E3 ubiquitin ligase SPOP regulates RLR signaling pathway and inhibits enterovirus 71 replication
10.3760/cma.j.cn112309-20230821-00047
- VernacularTitle:E3泛素连接酶斑点型锌指结构蛋白调控RLR信号通路抑制肠道病毒71型复制
- Author:
Xinyu YANG
1
;
Lichao ZANG
;
Yang PENG
;
Lijuan JIANG
;
Jinhong MA
;
Weifeng SHI
;
Wei ZHOU
Author Information
1. 苏州大学附属第三医院医学检验科，常州 213003
- Keywords:
SPOP;
Enterovirus 71;
RLR signaling pathway
- From:
Chinese Journal of Microbiology and Immunology
2024;44(8):706-712
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the role of speckle-type POZ(pox virus and zinc finger protein) protein (SPOP) in enterovirus 71 (EV71) infection.Methods:Immunoprecipitation analysis was employed to examine the impact of SPOP on the ubiquitin level of EV71 non-structural protein 2A protease (2A pro), while the phosphorylation level of IFR3 protein was assessed through Western blot. Cells were either overexpressed or knockdown of SPOP, followed by infection with EV71. RT-qPCR was utilized to analyze the transcription level of IFN-β, and the transcription level and protein level of EV71 structural protein VP1 were determined using RT-qPCR and Western blot, respectively. Results:The inhibition of EV71 infection in RD cells was observed following transfection with HA-SPOP. Additionally, it was found that the ubiquitin level of EV71-2A pro increased in a gradient-dependent manner. Subsequent transfection with shSPOP plasmid for endogenous SPOP knockdown resulted in a dose-dependent decrease in the levels of melanoma differentiation-associated gene 5 (MDA5), mitochondrial antiviral signaling (MAVS), and p-IRF3. Conversely, transfection with HA-SPOP plasmid led to a dose-dependent increase in the levels of MDA5, MAVS, and p-IRF3. The expression of SPOP, whether high or low, had an impact on the expression of IFN-β in cells. Additionally, the levels of VP1 mRNA or protein were found to be inhibited or increased. Conclusions:SPOP plays a role in increasing the ubiquitination level of EV71-2A pro, which in turn promotes the phosphorylation level of IRF3 and secretion of IFN-β. This effect is achieved by inhibiting the cleavage of 2A pro against key molecules MAVS and MDA5 in the RLR signaling pathway, ultimately leading to the inhibition of EV71 replication.
