捻转血矛线虫耐药虫株与敏感虫株circRNAs差异表达分析

Differential expression of circRNAs in resistant and sensitive strains of Haemonchus contortus

为分析捻转血矛线虫伊维菌素敏感虫株与耐药虫株之间差异表达的circRNAs在调控耐药性中的作用,本研究以捻转血矛线虫敏感虫株和耐药虫株为研究对象,采用Illumina Hiseq 2000平台进行高通量测序及cDNA文库的构建,利用相关生物信息学软件对测得的circRNAs进行差异性分析,同时对来源基因进行GO和KEGG pathway功能富集。运用TargetScan和Miranda软件预测关键circRNAs靶向的miRNAs,探究circRNAs-miRNAs的作用。随后又随机选取5个显著差异的circRNAs,进行RT-qPCR检测并与转录组测序结果进行一致性评定。结果显示:1)敏感虫株与耐药虫株共筛选出677个差异表达的circRNAs,其中47个显著差异,23个显著上调,24个显著下调。2)所有差异circRNAs的来源基因GO富集分析显示,共富集到生物过程336个、细胞组分59个和分子功能110个,主要与代谢过程、细胞膜结构和催化活性等相关,KEGG富集分析显示,富集到257条通路,主要与代谢(脂肪酸生物合成、甘油磷脂代谢和丙酸代谢)、胰高血糖素信号通路和AMPK信号通路等相关。3)circRNAs-miRNAs调控网络分析显示,627个差异circRNAs靶向结合187个miRNAs,组成了10602组调控关系。综上,本研究成功筛选了捻转血矛线虫伊维菌素敏感虫株和耐药虫株间差异表达的circRNAs,对所有差异circRNAs的来源基因进行功能注释和富集分析,并构建了部分显著差异circRNAs-miRNAs调控网络。本研究结果为捻转血矛线虫耐药性产生的分子机制研究提供了新思路。

The study was to investigate the role of differentially expressed circRNAs in the regulation of drug resistance between ivermectin sensitive and resistant strains of Haemonchus contortus.In this study,sensitive and resistant strains of H.contortus were taken as research objects.Illumina Hiseq 2000 platform was used for high-throughput sequencing and cDNA library construction,and relevant bioinformatics software was used to analyze the difference of circRNAs.Simultaneously,GO and KEGG pathway functions of the source genes were enriched.TargetScan and Miranda software were used to predict the miRNAs targeted by key circRNAs,and the role of circRNAs-miRNAs was explored.Subsequently,5 circRNAs with significant differences were randomly selected for RT-qPCR detection and consistency assessment with transcriptome sequencing result.The results showed that:1)A total of 677 differentially expressed circRNAs were screened between sensitive and resistant strains,of which 47 were significantly different,23 were significantly up-regulated,and 24 were significantly down-regulated.2)GO enrichment analysis of all differentially derived circRNAs showed that 336 biological processes,59 cell components and 110 molecular functions were enriched,which were mainly related to metabolic processes,cell membrane structure and catalytic activity.KEGG enrichment showed that 257 pathways were mainly enriched,most of which were related to metabolism(fatty acid biosynthesis,glycerophospholipid metabolism and propionate metabolism),glucagon signaling pathway and AMPK signaling pathway.3)circRNAs-miRNAs regulatory network analysis showed that 627 circRNAs target bound 187 miRNAs,forming 10602 regulatory relationships.In conclusion,this study successfully screened the differentially expressed circRNAs between ivermectin sensitive and resistant strains of H.contortus,functionally annotated and enriched the source genes of the all differentially expressed circRNAs,and constructed some significant differentially expressed circRNAs-miRNAs regulatory networks.The results of this study provids a new way to study the molecular mechanism of drug resistance in H.contortus.