猪瘟病毒非结构蛋白NS5A与Beclin1相互作用并促进病毒增殖

doi:10.11843/j.issn.0366-6964.2023.02.028

摘要/Abstract

摘要： 为探究宿主蛋白Beclin1在猪瘟病毒（classical swine fever virus，CSFV）非结构蛋白NS5A激活细胞自噬反应过程中的作用及具体分子机制，本研究在感染CSFV及表达NS5A蛋白的ST细胞中，利用qRT-PCR方法检测Beclin1、PI3K/Akt通路相关因子表达变化情况；利用激光共聚焦、Co-IP及GST-pulldown等方法研究Beclin1与NS5A相互作用关系；通过在ST细胞中过表达或敲低Beclin1，研究其对CSFV复制的影响。结果表明，ST细胞感染猪瘟病毒或外源表达NS5A蛋白，Beclin1转录和蛋白表达水平均显著升高，且PI3K/Akt通路相关因子表达水平与之呈正相关。此外，CSFV NS5A蛋白与Beclin1蛋白在细胞中存在共定位且具有相互作用。最后，作者发现在细胞中过表达Beclin1，对CSFV复制起到明显促进作用；反之，利用siRNA敲低Beclin1后，抑制PI3K/Akt通路活化，CSFV增殖表现出明显抑制效应。以上结果表明，Beclin1蛋白对CSFV复制具有促进作用，其机制是通过与NS5A的相互作用调控PI3K/Akt通路来实现。

关键词: PI3K/Akt信号通路, 猪瘟病毒, Beclin1, 蛋白互作, 自噬

Abstract: In order to explore the role of host protein Beclin1 in the process of autophagy activated by NS5A, a non structural protein of classical swine fever virus (CSFV), and the specific molecular mechanism, this study used qRT-PCR to detect the expression changes of Beclin1, PI3K/Akt signaling pathway related factors in ST cells infected with CSFV and expressing NS5A protein. The interaction between Beclin1 and NS5A was studied by laser confocal, Co IP and GST pulldown methods. By overexpressing or knocking down Beclin1 in ST cells, we studied its effect on CSFV replication. The results showed that ST cells infected with CSFV or expressed NS5A protein externally, the transcription and protein expression levels of Beclin1 were significantly increased, and the expression levels of PI3K/Akt signaling pathway related factors were positively correlated with them. In addition, CSFV NS5A protein and Beclin1 protein have co localization and interaction in cells. Finally, we found that overexpression of Beclin1 in cells significantly promoted CSFV replication; On the contrary, knockdown of Beclin1 with siRNA inhibited the activation of PI3K/Akt signaling pathway, and CSFV proliferation showed significant inhibitory effect. The above results indicate that Beclin1 protein can promote the replication of CSFV, and its mechanism is through the interaction with NS5A to regulate PI3K/Akt signaling pathway.

Key words: PI3K/Akt signaling pathway, CSFV, Beclin1, protein interaction, autophagy

中图分类号:

S852.659.6

张成成, 孙嘉豪, 王秀玲, 张小荣, 吴艳涛. 猪瘟病毒非结构蛋白NS5A与Beclin1相互作用并促进病毒增殖[J]. 畜牧兽医学报, 2023, 54(2): 715-725.

ZHANG Chengcheng, SUN Jiahao, WANG Xiuling, ZHANG Xiaorong, WU Yantao. Beclin1 Interacts with the Nonstructural Protein NS5A of CSFV and Promotes Virus Proliferation[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(2): 715-725.

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