Skip to main content
PMC home page
Elsevier - PMC COVID-19 Collection logoLink to Elsevier - PMC COVID-19 Collection
. 2004 Feb 9;169(2):354–364. doi: 10.1016/0042-6822(89)90161-X

Maturation of Japanese encephalitis virus glycoproteins produced by infected mammalian and mosquito cells

Peter W Mason 1,1
PMCID: PMC7125691  PMID: 2523178

Abstract

The Japanese encephalitis virus (JE) structural glycoprotein (E) and two nonstructural glycoproteins (NS1 and NS1′) were processed differently by JE-infected vertebrate and invertebrate cell lines. All three proteins were released slowly (t12 > 6 hr) from JE-infected monkey cells (Vero cells). Mosquito cell lines released E at a similar rate (t12 > 8 hr), while NS1 and NS1′ were retained in an undegraded form in the cell layer. The proteolytic processing of the three proteins appeared identical in both cell types, but some differences in Winked glycosylation were observed. E, NS1, and NS1′ found within the infected cells of both types contained high-mannose oligosaccharide groups for more than 8 hr after synthesis. Additional sugar residues were added to the single E protein oligosaccharide group prior to release from Vero cells, while sugar residues were trimmed from the E protein oligosaccharide group prior to release from mosquito cells. The forms of NS1 and NS1′ found in the culture fluid of infected Vero cells contained one complex and one high-mannose oligosaccharide. All three glycoproteins released from JE-infected Vero cells were associated with extracellular particles, the virion in the case of E and a low density particle in the case of NS1′ and NSV. Furthermore, E, NS1′ and NS1′ exhibited amphipathic properties in Triton X-114 extraction experiments. Taken together, these results suggest that both the structural (E) and nonstructural NS1′ and NSV) glycoproteins were accumulated within the secretory pathway of the infected Vero cells, assembled into particles, and then released into the extracellular fluid.

