Interferon-inducible MyD88 Protein Inhibits Hepatitis B Virus Replication

Interferon-inducible MyD88 Protein Inhibits Hepatitis B Virus ReplicationInfection of hepatitis B virus (HBV) continues to be a significant health problem. It is estimated that there are approximately 350 million chronic hepatitis B patients worldwide. These patients have a high risk of developing liver cirrhosis and hepatocellular carcinoma with high mortality rate. Interferon alpha (IFN-α) has been proven to be effective in the treatment of chronic hepatitis B patients, which can decrease viral tite and improve liver function. But long-term response of chronic hepatitis B for IFN-α treatment is low and the cost is expensive. The antiviral activation of IFN is the outcome of the interaction between HBV and host cells. Upon binding to its specific cell surface receptor, IFN-α activates the expression of a number of cellular genes, through the distinct, yet not completely understood pathways of signaling cascade in a rapid and direct manner. HBV may also decrease the expression of some IFN-α induced genes to escape the antiviral effect of the IFN system. Many studies have been focused on investigating the complex mechanisms responsible for the antiviral activity of IFN and the persistent existence of HBV in the cells, but only parts of them have been defined. To study the antiviral mechanisms of IFN and the global effect of HBV persistent existence on IFN induced cellular gene expression, cDNA microarrays dotted with 14112 human genes were used to examine the transcriptional changes between an HBV DNA transfected cell line (HepG2.2.15 cells) and its parental cell line (HepG2 cells) after the treatment of IFN-α for 6 h. The results showed that many genes related to cell cycle, proliferation, apoptosis and new ESTs were regulated by IFN-α. Many genes involved in kinase and signal transduction, transcription regulation, antigen presentation and processing were differentially regulated between these two cell lines post IFN-α treatment. For example, the transcription of myeloid differential primary response protein MyD88 was significantly up-regulated (over 3.0-fold) by IFN-α in HepG2 cells, but were only mildly increased in HepG2.2.15 cells. After inhibition of HBV replication by Lamivudine or Adefovir, the transcription of MyD88 gene induced by IFN-α was increased in HepG2.2.15 cells. These results imply that the reduction of the transcription of MyD88 gene in HBV existing cells may protect HBV from host cells' eradication, while the increasing expression of MyD88 gene may inhibit HBV replication.MyD88 is a critical component in the signaling cascade through Toll-like receptors, interleukin-1 and interleukin-18 receptors. But the role of MyD88 protein in IFN signaling pathway and its antiviral activity has not been reported. To examine the role of MyD88 in the antiviral activity of IFN-α against HBV, MyD88 stably expressing cell lines were established and the HBV replication in these lines after transient transfection of HBV replication competent plasmid was further studied. Results showed that the syntheses of HBV proteins and viral replicative intermediates in MyD88 expressing cells were effectively reduced by about 60% for HBV surface and E antigens, and 80% for HBV DNA replicative forms. A significant reduction of about 70% for total viral transcripts and 80% for cytoplasmic viral RNAs was also observed. Using nuclear factor-kappa B (NF-kB) dependent reporter assay, it was revealed that the activation of NF-kB could be induced by expression of MyD88 (about 2 fold) and further increased by co-expression of HBV (about 6-8 fold). The synergitic activation of NF-kB by N-domain of MyD88 protein and HBV or by HBV x protein and MyD88 protein was also revealed (about 5-6 fold). But C-domain of MyD88 protein and HBV core and surface antigen did not have any effect on the activation of NF-kB. Because the N-domain of MyD88 protein was also found to inhibit HBV replication, it is presumed that N-domain of MyD88 is the key region for the activation of NF-kB and the inhibition of HBV replication.In summa...