Influenza A Virus (H1N1) Facilitates Its Replication By Activating HIF-1 Pathway

BACKGROUND AND OBJECTIVE: High viral load and inflammatory cascade are important causes of mortality in patients with severe influenza A(H1N1)virus infection.Hypoxia-inducible factor 1(HIF-1)is a transcriptional activator of various genes related to cellular adaptive responses to hypoxia,is activated by a variety of pathogens and plays different roles in the pathogenesis of different pathogens.Whether HIF-1 signaling pathway is involved in the pathogenesis of influenza A virus remains unclear.This study explored the impact of influenza A(H1N1)virus on HIF-1 signaling pathway,as well as the role of HIF-1 in influenza A(H1N1)virus infection.It may provide a theoretical and experimental basis for a better understanding of virus-host interaction and finding potential therapeutic targets for severe influenza A virus infection.METHODS: Influenza A(H1N1)virus infected or mock infected mice model and A549 cells model were established.The expression of HIF-1α,factor inhibiting HIF-1(FIH-1),downstream target genes of HIF-1(VEGF and GLUT-1),the transcription of HIF-1α mRNA,degradation curve of HIF-1α,prolyl hydroxylation and ubiquitination of HIF-1α were detected by qRT-PCR,western blot,immunofluorescence and immunoprecipitation.A GFPu stable-expressing A549 cell line was established by lentivirus transfection technology to evaluate the impact of H1N1 infection on proteasome.HIF-1α and HIF-1β stable-knocking down and control A549 cell lines were established by lentivirus-mediated RNAi technology to detect the role of HIF-1 in H1N1 replication,the effect of H1N1 infection on glycometabolism,and the role of HIF-1 in the alteration of glycometabolism.Glycolysis inhibitors were used to observe their effects on H1N1 replication and cytopathy.RESULTS: 1.Influenza A(H1N1)virus infection up-regulates the expression of HIF-1α and activates the HIF-1 signaling pathway in vivo and in vitro.2.H1N1 infection does not increase HIF-1α mRNA transcription,nor impairs posttranslational prolyl hydroxylation or ubiquitination of HIF-1α,but inhibits the function of proteasome,resulting in HIF-1α accumulation.3.H1N1 infection also down-regulates the expression of FIH-1,which further activates HIF-1 signaling pathway.4.Knocking down HIF-1 significantly impairs the replication of H1N1 virus in alveolar epithelial cells.5.H1N1 infection enhances glycolysis in alveolar epithelial cells,and HIF-1 participates in this enhancement.6.Inhibition of glycolysis significantly reduces the replication of H1N1 virus and alleviates virus-induced cytopathy in alveolar epithelial cells.CONCLUSIONS: 1.Influenza A(H1N1)virus infection may stabilize HIF-1α by inhibiting the function of proteasome in host cells,down-regulate the expression of FIH-1,and activate the HIF-1 signaling pathway.2.Enhanced glycolysis after HIF-1 activation further promotes the replication of influenza A(H1N1)virus in alveolar epithelial cells,thus forming a cycle of "H1N1 infection-HIF-1 activation-glycolysis enhancement-viral replication".3.Inhibiting HIF-1 or glycolysis may become a new direction for the treatment of influenza A(H1N1)virus infection.