Fig. 7. Prestin YFP uses transferrin containing endosomes to transit from the Golgi to the basolateral membrane.

MDCK cells were transiently transfected with prestin-YFP and kept at 19°C to induce Golgi block. Cells were incubated with transferrin - Alexa 647 for 10 minutes. The incubation temperature was raised to 37°C after which cells were fixed at 0, 1 minute, 5 minutes, 10 minutes and 15 minutes. Cells were then imaged with confocal microscopy. Shown are confocal images in the X-Y plane at the basolateral Z axis of the cell that were fixed at different times after raising the temperature to 37°C. With increasing time there is co-localization of prestin-YFP exiting the Golgi with transferring - Alexa 647. The right hand panels shows merged and individual Alexa 647 and prestin-YFP images of an enlarged area at 15 minutes after raising the temperature to 37°C. Co-localization of prestin YFP and transferrin 647 is demonstrated. The scale bar is 10 microns.