1887

Abstract

Hepatitis delta virus (HDV) is a defective virus and requires hepatitis B virus (HBV) to supply envelope proteins (HBsAg) for maturation and secretion. It is known that two proteins produced by HDV, the small (SDAg) and large (LDAg) antigens, are located in the nucleolus, speckles and the cytoplasm and are involved in genome replication and virion packaging. However, little is known about how they are targeted to the specific sites where they act. A green fluorescence protein fused to LDAg (GFP–LD) has been shown previously to translocate from the nucleolus to SC-35 speckles in the presence of the casein kinase II inhibitor dichlororibofuranosyl benzimidazole. In this study, we determined which amino acids of GFP–LD were responsible for the translocation from the nucleolus to SC-35 speckles and created three GFP–LD derivatives, GFP–LDS2A, GFP–LDS123A and GFP–LDS2/123A. Fluorescence microscopy studies showed that Ser-123 mutants had a high tendency to target SC-35 speckles in both transfected HeLa and HuH-7 cells and suggested that Ser-123, but not Ser-2, plays a role in modulating LDAg translocation to the nucleolus or to SC-35 speckles. This study also demonstrated that HBsAg plays a role in facilitating the transportation of LDAg from the nucleus to cytoplasm. Compared with GFP–LD and GFP–LDS2A, mutants of Ser-123 were less efficiently transported to the cytoplasm and resulted in a lower level of secretion. In contrast, little or no isoprenylation mutant was observed in the cytoplasm of HuH-7 cells expressing HbsAg, suggesting that the isoprenylation of LDAg plays a role in export from the nucleus. Thus, the current study demonstrated that both and elements modulate HDAg translocation to various subcellular sites.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.19690-0
2004-06-01
2020-01-29
Loading full text...

Full text loading...

/deliver/fulltext/jgv/85/6/vir851685.html?itemId=/content/journal/jgv/10.1099/vir.0.19690-0&mimeType=html&fmt=ahah

