MicroRNAs regulate CYP3A4 expression via direct and indirect targeting

Yu Zhuo Pan, Wenqing Gao, Aiming Yu

Research output: Contribution to journalArticle

182 Citations (Scopus)

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. Downregulation 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.

Original languageEnglish (US)
Pages (from-to)2112-2117
Number of pages6
JournalDrug Metabolism and Disposition
Volume37
Issue number10
DOIs
StatePublished - Oct 2009
Externally publishedYes

Fingerprint

Cytochrome P-450 CYP3A
MicroRNAs
3' Untranslated Regions
Luciferases
Response Elements
Plasmids
Calcitriol Receptors
Cytoplasmic and Nuclear Receptors
Cyclophosphamide
Real-Time Polymerase Chain Reaction
Proteins
Down-Regulation
mouse MIRN298 microRNA

ASJC Scopus subject areas

  • Pharmacology
  • Pharmaceutical Science

Cite this

MicroRNAs regulate CYP3A4 expression via direct and indirect targeting. / Pan, Yu Zhuo; Gao, Wenqing; Yu, Aiming.

In: Drug Metabolism and Disposition, Vol. 37, No. 10, 10.2009, p. 2112-2117.

Research output: Contribution to journalArticle

Pan, Yu Zhuo ; Gao, Wenqing ; Yu, Aiming. / MicroRNAs regulate CYP3A4 expression via direct and indirect targeting. In: Drug Metabolism and Disposition. 2009 ; Vol. 37, No. 10. pp. 2112-2117.
@article{b9f5169bb43940229416483cf0540cbc,
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. Downregulation 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.",
author = "Pan, {Yu Zhuo} and Wenqing Gao and Aiming Yu",
year = "2009",
month = "10",
doi = "10.1124/dmd.109.027680",
language = "English (US)",
volume = "37",
pages = "2112--2117",
journal = "Drug Metabolism and Disposition",
issn = "0090-9556",
publisher = "American Society for Pharmacology and Experimental Therapeutics",
number = "10",

}

TY - JOUR

T1 - MicroRNAs regulate CYP3A4 expression via direct and indirect targeting

AU - Pan, Yu Zhuo

AU - Gao, Wenqing

AU - Yu, Aiming

PY - 2009/10

Y1 - 2009/10

N2 - 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. Downregulation 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.

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. Downregulation 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.

UR - http://www.scopus.com/inward/record.url?scp=70349268327&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=70349268327&partnerID=8YFLogxK

U2 - 10.1124/dmd.109.027680

DO - 10.1124/dmd.109.027680

M3 - Article

VL - 37

SP - 2112

EP - 2117

JO - Drug Metabolism and Disposition

JF - Drug Metabolism and Disposition

SN - 0090-9556

IS - 10

ER -