Fig. 2. Expression of C630 inhibits proliferation and delays progression through G2/M. HeLa cells were infected with either a control retrovirus or retroviruses encoding GFP, myc-tagged C630 and N-mBub 1. Puromycin-resistant cells were selected, expanded then analysed. (A) Timeline used to generate cells expressing the cDNA of interest. (B) Fluorescence histograms showing that 95% of the cells infected with GFP retroviruses express GFP. (C) Immunofluorescence analysis of HeLa cells infected with retroviruses encoding C630 immediately after or 12 hours after release from a G1/S block indicating that expression levels of C630 are low at G1/S but high in late G2. (D) Growth curves showing that expression of C630 results in a growth defect. (E) DNA content histograms of asynchronous cultures plus the number of cells in each phase of the cell cycle. The mitotic index was determined by quantitating MPM-2-positive cells. The values represent the average of three independent cultures indicating that expression of C630 results in accumulation of cells in G2/M. (F) DNA content histograms of infected HeLa cells following release from a G1/S block showing that expression of C630 delays progression through G2/M. The cells were analysed every 3 hours but only the key time points are shown.