INVOLVEMENT OF XRCC1 AND DNA-LIGASE-III GENE-PRODUCTS IN DNA-BASE EXCISION-REPAIR

DNA ligase III and the essential protein XRCC1 are present at greatly reduced levels in tile xrcc1 mutant CHO cell line EM-C11, Cell-free ex tracts prepared from these cells were used to examine the role of the XRCC1 gene product in DNA base excision repair is vitro. EM-C11 cell e xtract was partially defective in ligation of base excision repair pat ches, in comparison to wild type CHO-9 extracts, Of the two branches o f the base excision repair pathway, only the single nucleotide inserti on pathway was affected; no ligation defect was observed in the prolif erating cell nuclear antigen-dependent pathway. Full complementation o f the ligation defect in EM-C11 extracts was achieved by addition to t he repair reaction of recombinant; human DNA ligase III but not by XRC C1. This is consistent with the notion that XRCC1 acts as an important stabilizing factor of DNA ligase III. These data demonstrate for the first time that xrcc1 mutant cells are partially defective in ligation of base excision repair patches and that the defect is specific to th e polymerase beta-dependent single nucleotide insertion pathway.