Specificity of mutations induced by N-methyl-N-nitrosourea in a cDNA of the hprt gene

Abstract

N-Methyl-N-nitrosourea (MNU) reacts with DNA to produce a variety of lesions, of which O⁶-methylguanine (O⁶-MeG) has been implicated in the mutagenic and carcinogenic effects of this agent. The present study aimed to investigate the types and position specificities of mutations induced by MNU. Mutational sequence alterations were determined for 53 independent mutations induced by MNU in a cDNA of the human hprt gene, which is stably integrated into chromosomes of the mouse cell line VH12. The majority of the mutations induced by MNU were base substitutions (85%), mostly GC to AT transitions (41/43), and the remainder (15%) were frameshift or deletion mutations resulting from loss or addition of a few bases. The transition mutations occurred preferentially at the middle G in 5′-purine-G-N-3′ sequences in the non-transcribed strand, and were distributed non-randomly over the coding region of the gene. Analysis of the results suggests that, when interpreting mutational specificity and distribution, not only the nature of the mutational target sequence, but also the functional domains of the protein should be considered.