Analysis Of Hemagglutinin’s Sturcture Of Five Strains Of H9N2 AIVs And Proteomic Study Of A549 Cells Infected By H9N2 AIV

The avian influenza virus H9N2 subtype has been circulating in domestic poultry in mainland China since 1994. Although H9N2 AIV is low pathogenic, other pathogen infected animals afer it can cause high morbidity and mortality, which would cause huge losses to the poultry breeding. Besides, changes within the amino acid sequence of virus’ hemagglutinin would make prevalent strains become combine with mammalian cell receptor easily. Infected humans had keratitis or mild flu like symptoms. Thus the public pay more attention on the pathogen. Recently, increasing evidence has shown that avian H9N2 virus might act as a source of novel human influenza viruses. The internal genes of newly emerged human infections with AIV subtype H7N9 and H10N8 subtype AIV are derived from H9N2 AIV. It’s more urgent to understand the adaption of H9N2 AIV in mammalian.We found that A/Chicken/Shandong/ch/2011 is the most sensitive to A549 cells verificated by indirect immunofluorescence test.From the perspective of the protein tertiary structure and by homology modeling method, the hemagglutinin structure difference of strains possessed different sensitivity to human lung epithelial cells were compared. The protein’s 198 th and 234 th amino acids within the receptor binding sites in the primary structure were different, but in the senior structure the position were the same. The 336-342 amino acids in the primarystructure wer the same, but the senior structure are different. The senior structureformation of A/Chicken/Shandong/ch/2011 were alpha helix in this position, but A/Chicken/Shandong/W4/2012 and the other three strains of the viruses in this positon were ranndom coil. The difference of hemagglutinin protein glycosylation sites between the5 strains of virus were found in 218, 298 and 313 glycosylation sites.In order to reveal the change trend on a large number of strains in the glycosylation sites,we select the full-length sequence of hemagglutinin of avian influenza H9N2 subtype viruses virus within East China in NCBI databases and statisticsed the evolution of the glycosylation sites. Within the H9N2 subtype influenza virus hemagglutinin fragments,glycosylation sites were conserved in 29 NST, 141 NVS, 298 NTT, 305 NVS, and 492 NGT. After the year of 2006, most of the viruses added two new two glycosylation sites, namely 218 NRT, 313 NCS.To better understand the molecular and cellular basis of H9N2 avian influenza virus infection and adaptation in human airway cells, the differentially expressed proteins in A549cells(human lung adenocarcinoma epithelial cell line) infected with H9N2 AIV were investigated by two-dimensional electrophoresis. Seventeen spots were differentially expressed between the groups. Proteins located in the downstream of the NF-κB and IFN transcription factor pathways were identified, such as ISG15. Actin and keratin were also identified, which may suggest that the cytoskeleton plays an important role in the AIV infection of mammalian cells.