MicroRNAs Repress Lipogenesis In Hepatocytes By Downregulating LXRs

Non-alcoholic fatty liver disease (NAFLD), characterized by accumulation oftriglycerides in hepatocytes, is one of the most common causes of abnormal liver function.Lipid homeostasis is regulated by a family of transcription factors including the nuclearhormone receptor Liver X receptors (LXRs). LXRα (NR1H3) and LXRβ (NR1H2) are twoisoforms of LXRs, and are known to promote hepatic lipogenesis by activating the lipogenicgenes, such as sterol-regulatory element binding protein1c (SREBP-1c), fatty acid synthase(FAS), carbohydrate responsive element-binding protein (ChREBP) and acetyl-CoAcarboxylase (ACC). Therefore the inhibition of LXR expression and activity may protect theliver from injury inflicted by fat accumulation. MicroRNAs (miRs) are small noncodingRNAs (20–22nucleotide) which bind to the3’-untranslated region (3’-UTR) of mRNAs,thereby repressing gene expression. Recent advances in the understanding of lipid metabolismhave revealed that miRs play major roles in regulating cholesterol and fatty acid homeostasis.However whether miRs can affect hepatic lipogenesis by downregulation of LXRs isunknown.PartⅠ Molecular mechanism of LXRα suppression bymicroRNA-1/206/613and the following effect on hepatic lipogenesisObjective: The purpose of this part is to evaluate the molecular mechanism throughwhich miR-1/206suppress LXRα expression and the effect of miR-1/206on LXRα-inducedlipogenesis in human hepatocytes. Moreover, since miR-613has been reported todownregulate LXRα, so our study concentrate on its effect on LXRα-induced lipogenesis inhuman hepatocytes.Methods and results:1. Real-time PCR and Western blot analyses demonstrated that LXRα expression wasrepressed by miR-1/206/613at both mRNA and protein levels in hepatocytes. As for miR-613, our results were consistent with the former report.2. Bioinformatic tools (TargetScan and PicTar) revealed that miR-1/206/613shared thesame binding site in3’-UTR of human LXRα mRNA. And report assay showed thatmiR-1/206/613directly targeted this bingding site.3. Real-time PCR demonstrated that miR-1/206/613suppressed lipogenic genes（SREBP-1c、FAS、ChREBP andACC1）, which was reversed by LXRα-expressing plasmidpEX-LXRα (without3’-UTR in LXRα mRNA), suggesting that miR-1/206/613suppressedlipogenic genes through the downregulation of LXRα.4. Oil Red O staining showed that the lipid droplet accumulation was repressed bymiR-1/206/613in HepG2cells, which was reversed by LXRα-expressing plasmid pEX-LXRα(without3’-UTR in LXRα mRNA), suggesting that miR-1/206/613suppressed lipid dropletaccumulation through the downregulation of LXRα.Conclusion: Our findings firstly demonstrate that miR-1and miR-206suppress theexpression and activity of LXRα by directly targeting the binding site on LXRα3’-UTR, andmiR-1/206/613inhibit LXRα-mediated lipogenesis in hepatocytes. Therefore,“miR-1/206/613-LXRα” pathway may be a novel target for the treatment of lipogenesis-associated diseases.Part Ⅱ MiR-7/18directly suppress the expression of LXRβObjective: To evaluate the molecular mechanism through which miR-7/18suppress theexpression of LXRβ.Methods and results:1. Real-time PCR and Western blot analyses demonstrated that LXRβ expression wasrepressed by miR-7/18at both mRNA and protein levels.2. Report assay showed that miR-7/18directly targeted the predicted binding sites on3’-UTR of LXRβ mRNA.Conclusion: Our findings firstly demonstrate that miR-7/18directly reduce theexpression of LXRβ by targeting the binding site on LXRβ3’-UTR, which put new insightsinto the mechanism of LXRβ regulation.