Fat10 In The H5n1 Virus And Influenza A (h1n1) Virus Replication And The Role Of Human Airway Epithelial Cells Induced Apoptosis

The avian influenza H5N1virus causes severe disease and high mortality, which makes it a major worldwide public health concern. The virus is capable of using the host cellular machinery for aspects of its life cycle. In this report, we discovered that FAT10, a ubiquitin-like protein, was induced by the live H5N1virus infection in mice and in the human respiratory epithelial cell lines A549, BEAS-2B and CNE-2Z. Further studies demonstrated that FAT10was crucial for the replication of the H5N1virus and that FAT10ylation was involved. The knockdown of FAT10expression significantly reduced H5N1viral replication, while the overexpression of FAT10proteins markedly increased viral replication. Poly(IC), the analogue of viral RNA, could significantly increase FAT10expression through the RIG-1-p65pathway, suggesting that FAT10elevation might be crucial for viral evasion of the immune system. The p62protein, which was confirmed to be a substrate of FAT10, might be involved in this process. The knockdown of p62expression in A549cells significantly reduced the phosphorylation of STAT1during live H5N1virus infection, which suggests that p62might be involved in the innate immunity response against H5N1virus infection. Taken together, our data suggest that FAT10elevation and FAT10ylation might be crucial in viral evasion of the immune system. One of the pathogenesis of avian influenza H5N1virus is the virus-induced cell death of immune cells and respiratory tract epithelial cells. In this part of the thesis, we carried out preliminary exploration about the role of host factor FAT10in the cell death caused by H5N1virus. By measuring the cell viability through MTS, we found that the knockdown of FAT10could significantly improve the virus-induced death of respiratory tract epithelial cell, such as A549cell, BEAS-2B and CNE-2Z cell. Further study indicated that knockdown of FAT10could repress the virus-induced proteolytic cleavage of caspase-3, reduce the proportion of apoptotic cells and inhibit the mRNA level of pro-apoptotic factor TNFa and FASLG. Besides, by monitoring the protein level of LC3B, which is one of the main markers of autophagy, we found that the knockdown of FAT10could not inhibit the autophagy caused by H5N1virus infection.In conclusion, we found that the host factor FAT10plays an important role in the virus-induced respiritory tract epithelial cell death. The cellular apoptotic process but not the cellular autophagy process could be inhibited by the knockdown of FAT10. 2009pandemic H1N1virus possesses high transmissibilit and pathogenic potentiality, and its genome includes different segments from classic swine H1N1, Human H3N2and avian influenza. It could cause the acute respiratory distress syndrome (ARDS) and other severe diseases. A/Wenshan H1N1virus was studied as a strain of2009pandemic H1N1virus in our lab, causing severe apoptosis in CNE-2Z cells. Then we explored the role of FAT10in the apoptosis of CNE-2Z cells induced by A/Wenshan H1N1virus and also in the viral replication. In our research, we found that the knockdown of FAT10could inhibit the activation of caspase-3, reduce the apoptotic cell proportion, and repress the mRNA level of pro-apoptotic factor TNFa. The knockdown of FAT10could also repress the mRNA level of viral M1. Considering the first two parts of this thesis, we proposed that FAT10could be a general host factor during influenza virus infection.