microRNAs (miRNAs), a class of small ~22-nucleotide-long noncoding RNAs, emerge as key post-transcriptional regulators of gene expression in eukaryotic organisms and involve in a variety of cellular processes, such as cell proliferation, cell differentiation and apoptosis. In silico analysis of the tumor necrosis factor-a (TNF-α) and miR-369 showed that the 3'UTR region of porcine (Sus scrofa) TNF-α adenylate/uridylate-rich elements (AREs) contained two putative miR-369 target sites. The porcine TNF-α 3'UTR region was constructed into the dual-luciferase reporter vector, transfected in the porcine iliac artery endothelial cell (PIEC cell) line to measure the luciferase activity by dual-luciferase reporter assay. The results indicated that the luciferase activity of miR-369 mimics group was significantly lower than that of the control group (P<0.05). Nonetheless, there was not a significant differences between the inhibitors and the control groups (P=0.752). Our results demonstrated that the TNF-α should be the target gene of miR-369 and provides evidence for further research of post-transcriptional mechanisms of TNF-α which affect fat deposition.%microRNAs(miRNAs)是一类长约22 nt的非编码小RNA分子,作为关键的转录后调控因子调控真核生物的基因表达,广泛参与细胞的生长发育、分化和凋亡等生物学过程.本研究通过生物信息学预测发现,猪(Sus scrofa)肿瘤坏死抑制因子-α(TN F-α)的3'非编码区具有2个潜在的miR-369的靶位点.并进一步通过双荧光素酶报告系统分析,将猪TNF-α 3'非编码区构建到双荧光素酶载体中,转染至猪髋动脉内皮细胞系,荧光素酶活性测定显示,miR-369过表达组的荧光比值显著低于对照组(P＜0.05),而miR-369抑制组相比对照组没有显著差异(P=0.752).研究结果提示,猪miR-369能靶向调控TNF-α的表达,对深入研究脂肪沉积性状重要候选基因TNF-α的转录后调控机制具有重要价值.
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