Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1993 Jul;67(7):4264–4273. doi: 10.1128/jvi.67.7.4264-4273.1993

Assembly, processing, and infectivity of human immunodeficiency virus type 1 gag mutants.

C T Wang 1, E Barklis 1
PMCID: PMC237796  PMID: 7685414

Abstract

We studied the effects of gag mutations on human immunodeficiency virus type 1 (HIV-1) assembly, processing, and infectivity by using a replication-defective HIV expression system. HIV mutants were screened for infectivity by transduction of a selectable marker and were examined for assembly by monitoring particle release from transfected cells. Gag protein processing and reverse transcriptase activities of mutant particles were also assayed. Surprisingly, most Gag protein mutants were assembled and processed. The two exceptions to this rule were a myristylation-minus mutant, and one gag matrix domain mutant which expressed proteins that were trapped intracellularly. Interestingly, a mutant with a 56-amino-acid deletion within the HIV gag capsid domain still could assemble and process virus particles, exhibited a wild-type retrovirus particle density, and had wild-type reverse transcriptase activity. Indeed, although most HIV-1 gag mutants were noninfectious or poorly infectious, they produced apparently normal particles which possessed significant reverse transcriptase activities. These results strongly support the notion that the HIV-1 Gag proteins are functionally involved in post-assembly, postprocessing stages of virus infectivity.

