Vif induces an early G1-to-S transition in Vif-transfected HeLa cells. (A) Cell-cycle profiles of transfected HeLa cells at 5 different time points after release from cell-cycle synchronization. Cells were transfected with pcDNA (top) or wild-type Vif (bottom). Vertical axes denote relative cell number. Horizontal axes denote DNA content as measured by PI staining. Peaks in each plot denote different phases of the cell cycle as labeled in Ai-iii. Note that at 4 hours in Vif-transfected cells (Aix), the height of the S peak compared with the G1 peak is larger than in control cells (Aii). At 10 hours, the height of the G2 peak relative to the G1 peak is greater in Vif-transfected cells (Aviii) than in control cells (Aiv). (B) Proportion of cells in the S phase at time 0 and 3 hours after release from cell-cycle synchronization for the same types of cultures as in panel A after treatment with BrdU/7AAD. Vertical axes show the number of cells with incorporated BrdU. Horizontal axes show total DNA content by 7AAD staining. The top small box in each treatment condition denotes the cell number at the S phase of the cell cycle, while the lower small box denotes the cells at G1. The calculated S/G1 ratio is noted in the upper right of each of the 3 plots. (C) Combined results from multiple experiments of the types shown in panels A-B. Vertical axis denotes ratio of number of cells in S phase to number of cells in G1. Horizontal axis denotes time after release from synchronization in the S phase. The proportion of cells in S compared with G1 was obtained by analyzing the peaks of cell-cycle profiles such as those shown in (A). The proportion of cells in the S phase was increased in cells transfected with either wild-type Vif or the Vif mutant SLQ/AAA at 3, 6, and 9 hours after release from G1, compared with cells transfected with control vector. ■, control vector; ▲, wild-type Vif; and ●, Vif mutant SLQ/AAA.
