Transcriptional regulation of chicken cytochrome P450 2D49 basal expression by CCAAT/enhancer‐binding protein α and hepatocyte nuclear factor 4α

Abstract

Chicken cytochrome P450 (CYP)2D49 is structurally and functionally related to human CYP2D6, which is an important drug-metabolizing enzyme. To date, little is known about the transcriptional regulation of this cytochrome. Through deletion analysis of the CYP2D49 promoter, we identified two putative degenerate CCAAT/enhancer-binding protein (C/ EBP)-binding sites and an imperfect DR1 element (the site contains direct repeats of the hexamer AGGTCA separated by a one-nucleotide spacer motif) within regions -296/-274, -274/-226, and -226/-183, respectively, which may play critical roles in the transcriptional activation of the CYP2D49 gene. Electrophoretic mobility shift assays and chromatin immunoprecipitation assays showed that the putative C/EBP boxes and DR1 element in the CYP2D49 promoter are functional motifs that bind to C/ EBPa and hepatocyte nuclear factor 4a (HNF4a), respectively. Furthermore, we studied the functional importance and relationships of these transcription factor-binding sites by examining the effects of mutation and deletion of these regions on promoter activity. These studies revealed that the two C/EBP-binding sites show a compensatory relationship and work cooperatively with the DR1 element to modulate the transcription of CYP2D49. The results of overexpressing C/EBPa and HNF4a in culture cells further confirmed that both C/EBPa and HNF4a contribute significantly to sustaining a high level of CYP2D49 transcription. In conclusion, the data indicate that the constitutive hepatic expression of CYP2D49 is governed by both C/EBPa and HNF4a. Further studies will be required to fully characterize the molecular mechanisms that modulate CYP2D49 expression.

Proteomic analysis of eleven tissues in the Chinese giant salamander (Andrias davidianus)

“…Our analysis highlighted the importance of SRF in stomach protection. The activated transcription factor CEBPA regulates proteins related to metabolism and detoxification, especially cytochrome P450 26 , that were highly expressed in the liver of the CGS (Fig. 4D).…”

ACOX1, regulated by C/EBPα and miR-25-3p, promotes bovine preadipocyte adipogenesis

“…For example, C/EBPα regulates transcription of human fructose-1,6-bisphosphatase ( FBP1 ) gene via binding to the two overlapping C/EBPα binding sites located at nucleotide -228/-208 ( Wattanavanitchakorn et al 2018 ), C/EBPα binding to the human polo-like kinase 1 ( PLK1 ) promoter results in suppressed PLK1 expression ( Dasgupta et al 2017 ). Furthermore, there were two C/EBPα binding sites in the chicken cytochrome P450 (CYP) 2D49 promoter, and over-expression of C/EBPα significantly upregulated CYP2D49 transcription ( Yang et al 2014 ). In this study, we identified that C/EBPα binds to the ACOX1 promoter region and suppressed its transcription activity.…”

Coordinated Transcriptional Regulation of Cytochrome P450 3As by Nuclear Transcription Factor Y and Specificity Protein 1

“…Then, 59-biotin-labeled probes corresponding to putative NF-Y or Sp1 binding sites of porcine CYP3A29 and human CYP3A4 and CYP3A5 were prepared using an electrophoretic mobility shift assay (EMSA) Probe Biotin Labeling Kit (Beyotime). EMSA was performed with an EMSA Kit (Beyotime) according to a previously described method (Yang et al, 2014;Dong et al, 2015). Briefly, 5 mg nuclear extract protein from NF-YA/Sp1 overexpressed COS-7 or HepG2 cells were preincubated in binding buffer for 10 minutes at 20°C.…”