Regulation of a novel human phospholipase C, PLCε, through membrane targeting by Ras

Phosphoinositide-specific phospholipase C (PI-PLG) plays a pivotal role in regulation of intracellular signal transduction from various receptor molecules. More than 10 members of human PI-PLG isoforms have been identified and classified into three classes beta, gamma, and delta, which are regulated by distinct mechanisms. Here we report identification of a novel class of human PI-PLG, named PLG epsilon, which is characterized by the presence of a Res-associating domain at its C terminus and a CDC25-like domain at its N terminus. The Res-associating domain of PLGe specifically binds to the GTP-bound forms of Ha-Ras and Rap1A. The dissociation constant for Ha-Ras is estimated to be approximately 40 nM, comparable with those of other Res effecters. Co-expression of an activated Ha-Res mutant with PLGe induces its translocation from the cytosol to the plasma membrane. Upon stimulation with epidermal growth factor, similar translocation of ectopically expressed PLGe is observed, which is inhibited by co-expression of dominant-negative Ha-Res. Furthermore, using a liposome-based reconstitution assay, it is shown that the phosphatidylinositol 4,5-bisphosphate-hydrolyzing activity of PLC epsilon is stimulated in vitro by Ha-Ras in a GTP-dependent manner. These results indicate that Res directly regulates phosphoinositide breakdown through membrane targeting of PLCe.