Abstract
Multidrug resistance protein 4 (MRP4, ABCC4) is an efflux membrane transporter expressed in renal tubules, hepatocytes, brain capillaries, prostate and blood cells. MRP4 drives energy dependent efflux of important physiological and pharmacological compounds. MRP4 expression and function is highly variable but cannot be fully attributed to known mechanisms. The goal of this study was to characterize ABCC4 regulation by miRNAs and to assess the influence of ABCC4 3'-UTR polymorphisms on ABCC4 regulation by miRNAs. miR-124a and miR-506 decreased MRP4 protein levels in HEK293T/17 cells 20-30% and MRP4 function by 50%. These miRNAs did not affect ABCC4 mRNA expression. Moreover, miR-124a and miR-506 expression was negatively correlated with MRP4 protein expression in 26 human kidney samples (Spearman r=-0.62, P=0.007 and r=-0.41, P=0.03 for miR-124a and miR-506, respectively). To assess the effect of ABCC4 3'-UTR polymorphisms, six common 3'-UTR haplotypes were inferred in Caucasians, African Americans and Asians and tested in luciferase reporter assays. Multiple ABCC4 3'-UTR haplotypes caused significant reductions in luciferase activity; in the presence of miR-124a or miR-506 mimics the luciferase activity of all six ABCC4 3'-UTR haplotypes was further reduced. Mutation of the putative binding site for miR-124a and miR-506 in the ABCC4 3'-UTR eliminated the effect of these miRNAs. In conclusion, ABCC4 is directly regulated by miR-124a and miR-506 but polymorphisms in the ABCC4 3'-UTR have no significant effect on this miRNA regulation. Regulation of ABCC4 by miRNAs represents a novel mechanism for regulation of MRP4 function.