Abstract
Class switch recombination (CSR) produces secondary immunoglobulin isotypes and requires AID-dependent DNA deamination of intronic switch (S) regions within the immunoglobulin heavy chain (Igh) gene locus. Non-canonical repair of deaminated DNA by mismatch repair (MMR) or base excision repair (BER) creates DNA breaks that permit recombination between distal S regions. ATM-dependent phosphorylation of AID at serine-38 (pS38-AID) promotes its interaction with 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 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 to AtmD/Dand 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 to wildtype, AtmD/D, or Msh2-/- B cells. This data indicates that the absence of both ATM and MSH2 blocks non-homologous end joining (NHEJ), leading to inefficient CSR. We propose a model whereby ATM and MSH2 function cooperatively to regulate end-joining during CSR through pS38-AID.
Summary Loss of the DNA repair genes Atm and Msh2 produces a novel synthetic lethality in mice. B cell specific deletion of Atm on an Msh2-/- background reduces Ig CSR and inhibits NHEJ.
Competing Interest Statement
The authors have declared no competing interest.
Abbreviations
- AID
- activation induced cytidine deaminase
- A-EJ
- alternative end joining
- APE1
- apurinic/apyrimadinic endonuclease 1
- ATM
- ataxia telangiectasia mutated
- BER
- base excision repair pathway
- CSR
- Class switch recombination
- DNA-PKcs
- DNA-dependent protein kinase catalytic subunit
- DSB
- double stranded DNA break
- E13.5
- mouse embryonic day 13.5
- MMR
- mismatch repair pathway
- pS38-AID
- phosphorylation of serine 38 on AID
- MSH2
- mut S homolog 2
- MLH1
- mutL homology 1
- NHEJ
- Nonhomologous end joining
- P0
- newborn mouse pup
- P28
- Post natal day 28
- PMS2
- PMS1 homolog 2
- SSA
- single strand annealing
- SSB
- single stranded DNA break
- UNG
- uracil DNA glycosylase 1
- XRCC4
- X-ray repair cross complementing