FIG. 5.

Differential extraction of VP22 from cellular membranes. A7 cells were transfected with the plasmid encoding VP22-GFP and were metabolically labeled with EXPRE³⁵S³⁵S protein labeling mix for 2.5 h, prior to harvesting at 22.5 h posttransfection. Following incubation in hypotonic buffer, the swollen cells were disrupted by Dounce homogenization, and nuclei were removed by low-speed centrifugation. Postnuclear supernatants were either treated with 1 M NaCl, 0.1 M Na₂CO₃, or 1% Triton X-100 or mock treated for 1 h prior to membrane flotation gradient centrifugation. Membrane flotation gradient centrifugation was then performed as described in Materials and Methods. PhosphorImager quantification was performed as described in the legend to Fig. 4C, and results are displayed graphically.