与PRRSV nsp11互作的宿主细胞蛋白鉴定及生物信息学分析

【目的】研究猪繁殖与呼吸综合征病毒(Porcine reproductive and respiratory syndrome virus,PRRSV)nsp11与宿主细胞蛋白之间的相互作用,对于揭示nsp11在病毒复制过程中发挥的功能至关重要。【方法】在病毒感染细胞的基础上,利用nsp11的单克隆抗体,采用免疫沉淀结合串联质谱的方法,筛选与PRRSV nsp11相互作用的宿主细胞蛋白,并对所筛选出的宿主细胞蛋白进行了GO注释、COG注释和KEGG代谢通路注释;选取筛选出的宿主细胞蛋白IRAK1,利用免疫共沉淀技术和激光共聚焦技术鉴定其与nsp11之间的相互作用。【结果】与空白对照组相比,病毒感染组中出现3条差异带;经质谱分析共筛选得到了201个与nsp11相互作用的宿主细胞蛋白,分别与蛋白质代谢、细胞信号通路转导以及病原致病性等密切相关;在生物信息学分析的基础上,实验验证了nsp11确与宿主细胞蛋白IRAK1进行相互作用。【结论】鉴定出与PRRSV nsp11相互作用的宿主细胞蛋白,生物信息学分析显示它们在病毒的复制和致病过程中发挥重要作用。研究结果为探究nsp11的生物学功能指明了方向,也为研究宿主细胞蛋白与病毒蛋白间的相互作用及其调控病毒复制和致病性的分子机制奠定了基础。

Identification and bioinformatics analysis of host cellular proteins interacting with PRRSV nsp11

Objective] Exploring the interaction between porcine reproductive and respiratory syndrome virus (PRRSV) nsp11 and host cellular proteins is important for revealing the function of nsp11 in viral replication. Methods] The host cellular proteins that interact with nsp11 were screened by immunoprecipitation combined tandem mass spectrometry and analyzed by GO annotation, COG annotation as well as KEGG pathway annotation. The screened host cell protein IRAK1 was selected, and then the interaction between nsp11 and IRAK1 was determined by co-immunoprecipitation and confocal microscopy assays. Results] Compared with the control group, there were 3 differential bands in PRRSV-infected group and 201 host cellular proteins were identified on these differential bands by further LC-MS/MS analysis. These host cellular proteins are closely related to protein metabolism, transduction of cell signaling pathways and pathogenicity of pathogens. Based on the bioinformatics analysis, host cellular protein IRAK1 was identified to interact with the nsp11. Conclusion] This study identified the host cellular proteins that can interact with PRRSV nsp11, and bioinformatics analysis showed that these proteins play crucial role in virus replication and pathogenesis. The results indicate the direction of the study of nsp11, and also provide a foundation for elucidating the associated molecular mechanisms of the interaction of host cellular proteins with viral proteins in regulating the viral replication and pathogenesis.