References

  1. Bell, P., Brazas, R., Ganem, D. & Maul, G. G. ( 2000; ). Hepatitis delta virus replication generates complexes of large hepatitis delta antigen and antigenomic RNA that affiliate with and alter nuclear domain10. J Virol 74, 5329–5336.[CrossRef]
    [Google Scholar]
  2. Bichko, V. V. & Taylor, J. M. ( 1996; ). Redistribution of the delta antigens in cells replicating the genome of hepatitis delta virus. J Virol 70, 8064–8070.
    [Google Scholar]
  3. Casey, J. L. & Gerin, J. L. ( 1995; ). Hepatitis D virus RNA editing: specific modification of adenosine in the antigenomic RNA. J Virol 69, 7593–7600.
    [Google Scholar]
  4. Chang, J. & Taylor, J. ( 2002; ). In vivo RNA-directed transcription, with template switching, by a mammalian RNA polymerase. EMBO J 21, 157–164.[CrossRef]
    [Google Scholar]
  5. Chang, M.-F., Baker, S. C., Soe, L. H., Kamahora, T., Keck, J. G., Makino, S., Govindarajan, S. & Lai, M. M. C. ( 1988; ). Human hepatitis delta antigen is a nuclear phosphoprotein with RNA-binding activity. J Virol 62, 2403–2410.
    [Google Scholar]
  6. Chang, F.-L., Chen, P.-J., Tu, S.-J., Wang, C.-J. & Chen, D.-S. ( 1991; ). The large form of hepatitis delta antigen is crucial for assembly of hepatitis delta virus. Proc Natl Acad Sci U S A 88, 8490–8494.[CrossRef]
    [Google Scholar]
  7. Chang, M.-F., Chang, S. C., Chang, C.-I., Wu, K. & Kang, H.-Y. ( 1992; ). Nuclear localization signals, but not putative leucine zipper motif, are essential for nuclear transport of hepatitis delta antigen. J Virol 66, 6019–6027.
    [Google Scholar]
  8. Chao, M., Hsieh, S.-Y. & Taylor, J. ( 1990; ). Role of two forms of hepatitis delta antigen: evidence for a mechanism of self-limiting genome replication. J Virol 64, 5066–5069.
    [Google Scholar]
  9. Chen, P.-J., Kalpana, G., Goldberg, J., Mason, W., Werner, B., Gerin, J. & Taylor, J. ( 1986; ). Structure and replication of the genome of the hepatitis delta virus. Proc Natl Acad Sci U S A 83, 8774–8778.[CrossRef]
    [Google Scholar]
  10. Chen, P.-J., Wu, H.-L., Wang, C.-J., Chia, J.-H. & Chen, D.-S. ( 1997; ). Molecular biology of hepatitis D virus: research and potential for application. J Gastroenterol Hepatol 12, 188–192.[CrossRef]
    [Google Scholar]
  11. Fu, T.-B. & Taylor, J. ( 1993; ). The RNAs of hepatitis delta virus are copied by RNA polymerase II in nuclear homogenates. J Virol 67, 6965–6972.
    [Google Scholar]
  12. Glenn, J. S., Watson, J. A., Havel, C. M. & White, J. M. ( 1992; ). Identification of a prenylation site in delta virus large antigen. Science 256, 1331–1333.[CrossRef]
    [Google Scholar]
  13. Graham, E. L. & van der Eb, A. J. ( 1973; ). A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology 52, 456–467.[CrossRef]
    [Google Scholar]
  14. Hu, H.-M., Shih, K.-N. & Lo, S. J. ( 1996; ). Disulfide bond formation of the in vitro-translated large antigen of hepatitis D virus. J Virol Methods 60, 39–46.[CrossRef]
    [Google Scholar]
  15. Huang, W.-H., Yung, B. Y. M., Syu, W. J. & Lee, Y.-H. W. ( 2001; ). The nucleolar phosphoprotein B23 interacts with hepatitis delta antigens and modulates the hepatitis delta virus RNA replication. J Biol Chem 276, 25166–25175.[CrossRef]
    [Google Scholar]
  16. Huovila, A.-P. J., Eder, A. M. & Fuller, S. D. ( 1992; ). Hepatitis B surface antigen assembles in a post-ER, pre-Golgi compartment. J Cell Biol 118, 1305–1320.[CrossRef]
    [Google Scholar]
  17. Hwang, S. B. & Lai, M. M. C. ( 1993; ). Isoprenylation mediates direct protein–protein interactions between hepatitis large delta antigen and hepatitis B surface antigen. J Virol 67, 7659–7662.
    [Google Scholar]
  18. Jayan, G. C. & Casey, J. L. ( 2002; ). Increased RNA editing and inhibition of hepatitis delta virus replication by high-level expression of ADAR1 and ADAR2. J Virol 76, 3819–3827.[CrossRef]
    [Google Scholar]
  19. Kos, A., Dijkema, R., Arnberg, A. C., van de Meide, P. H. & Schelekens, H. ( 1986; ). The hepatitis delta (δ) virus possesses a circular RNA. Nature 323, 558–560.[CrossRef]
    [Google Scholar]
  20. Kuo, M. Y.-P., Chao, M. & Taylor, J. ( 1989; ). Initiation of replication of the human hepatitis delta virus genome from cloned DNA: role of delta antigen. J Virol 63, 1945–1950.
    [Google Scholar]
  21. Lai, M. M. C. ( 1995; ). The molecular biology of hepatitis delta virus. Annu Rev Biochem 64, 259–286.[CrossRef]
    [Google Scholar]
  22. Lee, C.-H., Chang, S. C., Wu, C. H. & Chang, M.-F. ( 2001; ). A novel chromosome region maintenance 1-independent nuclear export signal of the large form hepatitis delta antigen that is required for the viral assembly. J Biol Chem 276, 8142–8148.[CrossRef]
    [Google Scholar]
  23. MacNaughton, T. B., Shi, S. T., Modahl, L. E. & Lai, M. M. C. ( 2002; ). Rolling circle replication of hepatitis delta virus RNA carried out by two different cellular RNA polymerases. J Virol 76, 3920–3927.[CrossRef]
    [Google Scholar]
  24. Meisner, H. & Czech, M. P. ( 1991; ). Phosphorylation of transcriptional factors and cell-cycle-dependent proteins by casein kinase II. Curr Opin Cell Biol 3, 474–483.[CrossRef]
    [Google Scholar]
  25. Modahl, L. E., MacNaughton, T. B., Zhu, N., Johnson, D. L. & Lai, M. M. C. ( 2000; ). RNA-dependent replication and transcription of hepatitis delta virus RNA involve a distinct cellular RNA polymerase. Mol Cell Biol 73, 6030–6039.
