Figure 4.

Viral protein U blocks the degradation of Cactus. (A) Viral protein U (Vpu) affects Drosomycin (Drs) expression in S2 cells. Cells were co-transfected with 1 μg of an expression vector that contains a copper-inducible TollΔLRR construction (Tauszig et al., 2000), plus 1 μg of an empty expression vector (E), a vector that contains Vpu, or a vector that contains Vpu2/6, and 0.1 μg of a reporter plasmid that encodes luciferase under the control of the Drs promoter. Forty-eight hours after transfection, CuSO₄ (500 μM) was added to induce the expression of the TollΔLRR construct. After 60 or 120 min, the cells were harvested and luciferase activity was determined from cell extracts as described in the supplementary information online. C, control (no copper). (B) Vpu blocks Cactus degradation. S2 cells were stably transfected with a copper-inducible Torso–Pelle construct (Silverman et al., 2000). Cells were transfected with 1 μg of an empty expression vector (lanes 1 and 2) or with vectors that contain Vpu (lane 3) or Vpu2/6 (lane 4). Thirty-six hours after transfection, CuSO₄ was added to induce the expression of the Torso–Pelle construct. Cellular protein extracts were prepared and were analysed on a western blot using an anti-Cactus antibody. A nonspecific band (indicated with an asterisk) was used as a loading control.