Functional characterization of the promoter of human carbonyl reductase 1 (CBR1). Role of XRE elements in mediating the induction of CBR1 by ligands of the aryl hydrocarbon receptor.
Human carbonyl reductase 1 (CBR1) metabolizes a variety of substrates, including the anticancer doxorubicin and the antipsychotic haloperidol. The transcriptional regulation of CBR1 has been largely unexplored. Therefore, we first investigated the promoter activities of progressive gene-reporter constructs encompassing up to 2.4 kilobases upstream of the translation start site of CBR1. Next, we investigated whether CBR1 mRNA levels were altered in cells incubated with prototypical receptor activators (e.g., dexamethasone and rifampicin). CBR1 mRNA levels were significantly induced (5-fold) by the ligand of the aryl hydrocarbon receptor (AHR) beta-naphthoflavone. DNA sequence analysis revealed two xenobiotic response elements ((-122)XRE and (-5783)XRE) with potential regulatory functions. CBR1 promoter constructs lacking the (-122)XRE showed diminished (9-fold) promoter activity in AHR-proficient cells incubated with beta-naphthoflavone. Fusion of (-5783)XRE to the (-2485)CBR1 reporter construct enhanced its promoter activity after incubations with beta-naphthoflavone by 5-fold. Furthermore, we tested whether the potent AHR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induced Cbr1 expression in Ahr(+/-) and Ahr(-/-) mice. TCDD induced hepatic Cbr1 mRNA (TCDD, 2-fold) and Cbr1 protein levels (TCDD, 2-fold) in Ahr(+/-) mice compared with vehicle-injected controls. In contrast, no significant Cbr1 mRNA and Cbr1 protein induction was detected in livers from Ahr(-/-) mice treated with TCDD. These studies provide the first insights on the functional characteristics of the human CBR1 gene promoter. Our data indicate that the AHR pathway contributes to the transcriptional regulation of CBR1.
Lakhman
Sukhwinder S
SS
Department of Pharmaceutical Sciences, the State University of New York at Buffalo, Buffalo, New York 14260-1200, USA.
Chen
Xiaomin
X
Gonzalez-Covarrubias
Vanessa
V
Schuetz
Erin G
EG
Blanco
Javier G
JG
D000429:Alcohol Oxidoreductases / Q000235:genetics*; D000449:Aldehyde Reductase; D000074408:Aldo-Keto Reductases; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001943:Breast Neoplasms / Q000473:pathology; D006528:Carcinoma, Hepatocellular / Q000473:pathology; D045744:Cell Line, Tumor; D005260:Female; D017930:Genes, Reporter; D006801:Humans; D008024:Ligands; D008113:Liver Neoplasms / Q000473:pathology; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012333:RNA, Messenger / Q000032:analysis; D018336:Receptors, Aryl Hydrocarbon / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid
D008024:Ligands; D000072317:Polychlorinated Dibenzodioxins; D012333:RNA, Messenger; D018336:Receptors, Aryl Hydrocarbon; D000429:Alcohol Oxidoreductases; D000074408:Aldo-Keto Reductases; D000449:Aldehyde Reductase; C000614209:AKR7A5 protein, mouse; D008156:Luciferases