Molecular determinants within the C-termini of L-HDAg that regulate hepatitis D virus replication and assembly
Hongbo Guo,
Qiudi Li,
Chunyang Li,
Yao Hou,
Yibo Ding,
Dan Liu,
Yi Ni,
Renxian Tang,
Kuiyang Zheng,
Stephan Urban,
Wenshi Wang
Affiliations
Hongbo Guo
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China; Corresponding authors. Address: No. 209 Tongshan Road, Xuzhou Medical University, 221004, Xuzhou, China. Tel.: +86(0)516-83262124.
Qiudi Li
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China
Chunyang Li
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China
Yao Hou
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China
Yibo Ding
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China
Dan Liu
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China
Yi Ni
Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany; German Centre for Infection Research (DZIF), partner site Heidelberg, Heidelberg, Germany
Renxian Tang
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China
Kuiyang Zheng
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China
Stephan Urban
Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany; German Centre for Infection Research (DZIF), partner site Heidelberg, Heidelberg, Germany
Wenshi Wang
Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, China; Corresponding authors. Address: No. 209 Tongshan Road, Xuzhou Medical University, 221004, Xuzhou, China. Tel.: +86(0)516-83262124.
Background & Aims: Hepatitis D virus (HDV) is the causative agent of chronic hepatitis delta, the most severe form of viral hepatitis. HDV encodes one protein, hepatitis delta antigen (HDAg), in two isoforms: S- and L-HDAg. They are identical in sequence except that L-HDAg contains an additional 19-20 amino acids at its C-terminus, which confer regulatory roles that are distinct from those of S-HDAg. Notably, these residues are divergent between different genotypes. We aimed to elucidate the molecular determinants within the C-termini that are essential for the regulatory role of L-HDAg in HDV replication and assembly. Methods: Northern blot, reverse-transcription quantitative PCR, and a newly established HDV trans-complementary system were used in this study. Results: C-termini of L-HDAg, albeit with high sequence variation among different genotypes, are interchangeable with respect to the trans-inhibitory function of L-HDAg and HDV assembly. The C-terminus of L-HDAg features a conserved prenylation CXXQ motif and is enriched with proline and hydrophobic residues. Abolishment of the CXXQ motif attenuated the inhibitory effect of L-HDAg on HDV replication. In contrast, the enrichment of proline and hydrophobic residues per se does not modify the trans-inhibitory function of L-HDAg. Nevertheless, these residues are essential for HDV assembly. Mechanistically, prolines and hydrophobic residues contribute to HDV assembly via a mode of action independent of the prenylated CXXQ motif. Conclusions: Within the C-terminus of L-HDAg, the CXXQ motif and the enrichment of proline and hydrophobic residues are all essential determinants of L-HDAg’s regulatory roles in HDV replication and assembly. This intrinsic viral regulatory mechanism we elucidated deepens our understanding of the unique life cycle of HDV. Impact and implications: Hepatitis D virus (HDV) encodes one protein, hepatitis delta antigen (HDAg), in two isoforms: S- and L-HDAg. They are identical in sequence except that L-HDAg contains an additional 19-20 amino acids at its C-terminus. This C-terminal extension in L-HDAg confers regulatory roles in the HDV life cycle that are distinct from those of S-HDAg. Herein, we found that C-termini of L-HDAg, although with high sequence variation, are interchangeable among different HDV genotypes. Within the C-terminus of L-HDAg, the prenylation motif, and the enrichment of proline and hydrophobic residues are all essential determinants of L-HDAg’s regulatory roles in HDV replication and assembly.
