mRNA-蛋白质相互作用是基因表达过程的重要内容，与疾病发生、病毒-宿主相互作用等密切相关。原位高分辨可视解析mRNA-蛋白质相互作用能更好地理解基因表达调控和病毒侵染机制，但是目前对mRNA-蛋白质细胞内相互作用仍缺乏超高分辨成像研究方法。本申请拟结合蛋白片段互补技术和光控荧光蛋白mIrisFP的双重光活化特性，拆分并构建mIrisFP双分子和三分子荧光互补系统，建立mRNA-蛋白质相互作用STORM/PALM超高分辨成像分析新方法。并以甲型流感病毒mRNA-蛋白质相互作用为研究对象，将mIrisFP三分子荧光互补系统超高分辨成像技术用于细胞内流感病毒mRNA及宿主干扰素mRNA与流感病毒NS1蛋白相互作用研究。本申请将发展mRNA-蛋白质相互作用超高分辨成像新技术，深入解析甲型流感病毒重要mRNA-蛋白质相互作用行为，为理解流感病毒与宿主细胞相互作用关系以及病毒侵染与致病机制提供基础。

mRNA-protein interactions play key roles in gene expression. mRNA-protein interactions are also found being involved in several human diseases and virus-host interactions. Visulization mRNA-protein interactions directly in cells by super-resolution fluorescence imaging technique will help understanding the mechanism of gene expression and viral infection. However, so far, there is still no method for super-resolution fluorescence imaging of mRNA-protein interactions in cells. In this proposal, based on the technique of protein-fragment complementation and fluorescent protein mIrisFP with photoactivatable and photoswitchable properties, we will attempt to build new trimolecular fluorescence complementation (TriFC) systems for STORM/PALM super-resolution microscopy for imaging mRNA-protein interactions. Then, the newly constructed TriFC system will be used for imaging some of mRNA-protein interactions in influenza A virus infection. Interactions between some of influenza A virus mRNAs, cellular interferon mRNAs and the viral NS1 protein will be imaged and analyzed in cells. The planning study will develop new fluorescent protein-fragment complementation systems for imaging of mRNA-protein interactions with super-resolution.The study will also provide further understanding of influenza A virus mRNA-protein interactions, which will help to understand the virus-host interactions and pathogenicity of influenza A virus.