Skip to main content
NCBI home page
Journal of Virology logoLink to Journal of Virology
. 1996 Sep;70(9):6437–6441. doi: 10.1128/jvi.70.9.6437-6441.1996

Human immunodeficiency virus type 1 membrane fusion mediated by a laboratory-adapted strain and a primary isolate analyzed by resonance energy transfer.

V Litwin 1, K A Nagashima 1, A M Ryder 1, C H Chang 1, J M Carver 1, W C Olson 1, M Alizon 1, K W Hasel 1, P J Maddon 1, G P Allaway 1
PMCID: PMC190675  PMID: 8709277

Abstract

Previous studies of human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein-mediated membrane fusion have focused on laboratory-adapted T-lymphotropic strains of the virus. The goal of this study was to characterize membrane fusion mediated by a primary HIV-1 isolate in comparison with a laboratory-adapted strain. To this end, a new fusion assay was developed on the basis of the principle of resonance energy transfer, using HeLa cells stably transfected with gp120/gp41 from the T-lymphotropic isolate HIV-1LA1 or the macrophage-tropic primary isolate HIV-1JR-FL. These cells fused with CD4+ target cell lines with a tropism mirroring that of infection by the two viruses. Of particular note, HeLa cells expressing HIV-1JR-FL gp120/gp41 fused only with PM1 cells, a clonal derivative of HUT 78, and not with other T-cell or macrophage cell lines. These results demonstrate that the envelope glycoproteins of these strains play a major role in mediating viral tropism. Despite significant differences exhibited by HIV-1JR-FL and HIV-1LAI in terms of tropism and sensitivity to neutralization by CD4-based proteins, the present study found that membrane fusion mediated by the envelope glycoproteins of these viruses had remarkably similar properties. In particular, the degree and kinetics of membrane fusion were similar, fusion occurred at neutral pH and was dependent on the presence of divalent cations. Inhibition of HIV-1JR-FL envelope glycoprotein-mediated membrane fusion by soluble CD4 and CD4-IgG2 occurred at concentrations similar to those required to neutralize this virus. Interestingly, higher concentrations of these agents were required to inhibit HIV-1LAI envelope glycoprotein-mediated membrane fusion, in contrast to the greater sensitivity of HIV-1LAI virions to neutralization by soluble CD4 and CD4-IgG2. This finding suggests that the mechanisms of fusion inhibition and neutralization of HIV-1 are distinct.

Full Text

The Full Text of this article is available as a PDF (210.1 KB).

