In vivo selection of a hepatitis B virus mutant with abnormal viral protein expression

We have investigated the molecular basis for the in vivo selective advantage of a hepatitis B virus (HBV) mutant. We have determined the complete nucleotide sequences of the major HBV forms identified at the beginning (B1-83) and end (B1-89) of a 6 year follow-up of a chronically infected patient. The B1-89 sequence showed marked nucleotide rearrangements (a nucleotide divergence of 11.3 % compared with the adw2 subtype), but sequence comparison showed that both viral molecules were of common origin (62/138 mutations were found on both molecules, compared to adw2). In vitro transfection of Huh7 cells showed important modifications in B1-89 viral protein expression. We observed a decrease in B1-89 envelope protein expression associated with a modification of the migration pattern of the large envelope protein. For the B1-89 capsid protein, an insertion of 36 nucleotides at the 5' end of the C gene resulted in increased expression of a core-specific protein of abnormal size (24 kDa versus 22 kDa). Finally, our data also suggest an increase in the trans-complementation efficiency of the mutated B1-89 polymerase protein. Thus, we were able to demonstrate distinct intrinsic properties of HBV DNA molecules isolated from a chronic carrier with virus multiplication at different times during infection. Modifications of viral protein expression in the mutated form illustrate strategies used by the virus to prevent clearance and to contribute to viral persistence.