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. Author manuscript; available in PMC: 2009 Aug 18.
Published in final edited form as: Cancer Res. 2007 Jul 15;67(14):6685–6690. doi: 10.1158/0008-5472.CAN-07-0495

Fig. 5.

Fig. 5

ATRIP S224 is required for ATR-ATRIP-dependent G2/M checkpoint responses to DNA damage. (A) U2OS cells expressing siRNA resistant cDNAs for HA-ATRIP (WT), HA-ATRIP S224A or no cDNA (Vect) were transfected with non-specific (NS) or ATRIP (A4) siRNA. Cells were harvested three days after transfection and analyzed for ATRIP expression by immunoblotting. (B and C) Three days after transfection with ATRIP siRNA, U2OS cells containing wild type ATRIP, ATRIP S224A, or vector were treated with 25J/m2 UV radiation (B) or 4 Gy of IR (C). Nocodazole was added to the culture media to trap cells in mitosis. In (B) cells were harvested, fixed with Carnoy’s fixative, and mitotic spreads were analyzed 8 hours after exposure to UV. The percentage of mitotic cells was calculated based on counting at least 600 cells. Error bars represent the standard error. In (C) cells were harvested at the indicated time points following IR, fixed with ethanol, permeabilized and stained with anti-phospho-histone H3 S10 antibody and propidium idodine. The percentage of mitotic cells was determined by flow cytometry.