Tissue memory CD4+ T cells expressing IL-7 receptor-alpha (CD127) preferentially support latent HIV-1 infection
Fig 2
Relieving SAMHD1 restriction does not increase permissiveness of CD127+ Tm cells to productive infection by HIV-1.
A) The HLAC Tm subsets express low levels of phospho-SAMHD1. Amounts of SAMHD1 phosphorylated at Thr592 in THP1 cells, total HLACs, Tn cells, and the three Tm subsets, as determined by Western blot. Beta-actin served as a loading control. Both short and long exposures of the phospho-SAMHD1 blots are shown. B) Vpx degrades SAMHD1. Amounts of total SAMHD1 in unstimulated PBMCs that were left uninfected, or infected for 3 days with F4.HSA or F4.HSA harboring Vpx-Vpr fusion protein, as determined by Western blot. 2:1 and 1:2 refer to the ratios of F4.HSA provirus to Vpx-Vpr plasmid in the co-transfections. GAPDH served as a loading control. C) Vpx increases permissiveness of resting PBMCs to productive infection by F4.HSA. Unstimulated PBMCs were mock-treated or exposed for 3 days to F4.HSA lacking Vpx, or harboring different amounts of Vpx. Infection levels were then assessed by flow cytometry. Results are pre-gated on live, singlet CD3+CD8-CD45RO+CD45RA- cells. D) Vpx does not increase HIV infection of CD127+ Tm cells. Representative flow cytometric plots of HLACs mock-treated or exposed for 3 days to F4.HSA lacking or harboring Vpx as indicated. Expression levels of CD57 and CD127 were assessed in uninfected memory CD4+ T cells (plot on left), and HIV-infected (HSA+) memory CD4+ T cells (3 plots on right). Results are pre-gated on live, singlet CD3+CD8-CD45RO+CD45RA- cells. E) Vpx does not increase HIV infection of CD127+ Tm cells. Bar graphs comparing the absolute cell counts of CD57+, CD127+, and CD57-CD127- Tm cells amongst HIV-infected (HSA+) memory CD4+ T cells in two independent donors. Absolute cell counts were determined by normalizing the flow cytometric data to AccuCount beads run for each sample. Error bars correspond to experimental triplicates for each donor. *p<0.05, **p<0.01 as determined using a 2-tailed unpaired student’s t-test.