过表达非洲猪瘟病毒D1133L蛋白的MA-104细胞系建立及其初步应用

doi:10.11843/j.issn.0366-6964.2022.06.021

Abstract

Abstract: To study the function of D1133L gene encoded by African swine fever virus (ASFV), we constructed MA-104/D1133L cell line for overexpression of D1133L using lentivirus expression vector, and compared ASFV replication ability in MA-104/D1133L and MA-104 cells. The transcription and protein expression levels of p30 and p72 in ASFV infected MA-104/D1133L cells were evaluated by real-time qPCR and Western blot respectively, and the proliferation of ASFV was evaluated according to virus titer and virus copies number. The transcriptional and protein levels of ASFV p30 and p72 proteins in MA-104/D1133L cell line were higher than those in wild MA-104 cell, and the viral proliferation capacity of ASFV in MA-104/D1133L cell line was higher than that in wild MA-104 cell line. The results showed that overexpressing ASFV D1133L gene of MA-104 cell line was successfully constructed in this study, that compared with wild type cells, MA-104/D1133L cell line was more conducive to ASFV replication, and D1133L promoted ASFV proliferation. These results provide biomaterials for further study of the function of ASFV D1133L gene.

Key words: African swine fever virus, D1133L genes, MA-104 cells, gene function, overexpressed cell lines

CLC Number:

S852.659.6

ZHANG Ting, YANG Bo, CUI Huimei, YUAN Xingguo, ZHAO Dengshuai, YANG Jinke, HAO Yu, CHEN Xuehui, YAN Wenqian, SHEN Chaochao, SHI Xijuan, ZHANG Dajun, YANG Xing, LIU Xiangtao, ZHENG Haixue, ZHANG Keshan. Establishment and Preliminary Application of MA-104 Cell Line Overexpressing African Swine Fever Virus D1133L Protein[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(6): 1877-1885.

