EXPRESSION OF HIV ENV GENE IN A HUMAN T-CELL LINE FOR A RAPID AND QUANTIFIABLE CELL-FUSION ASSAY

Human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins pre sent at the surface of infected cells are known to mediate fusion with CD4-positive target cells. In this study we have developed a novel En v-expressing cell line for investigating the fusion process in a biolo gically significant system, Cell surface expression of the HIV-1 env g ene, isolated from the highly fusogenic strain SF33, was obtained in t he CD4-negative T cell line A2.01. To render the system versatile and efficient, HIV-1 regulatory proteins Tat and Rev were supplied in tran s. The presence of Env at the cell surface was shown by cytofluorometr y and immunofluorescence and precursor processing of gp160 to gp120/gp 41 was demonstrated by Western blot. The fusion capacity of A2.01-Env cells was assessed by coculture with CD4-positive T lymphocytes or the fusion indicator cell line, HeLa-CD4-LTR-beta-Gal. By coincubation wi th CD4-positive T cells such as SupT1, A2.01-Env cells were observed t o mediate rapidly numerous well-defined syncytia in a reproducible fas hion, By expressing Tat, they also had the capacity to trans-activate the LTR-linked reporter beta-Gal gene following fusion with HeLa-CD4-L TR-beta-Gal cells. The fusion-inhibiting anti-CD4 monoclonal antibodie s Q425 and Q428 were used to block specifically Env-mediated fusion wi th CD4-positive cells and to demonstrate application of this system to the search for potential fusion-blocking agents. Our system thus offe rs a biologically significant model for studying fusion events,vith th e advantages of being rapid, reproducible, and versatile.