PLATELET-DERIVED GROWTH FACTOR-DEPENDENT ACTIVATION OF PHOSPHATIDYLINOSITOL 3-KINASE IS REGULATED BY RECEPTOR-BINDING OF SH2-DOMAIN-CONTAINING PROTEINS WHICH INFLUENCE RAS ACTIVITY

Upon binding of platelet-derived growth factor (PDGF), the PDGF beta r eceptor (PDGFR) undergoes autophosphorylation on distinct tyrosine res idues and binds several SH2-domain-containing signal relay enzymes, in cluding phosphatidylinositol 3-kinase (PI3K), phospholipase C gamma (P LC gamma), the GTPase-activating protein of Ras (RasGAP), and the tyro sine phosphatase SHP-2, In this study, we have investigated whether PD GF-dependent PI3K activation is affected by the other proteins that as sociate with the PDGFR, We constructed and characterized a series of P DGFR mutants which contain binding sites for PI3K as well as one addit ional protein, either RasGAP, SHP-2, or PLC gamma, While all of the re ceptors had wild-type levels of PDGF-stimulated tyrosine kinase activi ty and associated with comparable amounts of PI3K activity, their abil ities to trigger accumulation of PI3K products in vivo differed dramat ically, The wild-type receptor, as well as receptors that recruited PI 3K or PI3K and SHP-2, were all capable of fully activating PI3K. In co ntrast, receptors that associated with PI3K and RasGAP or PI3K and PLC gamma displayed a greatly reduced ability to stimulate production of PI3K products, When this series of receptors was tested for their abil ity to activate Ras, we observed a strong positive correlation between Ras activation and PI3K activation, Further investigation of the rela tionship between Ras and PI3K indicated that Ras was upstream of PI3K. Thus, activation of PI3K requires not only binding of PI3K to the tyr osine-phosphorylated PDGFR but accumulation of GTP-bound Ras as well, Furthermore, PLC gamma and RasGAP negatively modulate PDGF-dependent P I3K activation, Finally, PDGF-stimulated signal relay can be regulated by altering the ratio of SH2-domain-containing enzymes that are recru ited to the PDGFR.