protein with a S2394L variation, appeared to induce phosphorylation at S1981, the epitope for the phospho-specific ATM antibody, but this increase was similar to non-irradiated (NR) background. Similarly, the ATM−/− Q3SA appeared to show a weaker trend of XR-induced pATM activity that was not different from NR. To determine if any evidence for functional ATM could be detected, we searched images for coincident pATM and γH2A.X staining (Figure 3C). Q3SC and CAR3 had robust, overlapping puncta, as expected. However, rare nuclei could be identified in both Q1SA and Q3SA that had not only pATM puncta but also overlapping γH2A.X puncta, indicating that ATM was expressed, autophosphorylated, and likely functional in triggering H2A.X phosphorylation. In the case of Q1SA, it is possible that the variation at S2394 was not sufficient to completely inhibit ATM activation in all cells, even though assay of γH2A.X in the absence of XR was negative (Figure 1F). However, the presence of a weak signal of pCHK2 (Figure 1E) along with the trend of occasional pATM and γH2A.X activation in Figure 3C suggests that the variant of ATM expressed in Q1SA is capable of phosphorylating downstream targets in a small number of cells. However, detection of pATM
