Functional role of miR-148a in oropharyngeal cancer: influence on pregnane X receptor and P-glycoprotein expression.

Abstract

MicroRNAs are short noncoding RNAs of about 19-25 nucleotides that usually target the 3' untranslated regions of mRNAs thus mediating post-transcriptional regulation of gene expression. Previous data indicate a role for miR-148a in the regulation of the pregnane X receptor (PXR/NR1I2), a nuclear receptor that regulates the expression of drug transporters like P-glycoprotein (P-gp/ABCB1). Our study investigated the effect of miR-148a on the post-transcriptional regulation of PXR and its target gene ABCB1 in oropharyngeal cancer cell lines (OPSCC). miR-148a was over-expressed and knocked-down in three OPSCC cell lines (HNO41, HNO206, and HNO413) by transfection with miR-148a mimic and miR-148a antagomir, respectively. Expression of miR-148a, NR1I2, and ABCB1 mRNA was quantified via real-time qPCR, protein expression of PXR was assessed by immunoblotting. Transfection of miR-148a mimic led to increased miR-148a levels in all cell lines and transfection of miR-148a antagomir reduced miR-148a expression in HNO206 and HNO413. Whereas these changes had no significant effect on PXR mRNA expression, protein expression was reduced in HNO41 by transfection with miR-148a and increased in HNO413 by transfection with miR-148a antagomir. Transfection of miR-148a downregulated ABCB1 mRNA in all cell lines, whereas antagonizing miR-148a had no significant effect. Our data demonstrate a modulation of PXR/NR1I2 and ABCB1 expression in OPSCC by miR-148a, however the effect was not uniform in all cell lines and depended on the range of expression of miR-148 and the genotype of rs1054190 SNP in NR1I2 3'UTR. Thus, our findings argue against an unequivocal association between miR-148a and PXR levels in OPSCC.