Reduced ligation during DNA base excision repair supported by BRCA2 mutantcells

The brest cancer predisposing genes BRCA1 and BRCA2 appear to be involved i n DNA repair. In particular, the sensitivity of BRCA2-deficient mouse embry onic fibroblasts to ionizing radiation and the demonstrated interaction of the BRCA2 protein with Rad51, a major factor in recombinational repair, ind icate that BRCA2 is important for double strand break repair. The human BRC A2-deficient human cell line Capan-1, whilst being sensitive to ionizing ra diation, is also sensitive to the alkylating agent methymethanesulfonate, T he major lesions induced by this agent are methylated bases which are remov ed primarily by the base excision repair (BER) pathway. We have investigate d the efficiency of BER in Capan-1 cells by an in vitro assay in which plas mid substrates containing a single lesion are repaired by mammalian cell ex tracts. In comparison to the control cell lines BxPC-3, T24 and MCF7, Capan -1 cells exhibited a reduced rate of DNA ligation during both the single-nu cleotide insertion and PCNA-dependent pathways of BER, The reduced rate of DNA ligation exhibited by Capan-1 cell extracts was complemented by additio n of bacteriophage T4 DNA ligase or human DNA ligase III, BXCA2-mutant Capa n-1 cells may possess reduced DNA ligase activity during BER.