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miRNA-486-5p对胃癌细胞SGC7901中NRP2表达的影响 被引量：1

Effect of miRNA-486-5p on expression of NRP2 in gastric carcinoma SGC7901 cells
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摘要 目的:预测并鉴定miRNA-486-5p在人胃癌细胞SGC7901中的靶基因及其表达。方法:采用生物信息学技术预测miRNA-486-5p的作用靶点,构建miRNA-486-5p过表达质粒(GV214-miR)并转染入SGC7901细胞(SGC7901-miR)中,以空质粒转染SGC7901细胞(SGC7901-miR-NC)为阴性对照,以SGC7901细胞为空白对照。Real-time PCR检测转染细胞中miRNA-486-5p及其靶基因神经纤毛蛋白2(neuropilin-2,NRP2)mRNA的表达,Western blotting检测NRP2的表达,双荧光素酶实验验证miRNA-486-5p对NRP2基因的调控机制。结果:经生物信息学预测,选择与胃癌生物学行为密切相关的NRP2作为miRNA-486-5p的靶基因。与空白组相比,GV214-miR转染后的SGC7901细胞miRNA-486-5p表达显著上调[(8.21±1.18)vs(1.02+0.26),P<0.01],NRP2 mRNA表达无明显变化(P>0.05),而NRP2蛋白表达则明显下调[(0.36±0.06)vs(0.76±0.05),P<0.05],双荧光素酶实验证实miRNA-486-5p可与NRP2 mRNA 3'-UTR直接结合,从而发挥对NRP2转录后翻译的抑制作用。结论:miRNA-486-5p在胃癌细胞SGC7901中可直接作用于NRP2 mRNA 3'UTR,从而抑制其表达。 Objective： To predict and identify the target genes of microRNA-486-5p（ miRNA-486-5p） in human gastric cancer SGC7901 cells. Methods： Possible target genes of miRNA-486-5p were predicted by bioinformatics techniques and accordingly miRNA-486-5p over-expressing plasmid（ GV214-miR） against the identified target gene,neuropilin-2（ NRP-2） was constructed. SGC7901 cells were transfected with a control miRNA and an NRP-2-specific miRNA-486-5p. In the transfectants and non-transfected control cells,miRNA-486-5p and NRP-2 mRNA levels and NRP-2 protein levels were analyzed by real-time PCR and Western blotting respectively,and the NRP-2 promoter activity was evaluated by a dual luciferase reporter assay. Results： The expression of miRNA-486-5p in miRNA-486-5p-transfected SGC7901 cells（ SGC7901-miR cells） was significantly up-regulated compared with that in the control group（ 8. 21 ± 1. 18 vs 1. 02 ± 0. 26,P 〈 0. 01）. No significant difference in NRP2 mRNA abundance was observed（ P 〉 0. 05）. However,the NRP2 protein level was significantly reduced in SGC7901-miR cells（ 0. 36 ± 0. 06） as compared with SGC7901 cells transfected with the control plasmid（ 0. 76 ± 0. 05,P 〈 0. 05）. Dual luciferase reporter assay demonstrated that miRNA-486-5p directly targeted the 3 ’-untranslated region（ UTR） of the NRP2 gene,resulting in inhibition of the post-transcriptional translation of NRP2. Conclusion： Sequence-specific miRNA-486-5p may suppress the expression of NRP2 at the protein level in human gastric cancer cells by binding to NRP2 mRNA 3’UTR directly.
作者 连海峰 李明 刘成霞 李锟 李丹
机构地区 滨州医学院附属医院消化内科 滨州医学院护理学院内科护理学教研室
出处 《中国肿瘤生物治疗杂志》 CAS CSCD 北大核心 2014年第3期298-302,共5页 Chinese Journal of Cancer Biotherapy
基金 山东省高校科技计划基金资助项目(No.J11LF75) 滨州医学院科研启动基金资助项目(No.BY2010KYQD02)~~
关键词 miRNA-486-5p 胃癌 生物信息学技术 神经纤毛蛋白2 转染 miR-486-5p gastric carcinoma bioinformatics technique neuropilin-2（NRP2） transfect
分类号 R735.2 [医药卫生—肿瘤] R730.54 [医药卫生—肿瘤]
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参考文献20

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同被引文献14

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中国肿瘤生物治疗杂志
2014年 第3期

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