Full text

PDF
4264

Images in this article

4267Image
on p.4267
4269Image
on p.4269
4270Image
on p.4270
4271Image
on p.4271

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aldovini A., Young R. A. Mutations of RNA and protein sequences involved in human immunodeficiency virus type 1 packaging result in production of noninfectious virus. J Virol. 1990 May;64(5):1920–1926. doi: 10.1128/jvi.64.5.1920-1926.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bess J. W., Jr, Powell P. J., Issaq H. J., Schumack L. J., Grimes M. K., Henderson L. E., Arthur L. O. Tightly bound zinc in human immunodeficiency virus type 1, human T-cell leukemia virus type I, and other retroviruses. J Virol. 1992 Feb;66(2):840–847. doi: 10.1128/jvi.66.2.840-847.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bolognesi D. P., Luftig R., Shaper J. H. Localization of RNA tumor virus polypeptides. I. Isolation of further virus substructures. Virology. 1973 Dec;56(2):549–564. doi: 10.1016/0042-6822(73)90057-3. [DOI] [PubMed] [Google Scholar]
  4. Bowerman B., Brown P. O., Bishop J. M., Varmus H. E. A nucleoprotein complex mediates the integration of retroviral DNA. Genes Dev. 1989 Apr;3(4):469–478. doi: 10.1101/gad.3.4.469. [DOI] [PubMed] [Google Scholar]
  5. Bryant M., Ratner L. Myristoylation-dependent replication and assembly of human immunodeficiency virus 1. Proc Natl Acad Sci U S A. 1990 Jan;87(2):523–527. doi: 10.1073/pnas.87.2.523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cann A. J., Karn J. Molecular biology of HIV: new insights into the virus life-cycle. AIDS. 1989;3 (Suppl 1):S19–S34. [PubMed] [Google Scholar]
  7. Cohen E. A., Terwilliger E. F., Sodroski J. G., Haseltine W. A. Identification of a protein encoded by the vpu gene of HIV-1. Nature. 1988 Aug 11;334(6182):532–534. doi: 10.1038/334532a0. [DOI] [PubMed] [Google Scholar]
  8. Crawford S., Goff S. P. Mutations in gag proteins P12 and P15 of Moloney murine leukemia virus block early stages of infection. J Virol. 1984 Mar;49(3):909–917. doi: 10.1128/jvi.49.3.909-917.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ehrlich L. S., Agresta B. E., Carter C. A. Assembly of recombinant human immunodeficiency virus type 1 capsid protein in vitro. J Virol. 1992 Aug;66(8):4874–4883. doi: 10.1128/jvi.66.8.4874-4883.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gheysen D., Jacobs E., de Foresta F., Thiriart C., Francotte M., Thines D., De Wilde M. Assembly and release of HIV-1 precursor Pr55gag virus-like particles from recombinant baculovirus-infected insect cells. Cell. 1989 Oct 6;59(1):103–112. doi: 10.1016/0092-8674(89)90873-8. [DOI] [PubMed] [Google Scholar]
  11. Gorelick R. J., Nigida S. M., Jr, Bess J. W., Jr, Arthur L. O., Henderson L. E., Rein A. Noninfectious human immunodeficiency virus type 1 mutants deficient in genomic RNA. J Virol. 1990 Jul;64(7):3207–3211. doi: 10.1128/jvi.64.7.3207-3211.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Graham F. L., van der Eb A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. doi: 10.1016/0042-6822(73)90341-3. [DOI] [PubMed] [Google Scholar]
  13. Göttlinger H. G., Dorfman T., Sodroski J. G., Haseltine W. A. Effect of mutations affecting the p6 gag protein on human immunodeficiency virus particle release. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3195–3199. doi: 10.1073/pnas.88.8.3195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Göttlinger H. G., Sodroski J. G., Haseltine W. A. Role of capsid precursor processing and myristoylation in morphogenesis and infectivity of human immunodeficiency virus type 1. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5781–5785. doi: 10.1073/pnas.86.15.5781. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hansen M., Jelinek L., Whiting S., Barklis E. Transport and assembly of gag proteins into Moloney murine leukemia virus. J Virol. 1990 Nov;64(11):5306–5316. doi: 10.1128/jvi.64.11.5306-5316.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hayashi T., Shioda T., Iwakura Y., Shibuta H. RNA packaging signal of human immunodeficiency virus type 1. Virology. 1992 Jun;188(2):590–599. doi: 10.1016/0042-6822(92)90513-o. [DOI] [PubMed] [Google Scholar]
