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1995 Fiscal Year Final Research Report Summary

cDNAs and antibodies for small GTP-binding proteins

Research Project

Project/Area Number 06557012
Research Category

Grant-in-Aid for Developmental Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field Pathological medical chemistry
Research InstitutionOsaka University

Principal Investigator

TAKAI Yoshimi  Osaka University Medical School, Professor, 医学部, 教授 (60093514)

Co-Investigator(Kenkyū-buntansha) KAWATA Sumio  Osaka University Medical School, Associate Professor, 医学部, 助教授 (90183285)
SASAKI Takuya  Osaka University Medical School, Research Associate, 医学部, 助手 (40241278)
TANAKA Kazuma  Osaka University Medical School, Assistant Professor, 医学部, 講師 (60188290)
Project Period (FY) 1994 – 1995
Keywordssmall GTP-binding protein / Ras / B-Raf / Rho / The ERM family / CD44 / Rab3A / Rabphilin-3A
Research Abstract

There is the small GTP-binding protein (G protein) superfamily wich consists of over fifty members. All members are monomeric proteins with Mrs between 20,000 and 30,000 which exhibit both GDP/GTP-binding and GTPase activities. In the present research project, we studied the functions and modes of activation and action of small G proteins and tried to prepare their cDNAs and antibodies as tools for diagnosis and treatment of various diseases. Ras is the representative of this superfamily and is well known to requlate gene expression at least through the MAP kinase cascade consisting of MAP kinase, MAP kinase kinase (MEK), and MEK kinase. We purified from bovine brain a MEK kinase which activated MEK in a GTPgammaS-Ki-Ras-dependent manner in a cell-free system and identified it as B-Raf complexed with 14-3-3 proteins. Rho regulates reorganization of actin filaments and is implicated in at least maintenance of cell morphology, cell motility, and cytokinesis in mammalian cells. We found t … More hat Rho is translocated to a membrane ruffling area when cells are stimulated by HGF and TPA.Rho was also translocated to the cell-cell adhesion area, which was induced by Ca^<2+>. Rho was also observed at the contractile ring produced during cytokinesis. The ERM family (Ezrin, Radixin, and Moesin), which directly interacts with actin filament through its C-terminal region and with a transmembrane protein, CD44, through its N-terminal region, is located at these areas.
Therefore, Rho may regulate reorganization of the ERM family-dependent association of actin filament with the plasma membrane. Rho also exists in the yeast Saccharomyces cerevisiae, and we found that RHO1 is essential for budding process. We genetically and biochemically isolated the regulatory proteins (RDI1 and ROM1/ROM2) and target proteins (PKC1, BNI1, glucan synthase) for RHO1.
Rab3A is a member of the Rab family which is particularly implicated in neurotransmitter release from the nerve termini. We had isolated its regulatory protein, named Rab GDI,and its target protein, named Rabphilin-3A.In this research project, we isolated other regulatory proteins for Rab3A,Rab3A GEP,and Rab3A GAP.The results obtained in this research project suggest that small G proteins play crucial roles in various cell functions and that their cDNAs and antibodies are useful as tools for diagnosis and treatment of various diseases. Less

  • Research Products

    (56 results)

All Other

All Publications (56 results)

