Abstract：
The NSs protein is a major virulence factor in bunyaviruses, crucial for viral pathogenesis. However, assessing NSs protein function can be challenging due to its inhibition of cellular RNA polymerase II, impacting NSs protein expression from plasmid DNA. The recombinant Rift Valley fever virus (RVFV) MP-12 strain (rMP-12), a highly attenuated vaccine strain, can be safely manipulated under biosafety level 2 conditions. Leveraging a reverse genetics system, we can engineer rMP-12 variants expressing heterologous NSs genes, enabling functional testing in cultured cells. Human macrophages hold a central role in viral pathogenesis, making them an ideal model for assessing NSs protein functions. Consequently, we can comprehensively compare and analyze the functional significance of various NSs proteins in human macrophages using rMP-12 NSs variants. In this chapter, we provide a detailed overview of the preparation process for rMP-12 NSs variants and introduce two distinct human macrophage models: THP-1 cells and primary macrophages. This research framework promises valuable insights into the virulence mechanisms of RVFV and other bunyaviruses and the potential for vaccine development.
摘要：
NSs蛋白是布尼亚病毒的主要毒力因子,对于病毒的发病机制至关重要。然而,评估NSs蛋白功能可能是具有挑战性的，因为它抑制了细胞RNA聚合酶II，影响来自质粒DNA的NSs蛋白表达。重组裂谷热病毒(RVFV)MP-12株(rMP-12),一种高度减毒的疫苗株，可以在生物安全2级条件下安全操纵。利用反向遗传学系统，我们可以设计表达异源NSs基因的rMP-12变体，能够在培养的细胞中进行功能测试。人巨噬细胞在病毒发病机制中具有重要作用，使它们成为评估NSs蛋白功能的理想模型。因此,我们可以使用rMP-12NSs变体全面比较和分析人巨噬细胞中各种NSs蛋白的功能意义。在这一章中,我们提供了rMP-12NSs变体的制备过程的详细概述,并介绍了两种不同的人类巨噬细胞模型:THP-1细胞和原代巨噬细胞.该研究框架承诺了对RVFV和其他布尼亚病毒的毒力机制以及疫苗开发潜力的宝贵见解。
