Effect of ionizing radiation on cell-cycle progression and cyclin B1 expression in human melanoma cells

In the present study we investigated the effect of gamma-irradiation (2.5 and 10 Gy) on cell-cycle progression of a human melanoma cell line, M14, characterized by a moderate radiosensitivity (SF2 = O.5). Flow cytometric analysis showed a dose-dependent S-phase accumulation, which was detectable 8 hr after treatment with 2 and 5 Gy and was still persistent at 12 hr after 10 Gy exposure. Such a delay in S-phase was paralleled or followed by an accumulation of cells in G2M, which was transient at the lowest radiation doses and still persistent at 72 hr after 10 Gy. Such an accumulation was, at least in part, due to a block in G2-M transition, as demonstrated by mitotic index analysis. Bivariate flow cytometric analysis of DNA content and cyclin B1 expression showed that, following 2 and 5 Gy, the fraction of cyclin B1-expressing cells was superimposable upon that of G2M cells. Conversely, in cells treated with 10 Gy, the fraction of cyclin B1-expressing cells was half the G2M cell fraction. Northern-blot analysis indicated that the radiation-induced decrease in cyclin B1 protein expression was accompanied by a reduced cyclin B mRNA level. On the whole, our results indicate a direct inhibitory effect of 10 Gy irradiation on cyclin B1 expression as a possible cause for the persistent G2 block in irradiated M14 cells.