猪繁殖与呼吸综合征病毒基因组RNA衣壳化机制的研究

猪繁殖与呼吸综合征病毒（PRRSV）基因组RNA衣壳化（encapsidation）是病毒组装过程中的关键步骤。这一过程涉及N蛋白RNA结合区（RNA-binding domain）与病毒基因组RNA包装信号（packaging signal）的相互作用。目前，PRRSV基因组RNA衣壳化的机制尚未清楚。本项目中，我们拟通过Northwestern blotting、电泳迁移率实验（EMSA）和基因操作等方法来鉴定病毒基因组RNA中的包装信号和N蛋白RNA结合区中的关键氨基酸，并进一步解析包装信号中与N蛋白相互作用的结构和功能位点。在此基础上，对N蛋白RNA结合区和基因组RNA包装信号进行反向遗传操作，拯救突变病毒。分析突变病毒的病毒学和分子生物学特性，研究病毒基因组RNA衣壳化在病毒复制过程中的作用。本项目为深入了解病毒基因组RNA衣壳化机制以及为发现潜在的抗病毒靶标奠定理论基础。

Porcine Reproductive and Respiratory Syndrome virus（PRRSV） genomic RNA encapsidation is a key step for virus assembly. This process involves the interaction between the RNA binding domain of N protein and RNA packaging signal locating in genomic RNA. Currently, the RNA packaging signal remains largely undescribed and little is known about the key motif or key amino acids or motif in RNA binding domain interacting with RNA packaging signal. In this proposal, to more precisely define the location of the PRRSV packaging signal within the genomic RNA and identify the motif or key amino acids of N protein interactive with genomic RNA sub-genomic RNA, electrophoretic mobility shift assay (EMSA), Northwestern blotting and genetic manipulation are performed to analyze the interaction between N protein and virus genomic RNA. Reverse genetic manipulation on RNA- binding region and packaging signal is carried out to analyze the virological and molecular biological characteristics of the mutated viruses. The finding of his study will lay a theoretical foundation for understanding the mechanism of PRRSV genomic RNA encapsidation and exploration of the potential antiviral targets.