Coordinated regulation of miR-27 by insulin/CREB/Hippo contributes to insulin resistance

Abstract MicroRNA-27 is a key non-coding metabolic gene and is often aberrantly overexpressed in non-alcoholic fatty livers (NAFLD). However, the pathogenic role of miR-27 in NAFLD remains largely unknown. In this study, we attempted to identify the mechanism by which miR-27 was regulated in the context of insulin resistance, a predisposed metabolic disorder in NAFLD. With cell culture experiments and animal studies, our results showed that miR-27 was coordinately regulated by insulin, CREB, and Hippo signalings. First, miR-27 was upregulated in palmitate-treated cells and high fat diet-fed mouse livers, both of which exhibited insulin resistance and increased CREB expression. Second, miR-27 was peaked in mouse liver at post-absorptive phase when CREB activity was increased. Also, miR-27 was increased rapidly in insulin-treated cell lines when CREB activity was decreased. Third, miR-27 was downregulated in cultured cells when CREB was decreased by siRNA or metformin treatment. In contrast, miR-27 was upregulated when CREB was activated by Forskolin. Fourth, miR-27 was repressed by Hippo signaling in a CREB-independent manner: miR-27 was reduced in cells at full confluence but was inhibited in cells transfected with siRNA against Lats2 and Nf2, which were two positive regulators of Hippo signaling. Lastly, bioinformatics and luciferase assay showed that miR-27 inhibited Akt phosphorylation by targeting Pdpk1 and Pik3r1. Overexpression of miR-27 impaired Akt phosphorylation in cell lines and primary mouse hepatocytes upon insulin stimulation. Taken together, our data suggest that insulin, CREB, and Hippo signalings contribute to aberrant miR-27 overexpression and eventually lead to insulin resistance in NAFLD.