Phospholipase Cε:: a novel Ras effector
Phospholipase Cε:: a novel Ras effector
复制标题
DOI：
10.1093/emboj/20.4.743
复制
发表时间：
2001-02-15
期刊：
影响因子：
11.4
通讯作者：
Smrcka, AV
中科院分区：
文献类型：
--
作者：
Kelley, GG;Reks, SE;Smrcka, AV
哺乳动物磷酸肌醇特异性磷脂酶C（PLC）的三个类别，PLC β，PLC γ和PLC δ，其差异调节异源三聚体G蛋白，酪氨酸激酶和钙的特点。在这里，我们描述了第四类，PLC β，除了保守的PLC结构域，包含一个GTP交换因子（GRF CDC 25）结构域和两个C-末端Ras结合（RA）结构域，RA 1和RA 2，RA 2结构域结合H-Ras在GTP依赖的方式，与Ras结合结构域的Raf-1相比，然而，RA 1结构域结合H-Ras与GTP无关的方式具有低亲和力。虽然G α（q）、G β γ或令人惊讶的H-Ras在体外不激活重组纯化蛋白，但组成型活性Q61 L H-Ras刺激在COS-7细胞中与Ras结合平行共表达的PLC β。RA 1或RA 2结构域的缺失抑制这种激活。RA 2结构域或Ras的定点突变证明了保守的Ras-效应子相互作用和Ras效应子结构域突变体的独特激活特征。这些研究鉴定了一种新的第四类哺乳动物PLC，其直接受Ras调节并连接两个关键信号通路。
Three classes of mammalian phosphoinositide-specific phospholipase C (PLC) have been characterized, PLC beta, PLC gamma and PLC delta, that are differentially regulated by heterotrimeric G-proteins, tyrosine kinases and calcium. Here we describe a fourth class, PLC epsilon, that in addition to conserved PLC domains, contains a GTP exchange factor (GRF CDC25) domain and two C-terminal Ras-binding (RA) domains, RA1 and RA2, The RA2 domain binds H-Ras in a GTP-dependent manner, comparable with the Ras-binding domain of Raf-1; however, the RA1 domain binds H-Ras with a low affinity in a GTP-independent manner. While G alpha (q), G beta gamma or, surprisingly, H-Ras do not activate recombinant purified protein in vitro, constitutively active Q61L H-Ras stimulates PLC epsilon co-expressed in COS-7 cells in parallel with Ras binding. Deletion of either the RA1 or RA2 domain inhibits this activation. Site-directed mutagenesis of the RA2 domain or Ras demonstrates a conserved Ras-effector interaction and a unique profile of activation by Ras effector domain mutants, These studies identify a novel fourth class of mammalian PLC that is directly regulated by Ras and links two critical signaling pathways.