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. 2013 Nov;19(11):1525–1536. doi: 10.1261/rna.041533.113

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© 2013 Wang et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society

This article is distributed exclusively by the RNA Society for the first 12 months after the full-issue publication date (see http://rnajournal.cshlp.org/site/misc/terms.xhtml). After 12 months, it is available under a Creative Commons License (Attribution-NonCommercial 3.0 Unported), as described at http://creativecommons.org/licenses/by-nc/3.0/.

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FIGURE 4. — Effects of A-to-I RNA editing at the U3 and U4 or U12 sites on the regulation of miR-30b-3p or miR-573. (A) Western blotting analysis of ARHGAP26 in MCF-10A cells transfected with miRNA negative control (NS-m) or miR-30b-3p mimics, miRNA inhibitor negative control (NS-i), or miR-30b-3p inhibitor. (B) Western blotting analysis of ARHGAP26 in MCF-10A cells transfected with miRNA negative control (NS-m) or miR-573 mimics, miRNA inhibitor negative control (NS-i), or miR-573 inhibitor, respectively. (C) Schematic representation of luciferase reporter constructs containing unedited (adenosine at the U3 and U4 editing sites) or edited (A → G at the U3 and U4 editing sites) form within the miR-30b-3p binding site. (D) Schematic representation of luciferase reporter constructs containing unedited (adenosine at the U12 editing sites) or edited (A → G at the U12 editing sites) formed within the miR-573 binding site. (E) The miR-30b-3p:target interaction was shown with the unedited or edited sites highlighted (upper panel). The effect of the unedited (A) or edited (G) form on the miR-30b-3p:target interaction by analysis of luciferase activity (lower panel). Relative luciferase activities for pGL3-26UTR-unedit or pGL3-26UTR-edit1 cotransfected with the predicted interacting miR-30b-3p and the negative control (=1); the results are presented as mean ± SD of three independent experiments performed in a total of six replicates. (**) Significant differences between unedited and edited targets (P < 0.01). (F) The miR-573:target interaction is shown with the unedited or edited sites highlighted (upper panel). The effect of the unedited (A) or edited (G) form on the miR-573:target interaction by analysis of luciferase activity (lower panel). Relative luciferase activities for pGL3-26UTR-unedit or pGL3-26UTR-edit2 cotransfected with the predicted interacting miR-573 and the negative control (=1); the results are presented as the mean ± SD of three independent experiments performed in a total of six replicates. (**) Significant differences between the unedited and edited targets (P < 0.01). (G) Immunoblotting of ARHGAP26 in MCF-10A cells transfected with control-miRNA, miR-30b-3p, or cotransfected with miR-30b-3p and ADAR1 expression vectors. (H) Immunoblotting of ARHGAP26 in MCF-10A cells transfected with control-miRNA, miR-573, or cotransfected with miR-573 and ADAR1 expression vectors. (I) qPCR analysis of miR-30b-3p in MCF-10A, MCF-7, Chang liver, and HepG2 cells. (J) qPCR analysis of miR-573 in MCF-10A, MCF-7, Chang liver, and HepG2 cells.