The CYP2B2 phenobarbital response unit contains binding sites for hepatocyte nuclear factor 4, PBX-PREP1, the thyroid hormone receptor beta and the liver X receptor.
A 163 bp enhancer in the CYP2B2 5' flank confers PB (phenobarbital) inducibility and constitutes a PBRU (PB response unit). The PBRU contains several transcription factor binding sites, including NR1, NR2 and NR3, which are direct repeats separated by 4 bp of the nuclear receptor consensus half-site AGGTCA, as well as an ER (everted repeat) separated by 7 bp (ER-7). Constitutive androstane receptor (CAR)-RXR (retinoic X receptor) heterodimers are known to bind to NR1, NR2 and NR3. Electrophoretic mobility-shift analysis using nuclear extracts from livers of untreated or PB-treated rats revealed binding of several other proteins to different PBRU elements. Using supershift analysis and in vitro coupled transcription and translation, the proteins present in four retarded complexes were identified as TRbeta (thyroid hormone receptor beta), LXR (liver X receptor), HNF-4 (hepatocyte nuclear factor 4) and heterodimers of PBX-PREP1 (pre-B cell homoeobox-Pbx regulatory protein 1). LXR-RXR heterodimers bound to NR3 and TRbeta bound to NR3, NR1 and ER-7, whereas the PBX-PREP1 site is contained within NR2. The HNF-4 site overlaps with NR1. A mutation described previously, GRE1m1, which decreases PB responsiveness, increased the affinity of this site for HNF-4. The PBRU also contains a site for nuclear factor 1. The PBRU thus contains a plethora of transcription factor binding sites. The profiles of transcription factor binding to NR1 and NR3 were quite similar, although strikingly different from, and more complex than, that of NR2. This parallels the functional differences in conferring PB responsiveness between NR1 and NR3 on the one hand, and NR2 on the other.
Beaudet
Marie-Josée
MJ
Centre de recherche en cancérologie de l'Université Laval, L'Hôtel-Dieu de Québec, Centre hospitalier universitaire de Québec, Québec G1R 2J6, Canada.
Desrochers
Marc
M
Lachaud
Antoine Amaury
AA
Anderson
Alan
A
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D001483:Base Sequence; D001665:Binding Sites; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D051557:Hepatocyte Nuclear Factor 4; D022781:Hepatocytes / Q000378:metabolism; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D000071518:Liver X Receptors; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D057093:Orphan Nuclear Receptors; D010634:Phenobarbital / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D020218:Response Elements; D013250:Steroid Hydroxylases / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D037042:Thyroid Hormone Receptors beta / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism
D004268:DNA-Binding Proteins; D051557:Hepatocyte Nuclear Factor 4; D018398:Homeodomain Proteins; D000071518:Liver X Receptors; D057093:Orphan Nuclear Receptors; D010750:Phosphoproteins; D018160:Receptors, Cytoplasmic and Nuclear; D037042:Thyroid Hormone Receptors beta; D014157:Transcription Factors; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C025619:steroid 16-beta-hydroxylase; D010634:Phenobarbital