Epoxide hydrolase (EH) is an important enzyme in the activation of a number of procarcinogens, e.g., polycyclic hydrocarbons. In this study, we propose to determine the levels of EH mRNA in the livers of fetal, neonatal, weanling and adult rats; in the livers of animals treated with trans-stilbene oxide, phenobarbital, and 2-acetylaminofluorene (AAF); in the hyperplastic nodules and hepatomas that result after administration of AAF. The processing of nuclei precursor mRNA will be ascertained in these various models. The ability of nuclei to transverse the EH mRNA under in vitro conditions will also be determined. Where this is to be so, we will establish if the induced can increase mRNA transcription under these conditions. We already have available a pBR322 recombinant DNA clone PB 17, which contains EH information. PB 17 plasmid DNA will be used for determining EH mRNA. The amount of mRNA will be ascertained by hybridization techniques under excess RNA conditions. Whether or not AAF treatment results in amplification of the EH gene will be ascertained. A full length cDNA will be produced using the chain extension method with pUC8 as the plasmid vector. The DNA sequence of the full length cDNA will be determined. Finally, a rat genomic library will be searched for EH gene information and the structural details will be established. The regulatory regions in the genomic DNA will be sought by incubation with the various inducers. These studies should give us information about the changes in EH mRNA during ontogeny, after administration of certain exogenous substances, during preneoplasia and hepatoma formation, as well as about the regulation of the genomic DNA.
