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. 2020 Mar 2;9:e51035. doi: 10.7554/eLife.51035

Figure 2. Combined effects of Nef and Vpr completely remove MR from a significant proportion of infected cells at early time points.

(A) Diagram of HIV NL4-3 ∆GPE-GFP. (B) Western blot analysis of whole cell lysates from 293T cells transfected with the indicated viral expression construct. (C) Flow cytometry plots indicating the gating strategy used to identify live GFP+ vs GFP- cells and the fraction of cells that are MRlow. (D) Representative flow cytometric analysis of MDM at five days post transduction by the indicated virus. The percentage of GFP+ cells that fell into the MRlow gate is indicated in each panel. (E) Summary graph depicting the percentage of GFP+ cells that fell into the MRlow gate in transduced MDM. For the uninfected column the results from GFP- cells are displayed. (Each dot indicates an independent donor, range 3–11). (F) Western blot analysis of whole cell lysates from 293T cells transfected with the indicated viral expression construct. (G) Summary graph depicting the frequency of transduced (GFP+) MDM at the time of harvest. (H) Representative flow cytometric plots of MDM transduced with the indicated adenoviral vector (n = 3 independent donors). For parts E and F mean +/- standard deviation is shown. Statistical significance was determined by a two-tailed, paired t-test. N.S. not significant, **p<0.01, ***p<0.001, ****p<0.0001.

Figure 2.

Figure 2—figure supplement 1. HIV Vpr reduces steady state levels of UNG2 but not MR in co-transfected 293T cells.

Figure 2—figure supplement 1.

(A) Flow cytometric plots of 293T cells co-transfected with NL4-3 ∆GPE-GFP, pCDNA3.1-hMR, and pMSCV 3x FLAG UNG2 IRES-GFP as indicated. (B) Western blot analysis of 293T cells co-transfected exactly as in A. Env-GFP indicates the location of the fusion protein containing the N terminus of Env followed by GFP as described in Methods.