Bimolecular Complementation to Visualize Filovirus VP40-Host Complexes in Live Mammalian Cells: Toward the Identification of Budding Inhibitors
Figure 5
Inhibition of HIV-1 Gag-Tsg101 interaction and VLP budding. (a) Human 293T cells grown on coverslips in 6-well plates were transfected with NYFP-Tsg101 + CYFP-Gag-WT (panels 1 and 2) or NYFP-Tsg101 + CYFP-Gag-ΔP6 (panel 3). Cells were washed, fixed, and examined for YFP fluorescence by confocal microscopy at 24 hours p.t. The image within the dotted square in panel 1 was enlarged and shown panel 2. (b) Human 293T cells grown on coverslips in 6-well plates were transfected with NYFP-Tsg101 + CYFP-Gag-WT in the presence of DMSO alone, or 25, 50, or 75 M of compound 5539-0062 as indicated. Cells were washed, fixed, and examined for YFP fluorescence by confocal microscopy at 24 hours p.t. (c) Standard budding assay and western blotting for Gag in VLPs and cell extracts from human 293T cells treated with DMSO alone, or the indicated concentrations of PTAP-inhibitor 5539-0062.
