Phosphoinositide-specific phospholipase C (PLC) plays a pivotal role in signal transduction from various receptor molecules on the plasma membrane. PLCvarepsilon is characterized by possession of two Ras/Rap-associating (RA) domains and a CDC25 homology domain acting as a guanine nucleotide exchange factor for Rap1. Our recent studies using PLCvarepsilon-deficient mice have suggested that PLCvarepsilon plays crucial roles in cardiac semilunar valvulogenesis downstream of the EGF receptor, as well as in chemical carcinogen-induced skin tumor development downstream of Ha-Ras. Stimulation of cultured mammalian cells with growth factors induces translocation of PLCvarepsilon from the cytoplasm to the plasma membrane and to the Golgi apparatus through direct association at its RA domains with the GTP-bound forms of Ras and Rap1, respectively. These results suggest that growth factor stimulation activates PLCvarepsilon by means of Ras and/or Rap1. However, growth factor-induced activation of the PLCvarepsilon lipase activity cannot be measured accurately because of simultaneous activation of PLCgamma through receptor-dependent phosphorylation. In this article, we introduce two methods to assay Ras- or Rap1-dependent activation of PLCvarepsilon lipase activity, with special emphasis on the use of cells expressing a mutant platelet-derived growth factor receptor lacking the PLCgamma-binding sites.
Ras and Rap1 Activation of PLCvarepsilon Lipase Activity. Publishing Authors By Initials
