DIFFERENTIAL ROLES OF THE SH2 DOMAINS OF PLC-GAMMA1 IN PDGF-INDUCED ACTIVATION

磷脂酶C-gamma1 （PLC-gamma1）包含两个串联SH结构域。在血小板衍生生长因子（PDGF）刺激细胞后，PLC-gamma1通过一个或两个SH2结构域与酪氨酸磷酸化的PDGF受体结合，被受体激酶磷酸化，从而被激活水解磷脂酰肌醇4,5-二磷酸。PLC-gamma1的两个SH2结构域在酶与PDGF受体相互作用中的单独作用现在已经通过功能性禁用每个结构域进行了研究。将每个SH2结构域的关键Arg残基突变为Ala，在PLC-gamma1缺失的成纤维细胞系中短暂表达野生型和突变型PLC-gamma1蛋白，并研究PDGF刺激转染细胞的影响。cooh末端SH2结构域失能的突变蛋白与PDGF受体结合，被受体磷酸化，并催化肌醇磷酸的产生，其程度与野生型酶相似。相反，nh2末端SH2结构域受损的突变体没有与PDGF受体结合，因此既没有磷酸化也没有激活。这些结果表明，pdgf诱导的PLC-gamma1激活需要PLC-gamma1的nh2末端SH2结构域，而不是cooh末端SH2结构域。-磷脂酶c - γ，血小板衍生生长因子，Src同源2域，蛋白酪氨酸激酶，磷酸肌肽

Phospholipase C-gamma1 (PLC-gamma1) contains two tandem SH domains. Upon stimulation of cells with platelet-derived growth factor (PDGF), PLC-gamma1 binds to the tyrosine-phosphorylated PDGF receptor through one or both of its SH2 domains, is phosphorylated by the receptor kinase, and is thereby activated to hydrolyze phosphatidylinositol 4,5- bisphosphate. The individual roles of the two SH2 domains of PLC-gamma1 in interaction of the enzyme with the PDGF receptor have now been investigated by functionally disabling each domain. A critical Arg residue of each SH2 domain was mutated to Ala, the wild-type and mutant PLC-gamma1 proteins were transiently expressed in a PLC-gamma1- deficient fibroblast cell line, and the effects of PDGF stimulation of the transfected cells were investigated. The mutant protein in which the COOH-terminal SH2 domain was disabled bound to the PDGF receptor, was phosphorylated by the receptor, and catalyzed the production of inositol phosphates to extents similar to those observed with the wild- type enzyme. In contrast, the mutant in which the NH2-terminal SH2 domain was impaired did not bind to the PDGF receptor and consequently was neither phosphorylated nor activated. These results suggest that the NH2-terminal SH2 domain, but not the COOH-terminal SH2 domain, of PLC-gamma1 is required for PDGF-induced activation of PLC-gamma1. - Phospholipase C-gamma, platelet-derived growth factor, Src homlogy 2 domain, protein tyrosine kinase, Phosphoinositide