References 31

[1]	TULMAN E R, DELHON G A, KU B K, et al. African swine fever virus[M]//VAN ETTEN J L. Lesser Known Large dsDNA Viruses. Berlin: Springer, 2009: 43-87.
[2]	TEKLUE T, SUN Y, ABID M, et al. Current status and evolving approaches to African swine fever vaccine development[J]. Transbound Emerg Dis, 2020, 67(2): 529-542.
[3]	WU K K, LIU J M, WANG L X, et al. Current State of Global African swine fever vaccine development under the prevalence and transmission of ASF in China[J]. Vaccines, 2020, 8(3): 531.
[4]	SÁNCHEZ E G, RIERA E, NOGAL M, et al. Phenotyping and susceptibility of established porcine cells lines to African Swine Fever Virus infection and viral production[J]. Sci Rep, 2017, 7(1): 10369.
[5]	GÓMEZ-VILLAMANDOS J C, BAUTISTA M J, SÁNCHEZ-CORDÓN P J, et al. Pathology of African swine fever: the role of monocyte-macrophage[J]. Virus Res, 2013, 173(1): 140-149.
[6]	RAI A, PRUITT S, RAMIREZ-MEDINA E, et al. Identification of a continuously stable and commercially available cell line for the identification of infectious African swine fever virus in clinical samples[J]. Viruses, 2020, 12(8): 820.
[7]	IYER L M, ARAVIND L, KOONIN E V. Common origin of four diverse families of large eukaryotic DNA viruses[J]. J Virol, 2001, 75(23): 11720-11734.
[8]	DIXON L K, ISLAM M, NASH R, et al. African swine fever virus evasion of host defences[J]. Virus Res, 2019, 266: 25-33.
[9]	YÁÑEZ R J, RODRÍGUEZ J M, BOURSNELL M, et al. Two putative African swine fever virus helicases similar to yeast 'DEAH' pre-mRNA processing proteins and vaccinia virus ATPases D11L and D6R[J]. Gene, 1993, 134(2): 161-174.
[10]	CACKETT G, SYKORA M, WERNER F. Transcriptome view of a killer: African swine fever virus[J]. Biochem Soc Trans, 2020, 48(4): 1569-1581.
[11]	ALEJO A, MATAMOROS T, GUERRA M, et al. A proteomic atlas of the African swine fever virus particle[J]. J Virol, 2018, 92(23): e01293-18.
[12]	DIXON L K, CHAPMAN D A G, NETHERTON C L, et al. African swine fever virus replication and genomics[J]. Virus Res, 2013, 173(1): 3-14.
[13]	JIA X X, LIU Y, MENG D M, et al. Screening and identification of Vero cell line to stably express capsid protein of peste des petits ruminants virus[J/OL]. Chinese Journal of Animal Infectious Diseases. (2020-02-12). S.20200211.1624.044.html." target="_blank">http://kns.cnki.net/kcms/detail/31.2031.S.20200211.1624.044.html. (in Chinese)贾雪霞, 刘莹, 孟德梅, 等. 稳定表达小反刍兽疫病毒衣壳蛋白的Vero细胞系的筛选和鉴定[J/OL]. 中国动物传染病学报. (2020-02-12). S.20200211.1624.044.html" target="_blank">http://kns.cnki.net/kcms/detail/31.2031.S.20200211.1624.044.html.
[14]	LI Z H, SONG Z A, YANG Z X, et al. Construction of BHK-21 cell line stably expressing VP6 protein of Bluetongue virus[J]. Journal of Agricultural Biotechnology, 2021, 29(4): 780-788. (in Chinese)李占鸿, 宋子昂, 杨振兴, 等. 稳定表达蓝舌病病毒VP6蛋白的BHK-21细胞系构建[J]. 农业生物技术学报, 2021, 29(4): 780-788.
[15]	WUDONG G W, WEN Y S, GUO H, et al. Construction of a Marc-145 cell line stably expressing PRRSV GP2 protein[J/OL]. Chinese Journal of Animal Infectious Diseases. (2021-04-27). S.20210427.1447.055.html" target="_blank">http://kns.cnki.net/kcms/detail/31.2031.S.20210427.1447.055.html. (in Chinese)乌东高娃, 温永胜, 郭昊, 等. 稳定表达PRRSV GP2蛋白的Marc-145细胞系的构建[J/OL]. 中国动物传染病学报. (2021-04-27). S.20210427.1447.055.html" target="_blank">http://kns.cnki.net/kcms/detail/31.2031.S.20210427.1447.055.html.
