Induction of protein kinase Cζ-related protein kinase by growth suppression in carcinogen-initiated epidermal cell-line WYF31 cells

• Published in: Cellular signalling Vol. 12; no. 1; pp. 15 - 22
• Main Authors: Nishikawa, Kiyotaka, Aoki, Junken, Yamamoto, Satoshi, Sameda, Hisamitsu, Kudo, Hiroyasu, Nagumo, Haruna, Wang, Jian Chun, Tsuzuki, Yasuhiko, Arai, Hiroyuki, Inoue, Keizo, Kato, Ryuichi
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 2000
• Subjects: Downregulation; Growth suppression; Initiated cell; Mouse epidermal cells; Overexpression; Phosphorylation; Protein kinase Cζ; WYF31 cells; l-PKCζ, low-salt eluted PKCζ; Phosphorylation; DG, diacylglycerol; PKC, protein kinase C; h-PKCζ, high-salt eluted PKCζ; Mouse epidermal cells; Growth suppression; Protein kinase Cζ; SDS, sodium dodecyl sulfate; PS, phosphatidylserine; peptide ϵ, protein kinase C-ϵ substrate; MEM, minimum essential medium; WYF31 cells; Overexpression; FCS, foetal calf serum; PAGE, polyacrylamide gel electrophoresis; PMSF, phenylmethylsulphonyl fluoride; TCA, trichloroacetic acid; Downregulation; Initiated cell; PMA, phorbol 12-myristate 13-acetate
• ISSN: 0898-6568; 1873-3913
• DOI: 10.1016/S0898-6568(99)00062-5

In primary cultured mouse epidermal cells, protein kinase C isozyme ζ (PKCζ) consists of multiple forms, for example, low-salt eluted PKCζ (l-PKCζ; 79 and 85 kDa) and high-salt eluted PKCζ (h-PKCζ; 79 and 85 kDa) on anion-exchange column chromatography. In this study, biochemical and biophysical differences between l-PKCζ and h-PKCζ were examined by using carcinogen-initiated mouse epidermal cell-line WYF31 cells, whose growth is stimulated by tumour promoter phorbol 12-myristate 13-acetate (PMA). The binding efficiency of h-PKCζ to anti-PKCζ antibody-affinity column was 10 times higher than that of l-PKCζ. T7-tagged rat PKCζ overexpressed in WYF31 cells was recovered only in the high-salt eluted area on the anion-exchange column. Furthermore, when rat PKCζ was stably overexpressed in WYF31 cells, the content of h-PKCζ increased 4 to 5 times compared to that of parental cells, but the content of l-PKCζ was not altered. All of these results indicate that h-PKCζ is the product of the PKCζ gene (referred to as PKCζ) and that l-PKCζ is closely related but different from PKCζ (referred to as PKCζ-related kinase). Interestingly, serum starvation of WYF31 cells caused a marked increase of the content of PKCζ-related kinase with a concomitant decrease of PKCζ content. These changes were reversed by stimulating the cell growth with 10% foetal calf serum. Prolonged treatment of starved cells with PMA, which induces the proliferation of WYF31 cells, also caused the downregulation of PKCζ-related kinase. These results suggest that the expression levels of PKCζ-related kinase and PKCζ are differently regulated, and that the increased expression of PKCζ-related kinase might play a significant role in the growth-suppression processes of WYF31 cells.