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. 1997 Apr;71(4):3178–3187. doi: 10.1128/jvi.71.4.3178-3187.1997

The cytoplasmic and transmembrane domains of the vaccinia virus B5R protein target a chimeric human immunodeficiency virus type 1 glycoprotein to the outer envelope of nascent vaccinia virions.

E Katz 1, E J Wolffe 1, B Moss 1
PMCID: PMC191450  PMID: 9060681

Abstract

The outer envelope of the extracellular form of vaccinia virus (EEV) is derived from the Golgi membrane and contains at least six viral proteins. Transfection studies indicated that the EEV protein encoded by the B5R gene associates with Golgi membranes when synthesized in the absence of other viral products. A domain swapping strategy was then used to investigate the possibility that the B5R protein contains an EEV targeting signal. We constructed chimeric genes encoding the human immunodeficiency virus (HIV) type 1 glycoprotein with the cytoplasmic and transmembrane domains replaced by the corresponding 42-amino-acid C-terminal segment of the B5R protein. Recombinant vaccinia viruses that stably express a chimeric B5R-HIV protein or a control HIV envelope protein with the original cytoplasmic and transmembrane domains were isolated. Cells infected with recombinant vaccinia viruses that expressed either the unmodified or the chimeric HIV envelope protein formed syncytia with cells expressing the CD4 receptor for HIV. However, biochemical and microscopic studies demonstrated that the HIV envelope proteins with the B5R cytoplasmic and transmembrane domains were preferentially targeted to the EEV. These data are consistent with the presence of EEV localization signals in the cytoplasmic and transmembrane domains of the B5R protein.

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Selected References

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  1. Ashorn P. A., Berger E. A., Moss B. Human immunodeficiency virus envelope glycoprotein/CD4-mediated fusion of nonprimate cells with human cells. J Virol. 1990 May;64(5):2149–2156. doi: 10.1128/jvi.64.5.2149-2156.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blasco R., Moss B. Extracellular vaccinia virus formation and cell-to-cell virus transmission are prevented by deletion of the gene encoding the 37,000-Dalton outer envelope protein. J Virol. 1991 Nov;65(11):5910–5920. doi: 10.1128/jvi.65.11.5910-5920.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blasco R., Moss B. Role of cell-associated enveloped vaccinia virus in cell-to-cell spread. J Virol. 1992 Jul;66(7):4170–4179. doi: 10.1128/jvi.66.7.4170-4179.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blasco R., Sisler J. R., Moss B. Dissociation of progeny vaccinia virus from the cell membrane is regulated by a viral envelope glycoprotein: effect of a point mutation in the lectin homology domain of the A34R gene. J Virol. 1993 Jun;67(6):3319–3325. doi: 10.1128/jvi.67.6.3319-3325.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Broder C. C., Dimitrov D. S., Blumenthal R., Berger E. A. The block to HIV-1 envelope glycoprotein-mediated membrane fusion in animal cells expressing human CD4 can be overcome by a human cell component(s). Virology. 1993 Mar;193(1):483–491. doi: 10.1006/viro.1993.1151. [DOI] [PubMed] [Google Scholar]
