(A) Mre11+/C, Mre11C/- and Mre11C/H129N immortalized MEFs were stably infected with retrovirus expressing ER-CRE- IRES-hCD2, MACS purified for hCD2+ cells and then induced with 4OHT (200 nM) for 48 hr. PCR confirmed the efficient conversion of the Mre11 conditional allele to the null alleles. PCRs were performed as previously described (Buis et al., 2008). (B and C) Independent Western blot analyses of Mre11+/-, Mre11-/- or Mre11H129N/- MEFs treated with CPT (C, 0.1 µM, 2 hr) or pretreated with ATM kinase inhibitor (CA, ATMi 15 µM, 1 hr, then together with CPT 0.1 µM, 2 hr). (D) Western blot analyses of Atm+/-, Atm-/- or AtmKD/- MEFs treated with CPT (C, 0.1 µM, 2 hr) or pretreated with Mre11 nuclease inhibitors (CM for Mirin at 500 µM, or CP for PFM01 (Shibata et al., 2014) (200 µM, for 1 hr) then with CPT (0.1 µM) together for 2 more hours. (E) Western blot analyses of cells treated with Aphidicolin (A) or CPT (C) or first Aphidicolin for 1 hr, then together with CPT for 2 hrs (AC). (F) Representative sensitivity assay to mitomycin C (MMC). (G) Western blot analyses of immortalized MEFs treated with CPT (C) or CPT and ATM kinase inhibitor (CA) together for 2 hr.