[16]	REN T W, QIN N, QUAN D Q, et al. Construction of a Marc-145 cell line stably expressing PRRSV N protein[J]. Chinese Journal of Veterinary Parasitology, 2022, 30(1): 15-19. (in Chinese)任同伟, 覃念, 全东群, 等. 稳定表达PRRSV N蛋白的Marc-145细胞系的构建[J]. 中国动物传染病学报, 2022, 30(1): 15-19.
[17]	JIA Y H, LIU W, XU Z K, et al. Construction of BHK-21 cell line stably expressing TIGAR gene and evaluation of its proliferation effect on Newcastle disease virus[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(2): 440-449. (in Chinese)栗永华, 刘伟, 徐智凯, 等. 稳定表达TIGAR基因的BHK-21细胞系的构建及其对新城疫病毒增殖效果的评价[J]. 畜牧兽医学报, 2021, 52(2): 440-449.
[18]	HOU J, SHEN C C, ZHANG D J, et al. Gene sequence analysis, protein structure prediction and subcellular localization of African swine fever virus helicase D1133L[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(7): 1953-1962. (in Chinese)侯景, 申超超, 张大俊, 等. 非洲猪瘟病毒解旋酶D1133L基因序列分析、蛋白结构预测及亚细胞定位[J]. 畜牧兽医学报, 2021, 52(7): 1953-1962.
[19]	BORCA M V, RAMIREZ-MEDINA E, SILVA E, et al. Development of a highly effective African swine fever virus vaccine by deletion of the I177L gene results in sterile immunity against the current epidemic Eurasia strain[J]. J Virol, 2020, 94(7): e02017-19.
[20]	YANG B, ZHANG D J, SHI X J, et al. Construction, identification and analysis of the interaction network of African swine fever virus MGF360-9L with host proteins[J]. Viruses, 2021, 13(9): 1804.
[21]	KING D P, REID S M, HUTCHINGS G H, et al. Development of a TaqMan® PCR assay with internal amplification control for the detection of African swine fever virus[J]. J Virol Methods, 2003, 107(1): 53-61.
[22]	TEKLUE T, WANG T, LUO Y Z, et al. Generation and evaluation of an African swine fever virus mutant with deletion of the CD2v and UK genes[J]. Vaccines, 2020, 8(4): 763.
[23]	PORTUGAL R, GOATLEY L C, HUSMANN R, et al. A porcine macrophage cell line that supports high levels of replication of OURT88/3, an attenuated strain of African swine fever virus[J]. Emerg Microbes Infect, 2020, 9(1): 1245-1253.
[24]	RAI A, PRUITT S, RAMIREZ-MEDINA E, et al. Detection and quantification of African swine fever virus in MA-104 cells[J]. Bio-protocol, 2021, 11(6): e3955.
[25]	WANG T, WANG L, HAN Y, et al. Adaptation of African swine fever virus to HEK293T cells[J]. Transbound Emerg Dis, 2021, 68(5): 2853-2866.
[26]	RODRIGUEZ J M, SALAS M L, SANTARÉN J F. African swine fever virus-induced polypeptides in porcine alveolar macrophages and in Vero cells: two-dimensional gel analysis[J]. Proteomics, 2001, 1(11): 1447-1456.
[27]	ZHAO D M, LIU R Q, ZHANG X F, et al. Replication and virulence in pigs of the first African swine fever virus isolated in China[J]. Emerg Microbes Infect, 2019, 8(1): 438-447.
[28]	TABARÉS E, OLIVARES I, SANTURDE G, et al. African swine fever virus DNA: deletions and additions during adaptation to growth in monkey kidney cells[J]. Arch Virol, 1987, 97(3-4): 333-346.
[29]	MONTEAGUDO P L, LACASTA A, LÓPEZ E, et al. BA71ΔCD2: a new recombinant live attenuated African swine fever virus with cross-protective capabilities[J]. J Virol, 2017, 91(21): e01058-17.
[30]	KRUG P W, HOLINKA L G, O'DONNELL V, et al. The progressive adaptation of a Georgian isolate of African swine fever virus to Vero cells leads to a gradual attenuation of virulence in swine corresponding to major modifications of the viral genome[J]. J Virol, 2015, 89(4): 2324-2332.
[31]	BORCA M V, RAI A, RAMIREZ-MEDINA E, et al. A cell culture-adapted vaccine virus against the current African swine fever virus pandemic strain[J]. J Virol, 2021, 95(14): e0012321.

