猪繁殖与呼吸综合征病毒N蛋白与核糖体蛋白S20相互作用对病毒复制的影响

doi:10.11843/j.issn.0366-6964.2023.01.027

畜牧兽医学报 ›› 2023, Vol. 54 ›› Issue (1): 293-303.doi: 10.11843/j.issn.0366-6964.2023.01.027

猪繁殖与呼吸综合征病毒N蛋白与核糖体蛋白S20相互作用对病毒复制的影响

王君君¹, 宋慧敏¹, 李倬伟¹, 姜平², 周双海¹, 李焕荣^1*, 刘雪威^1*

1. 北京农学院动物科学技术学院, 北京 100096;
2. 南京农业大学动物医学院, 南京 210014

收稿日期:2022-06-06 出版日期:2023-01-23 发布日期:2023-01-17
通讯作者: 刘雪威,主要从事猪病毒性疾病研究,E-mail:liuxuewei66@163.com;李焕荣,主要从事猪病毒性疾病研究,E-mail:lihuanrong1@126.com
作者简介:王君君(1997-),河南驻马店人,主要从事猪病毒性疾病研究,E-mail:1938817297@qq.com
基金资助:
北京农学院青年教师科研创新能力提升计划(QJKC2022016)；国家自然科学青年科学基金项目(32102663)

Effect of Interaction between N Protein and Ribosomal Protein S20 of Porcine Reproductive and Respiratory Syndrome Virus on Virus Replication

WANG Junjun¹, SONG Huimin¹, LI Zhuowei¹, JIANG Ping², ZHOU Shuanghai¹, LI Huanrong^1*, LIU Xuewei^1*

1. College of Animal Science and Technology Beijing University of Agriculture, Beijing 102206, China;
2. College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China

Received:2022-06-06 Online:2023-01-23 Published:2023-01-17

摘要/Abstract

摘要： 旨在筛选并鉴定与猪繁殖与呼吸综合征病毒(PRRSV)核衣壳蛋白(N蛋白)相互作用的宿主蛋白。本试验通过构建猪肺酵母cDNA文库、酵母双杂交、免疫共沉淀、激光共聚焦、过表达和干扰试验等手段，筛选与N蛋白互作的宿主蛋白并研究其对PRRSV复制的影响。结果显示：N蛋白和核糖体蛋白S20(RPS20)在酵母细胞内发生互作，免疫共沉淀和激光共聚焦试验进一步确定N蛋白和RPS20蛋白互作并共定位于细胞质中。过表达和干扰试验证实RPS20能够影响HP-PRRSV在细胞内的增殖。本研究丰富了PRRSV与宿主因子互作的网络，为进一步研究RPS20蛋白在PRRSV感染过程中的生物学功能奠定了基础。

关键词: 酵母双杂交, 猪繁殖与呼吸综合征病毒, N蛋白, 核糖体蛋白S20

Abstract: The present study aimed to screen and identify host proteins that interact with the nucleocapsid protein (N protein) of pathogenic porcine reproductive and respiratory syndrome virus (PRRSV). In this experiment, we screened host proteins that interact with N protein and investigated their effects on PRRSV replication by constructing a porcine lung yeast cDNA library, yeast two-hybrid, immunoprecipitation, laser confocal, overexpression and interference assays. The results showed that N protein and ribosomal protein S20 (RPS20) interacted with each other in yeast cells. The overexpression and interference assays also provided preliminary evidence that RPS20 can affect the proliferation of PRRSV in cells. This study enriches the network of PRRSV-host factor interactions and lays the foundation for further studies on the biological functions of RPS20 protein during PRRSV infection.

Key words: yeast two-hybrid, PRRSV, N protein, ribosomal protein S20

中图分类号:

S852.659.6

王君君, 宋慧敏, 李倬伟, 姜平, 周双海, 李焕荣, 刘雪威. 猪繁殖与呼吸综合征病毒N蛋白与核糖体蛋白S20相互作用对病毒复制的影响[J]. 畜牧兽医学报, 2023, 54(1): 293-303.

WANG Junjun, SONG Huimin, LI Zhuowei, JIANG Ping, ZHOU Shuanghai, LI Huanrong, LIU Xuewei. Effect of Interaction between N Protein and Ribosomal Protein S20 of Porcine Reproductive and Respiratory Syndrome Virus on Virus Replication[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(1): 293-303.