  17. Henderson L. E., Bowers M. A., Sowder R. C., 2nd, Serabyn S. A., Johnson D. G., Bess J. W., Jr, Arthur L. O., Bryant D. K., Fenselau C. Gag proteins of the highly replicative MN strain of human immunodeficiency virus type 1: posttranslational modifications, proteolytic processings, and complete amino acid sequences. J Virol. 1992 Apr;66(4):1856–1865. doi: 10.1128/jvi.66.4.1856-1865.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Jones T. A., Blaug G., Hansen M., Barklis E. Assembly of gag-beta-galactosidase proteins into retrovirus particles. J Virol. 1990 May;64(5):2265–2279. doi: 10.1128/jvi.64.5.2265-2279.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Klimkait T., Strebel K., Hoggan M. D., Martin M. A., Orenstein J. M. The human immunodeficiency virus type 1-specific protein vpu is required for efficient virus maturation and release. J Virol. 1990 Feb;64(2):621–629. doi: 10.1128/jvi.64.2.621-629.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kohl N. E., Emini E. A., Schleif W. A., Davis L. J., Heimbach J. C., Dixon R. A., Scolnick E. M., Sigal I. S. Active human immunodeficiency virus protease is required for viral infectivity. Proc Natl Acad Sci U S A. 1988 Jul;85(13):4686–4690. doi: 10.1073/pnas.85.13.4686. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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]
  22. Lapatto R., Blundell T., Hemmings A., Overington J., Wilderspin A., Wood S., Merson J. R., Whittle P. J., Danley D. E., Geoghegan K. F. X-ray analysis of HIV-1 proteinase at 2.7 A resolution confirms structural homology among retroviral enzymes. Nature. 1989 Nov 16;342(6247):299–302. doi: 10.1038/342299a0. [DOI] [PubMed] [Google Scholar]
  23. Le Grice S. F., Ette R., Mills J., Mous J. Comparison of the human immunodeficiency virus type 1 and 2 proteases by hybrid gene construction and trans-complementation. J Biol Chem. 1989 Sep 5;264(25):14902–14908. [PubMed] [Google Scholar]
  24. Le Grice S. F., Mills J., Mous J. Active site mutagenesis of the AIDS virus protease and its alleviation by trans complementation. EMBO J. 1988 Aug;7(8):2547–2553. doi: 10.1002/j.1460-2075.1988.tb03103.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Leis J., Baltimore D., Bishop J. M., Coffin J., Fleissner E., Goff S. P., Oroszlan S., Robinson H., Skalka A. M., Temin H. M. Standardized and simplified nomenclature for proteins common to all retroviruses. J Virol. 1988 May;62(5):1808–1809. doi: 10.1128/jvi.62.5.1808-1809.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Leuthardt A., Le Grice S. F. Biosynthesis and analysis of a genetically engineered HIV-1 reverse transcriptase/endonuclease polyprotein in Escherichia coli. Gene. 1988 Aug 15;68(1):35–42. doi: 10.1016/0378-1119(88)90596-3. [DOI] [PubMed] [Google Scholar]
  27. Lever A., Gottlinger H., Haseltine W., Sodroski J. Identification of a sequence required for efficient packaging of human immunodeficiency virus type 1 RNA into virions. J Virol. 1989 Sep;63(9):4085–4087. doi: 10.1128/jvi.63.9.4085-4087.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mervis R. J., Ahmad N., Lillehoj E. P., Raum M. G., Salazar F. H., Chan H. W., Venkatesan S. The gag gene products of human immunodeficiency virus type 1: alignment within the gag open reading frame, identification of posttranslational modifications, and evidence for alternative gag precursors. J Virol. 1988 Nov;62(11):3993–4002. doi: 10.1128/jvi.62.11.3993-4002.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Mulligan R. C., Berg P. Selection for animal cells that express the Escherichia coli gene coding for xanthine-guanine phosphoribosyltransferase. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2072–2076. doi: 10.1073/pnas.78.4.2072. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Navia M. A., Fitzgerald P. M., McKeever B. M., Leu C. T., Heimbach J. C., Herber W. K., Sigal I. S., Darke P. L., Springer J. P. Three-dimensional structure of aspartyl protease from human immunodeficiency virus HIV-1. Nature. 1989 Feb 16;337(6208):615–620. doi: 10.1038/337615a0. [DOI] [PubMed] [Google Scholar]
  31. Overton H. A., Fujii Y., Price I. R., Jones I. M. The protease and gag gene products of the human immunodeficiency virus: authentic cleavage and post-translational modification in an insect cell expression system. Virology. 1989 May;170(1):107–116. doi: 10.1016/0042-6822(89)90357-7. [DOI] [PubMed] [Google Scholar]
  32. Page K. A., Landau N. R., Littman D. R. Construction and use of a human immunodeficiency virus vector for analysis of virus infectivity. J Virol. 1990 Nov;64(11):5270–5276. doi: 10.1128/jvi.64.11.5270-5276.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Pal R., Reitz M. S., Jr, Tschachler E., Gallo R. C., Sarngadharan M. G., Veronese F. D. Myristoylation of gag proteins of HIV-1 plays an important role in virus assembly. AIDS Res Hum Retroviruses. 1990 Jun;6(6):721–730. doi: 10.1089/aid.1990.6.721. [DOI] [PubMed] [Google Scholar]