  6. Brown C. K., Turner P. C., Moyer R. W. Molecular characterization of the vaccinia virus hemagglutinin gene. J Virol. 1991 Jul;65(7):3598–3606. doi: 10.1128/jvi.65.7.3598-3606.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chakrabarti S., Brechling K., Moss B. Vaccinia virus expression vector: coexpression of beta-galactosidase provides visual screening of recombinant virus plaques. Mol Cell Biol. 1985 Dec;5(12):3403–3409. doi: 10.1128/mcb.5.12.3403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chakrabarti S., Robert-Guroff M., Wong-Staal F., Gallo R. C., Moss B. Expression of the HTLV-III envelope gene by a recombinant vaccinia virus. Nature. 1986 Apr 10;320(6062):535–537. doi: 10.1038/320535a0. [DOI] [PubMed] [Google Scholar]
  9. Cooney E. L., Collier A. C., Greenberg P. D., Coombs R. W., Zarling J., Arditti D. E., Hoffman M. C., Hu S. L., Corey L. Safety of and immunological response to a recombinant vaccinia virus vaccine expressing HIV envelope glycoprotein. Lancet. 1991 Mar 9;337(8741):567–572. doi: 10.1016/0140-6736(91)91636-9. [DOI] [PubMed] [Google Scholar]
  10. Cudmore S., Cossart P., Griffiths G., Way M. Actin-based motility of vaccinia virus. Nature. 1995 Dec 7;378(6557):636–638. doi: 10.1038/378636a0. [DOI] [PubMed] [Google Scholar]
  11. DALES S., SIMINOVITCH L. The development of vaccinia virus in Earle's L strain cells as examined by electron microscopy. J Biophys Biochem Cytol. 1961 Aug;10:475–503. doi: 10.1083/jcb.10.4.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. DALES S. The uptake and development of vaccinia virus in strain L cells followed with labeled viral deoxyribonucleic acid. J Cell Biol. 1963 Jul;18:51–72. doi: 10.1083/jcb.18.1.51. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Doms R. W., Blumenthal R., Moss B. Fusion of intra- and extracellular forms of vaccinia virus with the cell membrane. J Virol. 1990 Oct;64(10):4884–4892. doi: 10.1128/jvi.64.10.4884-4892.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Doms R. W., Lamb R. A., Rose J. K., Helenius A. Folding and assembly of viral membrane proteins. Virology. 1993 Apr;193(2):545–562. doi: 10.1006/viro.1993.1164. [DOI] [PubMed] [Google Scholar]
  15. Duncan S. A., Smith G. L. Identification and characterization of an extracellular envelope glycoprotein affecting vaccinia virus egress. J Virol. 1992 Mar;66(3):1610–1621. doi: 10.1128/jvi.66.3.1610-1621.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Earl P. L., Broder C. C., Long D., Lee S. A., Peterson J., Chakrabarti S., Doms R. W., Moss B. Native oligomeric human immunodeficiency virus type 1 envelope glycoprotein elicits diverse monoclonal antibody reactivities. J Virol. 1994 May;68(5):3015–3026. doi: 10.1128/jvi.68.5.3015-3026.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Earl P. L., Hügin A. W., Moss B. Removal of cryptic poxvirus transcription termination signals from the human immunodeficiency virus type 1 envelope gene enhances expression and immunogenicity of a recombinant vaccinia virus. J Virol. 1990 May;64(5):2448–2451. doi: 10.1128/jvi.64.5.2448-2451.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Earl P. L., Koenig S., Moss B. Biological and immunological properties of human immunodeficiency virus type 1 envelope glycoprotein: analysis of proteins with truncations and deletions expressed by recombinant vaccinia viruses. J Virol. 1991 Jan;65(1):31–41. doi: 10.1128/jvi.65.1.31-41.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Earl P. L., Moss B. Mutational analysis of the assembly domain of the HIV-1 envelope glycoprotein. AIDS Res Hum Retroviruses. 1993 Jul;9(7):589–594. doi: 10.1089/aid.1993.9.589. [DOI] [PubMed] [Google Scholar]
  20. Engelstad M., Howard S. T., Smith G. L. A constitutively expressed vaccinia gene encodes a 42-kDa glycoprotein related to complement control factors that forms part of the extracellular virus envelope. Virology. 1992 Jun;188(2):801–810. doi: 10.1016/0042-6822(92)90535-w. [DOI] [PubMed] [Google Scholar]
