Repair of double-strand breaks by homologous recombination in mismatch repair-defective mammalian cells

Chromosomal double-strand breaks (DSBs) stimulate homologous recombination by several orders of magnitude in mammalian cells, including murine embryon ic stem (ES) cells, but the efficiency of recombination decreases as the he terology between the repair substrates increases (B, Elliott, C, Richardson , J, Winderbaum, J, A. Nickoloff, and M, Jasin, Mel. Cell, Biol. 18:93-101, 1998), We have now examined homologous recombination in mismatch repair (M MR)-defective ES cells to investigate both the frequency of recombination a nd the outcome of events. Using cells with a targeted mutation in the msh2 gene, we found that the barrier to recombination between diverged substrate s is relaxed for both gene targeting and intrachromosomal recombination. Th us, substrates with 1.5% divergence are 10-fold more likely to undergo DSB- promoted recombination in Msh2(-/-) cells than in wild-type cells. Although mutant cells can repair DSBs efficiently, examination of gene conversion t racts in recombinants demonstrates that they cannot efficiently correct mis matched heteroduplex DNA (hDNA) that is formed adjacent to the DSB, As a re sult, >20-fold more of the recombinants derived from mutant cells have unco rrected tracts compared with recombinants from wild-type cells. The results indicate that gene conversion repair of DSBs in mammalian cells frequently involves mismatch correction of hDNA rather than double-strand gap formati on. In cells with MMR defects, therefore, aberrant recombinational repair m ay be an additional mechanism that contributes to genomic instability and p ossibly tumorigenesis.