miR-216a通过靶向调控蛋白激酶Cα抑制胶质瘤细胞增殖和侵袭

miR-216a suppresses cell proliferation and invasion by targeting PRKCA in glioma

背景与目的：MicroRNAs是一类19～25bp内源非编码的小分子RNA，其通过靶向抑制基因的转录和翻译水平。本研究旨在明确miR-216a是否通过靶向调控蛋白激酶Ca（PRKCA）表达而抑制胶质瘤细胞增殖和侵袭能力，从而进一步揭示miR-216a的抑瘤分子机制。方法：首先构建PRKCA3’UTR-荧光素酶报告载体，通过双荧光素酶报告检测观察miR-216a对PRKCA3’UTR-荧光素酶活性的影响；将miR-216amimics转染胶质瘤细胞U251，采用Westernblot检测PRKCA蛋白表达水平；将PRKCAsiRNA转染U251细胞，通过MTS细胞增殖活性检测和transwell侵袭实验观察PRKCA下调对U251细胞增殖和侵袭的影响。结果：双荧光素酶报告检测显示，miR-216a能特异性地与PRKCAmRNA的3’UTR结合，抑制其荧光素酶活性，下调41％。过表达miR-216a的U251细胞PRKCA蛋白表达水平降低。siRNA干扰PRKCA表达能抑制U251细胞的增殖和侵袭，它能部分模拟miR-216a的抑瘤功能。结论：miR-216a通过靶向PRKCAmRNA3’UTR而抑制胶质瘤细胞增殖和侵袭。

Background and purpose： MicroRNAs are 19-25-nucleotide noncoding RNA molecules that regulate gene expression at the level of transcription and translation. The study aimed to confirm whether miR-216a suppresses cell proliferation and invasion by targeting PRKCA, thus to reveal molecular mechanism that miR-216a functions as a tumor suppressor in glioma. Methods： PRKCA 3＇ untranslated region （UTR）-luciferase vector was constructed and duaMuciferase reporter gene assay was employed to examine the effect of miR-216a on luciferase activity. U251 cells were transfected with miR-216a mimics, and next Western blotting was performed to detect the expression of PRKCA protein. The effects of PRKCA downregulation on cell proliferation and invasion were observed after PRKCA siRNA was transfected into U251 cells. U251 cell proliferation assays were performed when cotransfected with miR-216a mimics. Results： The result demonstrated miR-216a could bind to the 3＇UTR of PRKCA and inhibited the luciferase activity by 41%. PRKCA protein expressions were significantly down-regulated when miR-216a was overexpressed in U251. siRNA-mediated downregulation of PRKCA could suppress the potentials of cell proliferation and invasion. Conclusion： miR-216a suppresses cell proliferation and invasion by targeting PRKCA in glioma.