References

  1. Adams G.A., Rose J.K. Structural requirements of a membrane-spanning domain for protein anchoring and cell surface transport. Cell. 1985;41:1007–1015. doi: 10.1016/s0092-8674(85)80081-7. [DOI] [PubMed] [Google Scholar]
  2. Bangs J.D., Andrews N.W., hart G.W., Englund P.T. Posttranslational modification and intracellular transport of a trypanosome variant surface glycoprotein. J. Cell Biol. 1986;103:255–263. doi: 10.1083/jcb.103.1.255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bordier C. Phase separation of integral membrane proteins in Triton X-1 14 solution. J. Biol. Chem. 1981;256:1604–1607. [PubMed] [Google Scholar]
  4. Chen S.S.-L., Huang A.S. Further characterization of the vesicular stomatitis virus temperature-sensitive 045 mutant: Intracellular conversion of the glycoprotein to a soluble form. J. Virol. 1986;59:210–215. doi: 10.1128/jvi.59.2.210-215.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Deubel V., Schlesinger J.J., Digoutte J.-P., Girard M. Comparative immunochemical and biological analysis of African and South American yellow fever viruses. Arch. Virol. 1987;94:331–338. doi: 10.1007/BF01310727. [DOI] [PubMed] [Google Scholar]
  6. Gould E.A., Buckley A., Barrett A.D.T., Cammack N. Neutralizing (54K) and non-neutralizing (54K and 48K) monoclonal antibodies against structural and non-structural yellow fever virus proteins confer immunity in mice. J. Gen. Virol. 1986;67:591–595. doi: 10.1099/0022-1317-67-3-591. [DOI] [PubMed] [Google Scholar]
  7. Hase T., Summers P.L., Eckels K.H., Baze W.B. An electron and immunoelectron microscope study of dengue-2 virus infection of cultured mosquito cells: Maturation events. Arch. Virol. 1987;92:273–291. doi: 10.1007/BF01317484. [DOI] [PubMed] [Google Scholar]
  8. Hase T., Summers P.L., Eckels K.H., Baze W.B. Maturation process of Japanese encephalitis virus in cultured mosquito cells in vitro and mouse brain cells in vivo. Arch. Virol. 1987;96:135–151. doi: 10.1007/BF01320956. [DOI] [PubMed] [Google Scholar]
  9. Helenius A., Simons K. Solubilization of membranes by detergents. Biochim. Biophys. Acta. 1975;415:29–79. doi: 10.1016/0304-4157(75)90016-7. [DOI] [PubMed] [Google Scholar]
  10. Hsieh P., Robbins P.W. Regulation of asparagine-linked oligosaccharide processing, oligosaccharide processing in Aedes albopictus mosquito cells. J. Biol. Chem. 1984;259:2375–2382. [PubMed] [Google Scholar]
  11. Igarashi A. Isolation of a Singh's Aedes albopictus cell clone sensitive to dengue and chikungunya viruses. J. Gen. Virol. 1978;40:531–544. doi: 10.1099/0022-1317-40-3-531. [DOI] [PubMed] [Google Scholar]
  12. Ishak R., Tovey D.G., Howard C.R. Morphogenesis of yellow fever 17D in infected cell cultures. J. Gen. Virol. 1988;69:325–335. doi: 10.1099/0022-1317-69-2-325. [DOI] [PubMed] [Google Scholar]
  13. Kornfeld R., Kornfeld S. Assembly of asparagine-linked oligosaccharides. Annu. Rev. Biochem. 1985;54:631–664. doi: 10.1146/annurev.bi.54.070185.003215. [DOI] [PubMed] [Google Scholar]
  14. Kuno G. A method for isolating continuous cell lines from Toxorhynchites amboinensis (Diptera: Culicidae) J. Med. Entomol. 1981;18:140–144. [Google Scholar]
  15. Laemmli U.K. Cleavage of structual proteins during the assembly of the head of bacteriophage T4. Nature (London) 1970;227:680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  16. Leary K., Blair C.D. Sequential events in the morphogenesis of Japanese encephalitis virus. J. Ultrastruct. Res. 1980;72:123–129. doi: 10.1016/s0022-5320(80)90050-7. [DOI] [PubMed] [Google Scholar]
  17. Madoff D.H., Lenard J. A membrane glycoprotein that accumulates intracellularly: Cellular processing of the large glycoprotein of LaCrosse virus. Cell. 1982;28:821–829. doi: 10.1016/0092-8674(82)90061-7. [DOI] [PubMed] [Google Scholar]
  18. Mason P.W., McAda P.C., Dalrymple J.M., Fournier M.J., Mason T.L. Expression of Japanese encephalitis virus antigens in Escherichia coli. Virology. 1987;158:361–372. doi: 10.1016/0042-6822(87)90208-x. [DOI] [PubMed] [Google Scholar]
  19. Mason P.W., McAda P.C., Mason T.L., Fournier M.J. Sequence of the dengue-1 virus genome in the region encoding the three structural proteins and the major non-structural protein NS1. Virology. 1987;161:262–267. doi: 10.1016/0042-6822(87)90196-6. [DOI] [PubMed] [Google Scholar]
  20. McAda P.C., Mason P.W., Schmauohn C.S., Dalrymple J.M., Mason T.L., Fournier M.J. Partial nucleotide sequence of the Japanese encephalitis virus genome. Virology. 1987;158:348–360. doi: 10.1016/0042-6822(87)90207-8. [DOI] [PubMed] [Google Scholar]
  21. Monath T.P. Pathobiology of the flaviviruses. In: Schlesinger S., Schlesinger M.J., editors. The Togaviridae and Flaviviridae. Plenum; New York/London: 1986. pp. 375–440. [Google Scholar]
  22. Ng M.L. Ultrastructural studies of Kunjin virus-infected Aedes albopictus cells. J. Gen. Virol. 1987;68:577–582. doi: 10.1099/0022-1317-68-2-577. [DOI] [PubMed] [Google Scholar]
  23. Pääbo S., Bhat B.M., Wold W.S.M., Peterson P.A. A short sequence in the COOH-terminus makes an adenovirus membrane glycoprotein a resident of the endoplasmic reticulum. Cell. 1987;50:311–317. doi: 10.1016/0092-8674(87)90226-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Pfeffer S.A., Rothman J.E. Biosynthetic protein transport and sorting by the endoplasmic reticulum and Golgi. Annu. Rev. Biochem. 1987;56:829–852. doi: 10.1146/annurev.bi.56.070187.004145. [DOI] [PubMed] [Google Scholar]