  • [Publications] Miki H.: "Association of Ash/Grb-2 with dynamin through the Src homology 3 domain." J.Biol.Chem.269. 5489-5492 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nishimura N.: "Molecular cloning and characterization of two rab GDI species from rat brain-Brain-specific and ubiquitous types." J.Biol.Chem.269. 14191-14198 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nakanishi H.: "Different functions of smq GDP dissociation stimulator and mammalian counterpart of yeast Cdc25." J.Biol.Chem.269. 15085-15091 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Masuda T.: "Molecular cloning and characterization of yeastrho GDP dissociation inhibitor." J.Biol.Chem.269. 19713-19718 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Shimizu K.: "Synergistic activation by Ras and 14-3-3 protein of a mitogen-activated protein kinase kinase kinase named Ras-dependent extracellular signal-regulated kinase kinase stimulator." J.Biol.Chem.269. 22917-22920 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Shirataki H.: "Rabphilin-3A is associated with synaptic vesicle through a vesicleprotein in a manner independent of Rab3A." J.Biol.Chem.269. 32717-32720 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nishiyama T.: "Rac p21 is involved in insulin-induced membrane ruffling and rho p21 is involved in hepatocyte growth factor-and 12-O-tetradecanoylphrbol-13-acetate (TPA)-induced membrane ruffling in KB cells." Mol.Cell.Biol.14. 2447-2456 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Yamochi W.: "Qrowth site-localization of Rho1 small GTP-binding protein and its involvement in bud formation in Saccharomyces cerevisiae." J.Cell.Biol.125. 1077-1093 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Huber L.A.: "Mapping of ras-related GTP-binding proteins by GTP overlay follwing two-dimensional gel electrophoresis." Proc.Natl.Acad.Sci.91. 7874-7878 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Takaishi K.: "Involvement of Rho p21 small GTP-binding protein and its regulator in the HGF-induced cell motility." Oncogene. 9. 273-279 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kuroda S.: "Purification and characterization of REKS from Xenopus eggs-Identification of REKS as a Ras-dependent mitogen-activated protein kinase kinase kinase" J.Biol.Chem.270. 2460-2465 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Bourgoin S.: "Low molecular weight GTP-binding proteins in HL-60 granulocytes." J.Biol.Chem.270. 3172-3178 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Yamamori B.: "Purification of a Ras-dependent mitogen-activated protein kinase kinase kinase from bovine brain cytosol and its identification as a complex of B-raf and 14-3-3 proteins." J.Biol.Chem.270. 11723-11726 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Chung S-H.: "Evidance that the Rab3A-binding protein,Rabphilin-3A,enhances requlated secretion-Studies in adrenal chromaffin cells." J.Biol.Chem.270. 16714-16718 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kuribara H.: "Synergistic activation of rat brain phospholipase D by ADP-ribosylation factor and rhoA p21,and its inhibition by clostridium botulinum C3 exoenzyme." J.Biol.Chem.270. 25667-25671 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] McGeady P.: "The farnesyl group of H-ras facilitates the activation of a soluble upstream activator of mitogen-activated protein kinase." J.Biol.Chem.270. 26347-26351 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Gotoh T.: "Idetification of Rapl as a target for the Crk SH3 Domain-binding guanine nucleotide-releasing factor C3G." Mol.Cell.Biol.15. 6746-6753 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nonaka H.: "A dowstream target of RHO1 small GTP-binding protein is PKC1,a homolog of protein kinase C,which leads to activation of the MAP kinase cascade in Saccharomyces cerevisiae." EMBO J.14. 5931-5938 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Takaishi K.: "Translocation of activated Rho from the cytoplasm to membrane ruffling area,cell-cell adhesion sites and cleavage furrows." Oncogene. 11. 39-48 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nishimura N.: "Cloning of a brain-type isoform of human rab GD1 and its expression in human neroblastoma cell lines and tumor specimens." Cancer Res.55. 5445-5450 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Takai Y.: "Rho as a regulator of the cytoskeleton." Trends in Biochemical Sciences. 20. 227-231 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Ohtsuka T.: "Activation of brain B-Raf protein kinase by RaplB small GTP-binding protein." J.Biol.Chem.271. 1258-1261 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Orita S.: "Doc2 enhances Ca2+-dependent exocytosis from PC12 cells." J.Biol.Chem.(in press).

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Fujita Y.: "Phosphorylation of Munc-18/n-Secl/rbSel by protein kinase C-Its implication in regulating the interaction of Munc-18/n-Secl/rbSec1 with Syntaxin." J.Biol.Chem.(in press).

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Drgonova J.: "RholP,a yeast protein at the juncture between cell polarization and morphogenesis." Science. (in press).

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Ozaki K.: "Romlp and Rom2p are GDP/GTP exchange proteins (GEPs) for the Rholp small GTP-binding protein in Saccharomyces cerevisiae." EMBO J.(in press).

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Matsuda S.: "A membrane-associated GDF/GTP exchange protein specific for Rho small GTP-binding protein-partial purification and characterization from rat brain." Oncogene. (in press).

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Taniguchi T.: "Cholecystokinin-B/gastrin receptors mediate rapid formation of actin stress fibers." Oncogene. (in press).

