Signal transduction of Small GTP-binding proteins

有一个小的GTP结合蛋白(G蛋白)超家族，由60多个成员组成。所有成员都是相对分子质量在20,000到30,000之间的单体蛋白，既具有GDP/GTP结合活性，又具有GTP酶活性。在本研究项目中，我们对小G蛋白的功能、激活和作用方式进行了研究。RAS是这一超家族的代表，众所周知，RAS至少通过由MAP、MAP-K和MEK-K组成的MAP-K级联来调节基因的表达。我们从牛脑中纯化了一种在无细胞体系中以GTPGammaS-Ki-Ras依赖的方式激活MEK的MEK激酶，并鉴定其为B-Raf与14-3-3蛋白的络合物。我们还发现，RAPL在其效应域具有与RAS相同的氨基酸序列，也参与了B-Raf依赖的MAP激酶级联的激活。RHO调节肌动蛋白细丝的重组，至少与维持…有关哺乳动物细胞中更多的细胞形态、细胞运动性和胞质分裂。我们发现，当细胞受到HGF和TPA刺激时，Rho被移位到一个膜皱褶区。在胞质分裂过程中产生的收缩环上也观察到了Rho。ERM家族(Ezrin、Radixin和Moesin)位于这些区域，通过其C-末端区域与肌动蛋白细丝直接相互作用，并通过其N-末端区域与跨膜蛋白CD44相互作用。因此，Rho可能调节ERM家族依赖的肌动蛋白细丝与质膜结合的重组。我们还发现Rho GDI直接与ERM相互作用，通过降低Rho GDI的活性来启动Rho的激活。Rho也存在于酿酒酵母中，我们发现Rho在发芽过程中是必不可少的。我们从基因和生物化学的角度分离了Rho的调控蛋白(RDI1和ROM1/ROM2)和靶蛋白(PKC1、BNI1/BNR1、葡聚糖合成酶)。BNI1和BNR1在FH1结构域直接与Profilin相互作用，提示Rho通过BNI1/BNR1-Profilin系统调节肌动蛋白细丝的重组。Rab3A是RAB家族的成员，尤其与神经末梢的神经递质释放有关。在本研究中，我们还分离了与Rab3A、Rab3GEP和Rab3GAP相关的其他调控蛋白。较少

There is the small GTP-binding protein (G protein) superfamily which consists of over sixty members. All the 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. Ras is the representative of this superfamily and is well known to regulate 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. We also found that Rapl, which has the same amino acid sequence at its effector domain as that of Ras, is also involved in the activation of the B-Raf-dependent MAP kinase cascade. Rho regulates reorganization of actin filaments and is implicated in at least maintenance … More of cell morphology, cell motility, and cytokinesis in mammalian cells. We found that Rho is translocated to a membrane ruffling area when cells are stimulated by HGF and TPA.Rho was also observed at the contractile ring produced during cytokinesis. The ERM family (Ezrin, Radixin, and Moesin), which directly interacts with actin filaments 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 filaments with the plasma membrane. We also found that Rho GDI directly interacts with ERM,initiating the activation of Rho by reducing the Rho GDI activity. Rho also exists in the yeast Saccharomyces cerevisiae, and we found that Rho is essential for budding process. We genetically and biochemically isolated the regulatory proteins (RDI1 and ROM1/ROM2) and target proteins (PKC1, BNI1/BNR1, glucan synthase) for Rho. BNI1 and BNR1 directly interact with profilin at the FH1 domains, suggesting that Rho regulates the reorganization of actin filaments through the BNI1/BNR1-profilin system. Rab3A is a member of the Rab family which is particularly implicated in neurotransmitter release from the nerve terminal. 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,Rab3 GEP and Rab3 GAP. Less