Fig. 8

HIV-1 transmitted/founder (T/F) viruses from subtypes B and C conserve the ability to antagonize autophagy. A, B HEK293T cells were transfected with the proviral DNA of HIV-1 NL4-3, HIV-1 NL4-3 Δnef or the selected T/F clones that belong to the pandemic HIV-1 subtype B (A) or subtype C (B). 24 h later, the cell medium was replaced and supplemented with rapamycin (6.5 μM) or DMSO. 18 h later, the percentage of maximal virus production was measured by the accumulation of HIV p24 in the culture supernatant relative to the DMSO treatment. Bottom blots: Cell lysates were also analyzed by western blot for p55 and ACTB. In each case, the percentage of maximal virus production is indicated as the mean and SEM from 3 independent biological replicates. C, D HEK293T cells were co-transfected with EGFP-LC3B and the proviral DNA of HIV-1 NL4-3, HIV-1 NL4-3Δnef or the selected T/F clones that belong to the pandemic HIV-1 subtype B (C) and subtype C (D). 48 h post-transfection, cells were treated with DMSO or rapamycin and subsequently analyzed by flow cytometry for autophagosome-associated EGFP-LC3. Data correspond to the mean and SEM from three independent replicates. Significantly different values are indicated by asterisks *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001