A novel approach for the mechanism of CYP2B induction: A study using mutant rats that lack response to the PB-mediated induction of CYP2B2 and the analyses of the gene structure and transcription factors

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The Qdj:SD rat is a mutant strain lacking in phenobarbital (PB)-mediated induction of CYP2B2. Presence of inter-individual differences in the hepatic content of CYP2B proteins and testosterone 16β-hydroxylase activity demonstrated that the breeding colony of Qdj :SD rats involves normal (+/+)-and intermediate (+/-)- phenotypes as well as mutant (-/-)-type rats. Analysis of regioselective metabolism of testosterone and 4-hydroxybiphenyl glucuronidation demonstrated normal catalytic activities associated with other forms of P450s, including CYP2A, 2C and 3A, as well as PB-inducible UGT-glucuronosyltransferase in Qdj:SD (-/-) rats. Hepatic CYP2B2 mRNA was detectable by reverse transcription - polymerase chain reaction in the Qdj:SD rats treated with phenobarbital (PB), but the content was far lesser (<1/10) than that in the drug-untreated Crj:SD rat, a reference animal with normal phenotype. The CYP2B2 gene in Qdj:SD (-/-) rats was same as those of the wild-type (+/+) rats in its length of the region containing all exon/introns and 5'-upstream up to -4.7 kbp. Malignant mutation such as stop codon formation was not observed in the exons, and no mutation was detected in the region containing the PB-responsive enhancer module (PBREM). The binding between 32P-labelled PBREM and hepatic nuclear proteins was not increased both in Qdj;SD (-/-) and Crj:SD rats by treatment of animals with PB, although PB-mediated increase in the binding was observed in mice. These results strongly suggest that the impaired induction of CYP2B2 in Qdj:SD (-/-) rats is due to the damage in basic expression of the gene. The impaired machinery for the expression is assumed to be either 1) mutation at the region different from PBREM and exons, or 2) absence or lowered expression of trans-acting factor(s) different from PBREM binding proteins.