Bta-miR-677靶向线粒体抗病毒信号蛋白影响Ⅰ型干扰素的生成

doi:10.11843/j.issn.0366-6964.2020.01.017

畜牧兽医学报 ›› 2020, Vol. 51 ›› Issue (1): 150-158.doi: 10.11843/j.issn.0366-6964.2020.01.017

Bta-miR-677靶向线粒体抗病毒信号蛋白影响Ⅰ型干扰素的生成

廖政^1,2, 嵇辛勤^1*, 李基棕^2*, 肖芳^1,2, 刘茂军², 毛立², 李文良², 孙敏²

1. 贵州大学动物科学学院, 贵阳 550025;
2. 江苏省农业科学院兽医研究所, 农业部兽用生物制品工程技术重点实验室, 南京 210014

收稿日期:2019-08-21 出版日期:2020-01-23 发布日期:2020-01-17
通讯作者: 嵇辛勤,主要从事动物传染病防治和诊断技术研究,E-mail:jxq972@aliyun.com;李基棕,主要从事动物传染病防治和诊断技术研究,Tel:025-84391152,E-mail:lijizong22@sina.com
作者简介:廖政(1993-),男,贵州黔东南苗族侗族自治州人,硕士,主要从事动物传染病防治和诊断技术研究,Tel:025-84391152,E-mail:320414112@qq.com
基金资助:
国家自然科学基金（31702272；31802196）；江苏省自然科学基金（BK2017059）

Bta-miR-677 Up-regulated the Expression of Type Ⅰ Interferon by Targeting Mitochondria Antiviral Signaling Protein

LIAO Zheng^1,2, JI Xinqin^1*, LI Jizong^2*, XIAO Fang^1,2, LIU Maojun², MAO Li², LI Wenliang², SUN Min²

1. College of Animal Science of Guizhou University, Guiyang 550025, China;
2. Key Laboratory of Veterinary Biological Engineering and Technology of the Ministry of Agriculture and Rural Affairs(MARA) of China, Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China

Received:2019-08-21 Online:2020-01-23 Published:2020-01-17

摘要/Abstract

摘要： 旨在探究bta-miR-677调控Ⅰ型干扰素（IFN）表达的分子机制。将bta-miR-677转染MDBK细胞，检测Ⅰ型IFN和干扰素刺激基因（ISGs）的转录水平；随后通过TargetScan预测bta-miR-677的靶基因，双荧光素酶报告基因系统、qRT-PCR和Western blot验证bta-miR-677的靶基因；利用siRNA敲减验证靶基因对Ⅰ型IFN转录的影响。结果显示，过表达bta-miR-677组IFN-α/β的转录水平高于对照组2~4倍（P<0.001），随后检测到6种ISGs（IFI6、OAS1Y、OAS1Z、RSAD2、MX1和MX2）转录量上调2~16倍（P<0.01或P<0.001）；反之，抑制表达bta-miR-677组IFN-α/β转录量下调（P<0.01或P<0.05），同时IFI6、OAS1Y、OAS1Z、RSAD2、MX1和MX2转录量下调（P<0.05或P<0.01）。经TargetScan预测和双荧光素酶报告基因系统、qRT-PCR和Western blot检测显示，bta-miR-677可靶向结合线粒体抗病毒信号蛋白（MAVS）的3'-UTR，抑制MAVS蛋白的表达；MAVS基因敲减后发现，IFN-α、IFN-β和6种ISGs转录量上调。研究结果表明，bta-miR-677通过靶向MAVS上调IFN-α/β转录水平，进而提高ISGs的表达量，为基于bta-miR-677研制抗病毒药物提供了重要资料。

关键词: bta-miR-677, Ⅰ型干扰素, 干扰素刺激基因, 线粒体抗病毒信号蛋白

Abstract: This study was conducted to investigate the molecular mechanism of bta-miR-677 regulating the expression of type Ⅰ interferon (IFN). Bta-miR-677 was transfected into MDBK cells to detect the transcription level of type Ⅰ IFN and interferon stimulating genes (ISGs). Then we predicted the target gene of bta-miR-677 by TargetScan, and the target gene of bta-miR-677 was verified by dual-luciferase report gene system, qRT-PCR and Western blot assay. The effect of the target gene on IFN transcription was verified by siRNA knockdown. The results showed that the transcription of IFN-α/β in bta-miR-677 group were 2-4 times higher than that of the control group (P<0.001), moreover, the transcription of six ISGs(IFI6, OAS1Y, OAS1Z, RSAD2, MX1 and MX2) were up-regulated by 2-16 times (P<0.01 or P<0.001). By contrast, the transcription of IFN-α/β in bta-miR-677 inhibitor group were down-regulated (P<0.01 or P<0.05), then the transcription of IFI6, OAS1Y, OAS1Z, RSAD2, MX1 and MX2 were down-regulated (P<0.05 or P<0.01). Subsequently, we demonstrated that bta-miR-677 could target the 3'-UTR of MAVS (mitochondria antiviral signaling protein) and the expression of MAVS is inhibited. Meanwhile, silencing of MAVS up-regulated the expression of IFN-α/β and the six ISGs. The results showed that bta-miR-677 up-regulated the transcription level of IFN-α/β by targeting MAVS, and then increased the transcription of ISGs. This study reveals that bta-miR-677 up-regulated type Ⅰ IFN by targeting MAVS to increase the expression of ISGs, which provided important data for the development of antiviral drugs based on bta-miR-677.

Key words: bta-miR-677, typeⅠ interferon, interferon stimulating genes, mitochondria antiviral signaling protein

中图分类号:

S852.659.5

廖政, 嵇辛勤, 李基棕, 肖芳, 刘茂军, 毛立, 李文良, 孙敏. Bta-miR-677靶向线粒体抗病毒信号蛋白影响Ⅰ型干扰素的生成[J]. 畜牧兽医学报, 2020, 51(1): 150-158.

LIAO Zheng, JI Xinqin, LI Jizong, XIAO Fang, LIU Maojun, MAO Li, LI Wenliang, SUN Min. Bta-miR-677 Up-regulated the Expression of Type Ⅰ Interferon by Targeting Mitochondria Antiviral Signaling Protein[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(1): 150-158.

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Bta-miR-677靶向线粒体抗病毒信号蛋白影响Ⅰ型干扰素的生成

Bta-miR-677 Up-regulated the Expression of Type Ⅰ Interferon by Targeting Mitochondria Antiviral Signaling Protein

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