Figure 6.

Artemis and DDB2 regulate cell cycle progression. (A) Depletion of Artemis causes a p27-dependent G₁ arrest. hTERT-RPE1 cells were transfected with Artemis and/or p27 specific siRNA or non-specific (NS) siRNA. Forty-eight h after transfection, BrdU was added to label S-phase cells. Thirty minutes after incubation with BrdU, cells were harvested and fixed. Cell cycle analysis was performed by FACS (upper panel), and results from three independent experiments are shown graphically (lower left panel). Samples collected in the cell cycle analysis were subjected to immunoblot analysis for p27 and Artemis (lower right panel). Upper numbers in the panels indicate the percentage of S-phase cells. (B) Artemis and DDB2 affect the G₁-G₀ transition via regulation of p27. hTERT-RPE1 cells were transfected with GST-Artemis or GST-DDB2 constructs. Thirty h after transfection, cells were placed into DMEM F-12 media containing 0.1% FBs. Thirty minutes prior to harvest at the indicated times, BrdU was added to the media. Cell cycle profiles were determined by FACS analysis (upper panel). Lower panel: Samples collected in the cell cycle analysis were subjected to immunoblot analysis for p27 (lower panel). (C) Artemis and DDB2 affect the G₁-G₀ transition via regulation of p27. Upper panel: hTERT-RPE1 cells were transfected with Artemis, DDB2, or non-specific siRNAs (NS). Twenty-four h after transfection cells were placed into media containing 0.1% FBS for the indicated time. BrdU was added into the culture 30 min before harvest, and cell cycle analysis was performed by FACS (upper panel). Samples collected in the cell cycle analysis were subjected to immunoblot analysis for p27 (lower panel).