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Figure 3


Fig. 3. Kinetics of GBF1 binding to and dissociation from Golgi and VTCs membranes in NRK cells stably expressing GFP-GBF1. Live NRK cells expressing GFP-GBF1 were examined by confocal microscopy at 37°C as described in Materials and Methods. (A) Cytosol FRAP was performed as described in Materials and Methods. Shown are still images at the indicated times. (B) Golgi FRAP. Experiment similar to that shown in 3A except that the ROI includes the Golgi complex. (C) Quantification of the Golgi FRAP experiment presented in 3B. The curve was obtained by fitting FRAP data to a single exponential corresponds to the equation: y=0.520 * (1-e-0.055t) + 0.381 with an R value of 0.991. (D) Golgi FRAP in absence of microtubules. FRAP analysis was performed as in 3B, except that cells were chilled on ice for 15 minutes and warmed to 37°C in the presence of 1 µg ml-1 NOZ. (E) VTCs FRAP. Similar to experiment shown in 3B except that the ROI was located in the cell periphery and contained GBF1-positive VTCs. Bars, 5 µm.





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