cDNAs and antibodies for small GTP-binding proteins

小 GTP 结合蛋白的 cDNA 和抗体

• 批准号: 06557012
• 负责人: TAKAI Yoshimi
• 金额: $ 11.84万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06557012/
• 关键词: small GTP-binding protein; Ras; B-Raf; Rho; The ERM family; CD44; Rab3A; Rabphilin-3A; 低分子量G蛋白質; Rap1; SmgGDS; SMAP; 14-3-3蛋白質; ERM-CD44系

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

有一个小的GTP结合蛋白（G蛋白）超家族由五十多名成员组成。所有成员都是单体蛋白质，MRS具有20,000至30,000之间，均表现出GDP/GTP结合和GTPase活动。在本研究项目中，我们研究了小G蛋白的激活和作用的功能和模式，并试图准备其cDNA和抗体作为诊断和治疗各种疾病的工具。 RAS是该超家族的代表，众所周知，可以至少通过MAP激酶级联反应基因表达，由MAP激酶，MAP激酶激酶（MEK）和MEK激酶组成。我们从牛脑中纯化了MEK激酶，该激酶在无细胞系统中以GTPGAMMAS-KI-RAS依赖性方式激活MEK，并将其确定为B-RAF与14-3-3-3蛋白复合的B-RAF。 RHO调节肌动蛋白丝的重组，并至少在哺乳动物细胞中至少维持细胞形态，细胞运动和细胞因子性。我们发现……当细胞被HGF和TPA刺激时，更多的HAT RHO被转化为膜荷叶边区域。在细胞因子期间产生的收缩环也观察到了Rho。 ERM家族（Ezrin，Radixin和Moesin）通过其C末端区域与肌动蛋白丝进行直接相互作用，并与跨膜蛋白CD44通过其N末端区域相互作用，因此位于这些区域。因此，Rho可能会调节ERM家族依赖性含量含量的ERM家族含量相关的重组。 Rho也存在于酿酒酵母的酵母糖疗法中，我们发现Rho1对于萌芽过程至关重要。我们将rho1.rab3a的一般和生化分离的调节蛋白（RDI1和ROM1/ROM2）和靶蛋白（PKC1，BNI1，葡聚糖合酶）是RAB家族的成员，在Neurotransmitter从Never Termini释放出来。我们已经隔离了其调节蛋白，称为Rab GDI，其靶蛋白名为Rabphilin-3A.在该研究项目中，我们隔离了Rab3a，Rab3a Gep和Rab3a Gap的其他调节蛋白。该研究项目中获得的结果表明，小型G蛋白在各种细胞功能中均可用于各种诊断和替代工具，这些蛋白质在各种CDEN和Antibs中都具有重要的作用。较少的

项目成果

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)

Takaishi, K.：“活化的 Rho 从细胞质转移到膜皱褶区域、细胞间粘附位点和裂解沟。”

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)

Nonaka, H.：“RHO1 小 GTP 结合蛋白的下游靶标是 PKC1，它是蛋白激酶 C 的同源物，可导致酿酒酵母中 MAP 激酶级联的激活。”

Ohtsuka, T.: "Activation of brain B-Raf protein kinase by Rap1B small GTP-binding protein." J.Biol.Chem.271. 1258-1261 (1996)

Ohtsuka, T.：“Rap1B 小 GTP 结合蛋白激活大脑 B-Raf 蛋白激酶。”

Nishiyama,T.: "Rac p21 is involved in insulin-induced mambrane 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)

Nishiyama,T.：“Rac p21 参与胰岛素诱导的膜波动，rho p21 参与肝细胞生长因子和 12-O-十四烷酰佛波醇-13-乙酸酯 (TPA) 诱导的 KB 细胞膜波动。”

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)

Nonaka H.：“RHO1 小 GTP 结合蛋白的下游靶标是 PKC1，它是蛋白激酶 C 的同源物，可导致酿酒酵母中 MAP 激酶级联的激活。”