Selected References

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

  1. Allaway G. P., Davis-Bruno K. L., Beaudry G. A., Garcia E. B., Wong E. L., Ryder A. M., Hasel K. W., Gauduin M. C., Koup R. A., McDougal J. S. Expression and characterization of CD4-IgG2, a novel heterotetramer that neutralizes primary HIV type 1 isolates. AIDS Res Hum Retroviruses. 1995 May;11(5):533–539. doi: 10.1089/aid.1995.11.533. [DOI] [PubMed] [Google Scholar]
  2. Bou-Habib D. C., Roderiquez G., Oravecz T., Berman P. W., Lusso P., Norcross M. A. Cryptic nature of envelope V3 region epitopes protects primary monocytotropic human immunodeficiency virus type 1 from antibody neutralization. J Virol. 1994 Sep;68(9):6006–6013. doi: 10.1128/jvi.68.9.6006-6013.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Broder C. C., Berger E. A. Fusogenic selectivity of the envelope glycoprotein is a major determinant of human immunodeficiency virus type 1 tropism for CD4+ T-cell lines vs. primary macrophages. Proc Natl Acad Sci U S A. 1995 Sep 12;92(19):9004–9008. doi: 10.1073/pnas.92.19.9004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Buchacher A., Predl R., Strutzenberger K., Steinfellner W., Trkola A., Purtscher M., Gruber G., Tauer C., Steindl F., Jungbauer A. Generation of human monoclonal antibodies against HIV-1 proteins; electrofusion and Epstein-Barr virus transformation for peripheral blood lymphocyte immortalization. AIDS Res Hum Retroviruses. 1994 Apr;10(4):359–369. doi: 10.1089/aid.1994.10.359. [DOI] [PubMed] [Google Scholar]
  5. Chesebro B., Buller R., Portis J., Wehrly K. Failure of human immunodeficiency virus entry and infection in CD4-positive human brain and skin cells. J Virol. 1990 Jan;64(1):215–221. doi: 10.1128/jvi.64.1.215-221.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Clapham P. R., Blanc D., Weiss R. A. Specific cell surface requirements for the infection of CD4-positive cells by human immunodeficiency virus types 1 and 2 and by Simian immunodeficiency virus. Virology. 1991 Apr;181(2):703–715. doi: 10.1016/0042-6822(91)90904-P. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Daar E. S., Li X. L., Moudgil T., Ho D. D. High concentrations of recombinant soluble CD4 are required to neutralize primary human immunodeficiency virus type 1 isolates. Proc Natl Acad Sci U S A. 1990 Sep;87(17):6574–6578. doi: 10.1073/pnas.87.17.6574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dimitrov D. S., Broder C. C., Berger E. A., Blumenthal R. Calcium ions are required for cell fusion mediated by the CD4-human immunodeficiency virus type 1 envelope glycoprotein interaction. J Virol. 1993 Mar;67(3):1647–1652. doi: 10.1128/jvi.67.3.1647-1652.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dimitrov D. S., Golding H., Blumenthal R. Initial stages of HIV-1 envelope glycoprotein-mediated cell fusion monitored by a new assay based on redistribution of fluorescent dyes. AIDS Res Hum Retroviruses. 1991 Oct;7(10):799–805. doi: 10.1089/aid.1991.7.799. [DOI] [PubMed] [Google Scholar]
  10. Dimitrov D. S., Hillman K., Manischewitz J., Blumenthal R., Golding H. Correlation between kinetics of soluble CD4 interactions with HIV-1-Env-expressing cells and inhibition of syncytia formation: implications for mechanisms of cell fusion and therapy for AIDS. AIDS. 1992 Mar;6(3):249–256. doi: 10.1097/00002030-199203000-00001. [DOI] [PubMed] [Google Scholar]
  11. Dragic T., Charneau P., Clavel F., Alizon M. Complementation of murine cells for human immunodeficiency virus envelope/CD4-mediated fusion in human/murine heterokaryons. J Virol. 1992 Aug;66(8):4794–4802. doi: 10.1128/jvi.66.8.4794-4802.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Frey S., Marsh M., Günther S., Pelchen-Matthews A., Stephens P., Ortlepp S., Stegmann T. Temperature dependence of cell-cell fusion induced by the envelope glycoprotein of human immunodeficiency virus type 1. J Virol. 1995 Mar;69(3):1462–1472. doi: 10.1128/jvi.69.3.1462-1472.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hsu K. C., Chao M. V. Differential expression and ligand binding properties of tumor necrosis factor receptor chimeric mutants. J Biol Chem. 1993 Aug 5;268(22):16430–16436. [PubMed] [Google Scholar]
  14. Keller P. M., Person S., Snipes W. A fluorescence enhancement assay of cell fusion. J Cell Sci. 1977 Dec;28:167–177. doi: 10.1242/jcs.28.1.167. [DOI] [PubMed] [Google Scholar]
  15. Klatzmann D. R., McDougal J. S., Maddon P. J. The CD4 molecule and HIV infection. Immunodefic Rev. 1990;2(1):43–66. [PubMed] [Google Scholar]
  16. Koito A., Harrowe G., Levy J. A., Cheng-Mayer C. Functional role of the V1/V2 region of human immunodeficiency virus type 1 envelope glycoprotein gp120 in infection of primary macrophages and soluble CD4 neutralization. J Virol. 1994 Apr;68(4):2253–2259. doi: 10.1128/jvi.68.4.2253-2259.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Koyanagi Y., Miles S., Mitsuyasu R. T., Merrill J. E., Vinters H. V., Chen I. S. Dual infection of the central nervous system by AIDS viruses with distinct cellular tropisms. Science. 1987 May 15;236(4803):819–822. doi: 10.1126/science.3646751. [DOI] [PubMed] [Google Scholar]
