Ataxia Telangiectasia Mutated and MSH2 Control Blunt DNA End Joining in Ig Class Switch Recombination

Abstract

Abstract Class-switch recombination (CSR) produces secondary Ig isotypes and requires activation-induced cytidine deaminase (AID)–dependent DNA deamination of intronic switch regions within the IgH (Igh) gene locus. Noncanonical repair of deaminated DNA by mismatch repair (MMR) or base excision repair (BER) creates DNA breaks that permit recombination between distal switch regions. Ataxia telangiectasia mutated (ATM)–dependent phosphorylation of AID at serine 38 (pS38-AID) promotes its interaction with apurinic/apyrimidinic endonuclease 1 (APE1), a BER protein, suggesting that ATM regulates CSR through BER. However, pS38-AID may also function in MMR during CSR, although the mechanism remains unknown. To examine whether ATM modulates BER- and/or MMR-dependent CSR, Atm−/− mice were bred to mice deficient for the MMR gene mutS homolog 2 (Msh2). Surprisingly, the predicted Mendelian frequencies of Atm−/−Msh2−/− adult mice were not obtained. To generate ATM and MSH2-deficient B cells, Atm was conditionally deleted on an Msh2−/− background using a floxed ATM allele (Atmf) and B cell–specific Cre recombinase expression (CD23-cre) to produce a deleted ATM allele (AtmD). As compared with AtmD/D and Msh2−/− mice and B cells, AtmD/DMsh2−/− mice and B cells display a reduced CSR phenotype. Interestingly, Sμ–Sγ1 junctions from AtmD/DMsh2−/− B cells that were induced to switch to IgG1 in vitro showed a significant loss of blunt end joins and an increase in insertions as compared with wild-type, AtmD/D, or Msh2−/− B cells. These data indicate that the absence of both ATM and MSH2 blocks nonhomologous end joining, leading to inefficient CSR. We propose a model whereby ATM and MSH2 function cooperatively to regulate end joining during CSR through pS38-AID.