IFN-α对山羊副流感病毒3型的抗病毒活性

doi:10.11843/j.issn.0366-6964.2023.02.030

畜牧兽医学报 ›› 2023, Vol. 54 ›› Issue (2): 736-743.doi: 10.11843/j.issn.0366-6964.2023.02.030

IFN-α对山羊副流感病毒3型的抗病毒活性

孙敏^1,2*, 郝飞¹, 张纹纹¹, 李文良^1,3,4, 杨蕾蕾¹, 毛立^1,2, 程子龙¹, 刘茂军^1,3,4

1. 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;
2. 江苏大学生命科学学院, 镇江 212013;
3. 江苏大学食品与生物工程学院, 镇江 212013;
4. 南京农业大学动物医学院, 南京 210095

收稿日期:2022-01-12 出版日期:2023-02-23 发布日期:2023-02-21
通讯作者: 孙敏,主要从事动物传染病防控研究,E-mail:sunmin9084@aliyun.com
作者简介:孙敏(1990-),女,安徽舒城人,副研究员,博士,主要从事动物传染病防控研究,E-mail:sunmin9084@aliyun.com
基金资助:
江苏省自然科学基金（BK20190272）；国家自然科学基金（31902269）

The Antiviral Activity of Goat Interferon Alpha to Caprine Parainfluenza Virus 3

SUN Min^1,2*, HAO Fei¹, ZHANG Wenwen¹, LI Wenliang^1,3,4, YANG Leilei¹, MAO Li^1,2, CHENG Zilong¹, LIU Maojun^1,3,4

1. Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences/Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing 210014, China;
2. School of Life Sciences, Jiangsu University, Zhenjiang 212013, China;
3. School of Food and Biological Engineering, Jiangsu University, Zhenjiang 212013, China;
4. College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China

Received:2022-01-12 Online:2023-02-23 Published:2023-02-21

摘要/Abstract

摘要： 旨在探讨原核表达纯化的山羊α干扰素（IFN-α）对山羊副流感病毒3型（CPIV3）的抗病毒活性。通过分析山羊IFN-α的序列特点，比对不同种属IFN-α的同源性，进而构建山羊IFN-α成熟蛋白编码基因（去除信号肽基因序列）的原核表达载体pET-30a-gIFN-α，将其转化至感受态细胞Rosetta （DE3），IPTG诱导后镍柱及亲和纯化获得山羊IFN-α。利用RT-qPCR测定山羊IFN-α作用于牛肾细胞（Madin-Darby bovine kidney cell，MDBK cell）后6种干扰素刺激基因（interferon-stimulated genes，ISGs）的相对表达水平；同时，利用TCID₅₀及Western blot测定其对CPIV3的抗病毒活性。结果表明，原核表达的山羊IFN-α蛋白含量为0.20 mg·mL^-1，Western blot结果表明表达产物相对分子质量约为20 ku，与预期结果相符。RT-qPCR结果显示，山羊IFN-α孵育MDBK细胞后，可显著刺激RSAD2、STAT1及ISG15等6种ISGs基因的转录上调。与对照组细胞相比，山羊IFN-α显著下调MDBK细胞中CPIV3的病毒滴度，Western blot结果显示，山羊IFN-α孵育显著下调CPIV3非结构蛋白C的表达水平。山羊IFN-α对CPIV3在MDBK细胞中的增殖具有显著抑制作用。本研究扩展了I型干扰素的研究范围，为CPIV3等反刍动物疫病的防控提供新的思路。

关键词: 山羊IFN-α, 原核表达, ISGs, 山羊副流感病毒3型, 抗病毒活性

Abstract: The aim of this study was to investigate the antiviral activity of goat interferon α (IFN-α) to caprine parainfluenza virus 3 (CPIV3). The sequence characteristics of goat IFN-α was analyzed, and the homology of IFN-α from different species was compared. Then the recombinant expression plasmid pET-30a-gIFN-α by inserting goat IFN-α gene (removing the signal peptide gene sequence) into vector pET-30a was constructed and transformed into Rosetta (DE3), after induced by IPTG, the goat IFN-α was expressed and purified. Then the relative expression level of interferon-stimulated genes (ISGs) was measured, and the anti-CPIV3 activity was measured after exogenous goat IFN-α treatment on Madin-Darby bovine kidney cell(MDBK cell) by TCID₅₀ and Western blot. The results showed that, the protein concentration of goat IFN-α was 0.20 mg·mL^-1, and Western blot analysis showed that the relative molecular weight was about 20 ku, which was consistent with expectations. The RT-qPCR result showed that the incubation of goat IFN-α on MDBK cells induced the up-regulation of the six ISGs including RSAD2, STAT1 and ISG15. Meanwhile, the goat IFN-α inhibited the CPIV3 replication, and down-regulated the expression level of the viral nonstructural protein C. All above results indicated that goat IFN-α could significantly inhibit the proliferation of CPIV3 on MDBK cells. This study extended the research type I interferon, and promoted the prevention and control of ruminant diseases.

Key words: goat IFN-α, prokaryotic expression, ISGs, caprine parainfluenza virus 3, antiviral activity

中图分类号:

S858.27

孙敏, 郝飞, 张纹纹, 李文良, 杨蕾蕾, 毛立, 程子龙, 刘茂军. IFN-α对山羊副流感病毒3型的抗病毒活性[J]. 畜牧兽医学报, 2023, 54(2): 736-743.

SUN Min, HAO Fei, ZHANG Wenwen, LI Wenliang, YANG Leilei, MAO Li, CHENG Zilong, LIU Maojun. The Antiviral Activity of Goat Interferon Alpha to Caprine Parainfluenza Virus 3[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(2): 736-743.

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IFN-α对山羊副流感病毒3型的抗病毒活性

The Antiviral Activity of Goat Interferon Alpha to Caprine Parainfluenza Virus 3

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