DIFFERENTIAL INDUCTION OF PHOSPHATIDYLCHOLINE HYDROLYSIS, DIACYLGLYCEROL FORMATION AND PROTEIN-KINASE-C ACTIVATION BY EPIDERMAL GROWTH-FACTOR AND TRANSFORMING GROWTH-FACTOR-ALPHA IN NORMAL HUMAN SKIN FIBROBLASTS AND KERATINOCYTES

We have investigated coupling between the epidermal growth factor (EGF ) receptor and the phospholipase C (PLC)/protein kinase C (PKC) signal -transduction system in normal skin fibroblasts and keratinocytes, for which EGF and transforming growth factor alpha (TGF-alpha) are mitoge nic. EGF and TGF-alpha induced a rapid increase in tyrosine phosphoryl ation of the EGF receptor, in both fibroblasts and keratinocytes, but failed to induce tyrosine phosphorylation of PLC-gamma1 or detectable phosphoinositide hydrolysis, as measured by two sensitive assays. In f ibroblasts, EGF induced phosphatidylcholine (PC) hydrolysis, resulting in increased diacylglycerol (DAG). In contrast, in keratinocytes, the re was no detectable PC hydrolysis or elevation of DAG in response to EGF or TGF-alpha. EGF and TGF-alpha activated PKC in fibroblasts, as e videnced by increased phosphorylation of a specific cellular PKC subst rate (myristoylated alanine-rich C-kinase substrate, 'MARCKS'). In ker atinocytes, TGF-alpha and EGF induced only a modest increase in MARCKS protein phosphorylation. This apparent modest activation of PKC, in t he absence of detectable DAG formation, may have been mediated by arac hidonic acid, which was released from keratinocytes in response to TGF -alpha, and has been shown to stimulate PKC activity in vitro. These d ata demonstrate that (1) in dermal fibroblasts and keratinocytes, whic h express normal levels of EGF receptors, EGF receptor activation is n ot coupled to tyrosine phosphorylation of PLC-gamma1 or PtdIns hydroly sis, suggesting that these events are not required for the mitogenic a ctivity of EGF or TGF-alpha in these cells, (2) coupling of EGF recept or to PC hydrolysis is cell-type specific, and (3) in skin fibroblasts , DAG, formed through EGF-induced PC hydrolysis, is capable of activat ing PKC.