  18. Liu J., Han J. R., Liu C. C., Suiko M., Liu M. C. Identification of a putative tyrosine-O-sulphate (TyrS) receptor possibly functioning in the biosynthetic transport of tyrosine-sulphated proteins in Madin-Darby canine kidney cells. Biochem J. 1993 Sep 1;294(Pt 2):407–417. doi: 10.1042/bj2940407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lusso P., Cocchi F., Balotta C., Markham P. D., Louie A., Farci P., Pal R., Gallo R. C., Reitz M. S., Jr Growth of macrophage-tropic and primary human immunodeficiency virus type 1 (HIV-1) isolates in a unique CD4+ T-cell clone (PM1): failure to downregulate CD4 and to interfere with cell-line-tropic HIV-1. J Virol. 1995 Jun;69(6):3712–3720. doi: 10.1128/jvi.69.6.3712-3720.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Maddon P. J., Dalgleish A. G., McDougal J. S., Clapham P. R., Weiss R. A., Axel R. The T4 gene encodes the AIDS virus receptor and is expressed in the immune system and the brain. Cell. 1986 Nov 7;47(3):333–348. doi: 10.1016/0092-8674(86)90590-8. [DOI] [PubMed] [Google Scholar]
  21. McDougal J. S., Nicholson J. K., Cross G. D., Cort S. P., Kennedy M. S., Mawle A. C. Binding of the human retrovirus HTLV-III/LAV/ARV/HIV to the CD4 (T4) molecule: conformation dependence, epitope mapping, antibody inhibition, and potential for idiotypic mimicry. J Immunol. 1986 Nov 1;137(9):2937–2944. [PubMed] [Google Scholar]
  22. Meier T., Arni S., Malarkannan S., Poincelet M., Hoessli D. Immunodetection of biotinylated lymphocyte-surface proteins by enhanced chemiluminescence: a nonradioactive method for cell-surface protein analysis. Anal Biochem. 1992 Jul;204(1):220–226. doi: 10.1016/0003-2697(92)90165-4. [DOI] [PubMed] [Google Scholar]
  23. Moore J. P., McKeating J. A., Huang Y. X., Ashkenazi A., Ho D. D. Virions of primary human immunodeficiency virus type 1 isolates resistant to soluble CD4 (sCD4) neutralization differ in sCD4 binding and glycoprotein gp120 retention from sCD4-sensitive isolates. J Virol. 1992 Jan;66(1):235–243. doi: 10.1128/jvi.66.1.235-243.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. O'Brien W. A., Koyanagi Y., Namazie A., Zhao J. Q., Diagne A., Idler K., Zack J. A., Chen I. S. HIV-1 tropism for mononuclear phagocytes can be determined by regions of gp120 outside the CD4-binding domain. Nature. 1990 Nov 1;348(6296):69–73. doi: 10.1038/348069a0. [DOI] [PubMed] [Google Scholar]
  25. Orloff S. L., Kennedy M. S., Belperron A. A., Maddon P. J., McDougal J. S. Two mechanisms of soluble CD4 (sCD4)-mediated inhibition of human immunodeficiency virus type 1 (HIV-1) infectivity and their relation to primary HIV-1 isolates with reduced sensitivity to sCD4. J Virol. 1993 Mar;67(3):1461–1471. doi: 10.1128/jvi.67.3.1461-1471.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Posner M. R., Cavacini L. A., Emes C. L., Power J., Byrn R. Neutralization of HIV-1 by F105, a human monoclonal antibody to the CD4 binding site of gp120. J Acquir Immune Defic Syndr. 1993 Jan;6(1):7–14. [PubMed] [Google Scholar]
  27. 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]
  28. Salter R. D., Howell D. N., Cresswell P. Genes regulating HLA class I antigen expression in T-B lymphoblast hybrids. Immunogenetics. 1985;21(3):235–246. doi: 10.1007/BF00375376. [DOI] [PubMed] [Google Scholar]
  29. Shioda T., Levy J. A., Cheng-Mayer C. Macrophage and T cell-line tropisms of HIV-1 are determined by specific regions of the envelope gp120 gene. Nature. 1991 Jan 10;349(6305):167–169. doi: 10.1038/349167a0. [DOI] [PubMed] [Google Scholar]
  30. Stefano K. A., Collman R., Kolson D., Hoxie J., Nathanson N., Gonzalez-Scarano F. Replication of a macrophage-tropic strain of human immunodeficiency virus type 1 (HIV-1) in a hybrid cell line, CEMx174, suggests that cellular accessory molecules are required for HIV-1 entry. J Virol. 1993 Nov;67(11):6707–6715. doi: 10.1128/jvi.67.11.6707-6715.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Trkola A., Pomales A. B., Yuan H., Korber B., Maddon P. J., Allaway G. P., Katinger H., Barbas C. F., 3rd, Burton D. R., Ho D. D. Cross-clade neutralization of primary isolates of human immunodeficiency virus type 1 by human monoclonal antibodies and tetrameric CD4-IgG. J Virol. 1995 Nov;69(11):6609–6617. doi: 10.1128/jvi.69.11.6609-6617.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Wanda P. E., Smith J. D. A general method for heterokaryon detection using resonance energy transfer and a fluorescence-activated cell sorter. J Histochem Cytochem. 1982 Dec;30(12):1297–1300. doi: 10.1177/30.12.7153501. [DOI] [PubMed] [Google Scholar]

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

RESOURCES