NDV感染的HeLa细胞的外泌体microRNA的表达谱及生物信息学分析

Detection of microRNA Expression Profiles in Exosomes Derived from HeLa Cells and Bioinformatics Analysis

分析经NDV感染HeLa细胞分泌外泌体microRNA表达变化,以期为外泌体microRNA在NDV感染中的具体作用机制提供初步的理论基础。采用经典差速离心法分离提取对照组与NDV感染组的HeLa细胞来源外泌体,经miRCURYTM Array芯片分析两组microRNA的表达谱后,通过Targetscan预测表达差异miRNA的可能靶基因,并利用DAVID、KEGG等在线预测工具对差异microRNA靶基因进行Go功能分析。相比于对照组,NDV感染组HeLa细胞外泌体中共有234个microRNA发生了2倍以上的表达差异(P<0.01),其中8个外泌体microRNA呈现出10倍以上的表达差异(P<0.01),分别为hsa-miR-3662、hsa-miR-3128、hsa-miR-345-5p、hsa-miR-376a-3p、hsa-miR-106b-3p、hsa-miR-133b、hsa-miR-410-3p和hsa-miR-454-3p。经Go功能分析发现,这8个microRNA主要分布在细胞外膜、高尔基膜、细胞核膜等部位,参与转录过程,负调节RNA聚合酶Ⅱ启动子转录过程,蛋白质均聚过程,正调节NF-kappaB转录因子活性过程、细胞缺氧反应过程、神经元分化过程等,并发挥着蛋白结合、金属离子结合、转录因子活性、染色体结合等功能。NDV感染会引发HeLa细胞的外泌体microRNA显著性差异表达,而其机制仍有待进一步研究。

The changes of microRNA expression in exosomes of HeLa cells infected with NDV (Newcastle disease virus) were analyzed in the present study in order to provide a preliminary theoretical basis for the specific mechanism of exosome microRNA during NDV infection.HeLa cell-derived exosomes were isolated by classical differential centrifugation.The expression profiles of two groups of microRNAs were analyzed by miRCURYTM Array.The possible target genes of differentially expressed microRNAs were predicted by Targetscan.The Go function of differentially expressed microRNAs was analyzed by online prediction tools such as DAVID and KEGG.Compared with the control group,234 microRNAs in HeLa cell exosomes of NDV infected group had more than twice expression difference (P<0.01).Eight of them showed more than 10 times expression difference (P<0.01).They were hsa-miR-3662,hsa-miR-3128,hsa-miR-345-5p,hsa-miR-376a-3p,hsa-miR-106b-3p,hsa-miR-133b,hsa-miR-410-3p,and hsa-miR-454-3p,respectively.The GO functional analysis showed that the eight mcroRNAs were mainly distributed in the outer membranes,Golgi membranes and nuclear membranes.They participated in the transcription,negative regulation of transcription from RNA polymerase II promoter,protein homooligomerization,positive regulation of NF-kappaB transcription factor activity,cellular response to hypoxia and neuronal differentiation,and played the roles in protein binding,metal ion binding,transcription factor activity,chromosome binding and so on.This study suggested that NDV infection could cause significant differences in the expression of microRNA in exosomes of HeLa cells and the underlying mechanism remains to be further investigated.