Flavin-containing oxidases regulate progression from G 1 to S phase of the cell cycle in normal human diploid fibroblasts

The effects of flavin-containing oxidase enzymes on cellular progression from G ₁ to S phase of the cell cycle were examined in normal human diploid fibroblasts by use of the cumulative labeling index method. Significant transient arrests in G ₁ occurred when quiescent cell populations incubated with diphenyliodonium (DPI) for a 20 h period at concentrations as low as 0.05 μM were stimulated to proliferate by subculturing to low density in control growth medium. Quiescent cells pre-treated with DPI and subcultured in DPI-containing growth medium were permanently arrested in G ₁. Induction of this G ₁ checkpoint was associated with activation of the p53/p21 ^Waf1 signaling pathway. Incubation of G ₀/G ₁ phase cells with DPI (0.05-0.2 μM) resulted in significant decreases in the expression levels of cyclin D1 and p21 ^Waf1; no effect was detected on cyclin A expression. Collectively, these results suggest that DPI-sensitive signaling primarily regulates early G ₁ events that lead to S-phase transition. Furthermore, they support a critical role for membrane originating oxidants in the regulation of cellular proliferation pathways.