流感病毒核输出蛋白NEP可以调控病毒vRNA、cRNA和mRNA合成，这种调控不依赖于其核输出功能，其具体调控机制未知。本项目以人、猪、禽流感病毒为研究对象，首先阐明病毒聚合酶活性高低与NEP调控作用之间的关系；并从对聚合酶活性增强效果差异巨大的两个NEP出发，寻找NEP发挥调控功能的关键位点。然后，从四个方面研究：1.利用BiFC及Co-IP研究NEP同聚合酶蛋白的互作，阐明NEP在聚合酶复合物形成中的作用；2.明确NEP对聚合酶复合物结合vRNA和cRNA模板的影响，阐明其调控作用是否通过改变聚合酶复制起始来实现；3.明确NEP关键位点及复制启动子区的突变对svRNA产生的影响及其在NEP调控中的作用；4.阐明不同聚合酶蛋白PA对svRNA-PA互作的影响及其在NEP调控中的作用，揭示NEP通过svRNA及PA调控病毒RNA合成的机制。最终解析NEP调控流感病毒RNA合成的分子机制。

The influenza virus nuclear export protein NEP can regulate the synthesis of viral vRNA, cRNA and mRNA and such regulation is independent on its nuclear export function. What’s more, the molecular mechanism of the regulation is unclear. Using human, pig, and avian influenza virus as subject, this study intends to clarify the inter-relationship between the level of polymerase activity and the regulation of NEP. With the two different NEPs with distinctive effects upon the regulation of polymerase activity, we also aim to discover the key function sites on NEP in regulation. What’s next are four aspects: 1. we will study the interaction between NEP and the polymerase proteins by employing the BiFC and Co-IP to make clear the function of NEP on the formation of polymerase complex; 2. by determining the effect of NEP on the binding between polymerase complex and vRNA or cRNA, the relationship between the NEP regulation and the viral polymerase replication initiation is to be revealed; 3. by determining the effect of the mutation of key sites on NEP and replication initiation domain on svRNA level, we will reveal its role in NEP regulation; 4. by clarifying the effect of different PA on svRNA-PA interaction, the role of PA and svRNA in the regulation on viral RNA synthesis by NEP is to be revealed. This project sets its ultimate aim at making a clear elucidation of the molecular mechanisms of the regulation of NEP on RNA synthesis of influenza A virus.