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Chinese Journal of Clinical Pharmacology and Therapeutics ›› 2018, Vol. 23 ›› Issue (8): 846-854.doi: 10.12092/j.issn.1009-2501.2018.08.002

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Effects of miR-1-3p/206/613 on the expression of LXRα and OATP1B1 proteins and its mechanism

YE Rongfang, LIU Mingyi, XU Yu, ZHANG Yanli, ZHU Dandan, DING Wei, XIONG Yuqing   

  1. Clinical Pharmacology Institute of Nanchang University，Nanchang 330006，Jiangxi，China
  • Received:2018-03-06 Revised:2018-07-04 Online:2018-08-26 Published:2018-08-28

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Abstract:

AIM: To explore the effect of miR-1-3p, miR-206 and miR-613 on the expression of LXRα and OATP1B1 proteins, and the effect of them on the transporting function of OATP1B1 and further to explore the molecular mechanisms underlying the above effects. METHODS: Bioinformatics databases were used to predict the potential miRNAs targeting on the 3'-UTR of LXRα and OATP1B1 mRNA; the levels of miR-1-3p/206/613,LXRα and OATP1B1 proteins were detected by RT-qPCR and Western blotting; the content of rosuvastatin in HepG2 cells was detected by LC-MS/MS; Dual-luciferase reporter assays was used to study the molecular mechanism in which miR-1-3p/206/613 affect the expression of LXRα and OATP1B1.RESULTS:The predicted results showed that miR-1-3p/206/613 has perfect complementary with 3′-UTR of LXRα and OATP1B1 mRNA, which has relatively high specificity,conservation and binding stability. Compared with the control group, miR-1-3p/206/613 mimics could significantly down-regulate LXRα protein level in HepG2 cells by 16.5%,16.0% and 25.1%, OATP1B1 protein level by 30.4%, 30.5% and 44.4%, respectively. On the contrary, miR-1-3p/206/613 inhibitors significantly upregulated LXRα protein level by 13.3%,13.3% and 16.0%,OATP1B1 protein level by 25.0%,25.6% and 30.4%. When the concentrations of RSV were 5,60 and 125 μmol/L, miR-1-3p/206/613 mimics significantly reduced the uptakes of RSV in HepG2 cells to 0.50,0.19,0.30 fold,0.49,0.24,0.23 fold,0.64,0.48,0.31 fold of the control group, respectively; In contrast,miR-1-3p/206/613 inhibitors upregulated the uptakes of RSV by 1.26,1.59,2.07 fold,1.97,2.44,2.63 fold,2.22,2.86,2.93 fold.Compared with the control group, miR-1-3p/206/613 mimics or miR-1-3p/206/613 inhibitors decreased the luciferase activities of pGL/LXRα-WT reporter gene by 38.5%,32.5%,2.8% or increased by 137.0%,75.8%,55.7%.But they had no obvious effect on the luciferase activity of pGL/LXRα-Mut reporter gene. As above, the luciferase activities of the pGL/OATP1B1-WT reporter gene decreased by 24.3%, 28.9%,32.1% or increased by 33.5%,48.3%,61.6%,respectively.But they had no obvious effect on the luciferase activity of the pGL/OATP1B1-Mut reporter gene. CONCLUSION: miR-1-3p/206/613 can regulate the expression of OATP1B1 protein and transporting function of OATP1B1 by directly targeting the 3'-UTR of OATP1B1 mRNA, and can also exert indirect regulation through targeting the 3'-UTR of LXRα mRNA.

Key words: miR-1-3p, miR-206, miR-613, LXRα, OATP1B1

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