Concerted Regulation of Wild-Type p53 Nuclear Accumulation and Activation by S100B and Calcium-Dependent Protein Kinase C
Christian Scotto, Christian Delphin, Jean Christophe Deloulme, and Jacques Baudier
The calcium ionophore ionomycin cooperates with the S100B protein to rescue a p53-dependent G1 check- point control in S100B-expressing mouse embryo fibroblasts and rat embryo fibroblasts (REF cells) which express the temperature-sensitive p53Val135 mutant (C. Scotto, J. C. Deloulme, D. Rousseau, E. Chambaz, and J. Baudier, Mol. Cell. Biol. 18:4272–4281, 1998). We investigated in this study the contributions of S100B and calcium-dependent PKC (cPKC) signalling pathways to the activation of wild-type p53. We first confirmed that S100B expression in mouse embryo fibroblasts enhanced specific nuclear accumulation of wild-type p53. We next demonstrated that wild-type p53 nuclear translocation and accumulation is dependent on cPKC activity. Mutation of the five putative cPKC phosphorylation sites on murine p53 into alanine or aspartic residues had no significant effect on p53 nuclear localization, suggesting that the cPKC effect on p53 nuclear translocation is indirect. A concerted regulation by S100B and cPKC of wild-type p53 nuclear translocation and activation was confirmed with REF cells expressing S100B (S100B-REF cells) overexpressing the temperature-sensitive p53Val135 mutant. Stimulation of S100B-REF cells with the PKC activator phorbol ester phorbol myristate acetate (PMA) promoted specific nuclear translocation of the wild-type p53Val135 species in cells positioned in early G1 phase of the cell cycle. PMA also substituted for ionomycin in the mediating of p53-dependent G1 arrest at the nonpermissive temperature (37.5°C). PMA-dependent growth arrest was linked to the cell apoptosis response to UV irradiation. In contrast, growth arrest mediated by a temperature shift to 32°C protected S100B-REF cells from apoptosis. Our results suggest a model in which calcium signalling, linked with cPKC activation, cooperates with S100B to promote wild-type p53 nuclear translocation in early G1 phase and activation of a p53-dependent G1 checkpoint control.