Hypoxia induces p53 accumulation in the S-phase and accumulation of hypophosphorylated retinoblastoma protein in all cell cycle phases of human melanoma cells

Hypoxia has been shown to induce accumulation of p53 and of hypophosphoryla ted retinoblastoma protein (pRb) in tumour cells. In this study, the cell c ycle dependence of p53 accumulation and pRb hypophosphorylation in four hum an melanoma cell lines that are wild type for p53 was investigated using tw o-parameter flow cytometry measurements of p53 or pRb protein content and D NA content. The hypoxia-induced increase in p53 protein was higher in S-pha se than in G(1) and G(2) phases in all cell lines. The accumulation of p53 in S-phase during hypoxia was not related to hypoxia-induced apoptosis or s ubstantial cell cycle specific cell inactivation during the first 24 h of r eoxygenation. pRb was hypophosphorylated in all cell cycle phases by hypoxi a treatment. The results did not support a direct link between p53 and pRb during hypoxia because p53 was induced in a cell cycle-specific manner, whe reas no cell cycle-dependent differences in pRb hypophosphorylation were de tected. Only a fraction of the cell populations (0.60 +/- 0.10) showed hypo phosphorylated pRb. Thus, pRb is probably not the only mediator of the hypo xia-induced cell cycle block seen in all cells and all cell cycle phases. M oreover, the cell cycle-dependent induction of p53 by hypoxia suggests that the primary function of p53 accumulation during hypoxia is other than to a rrest the cells.