title: Base excision repair intermediates are mutagenic in mammalian cells.
abstract: Base excision repair (BER) is the main pathway for repair of DNA damage in mammalian cells. This pathway leads to the formation of DNA repair intermediates which, if still unsolved, cause cell lethality and mutagenesis. To characterize mutations induced by BER intermediates in mammalian cells, an SV-40 derived shuttle vector was constructed carrying a site-specific lesion within the recognition sequence of a restriction endonuclease. The mutation spectra of abasic (AP) sites, 5'-deoxyribose-5-phosphate (5'dRp) and 3'-[2,3-didehydro-2,3-dideoxy-ribose] (3'ddR5p) single-strand breaks (ssb) in mammalian cells was analysed by RFLP/PCR and mutation frequency was estimated by quantitative PCR. Point mutations were the predominant events occurring at all BER intermediates. The AP site-induced mutation spectrum supports evidence for the 'A-rule' and is also consistent with the use of the 5' neighbouring base to instruct nucleotide incorporation (5'-rule). Preferential adenine insertion was also observed after in vivo replication of 5'dRp or 3'ddR5p ssb. We provide original evidence that not only the abasic site but also its derivatives 'faceless' BER intermediates are mutagenic, with a similar mutation frequency, in mammalian cells. Our findings support the hypothesis that unattended BER intermediates could be a constant threat for genome integrity as well as a spontaneous source of mutations.
authors.affiliation: Department of Environmental and Primary Prevention, Istituto Superiore di Sanità Viale Regina Elena 299, 00161 Roma, Italy.
authors.forename: Valeria
authors.initials: V
authors.lastname: Simonelli
authors.forename: Laura
authors.initials: L
authors.lastname: Narciso
authors.forename: Eugenia
authors.initials: E
authors.lastname: Dogliotti
authors.forename: Paola
authors.initials: P
authors.lastname: Fortini
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004260:DNA Repair; D045643:DNA Repair Enzymes / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000378:metabolism; D003855:Deoxyribose / Q000031:analogs & derivatives* / Q000378:metabolism*; D005822:Genetic Vectors; D016296:Mutagenesis; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D012267:Ribosemonophosphates / Q000378:metabolism*; D013539:Simian virus 40 / Q000235:genetics
chemical_list: C502792:2,3-didehydro-2,3-dideoxyribose; C502791:5'-deoxyribose-5-phosphate; D012267:Ribosemonophosphates; D003855:Deoxyribose; D004262:DNA Restriction Enzymes; D045643:DNA Repair Enzymes