  21. Engelstad M., Smith G. L. The vaccinia virus 42-kDa envelope protein is required for the envelopment and egress of extracellular virus and for virus virulence. Virology. 1993 Jun;194(2):627–637. doi: 10.1006/viro.1993.1302. [DOI] [PubMed] [Google Scholar]
  22. Gallaher W. R., Ball J. M., Garry R. F., Martin-Amedee A. M., Montelaro R. C. A general model for the surface glycoproteins of HIV and other retroviruses. AIDS Res Hum Retroviruses. 1995 Feb;11(2):191–202. doi: 10.1089/aid.1995.11.191. [DOI] [PubMed] [Google Scholar]
  23. Gerhard W., Yewdell J., Frankel M. E., Webster R. Antigenic structure of influenza virus haemagglutinin defined by hybridoma antibodies. Nature. 1981 Apr 23;290(5808):713–717. doi: 10.1038/290713a0. [DOI] [PubMed] [Google Scholar]
  24. Gong S. C., Lai C. F., Esteban M. Vaccinia virus induces cell fusion at acid pH and this activity is mediated by the N-terminus of the 14-kDa virus envelope protein. Virology. 1990 Sep;178(1):81–91. doi: 10.1016/0042-6822(90)90381-z. [DOI] [PubMed] [Google Scholar]
  25. Graham R. C., Jr, Karnovsky M. J. Glomerular permeability. Ultrastructural cytochemical studies using peroxidases as protein tracers. J Exp Med. 1966 Dec 1;124(6):1123–1134. doi: 10.1084/jem.124.6.1123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Green S., Issemann I., Sheer E. A versatile in vivo and in vitro eukaryotic expression vector for protein engineering. Nucleic Acids Res. 1988 Jan 11;16(1):369–369. doi: 10.1093/nar/16.1.369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Hiller G., Eibl H., Weber K. Characterization of intracellular and extracellular vaccinia virus variants: N1-isonicotinoyl-N2-3-methyl-4-chlorobenzoylhydrazine interferes with cytoplasmic virus dissemination and release. J Virol. 1981 Sep;39(3):903–913. doi: 10.1128/jvi.39.3.903-913.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Hiller G., Jungwirth C., Weber K. Fluorescence microscopical analysis of the life cycle of vaccinia virus in chick embryo fibroblasts. Virus-cytoskeleton interactions. Exp Cell Res. 1981 Mar;132(1):81–87. doi: 10.1016/0014-4827(81)90085-9. [DOI] [PubMed] [Google Scholar]
  29. Hiller G., Weber K. Golgi-derived membranes that contain an acylated viral polypeptide are used for vaccinia virus envelopment. J Virol. 1985 Sep;55(3):651–659. doi: 10.1128/jvi.55.3.651-659.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Hirt P., Hiller G., Wittek R. Localization and fine structure of a vaccinia virus gene encoding an envelope antigen. J Virol. 1986 Jun;58(3):757–764. doi: 10.1128/jvi.58.3.757-764.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Isaacs S. N., Wolffe E. J., Payne L. G., Moss B. Characterization of a vaccinia virus-encoded 42-kilodalton class I membrane glycoprotein component of the extracellular virus envelope. J Virol. 1992 Dec;66(12):7217–7224. doi: 10.1128/jvi.66.12.7217-7224.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Krijnse-Locker J., Schleich S., Rodriguez D., Goud B., Snijder E. J., Griffiths G. The role of a 21-kDa viral membrane protein in the assembly of vaccinia virus from the intermediate compartment. J Biol Chem. 1996 Jun 21;271(25):14950–14958. doi: 10.1074/jbc.271.25.14950. [DOI] [PubMed] [Google Scholar]
  33. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  34. Lai C. F., Gong S. C., Esteban M. Structural and functional properties of the 14-kDa envelope protein of vaccinia virus synthesized in Escherichia coli. J Biol Chem. 1990 Dec 25;265(36):22174–22180. [PubMed] [Google Scholar]
