Figure 5

Sensitivity vector metrics reveal differences in CD4/CCR5 usage efficiencies between Transmitter/Founder (T/F) and chronic envelopes. Normalized infection data using T/F and chronic Env clones were analyzed using VERSA. (A) Vector angle, (θ), (B) mean infectivity (M), and (C) vector amplitude (Δ) values were obtained for each Env clone. Each Env was profiled twice, in triplicate, across 25 combinations of CD4/CCR5 expression. Average metrics of 6 individuals from each group (T/F or chronic, N=12) are shown, each group consisting of 900 data points. The median value of each metric for the T/F and chronic Env cohorts is marked by a line. p values were generated by the non- parametric unpaired t test (***p = 0.0003; *p = 0.05). (D and E) The normalized infectivity for the chronic (blue line) and T/F envelopes (red line) are averaged, and compared as a group at (D) low and (E) high levels of CCR5 expression, across varying levels of CD4 as indicated. (F) Wedge plot of the average angle and amplitude (+/- S.D.) obtained for T/F (dark grey) versus chronic envelopes (light grey). (G) The infectivity profile of individual T/F and chronic Envs (from Additional file 5: Figure S3) were averaged to form their respective group profile. 2-D contour plots representing the averaged infectivity profiles of T/F and chronic envelopes are shown. (H) T/F Envs and macrophage tropic (YU2, ADA) and non-macrophage tropic (JRCSF) R5 Envs were used to produce Env pseudotyped luciferase reporter viruses, which were subsequently titrated on JC53 cells. Monocyte derived macrophages were inoculated with equivalent infectious units of each reporter virus, and luciferase activity measured in cell lysates at 72hrs post infection. Results of infection in 3 independent donors are shown. Results are means of triplicate wells, and error bars represent standard deviations.