1887

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Volume 77, Issue 7

Processing of human cytomegalovirus glycoprotein B in recombinant adenovirus-infected cells Free

Abstract

Intracellular processing of human cytomegalovirus (HCMV) glycoprotein B (gB; gpUL55) expressed by a recombinant adenovirus (Ad-gB) was studied in human A549 cells as processing events could affect immunogenicity when such viruses are used as live-recombinant vaccines. Cleavage of [S]methionine-labelled gp130 into gp93 and gp55 reached a maximum after a 3 h chase. Cleavage was completely inhibited by brefeldin A, suggesting that processing normally occurs as a late Golgi or post-Golgi event. Uncleaved gp130 remained completely sensitive to endo-β--acetylglucosaminidase H (Endo-H) in untreated cells following long chase periods, indicating high-mannose oligosaccharides at all of the 18 -linked glycosylation sites (-X-Ser/Thr) and retention in the endoplasmic reticulum. Endo-H analysis of gp55 from swainsonine-treated and untreated cells was consistent with glycosylation at all three potential sites, with two oligosaccharides remaining sensitive to Endo-H and one being processed to Endo-H resistance. The heavily glycosylated N-terminal gp93 subunit was not detected by [S]methionine-labelling but was easily detected along with gp55 after labelling with [H]mannose. No cleavage of gp130 was observed in analogous pulse-chase radiolabelling of Ad-gB-infected human fibroblasts, even though these cells are permissive for HCMV replication and can process the native gB molecule. Processing of gB in recombinant adenovirus-infected A549 cells was generally similar to that previously reported for native gB in HCMV-infected fibroblasts.

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© Journal of General Virology 1996
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1996-07-01
2024-04-20
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References

