FIG. 2.

Analysis of cell growth and survival. (A) The cell growth of primary MEFs of the indicated genotypes was analyzed using a modified 3T3 protocol. Nbs1^ΔB/ΔB Prkdc^scid/scid cell cultures showed little proliferation compared to WT, Prkdc^scid/scid, or Nbs1^ΔB/ΔB cell cultures. (B) Early-passage MEFs were stained for SA-β-Gal activity. p3 Nbs1^ΔB/ΔB Prkdc^scid/scid cultures showed increased SA-β-Gal activity compared to p3 Prkdc^scid/scid or p7 Nbs1^ΔB/ΔB MEFs, where little activity is detectable. (C) SV40-transformed EFs of the indicated genotypes were exposed to IR and assessed for colony formation. Nbs1^ΔB/ΔB Prkdc^scid/scid EFs were acutely sensitive to IR-induced damage, and this sensitivity was also observed in MEF cultures (data not shown). Enhanced sensitivity in Nbs1^ΔB/ΔB Art^−/− cell cultures was not observed (see Fig. S3 in the supplemental material). (D and E) SV40-transformed MEFs were exposed to the indicated doses of CPT for 24 h (D) or to the DNA cross-linker MMC for 2 h (E), and survival was assessed by monitoring colony formation. Double-mutant Nbs1^ΔB/ΔB Prkdc^scid/scid MEFs did not display increased sensitivity to either agent compared to that of single-mutant Nbs1^ΔB/ΔB MEFs.