  34. Peng C., Ho B. K., Chang T. W., Chang N. T. Role of human immunodeficiency virus type 1-specific protease in core protein maturation and viral infectivity. J Virol. 1989 Jun;63(6):2550–2556. doi: 10.1128/jvi.63.6.2550-2556.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. 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]
  36. Schwartzberg P., Colicelli J., Gordon M. L., Goff S. P. Mutations in the gag gene of Moloney murine leukemia virus: effects on production of virions and reverse transcriptase. J Virol. 1984 Mar;49(3):918–924. doi: 10.1128/jvi.49.3.918-924.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Sharova N., Bukrinskaya A. p17 and p17-containing gag precursors of input human immunodeficiency virus are transported into the nuclei of infected cells. AIDS Res Hum Retroviruses. 1991 Mar;7(3):303–306. doi: 10.1089/aid.1991.7.303. [DOI] [PubMed] [Google Scholar]
  38. Shioda T., Shibuta H. Production of human immunodeficiency virus (HIV)-like particles from cells infected with recombinant vaccinia viruses carrying the gag gene of HIV. Virology. 1990 Mar;175(1):139–148. doi: 10.1016/0042-6822(90)90194-v. [DOI] [PubMed] [Google Scholar]
  39. South T. L., Blake P. R., Sowder R. C., 3rd, Arthur L. O., Henderson L. E., Summers M. F. The nucleocapsid protein isolated from HIV-1 particles binds zinc and forms retroviral-type zinc fingers. Biochemistry. 1990 Aug 28;29(34):7786–7789. doi: 10.1021/bi00486a002. [DOI] [PubMed] [Google Scholar]
  40. Stephens E. B., Compans R. W. Assembly of animal viruses at cellular membranes. Annu Rev Microbiol. 1988;42:489–516. doi: 10.1146/annurev.mi.42.100188.002421. [DOI] [PubMed] [Google Scholar]
  41. Stewart L., Schatz G., Vogt V. M. Properties of avian retrovirus particles defective in viral protease. J Virol. 1990 Oct;64(10):5076–5092. doi: 10.1128/jvi.64.10.5076-5092.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Strebel K., Klimkait T., Martin M. A. A novel gene of HIV-1, vpu, and its 16-kilodalton product. Science. 1988 Sep 2;241(4870):1221–1223. doi: 10.1126/science.3261888. [DOI] [PubMed] [Google Scholar]
  43. Terwilliger E. F., Cohen E. A., Lu Y. C., Sodroski J. G., Haseltine W. A. Functional role of human immunodeficiency virus type 1 vpu. Proc Natl Acad Sci U S A. 1989 Jul;86(13):5163–5167. doi: 10.1073/pnas.86.13.5163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Towler D. A., Eubanks S. R., Towery D. S., Adams S. P., Glaser L. Amino-terminal processing of proteins by N-myristoylation. Substrate specificity of N-myristoyl transferase. J Biol Chem. 1987 Jan 25;262(3):1030–1036. [PubMed] [Google Scholar]
  45. Trono D., Feinberg M. B., Baltimore D. HIV-1 Gag mutants can dominantly interfere with the replication of the wild-type virus. Cell. 1989 Oct 6;59(1):113–120. doi: 10.1016/0092-8674(89)90874-x. [DOI] [PubMed] [Google Scholar]
  46. Wain-Hobson S., Sonigo P., Danos O., Cole S., Alizon M. Nucleotide sequence of the AIDS virus, LAV. Cell. 1985 Jan;40(1):9–17. doi: 10.1016/0092-8674(85)90303-4. [DOI] [PubMed] [Google Scholar]
  47. Wilcox C., Hu J. S., Olson E. N. Acylation of proteins with myristic acid occurs cotranslationally. Science. 1987 Nov 27;238(4831):1275–1278. doi: 10.1126/science.3685978. [DOI] [PubMed] [Google Scholar]
  48. Wlodawer A., Miller M., Jaskólski M., Sathyanarayana B. K., Baldwin E., Weber I. T., Selk L. M., Clawson L., Schneider J., Kent S. B. Conserved folding in retroviral proteases: crystal structure of a synthetic HIV-1 protease. Science. 1989 Aug 11;245(4918):616–621. doi: 10.1126/science.2548279. [DOI] [PubMed] [Google Scholar]
  49. Yu X., Yu Q. C., Lee T. H., Essex M. The C terminus of human immunodeficiency virus type 1 matrix protein is involved in early steps of the virus life cycle. J Virol. 1992 Sep;66(9):5667–5670. doi: 10.1128/jvi.66.9.5667-5670.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Yu X., Yuan X., Matsuda Z., Lee T. H., Essex M. The matrix protein of human immunodeficiency virus type 1 is required for incorporation of viral envelope protein into mature virions. J Virol. 1992 Aug;66(8):4966–4971. doi: 10.1128/jvi.66.8.4966-4971.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES