title: Mutation spectra analysis suggests that N-(2-chloroethyl)-N'-cyclohexyl-N-nitrosourea-induced lesions are subject to transcription-coupled repair in Escherichia coli.
abstract: To determine the influence of some bacterial DNA repair pathways on the mutagenic and the lethal effects of N-(2-chloroethyl)-N'-cyclohexyl-N-nitrosourea (CCNU), pZ189 plasmids treated in vitro with 2 mM CCNU were transfected into Escherichia coli strains with different repair capacities (uvr+ada+ogt+, uvr-ada+ogt+, and uvr-ada-ogt-). Despite the differences in repair capacities, no statistically significant difference in survival and mutability was observed among the tested strains. One hundred and sixty-six CCNU-induced supF mutants were isolated and sequenced. All mutants were characterized by single base-pair substitutions, most of which (more than 96%) were GC-->AT transitions (the mutated G being almost exclusively preceded 5' by a purine). Mutation distribution was not random. Position 160 (5'-GGT-3', nontranscribed (NT) strand) was a uvr+ada+ogt(+)-specific hot-spot. Position 123 (5'-GGG-3', NT strand) was a common hot-spot but significantly more mutable in repair-proficient strains than in repair-deficient strains. Conversely, position 168 (5'-GGA-3', transcribed (T) strand) was significantly more mutable in repair-deficient strains than in repair-proficient strains. By applying a computer program for comparison of mutational spectra, we found that the uvr+ mutational spectrum was significantly different from those obtained in uvr- strains, whereas in the uvr- background, no difference was observed between mutation spectra in ada+ogt+ versus ada-ogt- strains. Our results are consistent with the hypothesis that O6-alkylguanine is responsible for most mutations observed in all strains. The results also indicate that excision repair modulates the distribution of GC-->AT transitions. The fact that mutations at G lesions on the T strand were significantly less frequent in uvr+ than in uvr- strains suggests that CCNU-induced premutational lesions are susceptible to strand-preferential repair in E. coli.
authors.affiliation: Center for the Study of Tumours of Environmental Origins-Laboratory of Mutagenesis, National Institute for Research on Cancer (IST), Genova, Italy.
authors.forename: R
authors.initials: R
authors.lastname: Iannone
authors.forename: A
authors.initials: A
authors.lastname: Inga
authors.forename: F L
authors.initials: FL
authors.lastname: Luque-Romero
authors.forename: P
authors.initials: P
authors.lastname: Menichini
authors.forename: A
authors.initials: A
authors.lastname: Abbondandolo
authors.forename: N
authors.initials: N
authors.lastname: Abril
authors.forename: C
authors.initials: C
authors.lastname: Pueyo
authors.forename: G
authors.initials: G
authors.lastname: Fronza
mesh_terms: D001483:Base Sequence; D002273:Carcinogens / Q000633:toxicity*; D004249:DNA Damage; D004252:DNA Mutational Analysis; D004260:DNA Repair; D004269:DNA, Bacterial / Q000187:drug effects* / Q000378:metabolism; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D008130:Lomustine / Q000633:toxicity*; D008969:Molecular Sequence Data; D009153:Mutagens / Q000633:toxicity*; D014158:Transcription, Genetic
chemical_list: D002273:Carcinogens; D004269:DNA, Bacterial; D009153:Mutagens; D008130:Lomustine