PROTEIN-KINASE-C REGULATES KERATINOCYTE TRANSGLUTAMINASE (TG(K)) GENE-EXPRESSION IN CULTURED PRIMARY MOUSE EPIDERMAL-KERATINOCYTES INDUCED TO TERMINALLY DIFFERENTIATE BY CALCIUM

During the final stage of epidermal differentiation, activation of ker atinocyte transglutaminase results in covalent crosslinking of a varie ty of proteins to form highly protective cornified cell envelopes. We have studied the regulation of keratinocyte transglutaminase (TG(K)) g ene expression in murine epidermal keratinocytes induced to terminally differentiate in vitro by increasing the level of extracellular Ca+or treatment with the protein kinase C (PKC) activator 12-O-tetradecan oylphorbol- 13-acetate (TPA). Raising extracellular Catt induces squam ous differentiation of cultured keratinocytes and elicits a concentrat ion-dependent increase in expression of TG(K) mRNA; keratinocytes grow n for 24 h in 0.12 mM Ca++ medium express similar to 12 times as much TG(K) mRNA as basal cells (grown in 0.05 mM Ca++ medium), whereas cult ures exposed to 1.4 mM Ca++ express similar to 17 times as much. TPA i nduces squamous differentiation and TG(K) mRNA even in basal keratinoc yte cultures grown in 0.05 mM Ca++ medium, suggesting that expression of this differentiation marker is regulated by the PKC signaling pathw ay. Induction of TG(K) mRNA in response to TPA treatment is transient, reaching a peak at 6 - 8 h and returning to baseline by 24 h. In cont rast, elevation of TG(K) mRNA levels in response to Ca++ persists for at least 24 h. The increased abundance of TG(K) mRNA reflects increase d transcription of the TG(K) gene, based on nuclear run-on analysis of Ca++- and TPA-treated keratinocytes. Induction of TG(K) mRNA by eithe r TPA or Ca++ is blocked in the presence of cycloheximide, suggesting that a PKC-dependent protein factor is required for TG(K) gene express ion in response to both stimuli. Furthermore, the accumulation of TG(K ) mRNA in keratinocytes treated with TPA or Ca++ is blocked in cells t reated with the PKC inhibitor GF 109203X or bryostatin. These results suggest that the induction of TG(K) gene expression by Ca++ is depende nt on PKC, providing further support for the hypothesis that PKC plays a central role in regulating the late stages of epidermal differentia tion.