| 草鱼miR-462通过靶向cx32.2、slc9a3.1和tbk1调控嗜水气单胞菌感染诱导的免疫应答 |
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| 引用本文： | 王安琪,陶丽竹,周丰林,徐晓雁,沈玉帮,李家乐.草鱼miR-462通过靶向cx32.2、slc9a3.1和tbk1调控嗜水气单胞菌感染诱导的免疫应答[J].水产学报,2020,44(9):1467-1476. |
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| 作者姓名： | 王安琪 陶丽竹 周丰林 徐晓雁 沈玉帮 李家乐 |
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| 作者单位： | 上海海洋大学,农业农村部淡水水产种质资源重点实验室,上海 201306;上海海洋大学,水产科学国家级实验教学示范中心,上海 201306;上海海洋大学,农业农村部淡水水产种质资源重点实验室,上海 201306;上海海洋大学,水产科学国家级实验教学示范中心,上海 201306;上海海洋大学,水产养殖工程技术研究中心,上海 201306 |
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| 基金项目： | 现代农业产业技术体系专项(CARS-45-03)；上海市工程中心提升项目(19DZ2284300) |
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| 摘 要： | 为探究miR-462在嗜水气单胞菌感染草鱼肾脏细胞(Ctenopharyngodon idella kideny,CIK)后的调控机制,实验利用荧光定量技术检测了CIK细胞感染嗜水气单胞菌后miR-462表达水平的变化;运用RNAhybrid软件预测miR-462的靶基因,利用双荧光素酶报告基因系统进行确定;此外还分析了miR-462对靶基因下游基因的调控作用。结果显示,在CIK细胞感染嗜水气单胞菌的过程中,miR-462的表达发生显著变化;cx32.2、slc9a3.1和tbk1的表达先降低后升高,与miR-462的表达模式呈负相关。双荧光素酶报告系统显示,miR-462可靶向cx32.2、slc9a3.1和tbk1的3′非编码区抑制其表达,过表达miR-462可以显著抑制cx32.2、slc9a3.1和tbk1的表达。转染miR-462模拟物后,下游slc4a4a、tnfrsf5、cxcl9和cxcl11基因的表达受到抑制。研究表明,miR-462参与调控嗜水气单胞菌感染后草鱼CIK细胞中的免疫应答。cx32.2、slc9a3.1和tbk1被鉴定为miR-462的靶基因。miR-462可通过靶向slc9a3.1和tbk1影响下游基因的功能。
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| 关 键 词： | 草鱼肾脏细胞 嗜水气单胞菌 miR-462 免疫调节 |
| 收稿时间： | 2020/2/18 0:00:00 |
| 修稿时间： | 2020/4/13 0:00:00 |
miR-462 modulates cellular immune response by targeting cx32.2, slc9a3.1 and tbk1 in CIK cells infected with Aeromonas hydrophila |
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| WANG Anqi,TAO Lizhu,ZHOU Fenglin,XU Xiaoyan,SHEN Yubang,LI Jiale.miR-462 modulates cellular immune response by targeting cx32.2, slc9a3.1 and tbk1 in CIK cells infected with Aeromonas hydrophila[J].Journal of Fisheries of China,2020,44(9):1467-1476. |
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| Authors: | WANG Anqi TAO Lizhu ZHOU Fenglin XU Xiaoyan SHEN Yubang LI Jiale |
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| Institution: | Key Laboratory of Freshwater Aquatic Genetic Resources, Ministry of Agriculture and Rural Affairs, Shanghai Ocean University, Shanghai 201306, China;National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306, China; Key Laboratory of Freshwater Aquatic Genetic Resources, Ministry of Agriculture and Rural Affairs, Shanghai Ocean University, Shanghai 201306, China;National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306, China;Shanghai Engineering Research Center of Aquaculture, Shanghai Ocean University, Shanghai 201306, China |
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| Abstract: | Bacterial septicemia is a systemic inflammatory reaction mainly caused by the infection of Aeromonas hydrophila. Excessive development of inflammation may lead to septic shock or death in fish. A large number of studies have confirmed that miRNA is involved in the regulation of immune response after bacterial infection. To explore the regulatory mechanism of miR-462 in Ctenopharyngodon idella kidney (CIK) cells infected with A. hydrophila, the expression profiles of miR-462 upon A. hydrophila infection was detected by real-time quantitative PCR; the target genes of miR-462 were predicted by RNAhybrid software, and identified by dual-luciferase reporter assay system; in addition, the regulatory effect of miR-462 on downstream genes was analyzed. The results showed that the expression of miR-462 changed significantly after A. hydrophila infection, indicating that miR-462 participated in the regulation of immune response. Dual-luciferase reporter assay revealed that cx32.2, slc9a3.1 and tbk1 are the target genes of miR-462, which is further confirmed by the overexpression and inhibition experiments of miR-462. The expression of slc4a4a, tnfrs5, cxcl9 and cxcl11 were suppressed after miR-462 antagomir was transfected, which proved that miR-462 could affect the function of the downstream genes by targeting slc9a3.1 and tbk1. Our results may provide a theoretical basis for investigating the molecular mechanism of miR-462 regulating immune response in C. idella. |
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| Keywords: | Ctenopharyngodon idella kidney(CIK) Aeromonas hydrophila miR-462 immunomodulation |
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