Immunological demonstration of a calcium-unresponsive protein kinase C of the delta-type in different species and murine tissues. Predominance in epidermis

Abstract

An antiserum raised against a delta-protein kinase C (delta-PKC)-specific peptide recognized the purified calcium-unresponsive 76-kDa protein kinase of porcine spleen in the native and the denatured form. This antiserum was used to demonstrate the delta-PKC-like enzyme in spleen of different species, in various cell types and in murine tissues by immunoblotting of the respective extracts. Due to species differences, delta-PKC-like kinases with slightly different molecular weights were observed. The enzyme was found to be present in primary murine keratinocytes, primary bovine endothelial cells, and many cell lines originating from human, rat, and murine tissues. It was present also in all murine tissues tested, predominantly in epidermis, uterus, placenta, lung, brain, spleen, and kidney. In contrast to the conventional alpha, beta, gamma-PKC, it was located almost exclusively in the particulate fraction. The delta-like PKC could be demonstrated in the epidermis and brain of newborn mice, and in both tissues its concentration increased dramatically between day 7 and 14 after birth. The delta-PKC-like kinase of mouse epidermis (p82-kinase) was down-regulated after topical application of 12-O-tetradecanoylphorbol-13-acetate (TPA) to mouse skin. The amount of the enzyme decreased to less than 20% of the controls within 16 h and recovered almost completely within 72 h after TPA. The existence of the delta-PKC-like kinase in mouse skin, papillomas, and carcinomas could also be demonstrated by immunocytochemical staining of the respective sections. The enzyme was observed predominantly in epithelial layers. A remarkable immunostaining of nuclei in skin sections disappeared after TPA treatment of the animals.