Antagonistic effects of protein kinase C alpha and delta on both transformation and phospholipase D activity mediated by the epidermal growth factor receptor

Downregulation of protein kinase C delta (PKC delta) by treatment with the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) tr ansforms cells that overexpress the non-receptor class tyrosine kinase c-Sr c (Z. Lu ct al,, Mel. Cell. Biol. 17:3418-3428, 1997). We extended these st udies to cells overexpressing a receptor class tyrosine kinase, the epiderm al growth factor (EGF) receptor (EGFR cells); like c-Src, the EGF receptor is overexpressed in several human tumors. In contrast with expectations, do wnregulation of PKC isoforms with TPA did not transform the EGFR cells; how ever, treatment with EGF did transform these cells. Since TPA downregulates all phorbol ester-responsive PKC isoforms,,ve examined the effects of PKC delta- and PKC alpha-specific inhibitors and the expression of dominant neg ative mutants for both PKC delta and alpha. Consistent with a tumor-suppres sing function for PKC delta, the PKC delta-specific inhibitor rottlerin and a dominant negative PKC delta mutant transformed the EGFR cells in the abs ence of EGF. In contrast, the PKC alpha-specific inhibitor Go6976 and expre ssion of a dominant negative PKC or mutant blocked the transformed phenotyp e induced by both EGF and PKC delta inhibition. Interestingly, both rottler in and EGF induced substantial increases in phospholipase D (PLD) activity, which is commonly elevated in response to mitogenic stimuli, The elevation of PLD activity in response to inhibiting PKC delta, like transformation, was dependent upon PKC alpha and restricted to the EGFR cells. These data d emonstrate that PKC isoforms alpha and delta have antagonistic effects on b oth transformation and PLD activity and further support a tumor suppressor role for PKC delta that may be mediated by suppression of tyrosine kinase-d ependent increases in PLD activity.