  25. Rice C.M., Strauss E.G., Strauss J.H. Structure of the flavivirus genome. In: Schlesinger S., Schlesinger M.J., editors. The Togaviridae and Flaviviridae. Plenum; New York/London: 1986. pp. 279–326. [Google Scholar]
  26. Roehrig J.T. The use of monoclonal antibodies in studies of the structural proteins of togaviruses and flaviviruses. In: Schlesinger S., Schlesinger M.J., editors. The Togaviridae and Flaviviridae. Plenum; New York/London: 1986. pp. 251–278. [Google Scholar]
  27. Rose J.K., Bergmann J.E. Altered cytoplasmic domains affect intracellular transport of the vesicular stomatitis virus glycoprotein. Cell. 1983;34:513–524. doi: 10.1016/0092-8674(83)90384-7. [DOI] [PubMed] [Google Scholar]
  28. Russell P.K., Brandt W.E., Dalrymple J.M. Chemical and antigenic structure of flaviviruses. In: Schlesinger R.W., editor. The Togaviruses. Academic Press; New York/London: 1980. pp. 503–529. [Google Scholar]
  29. Schlesinger J.J., Brandriss M.W., Walsh E.E. Protection against 17D yellow fever encephalitis in mice by passive transfer of monoclonal antibodies to the nonstructural glycoprotein gp48 and by active immunization with gp48. J. Immunol. 1985;135:2805–2809. [PubMed] [Google Scholar]
  30. Schlesinger J.J., Brandriss M.W., Walsh E.E. Protection of mice against dengue 2 virus encephalitis by immunization with the dengue 2 virus non-structural glycoprotein NS1. J. Gen. Virol. 1987;68:853–857. doi: 10.1099/0022-1317-68-3-853. [DOI] [PubMed] [Google Scholar]
  31. Shope R.E. Medical significance of togaviruses: An overview of diseases caused by togaviruses in man and in domestic and wild vertebrate animals. In: Schlesinger R.W., editor. The Togaviruses. Academic Press; New York/London: 1980. pp. 47–82. [Google Scholar]
  32. Smith G.W., Wright P.J. Synthesis of proteins and glycoproteins in dengue type 2 virus-infected Vero and Aedes albopictus cells. J. Gen. Virol. 1985;66:559–571. doi: 10.1099/0022-1317-66-3-559. [DOI] [PubMed] [Google Scholar]
  33. Speight G., Coia G., Parker M.D., Westaway E.G. Gene mapping and positive identification of the nonstructural proteins NS2A, NS2B, NS3, NS4B and NS5 of the flavivirus Kunjin and their cleavage sites. J. Gen. Virol. 1988;69:23–34. doi: 10.1099/0022-1317-69-1-23. [DOI] [PubMed] [Google Scholar]
  34. Stephenson J.R., Crooks A.J., Lee J.M. The synthesis of immunogenic polypeptides encoded by tick-borne encephalitis virus. J. Gen. Virol. 1987;68:1307–1316. doi: 10.1099/0022-1317-68-5-1307. [DOI] [PubMed] [Google Scholar]
  35. Strous G.J.A.M., Lodish H.F. Intracellular transport of secretory and membrane proteins in hepatoma cells infected by vesicular stomatitis virus. Cell. 1980;22:709–717. doi: 10.1016/0092-8674(80)90547-4. [DOI] [PubMed] [Google Scholar]
  36. Tarentino A.L., Gómez C.M., Plummer T.H., Jr Deglycosylation of asparagine linked glycans by peptide:N-glycosidase F. Biochemistry. 1985;24:4665–4671. doi: 10.1021/bi00338a028. [DOI] [PubMed] [Google Scholar]
  37. Tarentino A.L., Trimble R.B., Maley F. Endo-β-N-acetylglucosamindase from Streptomyces plicatus. In: Ginsburg V., editor. Vol. 50. Academic Press; New York: 1978. pp. 574–580. (Methods in Enzymology). [DOI] [PubMed] [Google Scholar]
  38. Tesh R.B. A method for the isolation and identification of dengue viruses, using mosquito cell cultures. Amer. J. Trop. Med. Hyg. 1979;28:1053–1059. doi: 10.4269/ajtmh.1979.28.1053. [DOI] [PubMed] [Google Scholar]
  39. Tesh R.B., Duboise S.M. Viremia and immune response with sequential phlebovirus infections. Amer. J. Trop. Med. Hyg. 1987;36:662–668. doi: 10.4269/ajtmh.1987.36.662. [DOI] [PubMed] [Google Scholar]
  40. Tooze J., Tooze S.A., Fuller S.D. Sorting of progeny coronavirus from condensed secretory proteins at the exit from the trans-Golgi network of AtT20 cells. J. Cell Biol. 1987;105:1215–1226. doi: 10.1083/jcb.105.3.1215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Varma M.G.R., Pudney M., Leake C.J. Cell lines from larvae of Aedes (Stegomyia) malayensis Colless and Aedes (S) pseudoscutellaris (Theobald) and their infection with some arboviruses. Trans. R. Soc. Trop. Med. Hyg. 1974;68:374–382. doi: 10.1016/0035-9203(74)90152-7. [DOI] [PubMed] [Google Scholar]
  42. Wengler G., Castle E., Leidner U., Nowak T., Wengler G. Sequence analysis of the membrane protein V3 of the flavivirus West Nile virus and of its gene. Virology. 1985;147:264–274. doi: 10.1016/0042-6822(85)90129-1. [DOI] [PubMed] [Google Scholar]
  43. Westaway E. Replication of flaviviruses. In: Schlesinger R.W., editor. The Togaviruses. Academic Press; New York/London: 1980. pp. 531–581. [Google Scholar]
  44. Wieland F.T., Gleason M.L., Serafini T.A., Rothman J.E. The rate of bulk flow from the endoplasmic reticulum to the cell surface. Cell. 1987;50:289–300. doi: 10.1016/0092-8674(87)90224-8. [DOI] [PubMed] [Google Scholar]
  45. Winkler G., Heinz F.X., Kunz C. Studies on the glycosylation of flavivirus E proteins and the role of carbohydrate in antigenic structure. Virology. 1987;159:237–243. doi: 10.1016/0042-6822(87)90460-0. [DOI] [PubMed] [Google Scholar]
  46. Winkler G., Randolph V.B., Cleaves G.R., Ryan T.E., Stollar V. Evidence that the mature form of the flavivirus nonstructural protein NS1 is a dimer. Virology. 1988;162:187–196. doi: 10.1016/0042-6822(88)90408-4. [DOI] [PubMed] [Google Scholar]
  47. Wright P.J. Envelope protein of the flavivirus Kunjin is apparently not glycosylated. J. Gen. Virol. 1982;59:29–38. doi: 10.1099/0022-1317-59-1-29. [DOI] [PubMed] [Google Scholar]

Articles from Virology are provided here courtesy of Elsevier

RESOURCES