Figure 2

Vpx-VLPs or exogenous dN treatment greatly enhance HIV-1 infection in IFNα-treated MDMs. A. VSV-G pseudotyped HIV-1 derived GFP reporter virus was produced in 293T cells and used to infect control or IFNα-treated MDMs at different MOIs (0.08 to 10, as determined in 293T cells), in the presence or the absence of either Vpx-VLP or 0.5 mM or 2.5 mM deoxyribonucleosides (dN). Levels of infection were monitored using flow cytometry to measure the percentage of cells expressing GFP, and mean values from 4 independent experiments (using cells from 4 different donors) are shown. Notably, treatment of MDMs with the higher concentration of dN changed their morphology, with the cells becoming rounder and less adherent. We conclude that experiments using high dN doses must be analysed with caution. B. MDMs were treated or not with IFNα for 24 h and subsequently incubated or not with Ctrl-VLPs, Vpx-VLPs or dN (at 0.5 or 2.5 mM as indicated) for 16 h before lysis. Whole cell dATP was quantified using a single nucleotide quantification assay. Mean values from 4 independent experiments (using cells from 4 different donors) are shown. C. Western blot analysis. Cell lysates from MDMs (donor #4) were harvested 24 h post IFNα and Vpx-VLP treatment, and SAMHD1 expression was analysed by western blot; Hsp90 served as a loading control.