Figure 6.

Rab6 depletion inhibits trafficking of gD, gE and tsVSV-G, but not Gaussia luciferase. A) HeLa cells on coverslips were transfected with neg, Rab1 or Rab6 siRNAs and 2 days later transfected with plasmids expressing gD or gE. After 16 h the cells were processed for cell surface staining with the appropriate antibody, incubated with secondary antibody (red), and stained with DAPI (blue). B) HeLa cells on coverslips were transfected with neg, Rab1 or Rab6 siRNAs and 2 days later transfected with plasmid expressing GFP-tsVSV-G protein (green). The cells were incubated overnight at the non-permissive temperature of 39°C (pre-shift), then incubated at the permissive temperature of 32°C for 4 h (post-shift) before fixation and staining with DAPI (blue). C) HeLa cells treated as indicated were transfected with a plasmid expressing Gaussia luciferase and incubated for 16 h. Media was changed and 1 h later the level of secreted Gaussia luciferase was measured, with the amount secreted from cells transfected with the neg siRNA taken as 1. As a positive control BFA was added to cells at a concentration of 5 µg/mL before assay. Un, untransfected cells. Error bars indicate standard error from three experiments. Scale bar = 10 µm.