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GGA to GAA mutations in the 12th codon of the Hras gene are frequently observed in rat mammary tumors induced by N-nitroso-N-methylurea (NMU). We developed an assay to measure point mutations present in tissues at a frequency of 10(-5) and have now applied this assay to measure the specific G to A transition of the Hras gene in rat mammary epithelium. We find that (i) 70% of untreated rats contain detectable levels of Hras mutants; (ii) these mutants are clustered within the gland as sectors in a manner consistent with their origin as a mutation arising during early organ development; and (iii) treatment with a carcinogenic dose of NMU did not result in a significant increase in the number of such mutants, the fraction of organ sectors with mutant cells, or the fraction of animals containing detectable levels of ras mutants. We conclude that the NMU-induced mammary tumors carrying the G to A transition at the 12th codon of the Hras gene arose from preexisting ras mutants and that an independent effect of NMU was directly or indirectly responsible for tumor formation.

N-nitroso-N-methylurea-induced rat mammary tumors arise from cells with preexisting oncogenic Hras1 gene mutations.