Mitogen stimulation of Na⁺-H⁺ exchange: Differential involvement of protein kinase C

The mitogen-induced activation of Na⁺-H⁺ exchange was investigated in two cultured human fibroblast strains (HSWP and WI-38 cells), that, based on previous studies, differed in their response to the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) (L. M. Vincentini and M.L. Villereal, Proc. Natl. Acad. Sci. USA 82: 8053-8056, 1985). The role of protein kinase C in the activation of Na⁺-H⁺ exchange was investigated by comparing the effects of TPA on Na⁺ influx, in vitro phosphorylation, and in vivo phosphorylation in both cell types. Although both cell types have significant quantities of protein kinase C activity that can be activated by TPA in intact cells, the addition of TPA to intact cells stimulates Na⁺ influx in WI-38 cells but not in HSWP cells, indicating that in HSWP cells the stimulation of protein kinase C is not sufficient to activate the Na⁺-H⁺ exchanger. Cells were then depleted of protein kinase C activity by chronic treatment with high doses of TPA. Both HSWP and WI-38 cells were rendered protein kinase C deficient by this treatment as determined by in vitro and in vivo phosphorylation studies. Protein kinase C-deficient HSWP cells lose the ability for TPA to inhibit the serum-induced activation of Na⁺-H⁺ exchange, but there is no reduction in the stimulation of Na⁺ influx by serum, bradykinin, vasopressin, melittin, or vanadate, indicating that protein kinase C activity is not necessary for the mitogen-induced activation of Na⁺-H⁺ exchange in HSWP cells by agents known to stimulate phosphatidylinositol (G.A. Jamieson and M. Villereal. Arch. Biochem. Biophys. 252: 478-486, 1987). In contrast, depletion of protein kinase C activity in WI-38 cells significantly reduces both the TPA- and the serum-induced activation of the Na⁺-H⁺ exchange system, suggesting that protein kinase C activity is necessary for at least a portion of the mitogen-induced activation of the Na⁺-H⁺ exchangr in WI-38 cells. These results indicate that the mechanisms for regulating Na⁺-H⁺ exchange can differ dramatically between different types of fibroblasts.