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

Figure 7. Knockdown of MR enhances Env expression and spread to T cells in vpr-null infection of MDM.

(A) Western blot analysis of MDM from two independent donors treated with the indicated silencing vector and infected with the indicated HIV for 10 days. The shRNA sequences encoded by the negative control vector (shNC) and the MR silencing vector (shMR) are described in Methods. (B) Schematic diagram of experimental protocol used for silencing experiments. (C) Representative flow cytometric plots showing frequency of infected (Gag+) primary T cells following two days of co-culture with autologous, HIV 89.6 infected primary MDM. T cells were identified in co-culture by gating on CD3+ CD14- cells as shown in Figure 7—figure supplement 1B. (D) Summary graph displaying relative infection of MDM and T cells as measured in C (n = 5 independent donors). Data in the left panel are unnormalized. In the right panel the data have been normalized to the wild-type condition for each donor and shRNA.

Figure 7.

Figure 7—figure supplement 1. Cell-to-cell infection from macrophages to autologous CD4+ T cells is highly efficient and enhanced by Vpr.

Figure 7—figure supplement 1.

(A) Diagram of the MDM and T cell co-culture experiments depicted in parts B, D, and E. (B) Representative flow cytometric plots and gating strategy used to identify MDM and T cells in co-culture and the fraction of Gag+ cells of both types. (C) Flow cytometric histograms illustrating the PacBlue signal detected in the indicated cell type following treatment with the indicated antibody. (D) Summary graph of the percentage of cells of the indicated type that are Gag+ following infection by HIV-1 89.6. (E) Summary graph of the percentage of cells of the indicated type that are Gag+ following infection by HIV-1 T/F clone REJO.