FIGURE 8:

Sx16-ΔTM or Sx16 knockdown prevents the midbody accumulation of ALIX and causes reduced levels of Cep55. HeLa cells were infected with Sx16-ΔTM or Sx12-ΔTM virus as described. Cells were immunostained with antibodies against ALIX or Cep55. (A) Typical immunofluorescence staining of ALIX at the midbody of control (Sx12-ΔTM–infected) cells, revealed as a characteristic double ring–like structure. Cep55 also localized to the midbody of these cells. By contrast, ALIX did not localize to the midbody of Sx16-ΔTMxinfected cells, and Cep55 localization was impaired in more than half of the cells analyzed (see the text for explanation). The data shown are representative of three experiments of this type. (B) HeLa cells were transfected with an siRNA SmartPool designed to knock down Sx16 or a scrambled siRNA SmartPool control, then incubated for 48 h after transfection before fixation and staining with anti-ALIX as indicated. Data from a representative experiment, repeated three times with similar results. In all experiments, the extent of Sx16 knock down was >85% as determined by immunoblotting (unpublished data).