参考文献 26

[1]	DONE S, PATON D. Porcine reproductive and respiratory syndrome:clinical disease, pathology and immunosuppression[J]. Vet Rec, 1995, 136(2):32-35.
[2]	SCORTTI M, PRIETO C, ALVAREZ E, et al. Failure of an inactivated vaccine against porcine reproductive and respiratory syndrome to protect gilts against a heterologous challenge with PRRSV[J]. Vet Rec, 2007, 161(24):809-813.
[3]	HOLTKAMP D J, KLIEBENSTEIN J B, NEUMANN E J, et al. Assessment of the economic impact of porcine reproductive and respiratory syndrome virus on United States pork producers[J]. J Swine Health Prod, 2013, 21(2):72-84.
[4]	WOOTTON S K, YOO D. Homo-oligomerization of the porcine reproductive and respiratory syndrome virus nucleocapsid protein and the role of disulfide linkages[J]. J Virol, 2003, 77(8):4546-4557.
[5]	DOKLAND T. The structural biology of PRRSV[J]. Virus Res, 2010, 154(1-2):86-97.
[6]	LIN J S, LAI E M. Protein-protein interactions:yeast two-hybrid system[M]//JOURNET L, CASCALES E. Bacterial Protein Secretion Systems. New York:Humana Press, 2017:177-187.
[7]	PAIANO A, MARGIOTTA A, DE LUCA M, et al. Yeast two-hybrid assay to identify interacting proteins[J]. Curr Protoc Protein Sci, 2019, 95(1):e70.
[8]	LIVAK K J, SCHMITTGEN T D. Analysis of relative gene expression data using real-time quantitative PCR and the method[J]. Methods, 2001, 25(4):402-408.
[9]	ROSE N, RENSON P, ANDRAUD M, et al. Porcine reproductive and respiratory syndrome virus (PRRSv) modified-live vaccine reduces virus transmission in experimental conditions[J]. Vaccine, 2015, 33(21):2493-2499.
[10]	MUSIC N, GAGNON C A. The role of porcine reproductive and respiratory syndrome (PRRS) virus structural and non-structural proteins in virus pathogenesis[J]. Anim Health Res Rev, 2010, 11(2):135-163.
[11]	YOO D, WOOTTON S K, LI G, et al. Colocalization and interaction of the porcine arterivirus nucleocapsid protein with the small nucleolar RNA-associated protein fibrillarin[J]. J Virol, 2003, 77(22):12173-12183.
[12]	LIU L, LEAR Z, HUGHES D J, et al. Resolution of the cellular proteome of the nucleocapsid protein from a highly pathogenic isolate of porcine reproductive and respiratory syndrome virus identifies PARP-1 as a cellular target whose interaction is critical for virus biology[J]. Vet Microbiol, 2015, 176(1-2):109-119.
[13]	SONG Z B, BAI J, LIU X W, et al. S100A9 regulates porcine reproductive and respiratory syndrome virus replication by interacting with the viral nucleocapsid protein[J]. Vet Microbiol, 2019, 239:108498.
[14]	ZHAO K, LI L W, JIANG Y F, et al. Nucleocapsid protein of porcine reproductive and respiratory syndrome virus antagonizes the antiviral activity of TRIM25 by interfering with TRIM25-mediated RIG-I ubiquitination[J]. Vet Microbiol, 2019, 233:140-146.
[15]	ZHANG L X, LI R, GENG R, et al. Tumor susceptibility gene 101(TSG101) contributes to virion formation of porcine reproductive and respiratory syndrome virus via interaction with the nucleocapsid (N) protein along with the early secretory pathway[J]. J Virol, 2022, 96(6):e0000522.
[16]	王玉娥, 杨汉春. 酵母双杂交技术在病毒学研究中的应用[J]. 中国兽医学报, 2004, 24(3):307-309.WANG Y E, YANG H C. Applicating yeast two hybrid system in research of virology[J]. Chinese Journal of Veterinary Science, 2004, 24(3):307-309. (in Chinese)
[17]	CHANG C C, YOON K J, ZIMMERMAN J J, et al. Evolution of porcine reproductive and respiratory syndrome virus during sequential passages in pigs[J]. J Virol, 2002, 76(10):4750-4763.
[18]	BEYER J, FICHTNER D, SCHIRRMEIER H, et al. Porcine reproductive and respiratory syndrome virus (PRRSV):kinetics of infection in lymphatic organs and lung[J]. J Vet Med B Infect Dis Vet Public Health, 2010, 47(1):9-25.
[19]	LAI M D, XU J. Ribosomal proteins and colorectal cancer[J]. Curr Genomics, 2007, 8(1):43-49.
[20]	张元峰, 张昆丽, 薛娟, 等. 猪脑心肌炎病毒VP1蛋白与宿主RPS20蛋白相互作用研究[J]. 中国预防兽医学报, 2018, 40(8):715-719.ZHANG Y F, ZHANG K L, XUE J, et al. Encephalomyocarditis virus VP1 interacts with ribosomal protein 20[J]. Chinese Journal of Preventive Veterinary Medicine, 2018, 40(8):715-719. (in Chinese)
[21]	YAN X Y, XIE J G, LI J, et al. Screening and analysis on the protein interaction of the protein VP7 in grass carp reovirus[J]. Virus Genes, 2015, 50(3):425-433.
[22]	喻宏, 姚立红, 陈爱君, 等. 应用酵母双杂交系统筛选与乙型流感病毒BM2相互作用的蛋白质[J]. 中华实验和临床病毒学杂志, 2005, 19(2):182-184.YU H, YAO L H, CHEN A J, et al. Screening for new binding proteins which interact with BM2 of influenza B virus with yeast two-hybrid system[J]. Chinese Journal of Experimental and Clinical Virology, 2005, 19(2):182-184. (in Chinese)
[23]	DIGIUSEPPE S, ROLLINS M G, BARTOM E T, et al. ZNF598 plays distinct roles in interferon-stimulated gene expression and poxvirus protein synthesis[J]. Cell Rep, 2018, 23(5):1249-1258.
[24]	KRISHNAN R, BODDAPATI N, MAHALINGAM S. Interplay between human nucleolar GNL1 and RPS20 is critical to modulate cell proliferation[J]. Sci Rep, 2018, 8(1):11421.
[25]	COSTERS S, LEFEBVRE D J, DELPUTTE P L, et al. Porcine reproductive and respiratory syndrome virus modulates apoptosis during replication in alveolar macrophages[J]. Arch Virol, 2008, 153(8):1453-1465.
[26]	LABARQUE G, VAN GUCHT S, NAUWYNCK H, et al. Apoptosis in the lungs of pigs infected with porcine reproductive and respiratory syndrome virus and associations with the production of apoptogenic cytokines[J]. Vet Res, 2003, 34(3):249-260.

猪繁殖与呼吸综合征病毒N蛋白与核糖体蛋白S20相互作用对病毒复制的影响

Effect of Interaction between N Protein and Ribosomal Protein S20 of Porcine Reproductive and Respiratory Syndrome Virus on Virus Replication

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