EGF regulates plasminogen activator inhibitor-1 (PAI-1) by a pathway involving c-Src, PKCδ, and sphingosine kinase 1 in glioblastoma cells

Patients with gliomas expressing high levels of epidermal growth factor receptor ( EGFR) and plasminogen activator inhibitor-1 (PAI-1) have a shorter overall survival prognosis. Moreover, EGF enhances PAI-1 expression in glioma cells. Although multiple known signaling cascades are activated by EGF in glioma cells, we show for the first time that EGF enhances expression of PAI-1 via sequential activation of c-Src, protein kinase C delta (PKC delta), and sphingosine kinase 1 (SphK1), the enzyme that produces sphingosine-1-phosphate. EGF induced rapid phosphorylation of c-Src and PKC delta and concomitant translocation of PKC delta as well as SphK1 to the plasma membrane. Down-regulation of PKC delta abolished EGF-induced SphK1 translocation and up-regulation of PAI-1 by EGF; whereas, down-regulation of PKC alpha had no effect on the EGF-induced PAI-1 activation but enhanced its basal expression. Similarly, inhibition of c-Src activity by PP2 blocked both EGF-induced translocation of SphK1 and PKC delta to the plasma membrane and up-regulation of PAI-1 expression. Furthermore, SphK1 was indispensable for both EGF-induced c-Jun phosphorylation and PAI-1 expression. Collectively, our results provide a functional link between three critical downstream targets of EGF, c-Src, PKC delta, and SphK1 that have all been implicated in regulating motility and invasion of glioma cells.