Key Laboratory of Molecular Biophysics of the Ministry of Education, College of Life Science and Technology and Center for Human Genome Research, Huazhong University of Science and Technology, Wuhan, P. R. China.
Center for Cardiovascular Genetics, Department of Molecular Cardiology, Cleveland Clinic; Department of Molecular Medicine/CCLCM, Department of Genetics and Genome Sciences, Case Western Reserve University, Cleveland, OH 44195, USA.
Sci Rep. 2016 Oct 4;6:34034. doi: 10.1038/srep34034.
Hepatocellular carcinoma (HCC) is one of the most common malignant cancers. To elucidate new regulatory mechanisms for heptocarcinogenesis, we investigated the regulation of p21, a cyclin-dependent kinase (CDK) inhibitor encoded by CDKN1A, in HCC. The expression level of p21 is decreased with the progression of HCC. Luciferase assays with a luciferase-p21-3' UTR reporter and its serial deletions identified a 15-bp repressor element at the 3'-UTR of CDKN1A, which contains a binding site for miR-95-3p. Mutation of the binding site eliminated the regulatory effect of miR-95-3p on p21 expression. Posttranscriptional regulation of p21 expression by miR-95-3p is mainly on the protein level (suppression of translation). Overexpression of miR-95-3p in two different HCC cell lines, HepG2 and SMMC7721, significantly promoted cell proliferation, cell cycle progression and cell migration, whereas a miR-95-3p specific inhibitor decreased cell proliferation, cell cycle progression and cell migration. The effects of miR-95-3p on cellular functions were rescued by overexpression of p21. Overexpression of miR-95-3p promoted cell proliferation and tumor growth in HCC xenograft mouse models. Expression of miR-95-3p was significantly higher in HCC samples than in adjacent non-cancerous samples. These results demonstrate that miR-95-3p is a potential new marker for HCC and regulates hepatocarcinogenesis by directly targeting CDKN1A/p21 expression.
肝细胞癌（HCC）是最常见的恶性肿瘤之一。为了阐明肝癌发生的新调控机制，我们研究了细胞周期蛋白依赖性激酶（CDK）抑制剂 CDKN1A 编码的 p21 在 HCC 中的调控作用。p21 的表达水平随着 HCC 的进展而降低。带有荧光素酶-p21-3'UTR 报告基因及其系列缺失的荧光素酶检测，鉴定出 CDKN1A 3'-UTR 中的一个 15bp 抑制元件，该元件包含 miR-95-3p 的结合位点。结合位点的突变消除了 miR-95-3p 对 p21 表达的调控作用。miR-95-3p 对 p21 表达的转录后调控主要在蛋白质水平上（抑制翻译）。在两种不同的 HCC 细胞系 HepG2 和 SMMC7721 中过表达 miR-95-3p ，显著促进了细胞增殖、细胞周期进程和细胞迁移，而 miR-95-3p 的特异性抑制剂则降低了细胞增殖、细胞周期进程和细胞迁移。miR-95-3p 对细胞功能的影响可通过过表达 p21 得到挽救。miR-95-3p 的过表达促进了 HCC 异种移植小鼠模型中的细胞增殖和肿瘤生长。miR-95-3p 在 HCC 样本中的表达明显高于相邻非癌样本。这些结果表明，miR-95-3p 是 HCC 的一个潜在新标志物，通过直接靶向 CDKN1A/p21 表达来调节肝癌发生。
