MiR-20b Down-Regulates Intestinal Ferroportin Expression In Vitro and In Vivo
MiR-20b Down-Regulates Intestinal Ferroportin Expression In Vitro and In Vivo
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MiR-20b 下调体外和体内肠铁转运蛋白表达
DOI：
10.3390/cells8101135
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发表时间：
2019-10-01
期刊：
影响因子：
6
通讯作者：
Zhao, Ruqian
中科院分区：
文献类型：
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作者：
Jiang, Shuxia;Fang, Xi;Zhao, Ruqian
膜铁转运蛋白（FPN）是哺乳动物细胞中唯一已知的铁输出蛋白。然而，肠道FPN的转录后调控尚未完全了解。本研究应用生物信息学算法（TargetScan、PicTar、PITA和米兰达）预测、筛选和获得靶向FPN的microRNA-17家族成员（miR-17、miR-20 a、miR-20 b和miR-106 a）、“种子序列”和FPN 3′非翻译区（3′UTR）上的响应结合位点。双荧光素酶报告基因分析显示，miRNA-17家族成员的模拟物降低了荧光素酶活性，而其抑制剂增加了荧光素酶活性。与FPN 3′UTR野生型报告基因相比，FPN 3′UTR突变型报告基因与miRNA-17家族成员过表达质粒共转染可提高HCT 116细胞中荧光素酶的活性。转染miR-20 b过表达质粒可显著增强其表达，并抑制Caco-2细胞中内源性FPN蛋白的表达。此外，尾静脉注射miR-20 b导致十二指肠miR-20 b表达增加，十二指肠FPN蛋白表达减少，这与小鼠血浆铁水平降低密切相关。综上所述，这些数据表明，miR-20 b被鉴定为在体外和体内调节肠道FPN表达，这将为肠道铁输出提供潜在的靶点。
Ferroportin (FPN) is the only known cellular iron exporter in mammalian. However, post-transcriptional regulation of intestinal FPN has not yet been completely understood. In this study, bioinformatics algorithms (TargetScan, PicTar, PITA, and miRanda) were applied to predict, screen and obtain microRNA-17 family members (miR-17, miR-20a, miR-20b, and miR-106a) targeting FPN, ‘seed sequence’ and responding binding sites on the 3′untranslated region (3′UTR) region of FPN. Dual-luciferase reporter assays revealed miRNA-17 family members’ mimics decreased the luciferase activity, whereas their inhibitors increased the luciferase activity. Compared with the FPN 3′UTR wild type reporter, co-transfection of a miRNA-17 family members’ over-expression plasmids and FPN 3′UTR mutant reporters enhanced the luciferase activity in HCT116 cells. Transfection with miR-20b overexpression plasmid significantly enhanced its expression, and it inhibited endogenous FPN protein expression in Caco-2 cells. Additionally, tail-vein injection of miR-20b resulted in increasing duodenal miR-20b expression, decreasing duodenal FPN protein expression, which was closely related to lower plasma iron level in mice. Taken together, these data suggest that the miR-20b is identified to regulate intestinal FPN expression in vitro and in vivo, which will provide a potential target for intestinal iron exportation.