Hepatitis 6 virus (HDV) nucleotide 1012 is edited from uridine to cytidine in 10-40% of the RNA gonomes during replication. This editing event is an important control point in the HDV life cycle because it results in both the packaging of viral RNA and the inhibition of HDV replication. We find that the editing event is highly specific for both the sequences neighboring nucleotide 1012 and the base-paired context of position 1012 within the unbranched rod structure of HDV RNA. Prior studies identified the base transition at nucleotide 1012 but were unable to distinguish between editing of the genomic versus the antigenomic strands [Luo, G. X., Chao, M., Hsieh, S. Y., Sureau, C., Nishikura, K. & Taylor, J. (1990) J. Virol. 64, 1021-1027. In this study, comparisons of mutations that differentiate between base pairing in genomic and antigenomic RNAs indicate that the genomic strand of HDV is the actual editing substrate. We conclude that the virus uses a uridine to cytidine editing mechanism, which is provided by the host cell.Hepatitis 6 virus (HDV) is a subviral pathogen of humans. It requires concurrent infection with hepatitis B, which provides the viral coat protein (1, 2). Compared to infection with hepatitis B virus alone, coinfection or superinfection with HDV significantly increases the risk of more severe liver disease, including fulminant hepatitis (3). The HDV genome is an -1.7-kilobase, single-stranded, circular RNA molecule, which shares some unusual features with plant viroid RNAs (reviewed in ref. 4). The circular RNA molecule possesses significant intramolecular complementarity such that 70%o of the nucleotides can form base pairs in an unbranched rod structure (5, 6). Consistent with a rolling circle replication mechanism similar to that of the plant viroid agents, monomers and multimers of both genomic and antigenomic RNAs are found in infected and transfected cells (7-9). Unlike the viroids, HDV produces a single protein, the hepatitis 8 antigen (HDAg), which is translated from an antigenomic sense mRNA (9, 10). HDAg is an RNA binding protein (11-13) and is found as a mixture of 24-and 27-kDa species in both infected cells and virions (14, 15). The two forms of HDAg play central roles in the HDV replication cycle. p24 is necessary for HDV RNA replication in transfected cells in culture (8), while p27 both inhibits replication (16, 17) and enables packaging of the HDV RNA genome (18). As shown by others (19), the heterogeneity of HDAg likely arises via an RNA editing mechanism wherein the antigenomically encoded stop codon for p24 is changed from UAG to UGG; the resultant translation product, p27, contains an additional 19 amino acids at the C terminus. Because of the circular replication cycle of HDV RNA, the base change was found in both the genomic and antigenomic RNAs (19); it was therefore unclear whether the genomic or the antigenomic RNA is the actual substrate for editing. An A-to-G transition in the antigenomic strand, possibly involving a host doublestranded RNA modifying activi...

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Chimpanzees that were chronic hepatitis B virus carriers and that were superinfected with hepatitis D virus (HDV) apparently developed acute, self-limited type D hepatitis. Reevaluation with a sensitive hybridization-based assay using RNA probes specific for the HDV genome, however, demonstrated that greater than 50% of these animals still had detectable signs of ongoing HDV replication an average of 2.4 y (range, 1-6.4 y) after inoculation. The only positive marker for the presence of HDV was serum HDV RNA; HDV antigen was undetectable in both serum and the liver by an immunoblot assay and immunofluorescence, respectively. Because the detected amount of viral genome was very low, the previous failure to identify the chronic HDV carrier state in the chimpanzee can be attributed to the lower sensitivity of previously described assays.

Peripheral blood lymphocytes from a patient chronically infected with hepatitis D virus (HDV) were immortalized by Epstein-Barr virus transformation. Two stable monoclonal cell lines, derived from the same parent culture, were established and produced antibodies of the IgG isotype that were specific for the hepatitis delta antigen (HDAg). Both monoclonal antibodies (MAbs) recognized the major HDAg polypeptides of 24 kilodaltons and 27 kilodaltons that were previously detected by polyclonal antibodies to HDAg in both liver and serum from HDV-infected humans, chimpanzees, and woodchucks. This result indicates that the major polypeptides of HDAg share common epitopes. The MAbs also reacted with minor polypeptides of lower molecular weight, which were present in infected liver. In vitro translation products of HDV-specific RNA from infected liver were also detected by the MAbs; these polypeptides were 24 kilodaltons and 27 kilodaltons, respectively, and comigrated with liver- or serum-derived HDAg. In contrast, HDV RNA isolated from virions in serum was not translated into HDAg polypeptides in the in vitro system.

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