  35. Lifson J. D., Feinberg M. B., Reyes G. R., Rabin L., Banapour B., Chakrabarti S., Moss B., Wong-Staal F., Steimer K. S., Engleman E. G. Induction of CD4-dependent cell fusion by the HTLV-III/LAV envelope glycoprotein. Nature. 1986 Oct 23;323(6090):725–728. doi: 10.1038/323725a0. [DOI] [PubMed] [Google Scholar]
  36. Maa J. S., Rodriguez J. F., Esteban M. Structural and functional characterization of a cell surface binding protein of vaccinia virus. J Biol Chem. 1990 Jan 25;265(3):1569–1577. [PubMed] [Google Scholar]
  37. Machamer C. E. Targeting and retention of Golgi membrane proteins. Curr Opin Cell Biol. 1993 Aug;5(4):606–612. doi: 10.1016/0955-0674(93)90129-E. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. McIntosh A. A., Smith G. L. Vaccinia virus glycoprotein A34R is required for infectivity of extracellular enveloped virus. J Virol. 1996 Jan;70(1):272–281. doi: 10.1128/jvi.70.1.272-281.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Morgan C. Vaccinia virus reexamined: development and release. Virology. 1976 Aug;73(1):43–58. doi: 10.1016/0042-6822(76)90059-3. [DOI] [PubMed] [Google Scholar]
  40. Niles E. G., Seto J. Vaccinia virus gene D8 encodes a virion transmembrane protein. J Virol. 1988 Oct;62(10):3772–3778. doi: 10.1128/jvi.62.10.3772-3778.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Parkinson J. E., Smith G. L. Vaccinia virus gene A36R encodes a M(r) 43-50 K protein on the surface of extracellular enveloped virus. Virology. 1994 Oct;204(1):376–390. doi: 10.1006/viro.1994.1542. [DOI] [PubMed] [Google Scholar]
  42. Payne L. G. Characterization of vaccinia virus glycoproteins by monoclonal antibody precipitation. Virology. 1992 Mar;187(1):251–260. doi: 10.1016/0042-6822(92)90313-e. [DOI] [PubMed] [Google Scholar]
  43. Payne L. G. Significance of extracellular enveloped virus in the in vitro and in vivo dissemination of vaccinia. J Gen Virol. 1980 Sep;50(1):89–100. doi: 10.1099/0022-1317-50-1-89. [DOI] [PubMed] [Google Scholar]
  44. Payne L. Polypeptide composition of extracellular enveloped vaccinia virus. J Virol. 1978 Jul;27(1):28–37. doi: 10.1128/jvi.27.1.28-37.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Ratner L., Haseltine W., Patarca R., Livak K. J., Starcich B., Josephs S. F., Doran E. R., Rafalski J. A., Whitehorn E. A., Baumeister K. Complete nucleotide sequence of the AIDS virus, HTLV-III. Nature. 1985 Jan 24;313(6000):277–284. doi: 10.1038/313277a0. [DOI] [PubMed] [Google Scholar]
  46. Ravanello M. P., Hruby D. E. Characterization of the vaccinia virus L1R myristylprotein as a component of the intracellular virion envelope. J Gen Virol. 1994 Jun;75(Pt 6):1479–1483. doi: 10.1099/0022-1317-75-6-1479. [DOI] [PubMed] [Google Scholar]
  47. Ravanello M. P., Hruby D. E. Conditional lethal expression of the vaccinia virus L1R myristylated protein reveals a role in virion assembly. J Virol. 1994 Oct;68(10):6401–6410. doi: 10.1128/jvi.68.10.6401-6410.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Rodriguez J. F., Paez E., Esteban M. A 14,000-Mr envelope protein of vaccinia virus is involved in cell fusion and forms covalently linked trimers. J Virol. 1987 Feb;61(2):395–404. doi: 10.1128/jvi.61.2.395-404.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Rodríguez D., Esteban M., Rodríguez J. R. Vaccinia virus A17L gene product is essential for an early step in virion morphogenesis. J Virol. 1995 Aug;69(8):4640–4648. doi: 10.1128/jvi.69.8.4640-4648.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Roper R. L., Payne L. G., Moss B. Extracellular vaccinia virus envelope glycoprotein encoded by the A33R gene. J Virol. 1996 Jun;70(6):3753–3762. doi: 10.1128/jvi.70.6.3753-3762.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Sarov I., Joklik W. K. Studies on the nature and location of the capsid polypeptides of vaccinia virions. Virology. 1972 Nov;50(2):579–592. doi: 10.1016/0042-6822(72)90409-6. [DOI] [PubMed] [Google Scholar]
