MicroRNA-23a downregulates the expression of interferon regulatory factor-1 in hepatocellular carcinoma cells.
MicroRNA-23a downregulates the expression of interferon regulatory factor-1 in hepatocellular carcinoma cells.
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DOI：
10.3892/or.2016.4864
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发表时间：
2016-08
期刊：
影响因子：
4.2
通讯作者：
Geller DA
中科院分区：
文献类型：
--
作者：
Yan Y;Liang Z;Du Q;Yang M;Geller DA
干扰素调节因子 -1（IRF -1）是一种由干扰素 -γ（IFNγ）诱导的肿瘤抑制基因，在肝细胞癌（HCC）的细胞死亡中起重要作用。HCC肿瘤部分通过下调IRF -1的表达来逃避死亡，但HCC中导致IRF -1抑制的分子机制尚未明确。先前的研究表明，微小RNA -23a（miR -23a）可通过靶向IRF -1抑制细胞凋亡。因此，我们假设miR -23a通过下调IRF -1促进HCC生长。在体内研究中，分析了7例切除的HCC及相邻肝组织样本。在体外研究中，分别通过实时定量PCR和蛋白质印迹法检测了IFNγ刺激后HepG2和Huh -7 HCC细胞中的IRF -1 mRNA和蛋白质。为了确定miR -23a在调节IRF -1中的作用，用miR -23a模拟物或抑制剂转染HepG2细胞，并检测IRF -1的表达。通过将528bp的人IRF -1 3′ - 非翻译区（3′UTR）克隆到荧光素酶报告质粒pMIR - IRF -1 - 3′UTR中来评估miR -23a的结合。结果显示，与相邻的背景肝组织相比，人HCC肿瘤组织中IRF -1 mRNA表达下调。与原代人肝细胞相比，HepG2肿瘤细胞中IFNγ诱导的IRF -1蛋白较少。miR -23a的表达与IRF -1呈负相关，添加miR -23a抑制剂可增加基础IRF -1 mRNA和蛋白质。同样，miR -23a模拟物下调IFNγ诱导的IRF -1蛋白表达，而miR -23a抑制剂则增加IRF -1。此外，miR -23a模拟物抑制IRF -1 - 3′UTR报告基因活性，而miR -23a抑制剂则增加报告基因活性。这些结果表明，与背景肝组织相比，人HCC肿瘤中IRF -1表达下调。miR -23a下调HCC细胞中IRF -1的表达，并且IRF -1的3′UTR具有一个miR -23a结合位点，该位点可结合miR -23a并降低报告基因活性。这些发现表明，miR -23a对IRF -1的靶向作用可能是HCC中IRF -1下调的分子基础，并为微小RNA对HCC的调控提供了新的见解。
Interferon regulatory factor-1 (IRF-1) is a tumor-suppressor gene induced by interferon-γ (IFNγ) and plays an important role in the cell death of hepatocellular carcinoma (HCC). HCC tumors evade death in part by downregulating IRF-1 expression, yet the molecular mechanisms accounting for IRF-1 suppression in HCC have not yet been characterized. Previous studies have shown that microRNA-23a (miR-23a) can suppress apoptosis by targeting IRF-1. Therefore, we hypothesized that miR-23a promotes HCC growth by down-regulating IRF-1. For the in vivo studies, 7 cases of resected HCC and adjacent liver samples were analyzed. For the in vitro studies, IRF-1 mRNA and protein were examined in HepG2 and Huh-7 HCC cells after IFNγ stimulation by real-time PCR and western blotting, respectively. To determine the role of miR-23a in regulating IRF-1, HepG2 cells were transfected with an miR-23a mimic or inhibitor, and IRF-1 expression was examined. Binding of miR-23a was assessed by cloning the 528-bp human IRF-1 3′-untranslated region (3′UTR) into luciferase reporter plasmid pMIR-IRF-1-3′UTR. The results showed that IRF-1 mRNA expression was down-regulated in the human HCC tumor tissues compared to that in the adjacent background liver tissues. IFNγ-induced IRF-1 protein was less in the HepG2 tumor cells compared to that in the primary human hepatocytes. miR-23a expression was inversely correlated with IRF-1, and addition of the miR-23a inhibitor increased basal IRF-1 mRNA and protein. Likewise, the miR-23a mimic downregulated IFNγ-induced IRF-1 protein expression, while the miR-23a inhibitor increased IRF-1. Furthermore, the miR-23a mimic repressed IRF-1-3′UTR reporter activity, while the miR-23a inhibitor increased the reporter activity. These results demonstrated that IRF-1 expression is downregulated in human HCC tumors compared to that noted in the background liver. miR-23a downregulates the expression of IRF-1 in HCC cells, and the IRF-1 3′UTR has an miR-23a binding site that binds miR-23a and decreases reporter activity. These findings suggest that the targeting of IRF-1 by miR-23a may be the molecular basis for IRF-1 downregulation in HCC and provide new insight into the regulation of HCC by miRNAs.