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Study on the effect of host factor KRT2 on the replication of influenza A virus

WANG Yi-han;SHI Wen-jun;LI Qi-bing;HU Yu-zhen;ZHAO Yu-hui;WANG Guang-wen;YAN Ya;JIANG Li;CHEN Hua-lan;LI Cheng-jun;Animal Influenza Key Laboratory of the Ministry of Agriculture and Rural Affairs, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences;  
Host factor type II cytoskeleton epidermal protein 2(KRT2), which may be associated with influenza virus replication, was identified previously by mass spectrometry in our Laboratory. To further explore the effect of KRT2 on influenza virus replication, HEK293T cells were transfected with eukaryotic plasmid PCAGGs-KRT2-MyC for 24 hours and then infected with A/WSN/1933(H1N1) strain. The effect of overexpression of KTR2 protein on influenza virus replication was determined by plaque titration and confocal laser scanning microscopy(CLSM). The results showed that overexpression of KRT2 protein could downregulate influenza virus replication. In addition, the A549 cells were transfected with specific designed siRNA-1/2 targeting KRT2 gene to detect the effect of down-regulated expression of KRT2 protein on A549 cell viability by cell viability detection kit. The siRNA with optimal interference efficiency was screened by fluorescence quantitative PCR. The results showed that KRT2-specific siRNA did not affect A549 cell viability, and siRNA-2 could reduce KRT2 expression more effectively in comparation with the control siRNA. After the siRNA-2 transfected A549 cells were infected with A/WSN/1933(H1 N1) virus strain, the effect of downregulated KTR2 protein expression on influenza virus replication was observed by phage titration and CLSM. All the above results indicated that down-regulated KRT2 expression by siRNA interference could up-regulate influenza virus replication. The detection by using CLSM showed that influenza virus replication was inhibited at the early stage after down-regulation of KRT2 protein expression level. In conclusion, KRT2 is a host factor that negatively regulates influenza virus replication. This study enriches the network of interaction between influenza virus and host factors and deepens the understanding of the mechanism by which host factors IVA replication.
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