FIG. 4.

Analysis of the cell cycle and caspase activity of HeLa cells that expressed variants of C81. HeLa cells were transfected with pME18Neo that encoded Flag-tagged C81, C81/I60P, C81/L67P, C81/I74P, or C81/I81P or the control pME18Neo-Flag together with (C) or without (B) the GFP expression vector pEGFP-N1. (A) Mutations introduced into the leucine zipper-like domain of C81. Four Ile and Leu residues in the leucine zipper-like domain (represented by dark shading) were replaced by Pro to destroy the structure of the leucine zipper-like domain. Grey bars represent the Flag tag. (B) Thirty-six hours after, cell lysates were prepared and caspase-3 activity was determined as described in the legend to Fig. 2. Each column and error bar represent the mean ± standard deviation of results from three samples in two independent experiments. (C) Thirty-six hours after transfection, cells were stained with PI for analysis of DNA content and cells that were GFP positive were analyzed by flow cytometry as described in the text. Arrowheads indicate peaks of cells at the G₁ and G₂/M phases. The G₂/M:G₁ ratio is indicated in the upper right of each graph.