Vol. 59, Issue 4, 860-866, April 2001

Protein Kinase C- Coordinately Regulates Cytosolic Phospholipase A₂ Activity and the Expression of Cyclooxygenase-2 through Different Mechanisms in Mouse Keratinocytes

Hui Qin Wang, Michael P. Kim, Howard F. Tiano, Robert Langenbach, and Robert C. Smart

Cell Signaling and Cancer Group, Department of Environmental and Molecular Toxicology, North Carolina State University, Raleigh, North Carolina (H.Q.W., M.P.K., R.C.S.); and National Institute of Environmental Health Sciences, Laboratory of Experimental Carcinogenesis and Mutagenesis, Research Triangle Park, North Carolina (H.F.T., R.L.)

Transgenic mice (K5-PKC) in which the keratin 5 promoter directs the expression of protein kinase C- (PKC) to epidermal keratinocytes display a 10-fold increase in PKC protein in their epidermis and alterations in phorbol ester-induced cutaneous inflammation [J Cell Science 1999;112:3497-3506]. In the current study, we have used these K5-PKC mice to examine the role of PKC in keratinocyte phospholipid metabolism/eicosanoid production and cutaneous inflammation. Primary keratinocytes from wild-type and transgenic mice were prelabeled in culture with [³H]arachidonic acid (AA) and subsequently treated with TPA. Compared with wild-type keratinocytes, K5-PKC keratinocytes displayed a 2-fold increase in AA release. TPA treatment resulted in the phosphorylation of cPLA₂. PKC inhibitors GF-109203X or H7, but not mitogen-activated protein/extracellular signal-regulated protein kinase (MEK) inhibitor PD 98059, could inhibit phosphorylation and AA release. Topical 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment of K5-PKC mice resulted in a 5-fold increase in epidermal COX-2 induction and a 2- to 3-fold increase in prostaglandin (PG) E₂ levels above that observed in TPA-treated wild-type mice. PD 98059, GF-109203X, or H7 could block cyclooxygenase-2 (COX-2) induction by TPA. Because C/EBP, a basic leucine zipper transcription factor, can be activated via a PKC/mitogen-activated protein kinase pathway and can influence COX-2 expression, we examined whether C/EBP is involved in TPA-induced epidermal COX-2 expression. TPA-induced COX-2 expression was similar in C/EBP nullizygous and wild-type mice. In summary, our results indicate that epidermal PKC coordinately regulates cPLA₂ activity and COX-2 expression resulting in increased levels of AA and PGE₂. Furthermore, PKC-induced AA release and cPLA₂ phosphorylation are independent of MEK, whereas PKC-induced COX-2 expression and PGE₂ production are MEK-dependent and C/EBP-independent events.