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. 2004 Nov 8;167(3):531–543. doi: 10.1083/jcb.200408165

Figure 5.

Figure 5.

Inactivation of REs causes accumulation of VSV-G in the perinuclear region. MDCKT cells were infected to express VSV-G-YFP (third column, green) and incubated overnight at 40°C. Tfn-HRP (third column, blue) was internalized for 45 min at 40°C and was chased into REs by incubating cells in media without Tfn-HRP for 15 min at 40°C. Control cells were subject to only DAB while the samples were exposed to DAB and H2O2 for 1 h in the dark. Cells were washed in warm media and incubated at 31°C for 1.5 h in media/CHX to release VSV-G from the ER. Cells were washed in PBScmf, trypsinized, fixed, and processed for immunofluorescence. Cell surface VSV-G labeling (second column, red) using an antibody, TKG, against the ectodomain of VSV-G was performed on nonpermeabilized cells before permeabilization and internal labeling for HRP. Arrows denote accumulation of intracellular VSV-G in the perinuclear region.