CYTOCHROME P450 GENE EXPRESSION IN THE SPRAGUE-DAWLEY RAT (3-METHYLCHOLANTHRENE, PHENOBARBITAL, NUCLEAR RNA)

This dissertation attempted to investigate the mechanisms underlying cytochrome P-450 gene expression in the Sprague-Dawley rat. Various cDNA and genomic DNA clones containing information for cytochromes P-450c/d and a cytochrome P-450b/e-like protein were employed as probes to study the structure and/or expression of these genes in various tissues. Northern blot analysis revealed multiple nuclear RNAs homologous to each of these probes. The cytochrome P-450d and P-450b/e-like nuclear mRNAs were constitutively expressed in rat liver unlike the cytochrome P-450c nuclear mRNA. The cytochrome P-450c/d nuclear RNAs were inducible by 3-methylcholanthrene, isosafrole and Araclor 1254. The cytochrome P-450b/e-like nuclear RNAs were inducible by phenobarbital, 3-methylcholanthrene and Araclor 1254. The primary nuclear RNA transcript from the cytochrome P-450c gene was identified. A "superinduction" of cytochrome P-450c gene expression was observed in liver during the partial inhibition of protein synthesis in vivo. Cytochrome P-450c nuclear RNA was also inducible in fetal liver and adult lung, kidney and spleen. Cytochrome P-450c/d nuclear RNA was constitutively expressed in testes, and was not detected in brain. No cytochrome P-450d nuclear RNA was detected in lung and spleen. No differences were detected in a 9.2 kbp chromosomal domain encompassing the cytochrome P-450c gene in adult and fetal liver, and adult kidney, lung, spleen, and testes. A DNAase I-hypersensitive region at -3.2 to -5.1 kbp was observed in fetal and adult liver. Induction of cytochrome P-450c gene expression in adult liver resulted in a general increase in DNAase I-sensitivity that was most notable in the 5' regions, and that approached the sensitivity of the albumin gene in this tissue. The structural regions of the cytochrome P-450c gene were generally more resistant to DNAase I digestion than the albumin gene in control and "induced" fetal liver, and in control adult kidney and lung.