PHOSPHORYLATIONS OF MR 34,000 AND 40,000 PROTEINS BY PROTEIN KINASE-C IN MOUSE EPIDERMIS INVIVO

Abstract

Activation of protein kinase C (PKC) and the resulting phosphorylations of proteins in vivo were examined in mouse epidermis, a target tissue of tumor-promoting phorbol diesters, such as 12-O-tetradecanoylphorbol-13-acetate (TPA). Treatment of mouse skin with TPA caused rapid translocation of PKC from the cytosol to the membrane fraction of skin tissue, followed by its down regulation. Epidermal proteins were labeled locally with 32P using the ring-shaped forceps technique that economizes on the amount of 32P1 required. Treatment with TPA in vivo resulted in about 2-fold increases in the phosphorylations of epidermal proteins with molecular weights of 34,000 and 40,000 and isoelectric points of 4.7-5.1 and 5.2-6.2 (p34 and p40, respectively). The phosphorylations of these proteins were also stimulated by teleocidin B. Inhibitors of PKC, such as chlorpromazine, quercetin, and staurosporine inhibited these increases in phosphorylations of p34 and p40 on TPA treatment. Furthermore, p34 and p40 were phosphorylated by purified PKC in a cell-free system. These results indicate that p34 and p40 are phosphorylated by PKC in mouse epidermis in vivo and may be involved in tumor promotion.