Figure 3

Vpx-VLPs, exogenous dN treatment or SAMHD1 knock-down do not rescue the IFNα-induced block to HIV-1 infection in THP-1 cells. A. and B. VSV-G pseudotyped HIV-1 derived GFP reporter virus was produced in 293T cells and used to infect control or IFNα-treated PMA-differentiated THP-1 cells (A), or dividing THP-1 cells (B) at different MOIs (0.08 to 10), in the presence or the absence of either Vpx-VLP or 0.5 mM deoxyribonucleosides (dN). Levels of infection were monitored using flow cytometry to measure the percentage of cells expressing GFP, and mean values from 3 independent experiments are shown. C. Western blot analysis. Cell lysates from THP-1 cells were harvested 24 h post IFNα and Vpx-VLP treatment, and SAMHD1 expression was analysed by western blot; tubulin served as a loading control. D. THP-1 cells expressing a control shRNA or shRNA targeting SAMHD1 were differentiated with PMA or not in the presence or absence of IFNα for 24 h. Cells were infected with 3 different doses of VSV-G pseudotyped GFP reporter virus for 48 h, and GFP positive cells were enumerated by flow cytometry. The data are representative of 3 independent experiments with two independent shRNAs against SAMHD1. E. Western blot analysis of parallel samples from D. Protein levels of SAMHD1 and APOBEC3A (A3A) were determined and tubulin served as a loading control.