title: MicroRNAs regulate CYP3A4 expression via direct and indirect targeting.
abstract: CYP3A4 metabolizes many drugs on the market. Although transcriptional regulation of CYP3A4 is known to be tightly controlled by some nuclear receptors (NR) including vitamin D receptor (VDR/NR1I1), posttranscriptional regulation of CYP3A4 remains elusive. In this study, we show that noncoding microRNAs (miRNAs) may control posttranscriptional and transcriptional regulation of CYP3A4 by directly targeting the 3'-untranslated region (3'UTR) of CYP3A4 and indirectly targeting the 3'UTR of VDR, respectively. Luciferase reporter assays showed that CYP3A4 3'UTR-luciferase activity was significantly decreased in human embryonic kidney 293 cells transfected with plasmid that expressed microRNA-27b (miR-27b) or mouse microRNA-298 (mmu-miR-298), whereas the activity was unchanged in cells transfected with plasmid that expressed microRNA-122a or microRNA-328. Disruption of the corresponding miRNA response element (MRE) within CYP3A4 3'UTR led to a 2- to 3-fold increase in luciferase activity. Immunoblot analyses indicated that CYP3A4 protein was down-regulated over 30% by miR-27b and mmu-miR-298 in LS-180 and PANC1 cells. The decrease in CYP3A4 protein expression was associated with significantly decreased CYP3A4 mRNA levels, as determined by quantitative real-time PCR (qPCR) analyses. Likewise, interactions of miR-27b or mmu-miR-298 with VDR 3'UTR were supported by luciferase reporter assays. The mmu-miR-298 MRE site is well conserved within the 3'UTR of mouse, rat, and human VDR. Down-regulation of VDR by the two miRNAs was supported by immunoblot and qPCR analyses. Furthermore, overexpression of miR-27b or mmu-miR-298 in PANC1 cells led to a lower sensitivity to cyclophosphamide. Together, these findings suggest that CYP3A4 gene expression may be regulated by miRNAs at both the transcriptional and posttranscriptional level.
authors: Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, The State University of New York, Buffalo, NY 14260-1200, USA.
authors: Yu-Zhuo
authors: YZ
authors: Pan
authors: Wenqing
authors: W
authors: Gao
authors: Ai-Ming
authors: AM
authors: Yu
mesh_terms: D000818:Animals; D051544:Cytochrome P-450 CYP3A / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000502:physiology; D018390:Gene Targeting / Q000379:methods*; D006801:Humans; D008297:Male; D051379:Mice; D035683:MicroRNAs / Q000235:genetics / Q000494:pharmacology* / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D035683:MicroRNAs; D012333:RNA, Messenger; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human