  52. Schmelz M., Sodeik B., Ericsson M., Wolffe E. J., Shida H., Hiller G., Griffiths G. Assembly of vaccinia virus: the second wrapping cisterna is derived from the trans Golgi network. J Virol. 1994 Jan;68(1):130–147. doi: 10.1128/jvi.68.1.130-147.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Schmutz C., Payne L. G., Gubser J., Wittek R. A mutation in the gene encoding the vaccinia virus 37,000-M(r) protein confers resistance to an inhibitor of virus envelopment and release. J Virol. 1991 Jul;65(7):3435–3442. doi: 10.1128/jvi.65.7.3435-3442.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Schmutz C., Rindisbacher L., Galmiche M. C., Wittek R. Biochemical analysis of the major vaccinia virus envelope antigen. Virology. 1995 Oct 20;213(1):19–27. doi: 10.1006/viro.1995.1542. [DOI] [PubMed] [Google Scholar]
  55. Shida H. Nucleotide sequence of the vaccinia virus hemagglutinin gene. Virology. 1986 Apr 30;150(2):451–462. doi: 10.1016/0042-6822(86)90309-0. [DOI] [PubMed] [Google Scholar]
  56. Sodeik B., Cudmore S., Ericsson M., Esteban M., Niles E. G., Griffiths G. Assembly of vaccinia virus: incorporation of p14 and p32 into the membrane of the intracellular mature virus. J Virol. 1995 Jun;69(6):3560–3574. doi: 10.1128/jvi.69.6.3560-3574.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Sodeik B., Doms R. W., Ericsson M., Hiller G., Machamer C. E., van 't Hof W., van Meer G., Moss B., Griffiths G. Assembly of vaccinia virus: role of the intermediate compartment between the endoplasmic reticulum and the Golgi stacks. J Cell Biol. 1993 May;121(3):521–541. doi: 10.1083/jcb.121.3.521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Sodeik B., Griffiths G., Ericsson M., Moss B., Doms R. W. Assembly of vaccinia virus: effects of rifampin on the intracellular distribution of viral protein p65. J Virol. 1994 Feb;68(2):1103–1114. doi: 10.1128/jvi.68.2.1103-1114.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Stokes G. V. High-voltage electron microscope study of the release of vaccinia virus from whole cells. J Virol. 1976 May;18(2):636–643. doi: 10.1128/jvi.18.2.636-643.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Tooze J., Hollinshead M., Reis B., Radsak K., Kern H. Progeny vaccinia and human cytomegalovirus particles utilize early endosomal cisternae for their envelopes. Eur J Cell Biol. 1993 Feb;60(1):163–178. [PubMed] [Google Scholar]
  61. Wolffe E. J., Isaacs S. N., Moss B. Deletion of the vaccinia virus B5R gene encoding a 42-kilodalton membrane glycoprotein inhibits extracellular virus envelope formation and dissemination. J Virol. 1993 Aug;67(8):4732–4741. doi: 10.1128/jvi.67.8.4732-4741.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Wolffe E. J., Moore D. M., Peters P. J., Moss B. Vaccinia virus A17L open reading frame encodes an essential component of nascent viral membranes that is required to initiate morphogenesis. J Virol. 1996 May;70(5):2797–2808. doi: 10.1128/jvi.70.5.2797-2808.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  63. Wolffe E. J., Vijaya S., Moss B. A myristylated membrane protein encoded by the vaccinia virus L1R open reading frame is the target of potent neutralizing monoclonal antibodies. Virology. 1995 Aug 1;211(1):53–63. doi: 10.1006/viro.1995.1378. [DOI] [PubMed] [Google Scholar]

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