Role of the nucleotide excision repair gene ERCC1 in formation of recombination-dependent rearrangements in mammalian cells

Spontaneous recombination between direct repeats at the adenine phosphoribo syltransferase (APRT) locus in ERCC1-deficient cells generates a high frequ ency of rearrangements that are dependent on the process of homologous reco mbination, suggesting that rearrangements are formed by misprocessing of re combination intermediates. Given the specificity of the structure-specific Ercc1/Xpf endonuclease, two potential recombination intermediates are subst rates for misprocessing in ERCC1- cells: heteroduplex loops and heteroduple x intermediates with nonhomologous 3' tails. To investigate the roles of ea ch, we constructed repeats that would yield no heteroduplex loops during sp ontaneous recombination or that would yield two non-homologous 3' tails aft er treatment with the rare-cutting endonuclease I-Seel, Our results indicat e that misprocessing of heteroduplex loops is not the major source of recom bination-dependent rearrangements in ERCC1-deficient cells, Our results als o suggest that the Erccl/Xpf endonuclease is required for efficient removal of nonhomologous 3' tails, like its Rad1/Rad10 counterpart in yeast. Thus, it is likely that misprocessing of nonhomologous 3' tails is the primary s ource of recombination-dependent rearrangements in mammalian cells. We also find an unexpected effect of ERCC1 deficiency on I-Scel-stimulated rearran gements, which are not dependent on homologous recombination, suggesting th at the ERCC1 gene product may play a role in generating the rearrangements that arise after I-Scel-induced double-strand breaks.