Figure 3

The effect of CPSF6-358 on the infectivity of HIV-1 CA mutants correlates with the effect of TNPO3 KD. (A) Schematic representation of the protein domains of WT CPSF6 and the truncated mutant CPSF6-358. RNA recognition motif (RRM), proline-rich domain (P-rich), arginine/serine rich domain (RS). (B) Expression levels of CPSF6 in TZM-bl cells transduced with empty or CPSF6-358 vectors. Cell lysates were probed in western blots with anti-CPSF6 antibody (upper panel) and anti-β-actin antibody (lower panel). The upper panel shows the endogenous CPSF6 and the truncated form. (C) TZM-bl cells transduced with an empty vector or with a vector encoding CPSF6-358 were challenged with a panel of 27 HIV-1-GFP reporter vectors bearing either WT CA or the indicated CA mutants. At 72 hrs the percent GFP⁺ cells was determined by flow cytometry as an indication of infectivity. The ratio of HIV-1 infectivity in CPSF6-358 expressing cells and empty vector cells is shown. White bars show CA mutants inhibited to a similar extent as the WT virus by CPSF6-358, black bars shows CA mutants insensitive or slightly sensitive to CPSF6-358, and gray bars show CA mutants hypersensitive to the presence of CPSF6-358 in the cell. (D) Correlation between the infectivity ratios of the 27 CA mutants when infecting Ctrl KD vs TNPO3 KD [8] and Empty vector vs CPSF6-358 (R² = 0.8528).