FORMATION AND PERSISTENCE OF O-6-ETHYLGUANINE IN GENOMIC AND TRANSGENE DNA IN LIVER AND BRAIN OF LAMBDA-LACZ TRANSGENIC MICE TREATED WITH N-ETHYL-N-NITROSOUREA

LacZ transgenic mice are suitable for short-term mutagenicity studies in vivo. Mutagenicity in these mice is determined in the lacZ transgen e, Since the lacZ gene is of bacterial origin the question has been ra ised whether DNA-adduct formation and repair in the transgene are comp arable to those in total genomic DNA, Mice were treated with N-ethyl-N -nitrosourea (ENU) and killed at several time points following treatme nt, Some mice were pretreated with O-6-benzylguanine to inactivate the repair protein O-6-alkylguanine-DNA alkyltransferase (AGT), O-6-ethyl guanine (O-6-EtG) was determined in lacZ in liver and brain by means o f a monoclonal antibody-based immunoaffinity assay, In addition, O-6-E tG and N-7-ethylguanine (N7-EtG) were assayed in total genomic DNA of liver and brain with an immunoslotblot procedure, In liver, the initia l O-6-EtG level in total genomic DNA was 1.6 times that in lacZ, The e xtent of repair of O-6-EtG during the first 1.5 h after treatment was 2.1 times that in lacZ, At later time points, O-6-EtG repair was the s ame. N7-EtG repair in genomic DNA was evident. In contrast to the live r, little repair of O-6-EtG in total genomic and lacZ DNA occurred in the brain while N7-EtG was repaired, No initial difference in O-6-EtG levels were found in lacZ and genomic brain DNA, These findings indica te that in the liver, total genomic DNA is more accessible than lacZ t o ENU and/or the AGT protein, during the first 1.5 h following treatme nt, Because the difference in O-6-EtG levels in the transgene and geno mic DNA in the liver is restricted to the first 1.5 h after treatment, while the fixation of mutations occurs at later time points, O-6-EtG- induced mutagenesis most likely is also very similar in both types of DNA.