ALTERATIONS IN H-RAS1 PROMOTER CONFORMATION DURING N-NITROSO-N-METHYLUREA-INDUCED MAMMARY CARCINOGENESIS AND PREGNANCY

We previously demonstrated that H-ras1 oncogene mutations detected in N-nitroso-N-methylurea (NMU)-induced mammary tumors arose as backgroun d mutations within rat mammary cells (RMCs) and that NMU promoted the outgrowth of these preexisting mutants. We have now detected a putativ e DNA structure in the H-ras1 promoter of RMCs in vivo that was absent in NMU-induced mammary tumor cells. Analysis of the promoter in RMCs as a function of time after exposure to carcinogens indicated that NMU , but not 7,12-dimethylbenz(a)anthracene, initiated the loss of this s tructure with a half-life of 7 days. Although loss of the structure wa s irreversible in cells that gave rise to tumors, it was restored in n ormal RMCs by 120 days after exposure and was present in normal RMCs o f animals bearing tumors, even 1 year after NMU exposure. The structur e was also abrogated in RMCs during pregnancy and restored after lacta tion was terminated, suggesting that reversible regulation of the stru cture by hormones contributed to normal RMC growth. Thus, NMU may prom ote abnormal RMC growth by mimicking the effects of hormones on DNA co nformation. We hypothesize that the NMU-induced alterations in promote r conformation irreversibly deregulates H-ras1 expression in initiated cells, thereby increasing the phenotypic penetrance of the conditiona l H-ras1 mutations.