  1. Alford C. A., Britt W. J. 1993; Cytomegalovirus. In The Human Herpesviruses pp 227–255 Edited by Lopez C., Whitley R., Roizman B. New York: Raven Press;
     [Google Scholar]
  2. Alkhatib G., Richardson C., Shen S.-H. 1990; Intracellular processing, glycosylation, and cell-surface expression of the measles virus fusion protein (F) encoded by a recombinant adenovirus. Virology 175:262–270
     [Google Scholar]
  3. Ayata M., Sugano T., Murayama T., Sakamuro D., Takegami T., Matsumoto Y., Furukawa T. 1994; Different antibody response to a neutralizing epitope of human cytomegalovirus glycoprotein B among seropositive individuals. Journal of Medical Virology 43:386–392
     [Google Scholar]
  4. Berkner K. L. 1988; Development of adenovirus vectors for the expression of heterologous genes. Bio Techniques 6:616–629
     [Google Scholar]
  5. Britt W. J., Vugler L. G. 1989; Processing of the gp55–116 envelope glycoprotein complex of human cytomegalovirus. Journal of Virology 63:403–410
     [Google Scholar]
  6. Britt W. J., Vugler L. G. 1992; Oligomerization of the human cytomegalovirus major envelope glycoprotein complex gB (gp55–116). Journal of Virology 66:6747–6754
     [Google Scholar]
  7. Britt W. J., Vugler L., Butfiloski E. J., Stephens E. B. 1990; Cell surface expression of human cytomegalovirus (HCMV) gp55–116 (gB): use of HCMV–recombinant vaccinia virus-infected cells in analysis of the human neutralizing antibody response. Journal of Virology 64:1079–1085
     [Google Scholar]
  8. Chou S. 1992; Comparative analysis of sequence variation in gp116 and gp55 components of glycoprotein B of human cytomegalovirus. Virology 188:388–390
     [Google Scholar]
  9. Chou S., Marousek G. I. 1992; Homology of the envelope glycoprotein B of human herpesvirus-6 and cytomegalovirus. Virology 191:523–528
     [Google Scholar]
  10. Cranage M. P., Kouzarides T., Bankier A. T., Satchwell S., Weston K., Tomlinson P., Barrell B. G., Hart H., Bell S. E., Minson A. C., Smith G. L. 1986; Identification of the human cytomegalovirus glycoprotein B gene and induction of neutralizing antibodies via its expression in recombinant vaccinia virus. EMBO Journal 5:3057–3063
     [Google Scholar]
  11. Darlington J., Super M., Patel K., Grundy J. E., Griffiths P. D., Emery V. C. 1991; Use of the polymerase chain reaction to analyse sequence variation within a major neutralizing epitope of glycoprotein B (gp58) in clinical isolates of human cytomegalovirus. Journal of General Virology 72:1985–1989
     [Google Scholar]
  12. Dewar R. L., Natarajan V., Vasudevachari M. B., Salzman N. P. 1989; Synthesis and processing of human immunodeficiency virus type 1 envelope proteins encoded by a recombinant human adenovirus. Journal of Virology 63:129–136
     [Google Scholar]
  13. Doms R. W., Lamb R. A., Rose J. K., Helenius A. 1993; Folding and assembly of viral membrane proteins. Virology 193:545–562
     [Google Scholar]
  14. Gallichan W. S., Johnson D. C., Graham F. L., Rosenthal K. L. 1993; Mucosal immunity and protection after intranasal immunization with recombinant adenovirus expressing herpes simplex virus glycoprotein B. Journal of Infectious Diseases 168:622–629
     [Google Scholar]
  15. Gething M. J., Milammon K., Sambrook J. 1986; Expression of wild type and mutant forms of influenza hemagglutinin: the role of folding in intracellular transport. Cell 46:939–950
     [Google Scholar]
  16. Gönczöl E., Hudecz F., Lanacone J., Dietzschold B., Starr S., Plotkin S. A. 1986; Immune responses to isolated human cytomegalovirus envelope proteins. Journal of Virology 58:661–664
     [Google Scholar]
  17. Gönczöl E., Lanacone J., Ho W., Starr S., Meignier B., Plotkin S. A. 1990; Isolated gA/gB glycoprotein complex of human cytomegalovirus envelope induces humoral and cellular immune-responses in human volunteers. Vaccine 8:130–136
     [Google Scholar]
  18. Gönczöl E., de Taisne C., Hirka G., Berencsi K., Lin W., Paoletti E., Plotkin S. A. 1991; High expression of human cytomegalovirus (HCMV)-gB protein in cells infected with a vaccinia-gB recombinant: the importance of the gB protein in HCMV immunity. Vaccine 9:631–637
     [Google Scholar]
  19. Griffiths G., Simons K. 1986; The trans Golgi network – sorting at the exit site of the Golgi-complex. Science 234:438–443
     [Google Scholar]
  20. Hubbard S. C. 1988; Regulation of glycosylation: the influence of protein structure on N-linked oligosaccharide processing. Journal of Biological Chemistry 263:19303–19317
     [Google Scholar]
  21. Hunt L. A., Wright S. E. 1981; Rous sarcoma virus glycoproteins contain hybrid-type oligosaccharides. Journal of Virology 39:646–650
     [Google Scholar]
  22. Hunt L. A., Lamph W., Wright S. E. 1981; Transformationdependent alterations in the oligosaccharides of Prague C Rous sarcoma virus. Journal of Virology 37:207–215
     [Google Scholar]
  23. Hunt L. A., Brown D. W., Robinson H. L., Naeve C. W., Webster R.G. 1988; Retrovirus-expressed hemagglutinin protects against lethal influenza virus infections. Journal of Virology 62:3014–3019
     [Google Scholar]
  24. Hutchinson L., Browne H., Wargent V., Davis-Poynter N., Primorac S., Goldsmith K., Minson A. C., Johnson D. C. 1992; A novel herpes simplex virus glycoprotein, gL, forms a complex with glycoprotein H (gH) and affects normal folding and surface expression of gH. Journal of Virology 66:2240–2250
     [Google Scholar]
  25. Johnson D. C., Ghosh-Choudhury G., Smiley J. R., Fallis L., Graham F. L. 1988; Abundant expression of herpes simplex virus glycoprotein gB using an adenovirus vector. Virology 164:1–14
     [Google Scholar]
  26. Kang M. S., Elbein A. D. 1983; Alterations in the structure of the oligosaccharide of vesicular stomatitis virus G protein by swainsonine. Journal of Virology 46:60–69
     [Google Scholar]