Establishment and Preliminary Application of MA-104 Cell Line Overexpressing African Swine Fever Virus D1133L Protein

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References 31

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	LIU Chuanxia, WANG Xiao, LI Xuewen, BAO Miaofei, LI Tingting, CHEN Xin, WENG Changjiang, ZHENG Jun. Preparation of Monoclonal Antibody of African Swine Fever Virus pE120R [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 388-394.
[2]	FENG Yongzhi, GONG Ting, WU Dongdong, GAO Qi, ZHENG Xiaoyu, ZHANG Guihong, SUN Yankuo. Analysis of Factors Affecting the Infectivity of African Swine Fever Virus on Cultured Cells [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3406-3414.
[3]	LIU Taoxue, SU Bingqian, QI Yanli, GUO Jiangtao, LIU Zhonghu, CHU Beibei, WANG Jiang, ZENG Lei. Preparation of the Monoclonal Antibody against the African Swine Fever Virus p30 Protein and Identification of the Antigenic Epitope [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3415-3423.
[4]	DING Xiaoyan, HE Jiuxiang, ZHOU Xiaoyang, ZHOU Yuxin, LI Jintao. Preliminary Identification of Host Regulatory Genes and Virulence Genes during African Swine Fever Virus Infection [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(7): 2964-2971.
[5]	WANG Ying, ZHU Jiahong, ZHAO Jiakai, JI Pinpin, CHEN Xu, ZHANG Lu, LIU Baoyuan, SUN Yani, ZHAO Qin. Screening and Identification of Nanobodies against NP419L Protein of African Swine Fever Virus and Its Preliminary Application of Antibody Detection [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(6): 2509-2520.
[6]	LIU Wenhao, ZHU Yance, ZHANG Dongxuan, WANG Zhihao, ZHANG Chao. Construction of PK 15 Cell Line Stably Expressing African Swine Fever Virus E165R Protein [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(6): 2662-2666.
[7]	WANG Guochao, ZHAO Yaru, ZHANG Zhonghui, ZHANG Yulong, BAI Ge, GENG Shuxian, FAN Jie, YANG Jifei, GUAN Guiquan, YIN Hong, LUO Jianxun, NIU Qingli. Bioinformatics Analysis of RNA Polymerase Subunit D205R Gene of African Swine Fever Virus and Polyclonal Antibody Preparation [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 2042-2049.
[8]	ZHANG Ting, FENG Tao, YANG Jinke, HAO Yu, YANG Xing, ZHANG Dajun, SHI Xijuan, YAN Wenqian, CHEN Lingling, LIU Xiangtao, ZHENG Haixue, ZHANG Keshan. Construction and Growth Characteristics of Recombinant African Swine Fever Virus with Conditional Deletion of D1133L Gene [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(2): 706-714.
[9]	ZHANG Fangyuan, YANG Dawei, QIU Deyang, JIANG Guoqian, LI Guimei, SHAN Hu. Expression of ASFV P30 Protein and Development of ASFV Antibody Detection Method Based on x-MAP Technology [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(10): 4300-4310.
[10]	QI Yanli, LIU Taoxue, YU Haishen, ZHANG Chao, LU Weifei, WANG Jiang, CHU Beibei, ZHANG Gaiping. Preparation of the Monoclonal Antibody against the African Swine Fever Virus p54 Protein and Identification of the Antigenic Epitope [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(1): 281-292.
[11]	WANG Yang, CUI Shuai, XIN Ting, WANG Xixi, YU Hainan, CHEN Shiyu, JIANG Yajun, GAO Xintao, PANG Zhongbao, JIANG Yitong, GUO Xiaoyu, JIA Hong, ZHU Hongfei. ASFV MGF360-14L Interacts with MAVS and Inhibit the Expression of Type Ⅰ Interferon [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(9): 3272-3278.
[12]	XU Lifeng, JIA Chenyu, CHEN Fuzai, FANG Mengyuan, LIU Xiaodan, CHEN Jilong, LI Xunliang. Isolation, Identification and Whole Genome Analysis of a Mycoplasma synoviae Strain from Chicken [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(8): 2663-2676.
[13]	YUAN Xingguo, SHEN Chaochao, ZHAO Dengshuai, YANG Bo, CUI Huimei, HAO Yu, YANG Jinke, CHEN Xuehui, ZHANG Ting, ZHANG Keshan, LIU Xia, ZHENG Haixue, LIU Xiangtao. β-estradiol and Progesterone Promote Replication of African Swine Fever Virus in vitro [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(7): 2252-2259.
[14]	CHEN Xiaojun, MA Jun, CHEN Xiongnan, LIANG Yifan, GAO Qi, HUANG Zhao, XU Runda, ZHENG Jiachen, ZHENG Zezhong, ZHANG Guihong, WANG Heng, GONG Lang. Identification of the B Cell Epitopes Recognized by a Monoclonal Antibody against the p30 Protein of African Swine Fever Virus [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(7): 2239-2251.
[15]	ZHAO Xuyang, JIN Jiaxin, LU Wenlong, ZHANG Shuai, HUANG Li, ZHANG Gaiping, SUN Aijun, ZHUANG Guoqing. Advances in the Molecular Mechanism of Immune Escape of African Swine Fever Virus [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(7): 2074-2082.