文章摘要
草鱼Mst基因在免疫应答中的作用机制研究
Study on the Mechanism of MST Gene in Immune Response of Grass Carp (Ctenopharyngodon idellus)
投稿时间:2021-03-14  修订日期:2021-03-30
DOI:
中文关键词: 草鱼  Mst  CIK细胞  RNA-Seq  细胞凋亡
英文关键词: Grass carp  Mst  CIK cells  RNA-Seq  Cell apoptosis
基金项目:
作者单位邮编
李亚男 仲恺农业工程学院 501225
唐美珍 仲恺农业工程学院 
卢志杰 仲恺农业工程学院 
林蠡 仲恺农业工程学院 510225
秦真东 仲恺农业工程学院 
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中文摘要:
      为了初步阐明草鱼Mst基因在机体免疫中的作用机制,本文采用RNA-Seq技术对干扰了丝/苏氨酸蛋白激酶(Mammalian Sterile Twenty-like,Mst)基因后经脂多糖(lipopolysaccharide,LPS)应激的草鱼CIK细胞进行了转录组测序与验证分析。测序原始数据经de novo 拼接与组装后共获得22,374 unigenes,其中已知的基因为 21,199 个,预测的新基因为 1,175 个。干扰Mst基因后经LPS应激的unigenes 表达差异分析表明,对照组与试验组之间共存在38个差异基因(Differently Expressed Genes,DEGs),其中上调基因16个,下调基因22个。利用qPCR 对38个DEGs的RNA-Seq 结果进行了验证,结果显示,qPCR 和RNA-Seq 分析一致,说明RNA-Seq 分析结果可靠。干扰Mst基因经LPS处理后,CIK细胞转录组中差异表达基因参与免疫代谢的途径主要有MAPK信号通路(MAPK signaling pathway)、内吞作用途径(Endocytosis pathway)、自噬途径(Autophagy pathway)和细胞因子受体相互作用途径(Cytokine-cytokine receptor interaction pathway)。凋亡相关基因检测结果显示,干扰Mst基因经LPS处理后,促凋亡基因(Fas、Bad1、Bad2、caspase-3、caspase-8和caspase-9)转录水平上调,抗凋亡基因(Bcl2和Mcl1)与之相反;证明干扰Mst基因经LPS处理后会诱发细胞发生凋亡。综上所述,本研究结果初步阐明了草鱼Mst基因在参与机体免疫应答过程中的分子机理,而且为草鱼细菌性疾病的防控提供了一定的基础理论参考。
英文摘要:
      In order to elucidate the mechanism of grass carp Mst gene in immunity, RNA-Seq technique was used to analyze the transcriptome sequence of grass carp CIK cells stressed by LPS, after interfered with Mst (Mammalian Sterile Twenty-like) gene using siRNA.?Firstly, a total of 22374 unigenes were obtained from the original sequence data by de novo assembly, of which 21199 genes were known and 1175 new genes were predicted.?Secondly, after interfered with MST gene, the analysis of unigenes expression under LPS stress showed that there were 38 differential genes (Differently Expressed Genes, DEGs) between the control group and the experimental group, including 16 up-regulated genes and 22 down-regulated genes.?Thirdly, the RNA-Seq results of 38 DEGs were verified by qPCR. The results showed that the qPCR analysis was consistent with the RNA-Seq analysis, indicating that the RNA-Seq analysis was reliable.?Moreover, after interfered with Mst gene and then treated by LPS, the DEGs in CIK cells transcript group were mainly involved in immune metabolism pathway, containing MAPK signal pathway (MAPK signaling pathway), endocytosis pathway (Endocytosis pathway), autophagy pathway (Autophagy pathway) and cytokine receptor interaction pathway (Cytokine-cytokine receptor interaction pathway).?Interestingly, the detection of apoptosis-related genes showed that the transcriptional levels of pro-apoptotic genes (Fas, Bad1, Bad2, caspase3, caspase8 and caspase9) were up-regulated after interfering with Mst gene and treatment by LPS, while the anti-apoptotic genes (Bcl2 and Mcl1) were down-regulated. Therefore, it proved that interfering Mst gene could induce cell apoptosis after LPS treatment.?To sum up, the results of this study preliminarily clarify the molecular mechanism of grass carp Mst gene in the process of immune response, and provide some basic theoretical reference for the prevention and control of grass carp bacterial diseases.
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