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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

Transcriptional regulation of the chicken CYP2H1 gene. Localization of a phenobarbital-responsive enhancer domain.

The mechanism by which the drugs phenobarbital and 2-allyl-2-isopropylacetamide induce levels of chicken cytochrome P-450 (CYP) mRNAs has been investigated in primary hepatocyte cultures from 17-day-old chick embryos. It has been demonstrated that three CYP mRNAs of 3.5, 2.5, and 2.2 kilobases (kb) are strongly induced by phenobarbital in primary hepatocytes, as found previously in chick embryo liver in ovo (Hansen, A. J., Elferink, L. A., and May, B. K. (1989) DNA (NY) 8, 179-191), and that, at least for the 3.5-kb mRNA, this is predominantly a result of enhanced transcription of the corresponding gene, CYP2H1. Transient transfection assays were carried out in primary cultures using constructs containing different lengths of CYP2H1 gene 5'-flanking sequence fused to the reporter chloramphenicol acetyl-transferase (CAT) gene. These experiments established that cis-acting elements located in the first 0.5 kb of the CYP2H1 gene 5'-flanking region direct high basal expression of the CAT gene, but do not mediate phenobarbital inducibility. When constructs containing more than 1.1 kb of CYP2H1 gene 5'-flanking sequence were examined, phenobarbital induction of CAT expression was observed, and a drug-responsive domain between positions -5.9 and -1.1 kb was identified. This domain has the properties of an enhancer, since it is able to confer phenobarbital responsiveness to the enhancerless SV40 promoter when tested in either orientation or at different distances from the promoter. The enhancer domain also responds to 2-allyl-2-isopropylacetamide, but whether the action of the two drugs is mediated by a single nuclear receptor interacting with common DNA elements in the domain remains to be established.[1]


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