Specificity of bischloroethylnitrosourea-induced mutation in a Chinese hamster ovary cell line transformed to express human O6-alkylguanine-DNA alkyltransferase.
The human O6-alkylguanine-DNA alkyltransferase complementary DNA was transfected into the alkyltransferase-deficient Chinese hamster ovary cell line, D422, in an effort to dissect the underlying mechanisms of bischloroethylnitrosourea (BCNU)-induced mutations. The alkyltransferase-transformed cell line exhibited 100-fold protection against BCNU-induced toxicity and an overall decrease in mutation frequency to 25% of that observed in the parental cell line at the hemizygous adenine phosphoribosyl transferase gene target. The frequency of the predominant mutation in the parental cell line, the G:C-->T:A transversion, was reduced from 16 x 10(-6) to 0.7 x 10(-6) in the O6-alkyltransferase-transformed cell line. Likewise, the G:C-->A:T transitions, the second most common BCNU-induced mutation in the parental cell line, was reduced in frequency from 5.2 x 10(-6) to 0.9 x 10(-6) in the alkyltransferase-transformed Chinese hamster ovary cells. These findings suggest that both the G:C-->T:A transversions and G:C-->A:T transitions were O6-alkylguanine-mediated mutations. In the alkyltransferase-transformed Chinese hamster ovary cell line, T:A-->G:C transversions, comprising 45% (23 of 51) of the recovered mutations, emerged as the most common base substitution. In summation, in the absence of alkyltransferase-dependent DNA repair, mutations resulting from O6-alkylation of guanine underlie both the cytotoxic and mutagenic activity of BCNU. In cells expressing high levels of alkyltransferase activity, the cytotoxic and mutagenic actions of BCNU are greatly reduced and mutations resulting from A:T base pair modifications appear to be the major genotoxic lesions induced by the drug.
Minnick
D T
DT
Environmental Health Sciences, Case Western Reserve University, Cleveland, OH 44106.
Gerson
S L
SL
Dumenco
L L
LL
Veigl
M L
ML
Sedwick
W D
WD
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002330:Carmustine / Q000494:pharmacology*; D002461:Cell Line, Transformed; D006224:Cricetinae; D004260:DNA Repair; D017353:Gene Deletion; D006801:Humans; D008780:Methyltransferases / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D009154:Mutation; D019853:O(6)-Methylguanine-DNA Methyltransferase; D014162:Transfection
D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase; D002330:Carmustine