A cell-based system for hepatitis B virus replication: significance of clinically enhanced viral replication in relation to deletions in viral core promoter.

The core promoter of hepatitis B virus (HBV) is crucial in viral replication. Our previous investigation in clinical serum samples indicated that deletions in the viral core promoter (from nucleotide 1758 to 1777 and 1749 to 1768 respectively) may lead to increase in viral replication. We propose to characterize these deletion mutants in a cell-based system that supports HBV replication. Replicative HBV genome developed in our laboratory was used as template to generate deletions in viral core promoter corresponding to those found in clinical samples. These deletion constructs were used to transfect HepG2 cells, which then support the subsequent infection of cells from these deleted but replicative genomes. The effect of deletion on the viral replication was measured by comparing the amount of hepatitis B virus surface antigen (HBsAg) secreted in the culture medium with that from cells transfected with the wild type HBV genome. The amount of secreted HBsAg in cells transfected with deletion mutants was significantly lower than those transfected with the wild type genome. Our results provide a new system for molecular characterization of mechanism of HBV replication. Contrary to the increased replication observed in clinical samples, the reduced viral replication in genomes carrying deletions in the core promoter suggests a complex regulatory mechanism of viral replication which may involve other elements in the viral core promoter.