title: Translesion DNA synthesis-assisted non-homologous end-joining of complex double-strand breaks prevents loss of DNA sequences in mammalian cells.
abstract: Double strand breaks (DSB) are severe DNA lesions, and if not properly repaired, may lead to cell death or cancer. While there is considerable data on the repair of simple DSB (sDSB) by non-homologous end-joining (NHEJ), little is known about the repair of complex DSBs (cDSB), namely breaks with a nearby modification, which precludes ligation without prior processing. To study the mechanism of cDSB repair we developed a plasmid-based shuttle assay for the repair of a defined site-specific cDSB in cultured mammalian cells. Using this assay we found that repair efficiency and accuracy of a cDSB with an abasic site in a 5' overhang was reduced compared with a sDSB. Translesion DNA synthesis (TLS) across the abasic site located at the break prevented loss of DNA sequences, but was highly mutagenic also at the template base next to the abasic site. Similar to sDSB repair, cDSB repair was totally dependent on XrccIV, and altered in the absence of Ku80. In contrast, Artemis appears to be specifically involved in cDSB repair. These results may indicate that mammalian cells have a damage control strategy, whereby severe deletions are prevented at the expense of the less deleterious point mutations during NHEJ.
authors.affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Shay
authors.initials: S
authors.lastname: Covo
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: de Villartay
authors.forename: Penny A
authors.initials: PA
authors.lastname: Jeggo
authors.forename: Zvi
authors.initials: Z
authors.lastname: Livneh
mesh_terms: D034961:Antigens, Nuclear / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000096:biosynthesis* / Q000737:chemistry; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism; D003851:Deoxyribonucleases / Q000378:metabolism; D006801:Humans; D000072200:Ku Autoantigen; D010957:Plasmids
chemical_list: D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D004247:DNA; D003851:Deoxyribonucleases; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen