PIAS1对PRRSV N蛋白SUMO化修饰及病毒复制的影响

Effects of PIAS1 on sumoylation of PRRSV N protein and viral replication

摘要:

【目的】猪繁殖与呼吸综合征病毒(PRRSV)是一种危害全球养猪业的重要病原。SUMO (Small ubiquitin-like modifier)化修饰作为一种可逆的翻译后修饰在调节病毒复制方面发挥着重要功能。PIAS1 (Protein inhibitor of activated STAT1)是SUMO E3连接酶PIAS家族的一员，可以促进靶蛋白的SUMO化修饰，进而影响靶蛋白的功能，参与基因转录调控过程。探究PIAS1与PRRSV N蛋白相互作用的机制及其对N蛋白SUMO化修饰和病毒复制的影响，为进一步阐明PRRSV复制调控和致病的分子机制提供科学依据。【方法】利用酵母回复杂交、免疫共沉淀和激光共聚焦技术验证N蛋白与PIAS1的相互作用；以递增剂量外源性转染PIAS1观察其是否介导N蛋白SUMO化修饰；采用RNA干扰和慢病毒转导技术测定PIAS1对PRRSV复制的影响。【结果】PIAS1能与N蛋白相互作用，而且两者主要共定位于胞浆中；外源转染PIAS1并未增加N蛋白SUMO化修饰水平；在MARC-145细胞中，PIAS1的表达有利于PRRSV的复制。【结论】PIAS1可促进PRRSV的复制。

Abstract:

[Objective] Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most economically important pathogens that severely affect pork industry worldwide. As a reversible post-translational modification, sumoylation plays an important role in regulating viral replication. Protein inhibitor of activated STAT1 (PIAS1), as a member of SUMO E3 ligase PIAS family, can promote sumoylation of target protein, which affects the function of target protein and further participates in the transcriptional regulation process. This study was designed to study the mechanism of interaction between PIAS1 and PRRSV N protein, as well as its effect on N protein sumoylation and viral replication, to provide a scientific basis for further understanding the replication regulation and pathogenesis of PRRSV. [Methods] The interaction between PRRSV N protein and PIAS1 was first verified by yeast two-hybrid, Co-IP and confocal immunofluorescence assay. And then increased dose of PIAS1 were exogenously transfected to observe whether PIAS1 could mediate sumoylation of N protein. The effect of PIAS1 on PRRSV replication was investigated by testing the virus titer in cells with increased/reduced PIASI by using lentiviral transduction or RNA interference technology. [Results] PIAS1 could interact with the PRRSV N protein, and both of them predominantly located in the cytoplasm. Exogenous transfection of PIAS1 did not increase the sumoylation level of N protein. However, the overexpression of PIAS1 could promote PRRSV replication in MARC-145 cells. [Conclusion] PIAS1 has the function of promoting PRRSV replication.