Figure 4.
SAMHD1K11A has reduced sensitivity to VpxMAC mediated degradation. A) 293 T cells were transfected with plasmids encoding YFP, YFP-SAMHD1WT, or YFP-SAMHD1K11A along with plasmids encoding for Flag-tagged Vpx proteins from HIV-2ROD, SIVMAC or SIVRCM as indicated and percentages of YFP-positive cells were determined 24 h after transfection. A parallel sample was used for western blot analysis using an anti-Flag antibody (lower panel). Tubulin served as a loading control. B) YFP-tagged SAMHD1WT, SAMHD1K11A or SAMHD1K11A-SV40NLS were co-transfected in parallel with Flag-Vpx expression plasmids as in A) and percentages of YFP + cells were measured 24 h post transfection. Fold changes are indicated above the bars. C) Time course of VpxMAC VLP or control VLP treatment of THP-1 cells expressing YFP-SAMHD1WT or YFP-SAMHD1K11A. Western blot analysis was performed with an anti-SAMHD1 antibody detecting endogenous SAMHD1 as well as ectopically expressed YFP-SAMHD1 (arrow). Hsp90 served as loading control.