Fig. 5. Dispersion of the Golgi pool of palmitoylation-deficient GAD65-GFP but not wt GAD65-GFP following treatment with BFA. Selected single-layer confocal images obtained from a live-cell imaging time series recorded immediately after the addition of BFA (5 µg/ml) to COS-7 cells coexpressing GalT-mRFP1 and either the palmitoylation-deficient GAD65(C30,45A)-GFP (A) or wt GAD65-GFP (B). (A) BFA treatment has resulted in the complete dissociation of the Golgi pools of both GAD65(C30,45A)-GFP and GalT-mRFP1 at the 15 minute time-point following treatment with BFA (see also supplementary material Movie 4). Note that the cell in the lower right corner has protein aggregates because of overexpression, precluding analyses of the effect of BFA. (B) The Golgi pool of GalT-mRFP1 has redistributed to the ER at the 15 minute time-point following exposure to BFA (5 µg/ml). By contrast, only a fraction of the Golgi pool of wild-type GAD65-GFP has dispersed at 15 and 30 minutes, and a prominent fraction of the protein is still detected in punctate structures in the perinuclear region at those timepoints (see also supplementary material Movie 5). The effect of BFA is global and the cells shown in A and B are representative of cells transfected with palmitoylation-deficient GAD65-GFP and wt GAD65-GFP, respectively. Note that GAD65-containing cytosolic vesicles are not visible in the single confocal layer selected for imaging of the Golgi compartment.