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. 2018 Nov 10;10(11):620. doi: 10.3390/v10110620

Figure 5.

Figure 5

Effect of deoxynucleoside addition or viral protein X (Vpxmac239)-mediated sterile alpha motif (SAM) and histidine/aspartate (HD) domain-containing protein 1 (SAMHD1) degradation on early HIV-1 replication. (A) MDMs were either pre-incubated or not with nucleosides (100 µM each) 2 h prior to infection or transduced with HIV-1 virus-like particles (VLPs) lacking or carrying Vpxmac239 12 h prior to infection. Subsequently, cells were infected with R5- or VSV-G pseudotyped HIV-1NL4-3. Infectivity was scored at day 6 p.i. by CA immunostaining. Error bars represent SEM of infection rates from all the replicates of all the donors. Statistical significance was assessed by a non-paired two-tailed Mann–Whitney test; **** p < 0.0001; *** p = 0.0001; n.s.: not significant. (B) MDMs from four donors in two independent experiments were pre-incubated for 2 h in medium (dashed lines), or medium supplemented with 100 µM deoxynucleosides (solid lines) and then infected with R5-pseudotyped HIV-1NL4-3. At 24 h p.i., medium was changed and 5 µM MVC was added. Solutions of 5 µM EFV (red lines) or 5 µM RAL (blue lines), or the equivalent volume of DMSO (gray lines) were added to the cultures at the indicated time points and infectivity was scored at day 6 p.i. The graph summarizes mean infectivity values from four donors, each measured in quadruplicate. Error bars represent SEM. (C) MDMs from four donors in two independent experiments were transduced with HIV-1 VLPs lacking (grey lines) or carrying (black lines) Vpxmac239. After 12 h, medium was changed, and cells were infected in quadruplicates with R5-pseudotyped HIV-1NL4-3. Solutions of 5 µM EFV (left, solid lines), RAL (right, solid lines), or DMSO (dashed lines) were added at the indicated time points. At day 6 p.i., the proportion of infected cells was scored by CA immunostaining. The graph summarizes mean values from four donors, each measured in quadruplicate. Error bars represent SEM. The unexpected effect of the addition of DMSO to the cultures in the first time points under native conditions (−Vpx) was most likely caused by technical errors at the moment of infection.