FIG 6.

Activation by CD4mc of Env-mediated syncytium formation with CD4⁻ CCR5⁺ cells. (A) 293T cells expressing the HIV-1 Env and the α peptide of β-galactosidase were cocultivated with CD4-negative, CCR5-positive Cf2Th-CCR5 cells expressing the ω peptide of β-galactosidase in medium containing the indicated concentrations of CD4mc. After 6 h at 37°C, the cells were lysed, and the lysates were assayed for β-galactosidase activity. The results are reported as the percentage of β-galactosidase activity compared with that seen in the absence of added CD4mc. The results of a single experiment, with each point done in duplicate/triplicate, are shown. The experiment was repeated with comparable results. The error bars indicate standard deviations. (B) 293T cells expressing the HIV-1_JR-FL Env were radiolabeled with [³⁵S]cysteine and [³⁵S]methionine in medium alone or in the presence of DMSO, 20 μg/ml sCD4, or different concentrations of BNM-III-170 for 24 h at 37°C. The cell supernatants were then used for precipitation by a polyclonal antiserum from an HIV-1-infected individual. The precipitated gp120 was analyzed by SDS-PAGE, autoradiography, and densitometry. The background shedding of gp120 in the untreated control was subtracted from all of the other values. *, value less than or equal to the background of the assay.