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Fig. 6. The early secretory pathway is blocked in apoptotic cells. (A) (Top panel) HeLa cells were transfected with tsO45 VSV-G-GFP and incubated overnight at 39.5°C, then incubated in parallel for 2 hours at 39.5°C or 31°C as shown. They were stained without permeabilization, using anti-VSV-G, to score for transport to the cell surface. (Middle panel) Transfected cells were induced to enter apoptosis by UV irradiation and incubated for a further 4 hours at 39.5°C. Cells were either left at 39.5°C or shifted to 31°C for 2 hours, as indicated, then fixed in PFA. Apoptotic cells were identified using DAPI. (Bottom panel) As in middle panel, but with apoptosis induced by addition of 2 µM staurosporine. Results are presented as the mean ± s.e.m. (n=13-45). (B) Fluorescence microscopy of tsO45-VSV-G-GFP trafficking. UV-treated HeLa cells expressing VSV-G-GFP were treated as in A, and examined using confocal microscopy. The non-apoptotic cells show strong surface labelling (-VG), whilst the apoptotic cell (arrowhead) lacks surface label. Scale bar, 10 µm.

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