Temperature dependence of cell-cell fusion induced by the envelope glycoprotein of human immunodeficiency virus type 1.
We investigated cell-cell fusion induced by the envelope glycoprotein of human immunodeficiency virus type 1 strain IIIB expressed on the surface of CHO cells. These cells formed syncytia when incubated together with CD4-positive human lymphoblastoid SupT1 cells or HeLa-CD4 cells but not when incubated with CD4-negative cell lines. A new assay for binding and fusion was developed by using fluorescent phospholipid analogs that were produced in SupT1 cells by metabolic incorporation of BODIPY-labeled fatty acids. Fusion occurred as early as 10 min after mixing of labeled SupT1 cells with unlabeled CHO-gp160 cells at 37 degrees C. When both the fluorescence assay and formation of syncytia were used, fusion of SupT1 and HeLa-CD4 cells with CHO-gp160 cells was observed only at temperatures above 25 degrees C, confirming recent observations (Y.-K. Fu, T.K. Hart, Z.L. Jonak, and P.J. Bugelski, J. Virol. 67:3818-3825, 1993). This temperature dependence was not observed with influenza virus-induced cell-cell fusion, which was quantitatively similar at both 20 and 37 degrees C, indicating that cell-cell fusion in general is not temperature dependent in this range. gp120-CD4-specific cell-cell binding was found over the entire 0 to 37 degrees C range but increased markedly above 25 degrees C. The enhanced binding and fusion were reduced by cytochalasins B and D. Binding of soluble gp120 to CD4-expressing cells was equivalent at 37 and 16 degrees C. Together, these data indicate that during gp120-gp41-induced syncytium formation, initial cell-cell binding is followed by a cytoskeleton-dependent increase in the number of gp120-CD4 complexes, leading to an increase in the avidity of cell-cell binding. The increased number of gp120-CD4 complexes is required for fusion, which suggests that the formation of a fusion complex consisting of multiple CD4 and gp120-gp41 molecules is a step in the fusion mechanism.
Department of Biophysical Chemistry, Biozentrum of the University of Basel, Switzerland.
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Frey
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Marsh
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Günther
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Pelchen-Matthews
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Stephens
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Ortlepp
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Stegmann
D000818:Animals; D015704:CD4 Antigens / Q000378:metabolism; D016466:CHO Cells; D002459:Cell Fusion / Q000187:drug effects; D006224:Cricetinae; D003571:Cytochalasin B / Q000494:pharmacology; D015686:Gene Products, env / Q000502:physiology*; D015699:HIV Envelope Protein gp120 / Q000502:physiology*; D019302:HIV Envelope Protein gp160; D015497:HIV-1 / Q000472:pathogenicity*; D066298:In Vitro Techniques; D011498:Protein Precursors / Q000502:physiology*; D011994:Recombinant Proteins; D013696:Temperature
D015704:CD4 Antigens; D015686:Gene Products, env; D015699:HIV Envelope Protein gp120; D019302:HIV Envelope Protein gp160; D011498:Protein Precursors; D011994:Recombinant Proteins; D003571:Cytochalasin B