Inorganic phosphate (P-i) is a major regulator of cell metabolism. The P-i transport activity in the plasma membrane is a main determinant of the intracellular level of this ion. In bone-forming cells, P-i transport is important for the calcification of the bone matrix In this study, the effect of platelet-derived growth factor (PDGF) on P-i transport activity and the signaling mechanism involved in this cellular response were analyzed, The results indicate that PDGF is a potent and selective stimulator of sodium-dependent P-i transport in the mouse calvariaderived MC3T3-E1 osteoblast-like cells. The change in P-i transport induced by PDGF-BB was dependent on translational processes and affected the V-max of the P-i transport system, These observations suggested that enhanced P-i transport activity in response to PDGF resulted from insertion of newly synthesized P-i transporters in the plasma membrane. The role of activation of mitogen activated protein (MAP) kinase, phospholipase C (PLC)gamma or phosphatidylinositol 3-kinase (PI-3-kinase), in mediating this effect of PDGF, was investigated. A selective inhibitor of the PDGF receptor tyrosine kinase activity (CGP 53716) completely blocked PDGF-induced protein tyrosine phosphorylation of several proteins including the PDGF receptor, PLC gamma, MAP kinase, and association of the p85 subunit of PI-3'-kinase. Associated with this effect, the increase in P-i transport induced by PDGF was completely blunted by 5 mu M CGP 53716. Inhibition of MAP kinase activity by cAMP agonists did not influence P-i transport stimulation induced by PDGF. However, inhibitors of protein kinase C completely blocked this response. A selective inhibitor of PI-3-kinase, LY294002, also significantly reduced this effect of PDGF. In summary, these results indicate that PDGF is a potent and selective stimulator of P-i transport in osteoblastic cells. The mechanism responsible for this effect is not mediated by MAP kinase but involves tyrosine phosphorylation-dependent activation of PLC gamma and PI-3-kinase.
