Oligomerization of hepatitis delta antigen is required for both the trans-activating and trans-dominant inhibitory activities of the delta antigen.
Two forms of hepatitis delta antigen (HDAg) have different roles in the replication cycle of hepatitis delta virus (HDV); the small forms trans activates HDV RNA replication, whereas the large form suppresses it but is needed for virion assembly. To understand the mechanism of these regulatory activities, we studied the possible HDAg oligomerization and its role in HDV replication. In this report, we provide direct biochemical evidence for the in vitro and in vivo formation of homodimers and heterodimers between these two HDAg species. By deletion mutagenesis, we showed that this protein interaction is mediated by the leucine zipper-like sequence residing in the N-terminal one-third of HDAg. Furthermore, site-specific mutants with various substitutions on two of the leucine residues in this stretch of sequence had reduced or no ability to form HDAg dimers. Correspondingly, the small HDAg with mutations in the leucine zipper-like sequence had reduced abilities to trans activate HDV RNA replication. Similar mutations on the leucine zipper-like sequence of the large HDAg also resulted in loss of the ability of large HDAg to inhibit HDV RNA replication. The in vivo biological activities of both forms of HDAg (trans activation and trans-dominant inhibition of HDV RNA replication, respectively) correlated with the extent of HDAg oligomerization in vitro. Thus, we conclude that the small HDAg participates in HDV RNA replication as an oligomer form and that the large HDAg inhibits HDV RNA replication as a result of its complex formation with small HDAg. A "black sheep" model for the mechanism of trans-dominant inhibition by the large HDAg is presented.
Xia
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Department of Microbiology, University of Southern California, School of Medicine, Los Angeles 90033-1054.
Lai
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D000595:Amino Acid Sequence; D000956:Antigens, Viral / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D001483:Base Sequence; D004252:DNA Mutational Analysis; D003698:Hepatitis Delta Virus / Q000235:genetics*; D035923:Hepatitis delta Antigens; D016350:Leucine Zippers / Q000235:genetics; D008957:Models, Genetic; D008969:Molecular Sequence Data; D011487:Protein Conformation; D015533:Transcriptional Activation / Q000187:drug effects* / Q000235:genetics; D014779:Virus Replication
D000956:Antigens, Viral; D035923:Hepatitis delta Antigens; C442244:hepatitis delta virus large antigen