Wen Lianghai, Rong Fang, Dai Guo, Liu Yufu, Lv Yadi, Luo Qiong, Liu Ding Xiang, Chen Ruiai
College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China; Zhaoqing Branch Centre of Guangdong Laboratory for Lingnan Modern Agricultural Science and Technology, Zhaoqing 526238, China; Zhaoqing Institute of Biotechnology Co., Ltd., Zhaoqing 526238, China.
Zhaoqing Branch Centre of Guangdong Laboratory for Lingnan Modern Agricultural Science and Technology, Zhaoqing 526238, China.
Int J Biol Macromol. 2025 Mar;295:139218. doi: 10.1016/j.ijbiomac.2024.139218. Epub 2025 Jan 2.
Virus-host protein interaction is critical for successful completion of viral replication cycles. As the largest nonstructural protein (NSP) of porcine reproductive and respiratory syndrome virus (PRRSV), NSP2 plays multiple and critical roles in viral replication, antiviral immunity, cellular tropism and virulence. An interactome of this protein with host proteins would be instrumental in full understanding of these multifunctional roles. In this study, we report the identification of 120 NSP2-interacting host proteins by co-immunoprecipitation coupled liquid chromatography mass spectrometry, via rescuing and utilizing a recombinant PRRSV expressing an HA-tagged NSP2. By comparing and subtracting with cells infected with parental virus, a comprehensive interactome was constructed. Bioinformatics analysis revealed that these host factors are mainly involved in translation regulation, metabolism, signal transduction and innate immunity signaling pathways. Selection of five host proteins (CtBP1, CtBP2, HSPA2, PPP1CA, SH3KBP1) for further verification and characterization confirmed their interactions with NSP2 and differential effects on PRRSV replication. Intriguingly, interaction of NSP2 and SH3KBP1 led to specific cleavage of SH3KBP1, antagonizing its pro-apoptotic activity. Taken together, this study provides overarching views on the NSP2-host interactome, paving a solid foundation for functional studies of host proteins in PRRSV biology and their potential as targets for novel therapeutics development.
病毒与宿主蛋白的相互作用对于病毒复制周期的成功完成至关重要。作为猪繁殖与呼吸综合征病毒（PRRSV）最大的非结构蛋白（NSP），NSP2在病毒复制、抗病毒免疫、细胞嗜性和毒力方面发挥着多重关键作用。该蛋白与宿主蛋白的相互作用组有助于全面理解这些多功能作用。在本研究中，我们通过共免疫沉淀结合液相色谱质谱法，拯救并利用表达HA标签NSP2的重组PRRSV，报告了120种与NSP2相互作用的宿主蛋白的鉴定结果。通过与感染亲本病毒的细胞进行比较和减法分析，构建了一个全面的相互作用组。生物信息学分析表明，这些宿主因子主要参与翻译调控、代谢、信号转导和先天免疫信号通路。选择五种宿主蛋白（CtBP1、CtBP2、HSPA2、PPP1CA、SH3KBP1）进行进一步验证和表征，证实了它们与NSP2的相互作用以及对PRRSV复制的不同影响。有趣的是，NSP2与SH3KBP1的相互作用导致SH3KBP1的特异性切割，拮抗其促凋亡活性。综上所述，本研究提供了关于NSP2-宿主相互作用组的总体观点，为宿主蛋白在PRRSV生物学中的功能研究及其作为新型治疗靶点的潜力奠定了坚实基础。
