Study Of The Relationship Between Influenza A Viurs Infection And Cox2Expression

Influenza A virus (IAV) is a highly infectious RNA virus, which can infect the upper and lower airway epidermic cells. Cyclooxygenase2(COX2) is an important inflammatory factor related closely with many diseases including acute and chronic inflammation, cardiovascular disease and colon cancer, especially played an important role in the anti-viral reaction. At the same time, COX2also can cooperate with other inflammatory factors such as interleukin family and interferon family. In our research, we mainly focus on COX2up-regulation induced by IAV. We found the cooperation between epigenetic modification change and PKA signaling pathway activation plays important role in COX2up-regulation. And we also studied the expression change of DNMT3a and3b, which play important role in epigenetic modification in IAV infection. And next, we focus on a microRNA family-miR29family, and made a connection between miR29family and expression change of DNMT3a and3b. Finally, we found COX2may up-regulate a novel anti-viral gene IL29. to carry out the direct anti-viral activity.We proved IAV can cause COX-2up-regulation in both A549cell line and PBMC. Then we focused on the relationship between COX-2up-regulation and the expression change of three members of DNMT family:DNMT1. DNMT3a&3b. The results showed that DNMT3a and3b are down regulated sharply in early phage of virus infection, and COX-2up-regulation just happens in the same time, and tight pertinence between them is obvious.To find the unmethylated site in COX-2promoter, we analyzed COX-2promoter in A549infected by IAV. The results show that demethylation happens in COX-2promoter during IAV infection, and demethylation happens in a CpG site in an important transcriptional factor binding site CRE. Further more, to uncover the function of DNMTs in epigenetic modification and activation of COX-2promoter. Through ChIP assay, we found lower binding activity of DNMT3a and3b on COX-2promoter causes CpG demethylation and makes it easier for CREB binding to CRE site in COX-2promoter, and eventually leads to COX-2up-regulation.Next, in order to uncover the mechanism of DNMTs down-regulation, we found a miRNA family-miR29a, b&c directly located on DNMT3a and3b mRNA. In further research, we found that all the three members of miR29family can supress DNMT3a and3b expression. We also tested miR29expression in IAV infected A549cell line, PBMC and blood sample from patients and found miR29up regulated in all the above samples. This can partly tell the reason why DNMTs were down-regulated in early phage of virus infection.We also used specific miRNAs and signaling pathway inhibitors to screen the key signaling pathway related with COX-2infection in IAV infection. We found PKA-CREB pathway plays important role in COX-2up-regulation. And CREB phosphorylation is necessary for CREB binding to unmethylated CRE site in COX-2promoter and further activating COX-2expression.At last, we examined the serum samples of influenza patients. We found the downstream product of COX-2-PEG2and a newly found antiviral factor IL29are both up-regulated. To prove the relation between COX-2and IL29. we separately over-expressed COX-2and inhibited COX-2with COX-2inhibitor NS398. and then tested IL29expression. The results showed that COX-2expression contribute much to IL29expression.Generally speaking. IAV activating COX-2and IL29expression is a complex course, in which miR29family, DNMT3a&3b and CpG demethylation in CRE site in COX-2promoter are concerned. In the other hand. PKA-pCREB-CRE signaling pathway also plays important role in COX-2expression in influenza virus infection. We also found a newly found antiviral gene IL29can be up-regulated by COX-2expression, which may directly suppress virus replication.