Toxicity, mutagenicity, and mutational spectra of N-ethyl-N-nitrosourea in human cell lines with different DNA repair phenotypes.
We examined the toxicity, mutagenicity, and mutational spectra of N-ethyl-N-nitrosourea (ENU) in three Epstein-Barr virus-transformed human lymphoblastoid cell lines, each with a different DNA repair phenotype. One cell line lacks O6-alkylguanine-DNA alkyltransferase (AGT) activity; another, derived from a patient with xeroderma pigmentosum, complementation group A, lacks nucleotide exicision repair (NER) capability, and the third is competent in both repair functions. ENU-induced toxicity and mutagenicity at the hypoxanthine-guanine phosphoribosyltransferase locus were increased to a similar degree relative to the repair-competent cells in both AGT-deficient and NER-deficient cells. We determined the mutational spectra for ENU by identifying DNA sequence changes at the hypoxanthine-guanine phosphoribosyltransferase locus in at least 26 clones resistant to 6-thioguanine from each cell line. Of the characterized mutations, 89% were single-base pair substitutions. Transitions and transversions were found at AT and GC base pairs in all three cell lines. The biggest difference within the spectra was in the rate of transitions at GC base pairs. Compared to the repair-competent cell line, this mutation was elevated about 8-fold in the AGT-deficient cells and about 3-fold in the NER-deficient cells. We conclude that both AGT and NER play an important role in protecting human cells from the toxic and mutagenic effects of ENU. Furthermore, the mutational spectra suggest that both of these repair systems participate in the repair of O6-ethylguanine adducts.
Bronstein
S M
SM
Department of Pathology, Duke University, Durham, North Carolina 27710.
Cochrane
J E
JE
Craft
T R
TR
Swenberg
J A
JA
Skopek
T R
TR
D000293:Adolescent; D000328:Adult; D001483:Base Sequence; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects*; D002472:Cell Transformation, Viral; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004247:DNA / Q000187:drug effects*; D004260:DNA Repair / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D005038:Ethylnitrosourea / Q000494:pharmacology*; D005260:Female; D004854:Herpesvirus 4, Human; D006801:Humans; D066298:In Vitro Techniques; D008297:Male; D008780:Methyltransferases / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation / Q000187:drug effects*; D019853:O(6)-Methylguanine-DNA Methyltransferase; D016133:Polymerase Chain Reaction
D004247:DNA; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase; D005038:Ethylnitrosourea