Platelet-Derived Growth Factor-Dependent Activation of Phosphatidylinositol 3-Kinase Is Regulated by Receptor Binding of SH2-Domain-Containing Proteins Which Influence Ras Activity

Abstract

Upon binding of platelet-derived growth factor (PDGF), the PDGF β receptor (PDGFR) undergoes autophosphorylation on distinct tyrosine residues and binds several SH2-domain-containing signal relay enzymes, including phosphatidylinositol 3-kinase (P13K), phospholipase Cγ (PLCγ), the GTPase- activating protein of Ras (RasGAP), and the tyrosine phosphatase SHP-2. In this study, we have investigated whether PDGF-dependent P13K activation is affected by the other proteins that associate with the PDGFR. We constructed and characterized a series of PDGFR mutants which contain binding sites for P13K as well as one additional protein, either RasGAP, SHP-2, or PLCγ. While all of the receptors had wild-type lev els of PDGF-stimulated tyrosine kinase activity and associated with comparable amounts of P13K activity, their abilities to trigger accumulation of P13K products in vivo differed dramatically. The wild-type receptor, as well as receptors that recruited P13K or P13K and SHP-2, were all capable of fully activating P13K. In contrast, receptors that associated with P13K and RasGAP or P13K and PLCγ displayed a greatly reduced ability to stimulate production of P13K products. When this series of receptors was tested for their ability to activate Ras, we observed a strong positive correlation between Ras activation and P13K activation. Further investigation of the relationship between Ras and P13K indicated that Ras was upstream of P13K. Thus, activation of P13K requires not only binding of P13K to the tyrosine-phosphorylated PDGFR but accumulation of GTP-bound Ras as well. Furthermore, PLCγ and RasGAP negatively modulate PDGF-dependent P13K activation. Finally, PDGF-stimulated signal relay can be regulated by altering the ratio of SH2-domain-containing enzymes that are recruited to the PDGFR.