miRZip_124 anti_miR_124 microRNA construct productsSearch results for: miRZip_124 anti_miR_124 microRNA construct
#23648570 
2013/05/03
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Insulin suppresses distal-less homeobox 5 expression through the up-regulation of microRNA-124 in 3T3-L1 cells.
Distal-less homeobox 5 (Dlx5) is a pro-osteogenic but anti-adipogenic transcription factor that regulates lineage commitment in mesenchymal stem cells. Although the expression of Dlx5 is known to be decreased by adipogenic stimuli, the mechanism of Dlx5 down-regulation has not yet been clarified. MicroRNAs (miRNAs) are small regulatory RNAs that post-transcriptionally regulate many biological functions, including cell differentiation. In this study, we examined whether miRNAs are involved in down-regulation of Dlx5 following adipogenic stimuli. We screened candidate miRNAs that have a direct target site in the Dlx5 3'UTR using computational prediction programs, selected seven miRNA candidates with the highest binding score and observed their expression levels in 3T3-L1 murine pre-adipocytes. Among the miRNAs examined, only miR-124 was significantly up-regulated by 24-h incubation in adipogenic medium. Among the four components of adipogenic stimuli (1-methy-3-isobutyl xanthine, insulin, indomethacin and dexamethasone), insulin exhibited the highest stimulatory effect on miR-124 expression. Insulin significantly increased the expression of miR-124 precursors including pri-miR-124-1, pri-miR124-2 and pri-miR-124-3. LY294002, an inhibitor of phosphatidylinositol-3-kinase, prevented the regulatory effect of insulin on the expression levels of miR-124 and Dlx5. Over-expression of a miR-124 mimic decreased the expression of Dlx5 while increasing adipogenic differentiation in 3T3-L1 cells. Blocking miR-124 with anti-miR-124, a hairpin inhibitor of miR-124, increased the expression level of Dlx5 and suppressed adipogenic differentiation. When reporter assays were performed with a reporter construct containing the Dlx5 3'UTR sequence downstream of a luciferase gene, miR-124 mimic suppressed, but anti-miR-124 enhanced, luciferase activity in an miR-124 binding site-dependent manner. These results suggest that insulin-induced miR-124 plays a pivotal role in post-transcriptional regulation of Dlx5 during adipogenic differentiation and that miR-124 exerts pro-adipogenic effects by targeting Dlx5, at least in part.Abdul S Qadir, Kyung Mi Woo, Hyun-Mo Ryoo, Jeong-Hwa Baek
