Abstract
The biosynthesis and topology of the large envelope protein (L protein) of hepadnaviruses was investigated using the duck hepatitis B virus (DHBV) model, which also allows the study of hepadnavirus morphogenesis in experimentally infected hepatocytes. Results from proteolysis of virus particles and from the analysis of topology and posttranslational modification of L chains synthesized in vivo or in a cell-free system both support the presence of a mixed population of L-protein molecules with their N-terminal pre-S domain located either inside or outside the virus particle. During L biosynthesis and DHBV morphogenesis, pre-S, together with the neighboring transmembrane domain (TM-I), initially remained cytoplasmically disposed and was translocated only posttranslationally. Delayed pre-S translocation into a post-endoplasmic reticulum compartment is also indicated by the absence of glycosylation at a modification-competent pre-S glycosylation site. Major features of L-protein biosynthesis and of the resulting dual topology appear to be conserved between avian and mammalian hepadnaviruses, supporting the model that pre-S domains function in part either as an internal matrix for capsid envelopment or externally as a ligand for cellular receptor binding. However, differences in the mechanisms controlling pre-S translocation were revealed by the results of mutational analyses identifying and characterizing cis-acting determinants in pre-S that delay its cotranslational translocation. Our data from DHBV demonstrate the negative influence of a cluster of positively charged amino acid residues next to TM-I, a motif that is conserved among the avian but absent from mammalian hepadnaviruses. Additional control elements, which are apparently shared between both virus groups and which may serve in chaperone binding, were mapped by deletion analysis in the central part of pre-S.
Full Text
The Full Text of this article is available as a PDF (835.0 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bruss V., Lu X., Thomssen R., Gerlich W. H. Post-translational alterations in transmembrane topology of the hepatitis B virus large envelope protein. EMBO J. 1994 May 15;13(10):2273–2279. doi: 10.1002/j.1460-2075.1994.tb06509.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bruss V., Thomssen R. Mapping a region of the large envelope protein required for hepatitis B virion maturation. J Virol. 1994 Mar;68(3):1643–1650. doi: 10.1128/jvi.68.3.1643-1650.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bruss V., Vieluf K. Functions of the internal pre-S domain of the large surface protein in hepatitis B virus particle morphogenesis. J Virol. 1995 Nov;69(11):6652–6657. doi: 10.1128/jvi.69.11.6652-6657.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Condreay L. D., Aldrich C. E., Coates L., Mason W. S., Wu T. T. Efficient duck hepatitis B virus production by an avian liver tumor cell line. J Virol. 1990 Jul;64(7):3249–3258. doi: 10.1128/jvi.64.7.3249-3258.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Grgacic E. V., Anderson D. A. The large surface protein of duck hepatitis B virus is phosphorylated in the pre-S domain. J Virol. 1994 Nov;68(11):7344–7350. doi: 10.1128/jvi.68.11.7344-7350.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Guo J. T., Pugh J. C. Topology of the large envelope protein of duck hepatitis B virus suggests a mechanism for membrane translocation during particle morphogenesis. J Virol. 1997 Feb;71(2):1107–1114. doi: 10.1128/jvi.71.2.1107-1114.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hartmann E., Rapoport T. A., Lodish H. F. Predicting the orientation of eukaryotic membrane-spanning proteins. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5786–5790. doi: 10.1073/pnas.86.15.5786. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Junker M., Galle P., Schaller H. Expression and replication of the hepatitis B virus genome under foreign promoter control. Nucleic Acids Res. 1987 Dec 23;15(24):10117–10132. doi: 10.1093/nar/15.24.10117. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Klingmüller U., Schaller H. Hepadnavirus infection requires interaction between the viral pre-S domain and a specific hepatocellular receptor. J Virol. 1993 Dec;67(12):7414–7422. doi: 10.1128/jvi.67.12.7414-7422.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lippincott-Schwartz J., Donaldson J. G., Schweizer A., Berger E. G., Hauri H. P., Yuan L. C., Klausner R. D. Microtubule-dependent retrograde transport of proteins into the ER in the presence of brefeldin A suggests an ER recycling pathway. Cell. 1990 Mar 9;60(5):821–836. doi: 10.1016/0092-8674(90)90096-w. [DOI] [PubMed] [Google Scholar]
- Nassal M. Hepatitis B virus morphogenesis. Curr Top Microbiol Immunol. 1996;214:297–337. doi: 10.1007/978-3-642-80145-7_10. [DOI] [PubMed] [Google Scholar]
- Neurath A. R., Kent S. B., Strick N., Parker K. Identification and chemical synthesis of a host cell receptor binding site on hepatitis B virus. Cell. 1986 Aug 1;46(3):429–436. doi: 10.1016/0092-8674(86)90663-x. [DOI] [PubMed] [Google Scholar]
- Nilsson I., von Heijne G. Fine-tuning the topology of a polytopic membrane protein: role of positively and negatively charged amino acids. Cell. 1990 Sep 21;62(6):1135–1141. doi: 10.1016/0092-8674(90)90390-z. [DOI] [PubMed] [Google Scholar]
- Obert S., Zachmann-Brand B., Deindl E., Tucker W., Bartenschlager R., Schaller H. A splice hepadnavirus RNA that is essential for virus replication. EMBO J. 1996 May 15;15(10):2565–2574. [PMC free article] [PubMed] [Google Scholar]
- Ostapchuk P., Hearing P., Ganem D. A dramatic shift in the transmembrane topology of a viral envelope glycoprotein accompanies hepatitis B viral morphogenesis. EMBO J. 1994 Mar 1;13(5):1048–1057. doi: 10.1002/j.1460-2075.1994.tb06353.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Peterson D. L. The structure of hepatitis B surface antigen and its antigenic sites. Bioessays. 1987 Jun;6(6):258–262. doi: 10.1002/bies.950060604. [DOI] [PubMed] [Google Scholar]
- Prange R., Streeck R. E. Novel transmembrane topology of the hepatitis B virus envelope proteins. EMBO J. 1995 Jan 16;14(2):247–256. doi: 10.1002/j.1460-2075.1995.tb06998.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pugh J. C., Di Q., Mason W. S., Simmons H. Susceptibility to duck hepatitis B virus infection is associated with the presence of cell surface receptor sites that efficiently bind viral particles. J Virol. 1995 Aug;69(8):4814–4822. doi: 10.1128/jvi.69.8.4814-4822.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rigg R. J., Schaller H. Duck hepatitis B virus infection of hepatocytes is not dependent on low pH. J Virol. 1992 May;66(5):2829–2836. doi: 10.1128/jvi.66.5.2829-2836.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
- Weber M., Bronsema V., Bartos H., Bosserhoff A., Bartenschlager R., Schaller H. Hepadnavirus P protein utilizes a tyrosine residue in the TP domain to prime reverse transcription. J Virol. 1994 May;68(5):2994–2999. doi: 10.1128/jvi.68.5.2994-2999.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Yuasa S., Cheung R. C., Pham Q., Robinson W. S., Marion P. L. Peptide mapping of neutralizing and nonneutralizing epitopes of duck hepatitis B virus pre-S polypeptide. Virology. 1991 Mar;181(1):14–21. doi: 10.1016/0042-6822(91)90465-n. [DOI] [PubMed] [Google Scholar]
- von Heijne G. Control of topology and mode of assembly of a polytopic membrane protein by positively charged residues. Nature. 1989 Oct 5;341(6241):456–458. doi: 10.1038/341456a0. [DOI] [PubMed] [Google Scholar]