VP1 protein of encephalomyocarditis virus inhibits type I IFN signaling pathway and promotes virus proliferation in vitro

doi:10.3969/j.issn.1004-1524.2021.01.03

Acta Agriculturae Zhejiangensis ›› 2021, Vol. 33 ›› Issue (1): 18-26.DOI: 10.3969/j.issn.1004-1524.2021.01.03

• Animal Science • Previous Articles Next Articles

VP1 protein of encephalomyocarditis virus inhibits type I IFN signaling pathway and promotes virus proliferation in vitro

HAN Yumei¹^,²(), XIE Jingying³, BI Yingjie¹^,², XU Shujuan¹^,², FENG Ruofei¹^,^*()

1. Key Laboratory of Biotechnology and Bioengineering of State Ethnic Affairs Commission, Biomedical Research Center, Northwest Minzu University, Lanzhou 730030, China
2. College of Life Science and Engineering, Northwest Minzu University, Lanzhou 730030, China
3. College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China

Received:2020-04-23 Online:2021-01-25 Published:2021-01-25
Contact: FENG Ruofei

Abstract

Abstract:

Encephalomyocarditis virus (EMCV) is an important zoonotic pathogen. In order to preliminarily study the molecular mechanism of EMCV mediated innate immune response, this study cloned VP1 protein into eukaryotic expression vector pCMV-HA, transfected it into HEK293 cells to make it overexpress in cells. The effects of VP1 overexpression on IFN-β expression and RLRs signaling pathway-related factors and downstream stimulation gene expression were detected by ELISA and real time fluorescence quantitative PCR. Results showed VP1 could significantly inhibit IFN-β expression. Further study found that VP1 could significantly inhibit the mRNA level of EMCV-induced RLRs signaling pathway-related factors, and with the expression of VP1, MAVS protein expression was significantly reduced, but had no effect on the expression of MDA5. Therefore, the data of this experiment preliminarily showed that VP1 protein could inhibit the expression of type Ⅰ IFN gene expression and block its downstream signaling pathway by inhibiting the expression of MAVS.

Key words: encephalomyocarditis virus, structural protein VP1, Type Ⅰ IFN signaling pathway

CLC Number:

S855.99

HAN Yumei, XIE Jingying, BI Yingjie, XU Shujuan, FENG Ruofei. VP1 protein of encephalomyocarditis virus inhibits type I IFN signaling pathway and promotes virus proliferation in vitro[J]. Acta Agriculturae Zhejiangensis, 2021, 33(1): 18-26.

Figures/Tables 9

Table 1 TaqMan real time fluorescence quantitative PCR primers and probe for EMCV detection

引物 Primer	引物序列Primer sequence(5'-3')
EMCV-probe	(FAM)CACTTCGATCACTATGCTTGCCGTT(Eclipse)
EMCV-qF	GTCATACTATCGTCCAGGGACTCTAT
EMCV-qR	CATCTGTACTCCACACTCTCGAATG

Table 2 Primer sequences for real time fluorescence quantitative PCR

引物名称Primer name	引物序列Primer sequence(5'-3')
IFN-β-F	TTGTTGAGAACCTCCTGGCT
IFN-β-R	TGACTATGGTCCAGGCACAG
RIG-I-F	CACCTCAGTTGCTGATGAAGGC
RIG-I-R	GTCAGAAGGAAGCACTTGCTACC
MDA5-F	GCTGAAGTAGGAGTCAAAGCCC
MDA5-R	CCACTGTGGTAGCGATAAGCAG
MAVS-F	ATGGTGCTCACCAAGGTGTCTG
MAVS-R	TCTCAGAGCTGCTGTCTAGCCA
TBK1-F	CAACCTGGAAGCGGCAGAGTTA
TBK1-R	ACCTGGAGATAATCTGCTGTCGA
IRF3-F	TCTGCCCTCAACCGCAAAGAAG
IRF3-R	TACTGCCTCCACCATTGGTGTC

Fig.1 Real time fluorescence quantitative PCR detected the mRNA level of type Ⅰ IFN signaling pathway related factor during EMCV infection *,P<0.05;**,P<0.01;***,P<0.001.

Fig.2 Expression of VP1 in different cell lines and cell viability detection

Fig.3 EMCV virus copy number and virus titer were detected by real time fluorescence quantitative PCR and TCID50 assay

Fig.4 Effect of VP1 on type Ⅰ IFN signaling pathway related factors mRNA expression

Fig.5 Effect of VP1 on EMCV induced type Ⅰ IFN signaling pathway related factors mRNA expression

Fig.6 Effects of VP1 on the expression of MDA5 and MAVS by Western-blot

Fig.7 Pattern of EMCV VP1 suppressing IFN-β signaling pathway

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VP1 protein of encephalomyocarditis virus inhibits type I IFN signaling pathway and promotes virus proliferation in vitro

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