  27. Kornfeld R., Kornfeld S. 1985; Assembly of asparagine-linked oligosaccharides. Annual Review of Biochemistry 54:631–664
     [Google Scholar]
  28. Li Y., Luo L., Thomas D. Y., Kang C. Y. 1994; Control of expression, glycosylation, and secretion of HIV-1 gp120 by homologous and heterologous signal sequences. Virology 204:266–278
     [Google Scholar]
  29. Lippincott-Schwartz J., Yuan L. C., Bonifacino J. S., Klausner R. D. 1989; Rapid redistribution of Golgi proteins into the ER in cells treated with brefeldin-A: evidence for membrane cycling from Golgi to ER. Cell 56:801–813
     [Google Scholar]
  30. Machamer C. E., Florkiewicz R. Z., Rose J. K. 1985; A single N- linked oligosaccharide at either of the two normal sites is sufficient for transport of vesicular stomatitis virus G protein to the cell surface. Molecular and Cellular Biology 5:3074–3083
     [Google Scholar]
  31. Marshall G. S., Plotkin S. A. 1993; Cytomegalovirus Vaccines. In The Human Herpesviruses pp 381–395 Edited by Lopez C., Whitley R., Roizman B. New York: Raven Press;
     [Google Scholar]
  32. Marshall G. S., Ricciardi R. P., Rando R. F., Puck J., Ge R., Plotkin S. A., Gonczol E. 1990; An adenovirus recombinant that expresses the human cytomegalovirus major envelope glycoprotein and induces neutralizing antibodies. Journal of Infectious Diseases 162:1177–1181
     [Google Scholar]
  33. Marshall G. S., Rabalais G. P., Stout G. G., Waldeyer S. L. 1992; Antibodies to recombinant-derived glycoprotein B after natural human cytomegalovirus infection correlate with neutralizing activity. Journal of Infectious Diseases 165:381–384
     [Google Scholar]
  34. Marshall G. S., Stout G. G., Knights M., Rabalais G. P., Ashley R., Miller H., Rossier E. 1994; Ontogeny of glycoprotein gB-specific antibody and neutralizing activity during cytomegalovirus infection. Journal of Medical Virology 43:77–83
     [Google Scholar]
  35. Meyer H., Sundqvist V.-A., Pereira L., Mach M. 1992; Glycoprotein gpll6 of human cytomegalovirus contains epitopes for strain-common and strain-specific antibodies. Journal of General Virology 73:2375–2333
     [Google Scholar]
  36. Ohlin M., Sundqvist V.-A., Mach M., Wahren B., Borrebaeck C. A. K. 1993; Fine specificity of the human immune response to the major neutralization epitopes expressed on cytomegalovirus gp58/116 (gB), as determiend with human monoclonal antibodies. Journal of Virology 67:703–710
     [Google Scholar]
  37. Plotkin S. A., Starr S. E., Friedman H. M., Brayman K., Harris S., Jackson S., Tustin N. B., Grossman R., Dafoe D., Barker C. 1991; Effect of Towne live virus vaccine on cytomegalovirus disease after renal transplant. A controlled trial. Annals of Internal Medicine 114:525–531
     [Google Scholar]
  38. Qadri I., Navarro D., Paz P., Pereira L. 1992; Assembly of conformation-dependent neutralizing domains on glycoprotein B of human cytomegalovirus. Journal of General Virology 73:2913–2921
     [Google Scholar]
  39. Rapp M., Messerle M., Buhler B., Tannheimer M., Keil G. M., Koszinowski U. H. 1992; Identification of the murine cytomegalovirus glycoprotein B gene and its expression by recombinant vaccinia virus. Journal of Virology 66:4399–4406
     [Google Scholar]
  40. Rasmussen L., Matkin C., Spaete R., Pachl C., Merigan T. C. 1991; Antibody response to human cytomegalovirus glycoproteins gB and gH after natural infection in humans. Journal of Infectious Diseases 164:835–842
     [Google Scholar]
  41. Reis B., Bogner E., Reschke M., Richter A., Rubin Th., Radsak K. 1993; Stable constitutive expression of glycoprotein B (gpUL55) of human cytomegalovirus in permissive astrocytoma cells. Journal of General Virology 74:1371–1379
     [Google Scholar]
  42. Rubin B. A., Rorke L. B. 1994; Adenovirus Vaccines. In Vaccines pp 475–502 Edited by Plotkin S. A., Mortimer E. A. Philadelphia: W. B. Saunders;
     [Google Scholar]
  43. Spaete R. R., Thayer R. M., Probert W. S., Masiarz F. R., Chamberlain S. H., Rasmussen L., Merigan T. C., Pachl C. 1988; Human cytomegalovirus Towne glycoprotein B is processed by proteolytic cleavage. Virology 167:207–225
     [Google Scholar]
  44. Spaete R. R., Saxena A., Scott P. I., Song G. J., Probert W. S., Britt W. J., Gibson W., Rasmussen L., Pachl C. 1990; Sequence requirements for proteolytic processing of glycoprotein B of human cytomegalovirus strain Towne. Journal of Virology 64:2922–2931
     [Google Scholar]
  45. Tomiyama T., Matsumoto Y.-I., Sugano T., Fujinaga S., Masuho Y. 1990; Preparation of human monoclonal antibodies against a cytomegalovirus glycoprotein complex of 130 and 55 kDa. Journal of Immunological Methods 131:249–255
     [Google Scholar]
  46. Vey M., Schäfer W., Reis B., Ohuchi R., Britt W., Garten W., Klenk H.-D., Radsak K. 1995; Proteolytic processing of human cytomegalovirus glycoprotein B (gpUL55) is mediated by the human endoprotease furin. Virology 206:746–749
     [Google Scholar]
  47. Wagner B., Dropff B., Kalbacher H., Britt W., Sundqvist V.-A., Ostberg L., Mach M. 1992; A continuous sequence of more than 70 amino acids is essential for antibody binding to the dominant antigenic site of glycoprotein gp58 of human cytomegalovirus. Journal of Virology 66:5290–5297
     [Google Scholar]
  48. Whealy M. E., Card J. P., Meade R. P., Robbins A. K., Enquist L. W. 1991; Effect of brefeldin A on alphaherpesvirus membrane protein glycosylation and virus egress. Journal of Virology 65:1066–1081
     [Google Scholar]
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Processing of human cytomegalovirus glycoprotein B in recombinant adenovirus-infected cells
J. Gen. Virol. 77, 1549 (1996); https://doi.org/10.1099/0022-1317-77-7-1549
/content/journal/jgv/10.1099/0022-1317-77-7-1549
/content/journal/jgv/10.1099/0022-1317-77-7-1549
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