Figure 5. vCyclin is required to maintain accelerated G1/S transition at contact-inhibited condition. (A and B) Deletion of vCyclin does not affect cell cycle progression at low-density as shown by representative histograms (A) and results of averages from three repeats (B). Cells seeded at a low-density at 2.5 × 105 cells/flask in T25 flasks overnight were analyzed for cell cycle. There was no difference in cell cycle profiles among WT, mutant, and revertant cells. However, all 3 KSHV-transformed cells had more cells in S phase and less cells in G1 phase than mock cells had. (C and D) Deletion of vCyclin compromises cell cycle progression at a high-density as shown by representative histograms (C) and results of averages from 3 repeats (D). Cells seeded at a high-density at 106 cells/flask in T25 flasks overnight were analyzed for cell cycle. WT and revertant cells had more cells in S-phase and less cells in G1-phase than mutant and mock cells had. (E and F) Deletion of vCyclin does not affect BrdU incorporation at low-density as shown by representative FACS images (E) and results of averages from 3 repeats (F). Cells seeded at a low-density at 2.5 × 105 cells/flask in T25 flasks overnight were pulsed with BrdU and analyzed by FACS. There was no difference in BrdU incorporation among WT, mutant, and revertant cells. However, all 3 KSHV-transformed cells had more BrdU incorporation than mock cells had. (G and H) Deletion of vCyclin decreases BrdU incorporation at high-density as shown by representative FACS images (G) and results of averages from 3 repeats (H). Cells seeded at a high-density at 106 cells/flask in T25 flasks overnight were pulsed with BrdU and analyzed by FACS. WT and revertant cells had more BrdU incorporation than mutant and mock cells had. (I and J) Deletion of vCyclin has no effect on apoptosis at a low-density (I) or a high-density condition (J). Cells were seeded at 2.5 × 105 cells/flask (I) or 106 cells/flask (J) in T25 flasks overnight and analyzed for apoptosis by Annexin V staining.