Effects of mutations in the X gene of hepatitis B virus on the virus replication

Previously, we have found a new mutation at nt 1726-1730 that is associated with lower hepatitis B virus (HBV) DNA levels in the liver, and mutations at nt 1762/1764 that are correlated with higher HBV DNA levels. To confirm the effects of these mutations on the virus replication efficiency, substitutions nt 1726-1730 CTGAG and A1762T/G1764A in the HBV X (HBX) gene region were investigated alone or in combination. Cells Huh-7 or HepG2 were transfected with these constructs. The effects of these mutations on HBV were investigated at the gene and protein levels. The double mutation A1762T/G1764A increased whereas the nt 1726-1730 CTGAG mutations decreased the levels of released virion-associated and intracellular HBV DNA. The combined mutations had no appreciable effect on the replication capacity of the virus. Cells bearing the constructs with double mutations A1762T/G1764A contained the lowest levels of hepatitis B e antigen (HBeAg). Lowest expression of HBV X protein was in constructs that had both A1762T/G1764A and 1726-1730 CTGAG mutations. We think that changes in secondary RNA structure that were caused by these mutations might have been responsible for those results.hepatitis B virus; X gene; mutants; replication.