title: Effect of nucleotide excision repair on hprt gene mutations in rodent cells exposed to DNA ethylating agents.
abstract: The role of the nucleotide excision repair (NER) pathway in removal of DNA ethylation damage was investigated by means of hprt mutational spectra analysis in the NER-deficient Chinese hamster ovary cell line UV5, which lacks ERCC2/XPD, and in its parental cell line AA8. Both cell lines were exposed to ethyl methanesulfonate (EMS) or N-ethyl-N-nitrosourea (ENU). EMS gave a similar dose-dependent increase in hprt mutants in UV5 compared with AA8. In both cell lines EMS-induced mutations in the hprt coding region consisted almost exclusively of GC-->AT transitions, probably due to the direct miscoding lesion O6-ethylguanine. ENU, an agent that in addition to O6-ethylguanine also induces other O-alkylation products, was significantly more mutagenic in UV5 than in AA8. Mutational spectra analysis showed that the proportions of ENU-induced GC-->AT, AT-->TA and AT-->GC base pair changes were similar for both cell lines. ENU-induced DNA lesions that may be involved in GC-->AT transitions are O6-ethylguanine and O2-ethylcytosine, the latter being a chemically stable DNA lesion of which the miscoding properties and repair characteristics are largely unknown. ENU-induced AT-->TA transversions are probably caused by O2-ethylthymine, which mispairs with thymine. In AA8 thymines in ENU-induced AT-->TA transversions were exclusively located in the non-transcribed strand of the hprt gene, whereas in UV5 30% of these thymines were found in the transcribed strand. Together, these results indicate that O6-ethylguanine is a poor substrate for NER in rodent cells and that O2-ethylpyrimidines are preferentially removed from the transcribed strand of the hprt gene by NER.
authors.affiliation: MGC-Department of Radiation Genetics and Chemical Mutagenesis, Leiden University, The Netherlands.
authors.forename: C W
authors.initials: CW
authors.lastname: Op het Veld
authors.forename: S
authors.initials: S
authors.lastname: van Hees-Stuivenberg
authors.forename: A A
authors.initials: AA
authors.lastname: van Zeeland
authors.forename: J G
authors.initials: JG
authors.lastname: Jansen
mesh_terms: D000477:Alkylating Agents / Q000633:toxicity*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D004252:DNA Mutational Analysis; D004260:DNA Repair; D005020:Ethyl Methanesulfonate / Q000633:toxicity*; D005038:Ethylnitrosourea / Q000633:toxicity*; D005796:Genes / Q000187:drug effects*; D006147:Guanine / Q000378:metabolism; D007041:Hypoxanthine Phosphoribosyltransferase / Q000187:drug effects* / Q000235:genetics*; D009152:Mutagenicity Tests; D009153:Mutagens / Q000633:toxicity*; D009154:Mutation; D013045:Species Specificity; D013941:Thymine / Q000378:metabolism
chemical_list: D000477:Alkylating Agents; D009153:Mutagens; D006147:Guanine; D004247:DNA; D005020:Ethyl Methanesulfonate; D007041:Hypoxanthine Phosphoribosyltransferase; D005038:Ethylnitrosourea; D013941:Thymine