TRANSCRIPTION OF HEPATITIS-DELTA ANTIGEN MESSENGER-RNA CONTINUES THROUGHOUT HEPATITIS-DELTA VIRUS (HDV) REPLICATION - A NEW MODEL OF HDV RNA-TRANSCRIPTION AND REPLICATION

Hepatitis delta virus (HDV) replicates by RNA-dependent RNA synthesis according to a double rolling circle model. Also synthesized during re plication is a 0.8-kb, polyadenylated mRNA encoding the hepatitis delt a antigen (HDAg), It has been proposed that this mRNA species represen ts the initial product of HDV RNA replication; subsequent production o f genomic-length HDV RNA relies on suppression of the HDV RNA polyaden ylation signal by HDAg. However, this model was based on studies which required the use of an HDV cDNA copy to initiate HDV RNA replication in cell culture, thus introducing an artificial requirement for DNA-de pendent RNA synthesis. We have now used an HDV cDNA-free RNA transfect ion system and a method that we developed to detect specifically the m RNA species transcribed from the HDV RNA template. We established that this polyadenylated mRNA is 0.8 kb in length and its 5' end begins at nucleotide 1631. Surprisingly, kinetic studies showed that this mRNA continued to be synthesized even late in the viral replication cycle a nd that the mRNA and the genomic-length RNA increased in parallel, eve n in the presence of HDAg, Thus, a switch from production of the HDAg mRNA to the full-length HDV RNA does not occur in this system, and sup pression of the polyadenylation site by HDAg may not significantly reg ulate the synthesis of the HDAg mRNA, as previously proposed. These fi ndings reveal novel insights into the mechanism of HDV RNA replication . A new model of HDV RNA replication and transcription is proposed.