miR-145 regulates IGF1R expression.

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miR-145 regulates IGF1R expression. miR-145 regulates IGF1R expression. (A,B) Ishikawa cells were transfected with a non-targeting (control mimic) or miR-145 specific miR mimic (10–100 nM) and appropriate controls. (A) IGF1R mRNA expression was quantified by QPCR and normalized to β-actin mRNA. Data are presented as a fold change in comparison to control (n = 6). There is no observed change in mRNA expression. (B) Western blotting revealed decreased expression of IGF1R protein following transfection with miR-145 mimic at 100 nM in comparison to control. Blots were stripped and re-probed with anti-GAPDH antibodies to correct for protein loading. Blots represent at least three individual sample sets. Data are expressed as median and interquartile range. (C) Synthetic oligonucleotides for the specific (IGF1R 3804–3810) or mutated (IGF1R Δ3804–3810) miR-145-binding sites in IGF1R mRNA, were cloned in to pmiR-GLO luciferase reporter vector and transfected in to Ishikawa cells in the presence of non-targeting (control mimic) or miR-145 specific miR-mimics. Direct interaction between miR-145 and IGF1R in Ishikawa cells was assessed by luciferase reporter assay. Levels of firefly luciferase were assessed and normalized to levels of β-galactosidase. *P<0.05 (n = 5, Kruskal–Wallis test). For A and C, the box represents the 25–75th percentiles, and the median is indicated. The whiskers show the range. Youn-Jung Kang et al. J Cell Sci 2015;128:804-814 © 2015. Published by The Company of Biologists Ltd