Fig 1.
Role for Vpu in disruption of IRF3-dependent signaling. (A) Proviral constructs for HIV strains NL4-3, JR-CSF, and YU2 or control plasmid were transfected in 293 cells along with the IFN-β promoter luciferase construct and challenged with SeV to drive IRF3-dependent signaling. (B) (Top) Alignment of LAI, NL4-3, JR-CSF, and YU2 highlighting the start codon (arrow) for Vpu and the A-to-C transversion mutation found in the YU2 strain. *, upstream, nonproductive ATG. (Bottom) Amino acid alignment of Vpu from the same strains. (C) Vpu or Vif overexpression constructs tested as described for panel A. Vpu expression was titrated by expressing 50 ng to 375 ng of expression plasmid within equal DNA dosage transfections using control plasmid as filler; Vif expression from 375 ng of expression plasmid is shown. (D) Immunoblot analysis of either mock-infected HIV-1LAI-infected SupT1 cells or HIV-1LAI-infected SupT1 cells at 8, 24, or 48 h postinfection. Cell lysates were probed for IRF3, Vpu, HIV-1 Gag, or actin as a loading control. Luciferase reporter gene experiments were repeated 3 or more times, and representative immunoblot analyses are shown.