© 1992 Oxford University Press
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DNA sequence analysis of 1-nitrosopyrene-induced mutations in the hprt gene of Chinese hamster ovary cells
Division of Genetic Toxicology, National Center for Toxicological Research Jefferson, AR 72079, USA
DNA sequence was determined in 21 mutants induced at the hprt locus of Chinese hamster ovary (CHO) cells by 1-nitrosopyrene, a metabolite of the tumorigenic environmental pollutant 1-nitropyrene. Following cDNA synthesis using RNA from each of the mutants, the hprt protein-coding region was amplified by the potymerase chain reaction (PCR) and subjected to direct DNA sequence analysis. Sixteen primary mutations were found: seven were G:C 
T:A transversions, five were G:C A:T transitions, two were single basepair insertions, one was a single basepair deletion, and one was a complex mutation involving substitutions at two A:T basepairs. The simple basepair substitution mutations preferentially occurred with one or two purines 3' to the mutated dG, and mutations in exons 1–4 disproportionately occurred with the mutated dG on the non-transcribed DNA strand. In addition, 12 of the mutants produced one or more cDNA PCR products with partial or complete exon deletions. Seven mutants with multiple PCR products had point mutations in one of the products; exon deletions in the other product(s) removed these point mutations. A group of solvent control mutants had a different distribution of basepair substitution mutations and a lower proportion of cDNAs with exon deletions than that found for the 1-nitrosopyrene-induced mutants. The results indicate a specificity for the induction of mutations in the hprt gene of CHO cells by 1-nitrosopyrene with respect to both the types of mutations produced and their location in the hprt gene. Also, the elimination of point mutations in many of the cDNA PCR products with exon deletions suggests that mutations in the protein-coding sequence affect hprt mRNA processing.
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