Figure 3.

ATM- and MRN-dependent responses to IR-induced breaks in Art-P70 mutant cells. (A) Art-P70 mutation does not impair ATM-dependent DNA damage responses to IR. Art^+/+, Art^−/−, Art^P70/P70 and ATM^−/− MEFs, as indicated, were unirradiated (−) or exposed to IR (10 Gy, +) and then harvested at 1 h post-irradiation. Whole-cell lysates were analyzed by western blotting using the indicated antibodies. Tubulin was used as a loading control. (B) The MRN complex is intact in Art-P70 mutant cells. The MRN complex was co-immunoprecipitated from whole-cell lysates generated from wild-type or Art^P70/P70 MEFs using α-Mre11 antibodies. The immunoprecipitates were washed with 300 mm KCl, and the proteins were analyzed on a 8% SDS–PAGE followed by western blotting with the indicated antibodies. WCL, whole-cell lysate; −, no antibody; +, α-Mre11 antibody. (C) Mre11 foci formation is not impaired in Art-P70 mutant cells. Art^+/+, Art^−/− and Art^P70/P70 MEFs were grown on coverslips then irradiated (10 Gy). The irradiated cells and unirradiated controls were fixed in paraformaldehyde and then stained with α-Mre11 antibodies. The number of cells containing >20 Mre11 foci were scored. Left panels, representative images of nuclei. Right panels, quantitative results of Mre11 foci-positive cells. Average of two independent experiments is shown.