Transcriptional regulation of rat alpha 1-acid glycoprotein gene by phenobarbital.

T Fournier,N Mejdoubi,C Lapoumeroulie,J Hamelin,J Elion,G Durand,D Porquet

doi:10.1016/s0021-9258(18)46964-5

Abstract

Phenobarbital induces gene transcription of both cytochrome P450IIB (the barbiturate-inducible cytochrome P450 in mammals) and alpha 1-acid glycoprotein, one of the major acute-phase proteins in rats and humans. Analysis of the 5'-regulatory sequences of cytochrome P450IIB and alpha 1-acid glycoprotein genes in rats revealed the presence of a consensus sequence of 10 base pairs, termed the phenobarbital-responsive element or Barbie box, located in a region extending from positions -136 to -127 from the transcription start site of the alpha 1-acid glycoprotein gene. A 17-base pair oligonucleotide probe specific for alpha 1-acid glycoprotein and including the consensus sequence showed, in mobility shift assays, slight binding to liver nuclear protein from untreated animals. This binding was strongly and specifically increased with protein extracts from phenobarbital-treated rats. Transfection of rat primary hepatocytes with the pAGPcat construct induced basal expression of chloramphenicol acetyltransferase activity, which was increased by phenobarbital and dexamethasone treatment of cells. Induction of chloramphenicol acetyltransferase activity by phenobarbital was abolished when hepatocytes were transfected by constructs with a mutation or deletion of the Barbie box sequence. These results strongly suggest that the Barbie box sequence is involved in alpha 1-acid glycoprotein gene regulation by phenobarbital.

Highlights

We have shownin the rat that PB induces an increase in the transcription rateof gene of a protein thatt,o our knowledge, is Phenobarbital induces gene transcription of both cy-not involved in steroid or drug metabolism, uiz. a,acid glycotochromPe450IIB (thbearbiturate-induciblceytoprotein (AGP) (16), oneof the major positive acute-phase prochrome P450in mammals) and a,acid glycoprotein, onteeins in rodentsandhumans (17)
17-bp PBRE sequence from positions -140 to -124; construct 3), the protein extracted from turpentine-treated rats did not bind to EcoRIIXhoI fragment of pAGPcat was subcloned into bacteriophage the probe
Phenobarbital hasprofound inductive effects on a variety of biotransformation systems, and severmalolecular mechanisms have been proposed to account for transcriptional activation of the corresponding genes(see Ref. 2 for a review)

Summary

Introduction

This binding was strongly and increased in 1992 (2)) has been evidenced for genes encoding CYPBB in with protein extracts from phenobarbital-treated rats. Basis of this observation, a labeled oligonucleotide probe corresponding to thePBRE sequence was synthesized and tested in gel retardation assays with nuclear protein extrfarcotms livers of control and PB-treatedrats.

Results

Conclusion