FIG 9.

The Q4R mutation accelerates initiation of uncoating of the RGDA/Q112D virus. (A) (Top) Time-lapse images of iGFP-tdTomato-Vpr infection in Jurkat cells (a bright-field cell reference is shown in gray). The GFP signal (green) detected in viral particles, as reported by the tdTomato-Vpr signal (red), was reduced over time until it completely disappeared. (Bottom) Display of the mean particle intensity of the GFP signal (green) of the particle shown in the images at the top. When fusion occurs, there is first a drop of the GFP signal (52.5 min), and when the capsid integrity is compromised, there is a complete loss of the GFP signal (82.5 min). A.U., absorbance units. (B) The difference in the time (t) of initiation of uncoating (integrity loss) and the time of fusion (Δt) was calculated for each individual tracked particle. A representation of all iGFP-tdTomato-Vpr particles that fused into the cell is shown. The results for particles that kept HIV-iGFP until the end of the time lapse (>120 min), particles that lost all GFP at fusion (leaky capsid), and particles that showed two signal losses (violin density/probability plots) are shown. For particles with dual drop events, respective medians are shown with white dots, boxes show the interquartile ranges, and vertical lines show the range of all dual drop events. P values were determined by the Kruskal-Wallis test followed by Dunn’s multiple comparison. ****, P < 0.0001.