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Miki, H.: "Association of Ash/Grb-2 with dynamin through the Src homology 3 domain." J.Biol.Chem.269. 5489-5492 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nishimura, N.: "Molecular cloning and characterization of two rab GDI species from rat brain-Brain-specific and ubiquitous types." J.Biol.Chem.269. 14191-14198 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nakanishi, H.: "Different functions of smg GDP dissociation stimulator and mammalian counterpart of yeast Cdc25." J.Biol.Chem.269. 15085-15091 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Masuda, T.: "Molecular cloning and characterization of yeast rho GDP dissociation inhibitor." J.Biol.Chem.269. 19713-19718 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Shimizu, K.: "Synergistic activation by Ras and 14-3-3 protein of a mitogen-activated protein kinase kinase kinase named Ras-dependent extracellular signal-regulated kinase kinase stimulator." J.Biol.Chem.269. 22917-22920 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Shirataki, H.: "Rabphilin-3A is associated withsynaptic vesicle through a vesicle protein in a manner independent of Rab3A." J.Biol.Chem.269. 31717-32720 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nishiyama, T.: "Rac p21 is involved in insulin-induced membrane ruffling and rho p21 is involved in hepatocyte growth factor- and 12-O-tetradecanoylphorbol-13-acetate (TPA) -induced membrane ruffling in KB cells." Mol.Cell.Biol.14. 2447-2456 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yamochi, W.: "Growth site-localization of Rho1 small GTP-binding protein and its involvement in bud formation in Saccharomyces cerevisiae." J.Cell Biol.125. 1077-1093 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Huber, L.A.: "Mapping of ras-related GTP-binding proteins by GTP overlay following two-dimensional gel electrophoresis." Proc.Natl.Acad.Sci.USA. 91. 7874-7878 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Takaishi, K.: "Involvement of Rho p21 small GTP-binding protein and its regulator in the HGF-induced cell motility." Oncogene. 9. 273-279 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kuroda, S.: "Purification and characterization of REKS from Xenopus eggs-Identification of REKS as a Ras-dependent mitogen-activated protein kinase kinase kinase." J.Biol.Chem.270. 2460-2465 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Bourgoin, S.: "Low molecular weight GTP-binding proteins in HL-60 granulocytes." J.Biol.Chem.270. 3172-3178 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yamamori, B.: "Purification of a Ras-dependent mitogen-activated protein kinase kinase kinase from bovine brain cytosol and its identification as a complex of B-raf and 14-3-3 proteins." J.Biol.Chem.270. 11723-11726 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Chung, S-H.: "Evidence that the Rab3A-binding protein, Rabphilin-3A,enhances regulated secretion-Studies in adrenal chromaffin cells." J.Biol.Chem.270. 16714-16718 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kuribara, H.: "Synergistic activation of rat brain phospholipase D by ADP-ribosylation factor and rhoA p21, and its inhibition by clostridium botulinum C3 exoenzyme." J.Biol.Chem.270. 25667-25671 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] McGeady, P.: "The farnesyl group of H-ras facilitates the activation of a soluble upstream activator of mitogen-activated protein kinase." J.Biol.Chem.270. 26347-26351 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Gotoh, T.: "Identification of Rap1 as a target for the Crk SH3 Domain-binding guanine nucleotide-releasing factor C3G." Mol.Cell.Biol.15. 6746-6753 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nonaka, H.: "A downstream target of RHO1 small GTP-binding protein is PKC1, a homolog of protein kinase C,which leads to activation of the MAP kinase cascade in Saccharomyces cerevisiae." EMBO J.14. 5931-5938 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Takaishi, K.: "Translocation of activated Rho from the cytoplasm to membrane ruffling area, cell-cell adhesion sites and cleavage furrows." Oncogene. 11. 39-48 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nishimura, N.: "Cloning of a brain-type isoform of human rab GDI and its expression in human neroblastoma cell lines and tumor specimens." Cancer Res.55. 5445-5450 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Takai, Y.: "Rho as a regulator of the cytoskeleton." Trends in Biochemical Sciences. 20. 227-231 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Ohtsuka, T.: "Activation of brain B-Raf protein kinase by Rap1B small GTP-binding protein." J.Biol.Chem.271. 1258-1261 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Orita, S.: "Doc2 enhances Ca^<2+>-dependent exocytosis from PC12 cells." J.Biol.Chem.(in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Fujita, Y.: "Phosphorylation of Munc-18/n-Sec1/rbSec1 by protein kinase C-Its implication in regulating the interaction of Munc-18/n-Sec1/rbSec1 with Syntaxin." J.Biol.Chem.(in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Drgonova, J.: "Rho1P,a yeast protein at the juncture between cell polarization and morphogenesis." Science. (in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Ozaki, K.: "Rom1p and Rom2p are GDP/GTP exchange proteins (GEPs) for the Rho1p small GTP-binding protein in Saccharomyces cerevisiae." EMBO J.(in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Matsuda, S.: "A membrane-associated GDP/GTP exchange protein specific for Rho small GTP-binding protein-partial purification and characterization from rat brain." Oncogene. (in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Taniguchi, T.: "Cholecystokinin-B/gastrin receptors mediate rapid formation of actin stress fibers." Oncogene. (in press). (1996)

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 1997-03-04  

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