Characterizing Chromosomal Aberrations in Cells Deficient for Both ATM and MSH2

Ataxia telangiectasia mutated (ATM) and mutS homologue 2 (MSH2) are important DNA repair proteins that participate in DNA repair pathways to maintain genomic integrity. Mice deficient for ATM and MSH2 mice are viable. However, ATM-/- mice show growth retardation, neurological defects, and spontaneous lymphomagenesis. MSH2-/- mice suffer from aggressive lymphoid tumors between two to five months of age and have increased microsatellite instability, which predisposes MSH2-/- mice to carcinomas. However, mice deficient in both ATM and MSH2 are unable to survive beyond postnatal day 21 (P21). The observed lethality in ATM-/-MSH2-/- mice may result from the accumulation of genomic instability. To address this hypothesis, metaphase spreads were examined for chromosomal aberrations in B cells, which undergo genetically programmed DNA recombination and mutation. To bypass the lethality in ATM-/-MSH2-/- mice, a conditional knockout of ATM was bred onto an MSH2-/- background strain to generate CD21cre+ATMF/FMSH2-/- mice, which are referred to as ATM∆/∆MSH2-/-. In these mice, Cre expression, which is regulated by the CD21 promoter, deletes floxed ATM alleles in naïve mature MSH2-/- B-cells. ATM∆/∆ B-cells showed low levels of chromosomal aberrations whereas ATM∆/∆MSH2-/-B-cells had levels of chromosomal aberrations that was comparable to MSH2-/- B-cells. The latter result suggested that ATM was incompletely deleted in ATM∆/∆ B-cells. Analysis of ATM∆/∆ B cell lysates showed residual ATM protein that may account for the comparable chromosomal aberrations in ATM∆/∆MSH2-/-and MSH2-/- metaphase spreads. ATM-/-MSH2-/- embryos die in utero at approximately E16-E20. Thus, mouse embryonic fibroblasts (MEFs) were isolated from E13.5-15.5 ATM-/-MSH2-/- embryos and examined for chromosomal aberrations. The ATM-/-MSH2-/- MEF metaphase spreads displayed a similar pattern of chromosomal aberrations as ATM-/- MEFs, which does not support the hypothesis that ATM-/-MSH2-/- mice die from increased genomic instability; however, additional metaphase analyses are required to conclusively determine whether genomic instability does or does not impair ATM-/-MSH2-/- embryonic development