PROTEIN-KINASE-B ACTIVATION AND LAMELLIPODIUM FORMATION ARE INDEPENDENT PHOSPHOINOSITIDE 3-KINASE-MEDIATED EVENTS DIFFERENTIALLY REGULATED BY ENDOGENOUS RAS

Regulation of phosphoinositide 3-kinase (PI 3-kinase) can occur by bin ding of the regulatory p85 subunit to tyrosine-phosphorylated proteins and by binding of the p110 catalytic subunit to activated Ras. Howeve r, the way in which these regulatory mechanisms act to regulate PI 3-k inase in vivo is unclear, Were we show that several growth factors (ba sic fibroblast growth factor [bFGF], platelet-derived growth factor [P DGF], and epidermal growth factor [EGF; to activate an EGF receptor-Re t chimeric receptor]) all activate PI3-kinase in vivo in the neuroecto derm-derived cell line SKF5. However, these growth factors differ in t heir ability to activate PI 3-kinase-dependent signaling. PDGF and EGF (Ret) treatment induced PI 3-kinase-dependent lamellipodium formation and protein kinase B (PKB) activation. In contrast, bFGF did not induc e lamelli-podium formation but activated PKB, albeit to a small extent . PDGF and EGF(Ret) stimulation resulted in binding of p85 to tyrosine -phosphorylated proteins and strong Ras activation, bFGF, however, ind uced only strong activation of Ras, In addition, while Ras(Asn17) abol ished bFGF activation of PKB, PDGF-and EGF(Rea)-induced PKB activation was only partially, inhibited and lamelli-podium formation was unaffe cted. Interestingly, in contrast to activation of only endogenous Ras (bFGF), ectopic expression of activated Ras did result in lamelli-podi um formation, From this we conclude that, in vivo, p85 and Ras synergi ze to activate PI 3-kinase and that strong activation of only endogeno us Bas exerts a small effect on PI 3-kinase activity, sufficient for P KB activation but not lamelli-podium formation. This differential sens itivity to PI3-kinase activation could be explained by our Ending that PKB activation and lamelli-podium formation are independent PI 3-kina se-induced events.