulum and Is Associated with GRP78 and GRP94." Blood. 87(10). 4164-4175 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] T.Yamazaki,H.Saito,et al.: "Molecular Basis of a Hereditary Type I Protein S Deficiency Caused By a Substitution of Cys for Arg474." Blood. 87(11). 4643-4650 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Y.Okamoto,T.Kojima,et al.: "A Novel Nonsense Mutation Associated with an Exon Skipping in a Patient with Protein S Deficiency Type I" Thromb.Haemost.75(6). 877-882 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] H.Saito,J.Takamatsu: "Disorders of prothrombin conversion. Gross S.and Roath S edits ; Hematology,A problem-oriented approach." Williams & Wilkins,Baltimore, 569-578 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] H.Saito et al.: "Depolymeriged Holothurian Glycosaminoglycan (DHG) with Novel Anticoagulant Actions Antithrombin III-and Heparin Cofador II-Independent Inhibition of Factor X Activation by……" Blood. 85. 1527-1534 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] H.Saito et al.: "Diagnosis of Hemophilia B Carriers Using Two Novel Dinucleotide Polymorphisms and Hha I RFLP of the Factor IX gene in Japanese Subjects." Thromb Haemost. 74. 1009-1014 (1975)

    • Related Report
      1995 Annual Research Report
  • [Publications] H.Saito et al.: "A Quanfitafloc Protein S Deficieucy Associated with a Novel Nonseuse Mutation and Markedly Reduced Level of Mutated mRNA." Thromb Haemost. 74. 590-595 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] H.Saito et al.: "Protein C Nagoya,an Elongated Mutant of Protein C,is Retainod within the Eudoplasuic Reticuluns and is Associated with GRP78 and GRP94." Blood. (in press). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] H.Saito et al: "Molecular Basis of a Hereditavy Type I Protein S Doficiency Cansat by a Substitution of Cys for Arg 474." Blood. (in press). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] H.Saito et al.: "Human Ryudocan from Endothelial-like Cells binds Basic Fibroblast Growth Factor,Midkine,and Tissue Factor Pithway Inhibitor" J.Biol Chem.(in press). (1996)

    • Related Report
      1995 Annual Research Report
  • [Publications] H.Saito et al.: "Factor-XII,Prekallikrein,and High-Mdecula-Weight Kininogen In htolccular Basis of Thronbosis and Hemostasis." K.H.High and HR.Roberts (eds) . Marcel Dekker Inc., 17 (1995)

    • Related Report
      1995 Annual Research Report

URL: 

Published: 1995-04-01   Modified: 2016-04-21  

Information User Guide FAQ News Terms of Use Attribution of KAKENHI

Powered by NII kakenhi