Figure 1.

Description and characterization of the pMorpheus-V5 viral reporter vector

(A) Representation of the HIV reporter viral vector pMorpheus-V5 (pLAI2-V5-NGFR HSA-mCherry-IRES-Nef).

(B) Gating strategy to identify cells latently or productively infected by pMorpheus-V5 using flow cytometry or CyTOF is depicted.

(C) Latently (V5+, NGFR+) or productively (HSA_POS, mCherry_POS, V5_POS, NGFR_POS, p24_POS) infected, IL-2 + α-CD3/CD28-stimulated CD4⁺ T cells were identified using flow cytometry after infection with pMorpheus-V5. The data show one representative example of four independent experiments using cells from different healthy donors. The percentage of CD4⁺ T cells in each quadrant is indicated.

(D) Nef down-regulates CD4 levels during pMorpheus-V5 infection. CD4⁺ T cells from two different donors were stimulated with IL-2 + α-CD3/CD28 and infected with pMorpheus-V5. CyTOF was performed 5 days post-infection (dpi). Histogram plots show the CD4 expression for uninfected cells (shaded light gray), p24_POS cells (orange), and p24_NEG cells (light blue). The data were analyzed using FlowJo.

(E) Cells were stimulated with IL-2 + α-CD3/CD28 prior to infection with p-Morpheus-V5. Histogram analysis of CD4 expression levels of NGFR (magenta), V5 (blue), mCherry (red), or HSA (green) positive cells. The data were analyzed using FlowJo. Data from two independent donors.