PT  - JOURNAL ARTICLE
AU  - Pan, Yu-Zhuo
AU  - Gao, Wenqing
AU  - Yu, Ai-Ming
TI  - MicroRNAs Regulate CYP3A4 Expression via Direct and Indirect Targeting
AID  - 10.1124/dmd.109.027680
DP  - 2009 Oct 01
TA  - Drug Metabolism and Disposition
PG  - 2112--2117
VI  - 37
IP  - 10
4099  - http://dmd.aspetjournals.org/content/37/10/2112.short
4100  - http://dmd.aspetjournals.org/content/37/10/2112.full
SO  - Drug Metab Dispos2009 Oct 01; 37
AB  - 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. Copyright © 2009 by The American Society for Pharmacology and Experimental Therapeutics