Osteopontin stimulates cell motility and nuclear factor kappaB-mediated secretion of urokinase type plasminogen activator through phosphatidylinositol 3-kinase/Akt signaling pathways in breast cancer cells.

We have recently reported that osteopontin (OPN) induces nuclear factor kappaB (NFkappaB)-mediated promatrix metalloproteinase-2 activation through IkappaBalpha/IKK signaling pathways and that curcumin (diferulolylmethane) down-regulates these pathways (Philip, S., and Kundu, G. C. (2003) J. Biol. Chem. 278, 14487-14497). However, the molecular mechanism by which upstream kinases regulate the OPN-induced NFkappaB activation and urokinase type plasminogen activator (uPA) secretion in human breast cancer cells is not well defined. Here we report that OPN induces the phosphatidylinositol 3'-kinase (PI 3'-kinase) activity and phosphorylation of Akt in highly invasive MDA-MB-231 and low invasive MCF-7 cells. The OPN-induced Akt phosphorylation was inhibited when cells were transfected with a dominant negative mutant of the p85 domain of PI 3-kinase (Deltap85) and enhanced when cells were transfected with an activated form of PI 3-kinase (p110CAAX), indicating that PI 3'-kinase is involved in Akt phosphorylation. OPN enhances the interaction between IkappaBalpha kinase (IKK) and phosphorylated Akt. OPN also induces NFkappaB activation through phosphorylation and degradation of IkappaBalpha by inducing the IKK activity. However, both pharmacological (wortmannin and LY294002) and genetic (Deltap85) inhibitors of PI 3'-kinase inhibited OPN-induced Akt phosphorylation, IKK activity, and NFkappaB activation through phosphorylation and degradation of IkappaBalpha. OPN also enhances uPA secretion, cell motility, and extracellular matrix invasion. Furthermore, cells transfected with Deltap85 or the super-repressor form of IkappaBalpha suppressed the OPN-induced uPA secretion and cell motility, whereas cells transfected with p110CAAX enhanced these effects. Pretreatment of cells with PI 3-kinase inhibitors or NFkappaB inhibitory peptide (SN-50) reduced the OPN-induced uPA secretion, cell motility, and invasion. To our knowledge, this is first report that OPN induces NFkappaB activity and uPA secretion by activating PI 3'-kinase/Akt/IKK-mediated signaling pathways and further demonstrates a functional molecular link between OPN-induced PI 3'-kinase-dependent Akt phosphorylation and NFkappaB-mediated uPA secretion, and all of these ultimately control the motility of breast cancer cells.