DNA-dependent activator of interferon-regulatory factors inhibits hepatitis B virus replication

AIM： To investigate whether DNA-dependent activator of interferon-regulatory factors （DAI） inhibits hepatitis B virus （HBV） replication and what the mechanism is. METHODS： After the human hepatoma cell line Huh7 was cotransfected with DAI and HBV expressing plas- mid, viral protein （HBV surface antigen and HBV e an- tigen） secretion was detected by enzyme-linked immu- nosorbent assay, and HBV RNA was analyzed by real- time polymerase chain reaction and Northern blotting, and viral DNA replicative intermediates were examined by Southern blotting. Interferon regulatory factor 3 （IRF3） phosphorylation and nuclear translocation were analyzed via Western blotting and immunofluorescence staining respectively. Nuclear factor-KB （NF-KB） activity induced by DAI was detected by immunofluorescence staining of P65 and dual luciferase reporter assay. Tran- swell co-culture experiment was performed in order to investigate whether the antiviral effects of DAI were dependent on the secreted cytokines. RESULTS： Viral protein secretion was significantly re- duced by 57% （P 〈 0.05）, and the level of total HBV RNA was reduced by 67% （P 〈 0.05）. The viral core particle-associated DNA was also dramatically down- regulated in DAI-expressing Huh7 cells. Analysis of involved signaling pathways revealed that activation of NF-KB signaling was essential for DAI to elicit antivi- ral response in Huh7 cells. When the NF-KB signaling pathway was blocked by a NF-KB signaling suppressor （I~：B^-SR）, the anti-HBV activity of DAI was remarkably abrogated. The inhibitory effect of DAI was indepen- dent of IRF3 signaling and secreted cytokines. CONCLUSION： This study demonstrates that DAI can inhibit HBV replication and the inhibitory effect is asso- ciated with activation of NF-KB but independent of IRF3 and secreted cytokines.

AIM: To investigate whether DNA-dependent activator of interferon-regulatory factors (DAI) inhibits hepatitis B virus (HBV) replication and what the mechanism is. METHODS: After the human hepatoma cell line Huh7 was cotransfected with DAI and HBV expressing plasmid, viral protein (HBV surface antigen and HBV e antigen) secretion was detected by enzyme-linked immunosorbent assay, and HBV RNA was analyzed by real-time polymerase chain reaction and Northern blotting, and viral DNA replicative intermediates were examined by Southern blotting. Interferon regulatory factor 3 (IRF3) phosphorylation and nuclear translocation were analyzed via Western blotting and immunofluorescence staining respectively. Nuclear factor-B (NF- B) activity induced by DAI was detected by immunofluorescence staining of P65 and dual luciferase reporter assay. Transwell co-culture experiment was performed in order to investigate whether the antiviral effects of DAI were dependent on the secreted cytokines. RESULTS: Viral protein secretion was significantly reduced by 57% (P < 0.05), and the level of total HBV RNA was reduced by 67% (P < 0.05). The viral core particle-associated DNA was also dramatically down-regulated in DAI-expressing Huh7 cells. Analysis of involved signaling pathways revealed that activation of NF-B signaling was essential for DAI to elicit antiviral response in Huh7 cells. When the NF-B signaling pathway was blocked by a NF-B signaling suppressor (I B -SR), the anti-HBV activity of DAI was remarkably abrogated. The inhibitory effect of DAI was independent of IRF3 signaling and secreted cytokines. CONCLUSION: This study demonstrates that DAI can inhibit HBV replication and the inhibitory effect is associated with activation of NF-B but independent of IRF3 and secreted cytokines.