miRNA在乌鳢水泡病毒感染月鳢细胞(SSN-1)中的表达变化及其调控作用  

Effects of miRNA on snakehead vesiculovirus(SHVV) infection in stripped snakehead(SSN-1) cells

作　　者：伊丽竹 徐镇[1] 林蠡[1,2] 涂加钢 YI Lizhu;XU Zhen;LIN Li;TU Jiagang(College of Fishery,Huazhong Agricultural University,Wuhan 430070,China;College of Animal Sciences and Technology,Zhongkai University of Agriculture and Engineering,Guangzhou 510225,China)

机构地区：[1]华中农业大学水产学院,湖北武汉430070 [2]仲恺农业工程学院动物科技学院,广东广州510225

出　　处：《水产学报》2019年第5期1288-1297,共10页Journal of Fisheries of China

基　　金：国家自然科学基金(31602195)~~

摘　　要：为了探究microRNA(miRNA)对乌鳢水泡病毒(snakehead vesiculovirus, SHVV)感染月鳢细胞(striped snakehead cell, SSN-1)的影响,本研究对SHVV感染的SSN-1细胞及未感染的细胞进行miRNA高通量测序。分别用U6、β-actin和5S rRNA作为内参基因研究SHVV感染对细胞内22个高丰度(transcripts per million, TPM≥1 000) miRNA表达水平的影响,结果显示,U6基因作为内参时,SHVV感染对β-actin没有显著影响,但是5S rRNA显著上调表达,说明U6和β-actin基因适合作为内参基因研究SHVV感染对miRNA表达水平的影响。此外,我们研究了14种miRNA对SHVV增殖的影响。结果发现,miR-27a-3p、miR-26a-5p、miR-30e-3p等11种miRNA显著促进SHVV增殖,miR-150和miR-216b显著抑制SHVV增殖。进一步研究发现高丰度的miR-100-5p在SHVV感染SSN-1细胞早期上调表达,晚期下调表达。过表达miR-100-5p可以显著抑制SHVV的增殖,而抑制表达miR-100-5p可以显著促进SHVV的增殖。本研究为开发抗SHVV的核酸药物提供了理论基础。In order to study the effects of miRNAs in stripped snakehead (SSN-1) cell line upon snakehead vesic- ulovirus (SHVV) infection, SHVV-infected and mock-infected SSN-1 cells were analyzed by miRNA high- throughput sequencing. The expression levels of twenty-two miRNAs with levels equal to or higher than 1 000 reads per million (TPM≥1 000) were determined by qRT-PCR with U6,β-actin, and 5S rRNA as internal control. When U6 gene was used as internal control, no significant changes were observed for β-actin upon SHVV infection, while 5S rRNA was upregulated in SSN-1 cells upon SHVV infection. The results showed that U6 gene or β-actin could be used as internal control genes for miRNAs in SSN-1 cells. Furthermore, fourteen miRNAs were selected and their effects on SHVV replication were analyzed. The results showed that eleven miRNAs (miR- 27a-3p, miR-26a-5p, miR-30e-3p, etc.) promoted, whereas miR-150 and miR-216b suppressed, SHVV replication. Next, we found that high-expression miR-100-5p was upregulated at early stage of SHVV infection, while downregulated at later stage. Overexpression of miR-100-5p exhibited significant suppression of SHVV repli- cation, while inhibition of expression of miR-100-5p promoted SHVV replication. Our findings presented valuable information for the study of molecular drugs against SHVV.

关 键 词：乌鳢水泡病毒 SSN-1细胞 miR-100-5p 抗病毒 

分 类 号：S941.41[农业科学—水产养殖] Q786[农业科学—水产科学]