EFFECTS OF MUTATIONS WITHIN AND ADJACENT TO THE TERMINAL REPEATS OF HEPATITIS-B VIRUS PREGENOMIC RNA ON VIRAL-DNA SYNTHESIS

The viral polymerase and several cis-acting sequences are essential fo r hepadnaviral DNA replication, but additional host factors are likely to be involved in this process. We previously identified two sequence s, UBS and DBS (upstream and downstream binding sites), present in mul tiple copies in and adjacent to the pregenomic RNA (pgRNA) terminal re dundancy, that were specifically recognized by a 65-kDa host factor, p 65. The possible roles of these two sequences in hepatitis B virus (HB V) replication were investigated in the context of the intact viral ge nome. UBS is contained within the terminal redundancy of pgRNA, and th e 5' copy of this sequence is essential for viral replication. Mutatio ns within the central core of UBS ablate p65 binding and selectively b lock synthesis of plus-strand DNA, without affecting RNA packaging or minus-strand synthesis. The DBS sequence, which is located downstream of the pgRNA polyadenylation site, overlaps the core (C) protein codin g region. All mutations introduced into this site severely affected vi ral replication. However, these effects were shown to result from domi nant negative effects of mutant core polypeptides rather than from cis -acting effects on RNA recognition. Thus, the 5' UBS but not DBS sites play important cis-acting roles in HBV DNA replication; however, the involvement of p65 in these roles remains a matter for investigation.