ASSOCIATION OF PROTEIN-KINASE-C ACTIVATION WITH INDUCTION OF ORNITHINE DECARBOXYLASE IN MURINE BUT NOT HUMAN KERATINOCYTE CULTURES

The goal of this study was to compare the response of mouse epidermal keratinocytes (MEKs) and human epidermal keratinocytes (HEKs) to 12-O- tetradecanoylphorbol-13-acetate (TPA) with respect to the activation a nd downregulation of protein kinase C (PKC), the expression of c-jun a nd c-fos, and the expression and induction of ornithine decarboxylase (ODC) activity. Keratinocytes from adult CD-1 mice and from discarded adult human skin were grown in primary culture in a high-calcium serum -free medium that supported proliferation and differentiation. Immunob lotting of freshly isolated and cultured MEKs and HEKs for isozymes of protein kinase C revealed that fresh HEKs contained PKCalpha, PKCbeta , and PKCdelta; no PKC-gamma, PKCepsilon, or PKCzeta were detected. In fresh MEKs, PKCalpha, PKCbeta, PKCDELTA, and PKCzeta were observed, b ut not PKCgamma or PKCepsilon. After 2 wk in culture, the isozyme prof iles of MEKs and HEKs were similar except that PKCgamma was noticeably present in HEK cultures. Activation of partially purified total PKC b y TPA was similar in freshly isolated and cultured MEKs and HEKs, indi cating that the two species were similar in this regard and that 2 wk of culture did not alter this characteristic. When MEK and HEK culture s were treated with TPA for 3 h, less than 30% of the control level of PKC activity was detected, indicating that TPA-induced downregulation of PKC was similar in MEKs and HEKs. After treatment with TPA, MEK cu ltures produced a large induction of both c-jun and c-fos mRNA by 60 m in, as determined by northern blot analysis, and a large induction of ODC mRNA and enzyme activity by 6 h. TPA treatment of cultured HEKs, h owever, did not induce ODC activity; in fact, less activity, compared with that of control cultures, was observed. Northern blot analysis al so revealed no increase in c-jun, c-fos, and ODC mRNA in HEKs. However , c-jun and c-fos mRNA and both ODC mRNA and enzyme activity were indu ced in HEKs fed growth factors after several days of deprivation. This suggests that the lack of ODC induction by TPA in HEKs is probably du e to species differences in downstream steps in PKC signal transductio n. (C) 1993 Wiley-Liss, Inc.