    [Google Scholar]
  26. Mu, J.-J., Wu, H.-L., Chiang, B.-L., Chang, R.-P., Chen, D.-S. & Chen, P.-J. ( 1999; ). Characterization of phosphorylated forms and the phosphorylated residues of hepatitis delta virus delta antigens. J Virol 73, 10540–10545.
    [Google Scholar]
  27. Mu, J.-J., Chen, D.-S. & Chen, P.-J. ( 2001; ). The conserved serine 177 in the delta antigen of hepatitis delta virus is one putative phosphorylation site and is required for efficient viral RNA replication. J Virol 75, 9087–9095.[CrossRef]
    [Google Scholar]
  28. Nickel, W. ( 2003; ). The mystery of non-classical protein secretion: a current review on cargo proteins and potential export routes. Eur J Biochem 270, 2109–2119.[CrossRef]
    [Google Scholar]
  29. Rizzetto, M., Canese, M. G., Gerin, J. L., London, W. T., Sly, D. L. & Purcell, R. H. ( 1980; ). Transmission of the hepatitis B virus-associated delta antigen to chimpanzees. J Infect Dis 141, 590–602.[CrossRef]
    [Google Scholar]
  30. Ryu, W. S., Bayer, M. & Taylor, J. ( 1992; ). Assembly of hepatitis delta virus particles. J Virol 66, 2310–2315.
    [Google Scholar]
  31. Sato, S., Wong, S. K. & Lazinski, D. W. ( 2001; ). Hepatitis delta virus minimal substrates competent for editing by ADAR1 and ADAR2. J Virol 75, 8547–8555.[CrossRef]
    [Google Scholar]
  32. Sheu, S. Y. & Lo, S. J. ( 1994; ). Biogenesis of the hepatitis B viral middle (M) surface protein in a human hepatoma cell line: demonstration of an alternative secretion pathway. J Gen Virol 75, 3031–3039.[CrossRef]
    [Google Scholar]
  33. Sheu, S. Y., Chen, K.-L., Lee, Y.-H. W. & Lo, S. J. ( 1996; ). No intermolecular interaction between the large hepatitis delta antigens is required for the secretion with hepatitis B surface antigen: a model of empty HDV particle. Virology 218, 275–278.[CrossRef]
    [Google Scholar]
  34. Shih, K.-N. & Lo, S. J. ( 2001; ). The HDV large-delta antigen fused with GFP remains functional and provides for studying its dynamic distribution. Virology 285, 138–152.[CrossRef]
    [Google Scholar]
  35. Shih, K.-N., Chuang, Y.-T., Liu, H. & Lo, S. J. ( 2004; ). Hepatitis D virus RNA editing is inhibited by a GFP fusion protein containing a C-terminally deleted delta antigen. J Gen Virol 85, 947–957.[CrossRef]
    [Google Scholar]
  36. Stetler, D. A. & Rose, K. M. ( 1982; ). Phosphorylation of deoxyribonucleic acid dependent RNA polymerase II by nuclear protein kinase NII: mechanism of enhanced ribonucleic acid synthesis. Biochemistry 21, 3721–3728.[CrossRef]
    [Google Scholar]
  37. Sureau, C., Fournier-Wirth, C. & Maurel, P. ( 2003; ). Role of N glycosylation of hepatitis B virus envelope proteins in morphogenesis and infectivity of hepatitis delta virus. J Virol 77, 5519–5523.[CrossRef]
    [Google Scholar]
  38. Taylor, J. M. ( 2003; ). Replication of human hepatitis delta virus: recent developments. Trends Microbiol 11, 185–190.[CrossRef]
    [Google Scholar]
  39. Wang, K.-S., Choo, Q.-L., Weiner, A. J. & 7 other authors ( 1986; ). Structure, sequence and expression of the hepatitis delta (δ) viral genome. Nature 323, 508–514.[CrossRef]
    [Google Scholar]
  40. Wang, C.-J., Sung, S.-Y., Chen, D.-S. & Chen, P.-J. ( 1996; ). N-glycosylation of hepatitis B surface antigen is involved but not essential in the assembly of hepatitis delta virus. Virology 220, 28–36.[CrossRef]
    [Google Scholar]
  41. Weiner, A. J., Choo, Q.-L., Wang, K.-S., Govindarajan, S., Redeker, A. G., Gerin, J. L. & Houghton, M. ( 1988; ). A single antigenomic open reading frame of the hepatitis delta virus encodes the epitope(s) of both hepatitis delta antigen polypeptides p24δ and p27δ. J Virol 62, 594–599.
    [Google Scholar]
  42. Wu, J. C., Chen, C.-L., Lee, S.-D., Sheen, I.-J. & Ting, L.-P. ( 1992; ). Expression and localization of the small and large delta antigens during the replication cycle of hepatitis D virus. Hepatology 16, 1120–1127.[CrossRef]
    [Google Scholar]
  43. Xia, Y.-P., Yeh, C.-T., Ou, J.-H. & Lai, M. M. C. ( 1992; ). Characterization of nuclear targeting signal of hepatitis delta antigen: nuclear transport as a protein complex. J Virol 66, 914–921.
    [Google Scholar]
  44. Xu, Z., Jensen, G. & Yen, T. S. ( 1997; ). Activation of hepatitis B virus S promoter by the viral large surface protein via induction of stress in the endoplasmic reticulum. J Virol 71, 7387–7392.
    [Google Scholar]
  45. Yamaguchi, Y., Filipovska, J., Yano, K. & 7 other authors ( 2001; ). Stimulation of RNA polymerase II elongation by hepatitis D antigen. Science 293, 124–127.[CrossRef]
    [Google Scholar]
  46. Yeh, T.-S. & Lee, Y.-H. W. ( 1998; ). Assembly of hepatitis delta virus particles: package of multimeric hepatitis delta virus genomic RNA and the role of phosphorylation. Virology 249, 12–20.[CrossRef]
    [Google Scholar]
  47. Yeh, T.-S., Lo, S. J., Chen, P.-J. & Lee, Y.-H. W. ( 1996; ). Casein kinase II and protein kinase C modulate hepatitis delta virus RNA replication but not empty viral particle assembly. J Virol 70, 6190–6198.
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.19690-0
Loading
/content/journal/jgv/10.1099/vir.0.19690-0
Loading

Data & Media loading...

Most cited articles

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error