Early Delivery of Misfolded PrP from ER to Lysosomes by Autophagy
Figure 8
Autophagy participates in the delivery of Mut-PrP to LysoTracker labeled vesicles. (a) Three hours after transfection, HeLa cells expressing GFP-tagged Wt-PrP or Mut-PrP were treated for 16 h with 2 mM 3-MA, 2 μM rapamycin, or fresh media alone (untreated), followed by incubation with 50 nM LysoTracker Red for 30 min prior to live cell confocal fluorescence microscopy. Individual channels are displayed above merged images. Yellow indicates colocalization. Scale bars = 10 μm. Graphic display of Li’s correlation coefficient between Mut-PrP and LysoTracker for each treatment is shown to the right (***, student’s paired -test, difference from untreated, each). (b) Normal (ATG5+/+) and autophagy-deficient (ATG5−/−) mouse embryonic fibroblasts (MEFs) transiently transfected with GFP-tagged Wt-PrP or Mut-PrP were allowed to express for 16 h prior to confocal microscopy, performed 30 min following the addition of 50 nM LysoTracker Red. Single 0.5 μm slice images are shown. Each channel is displayed to the left of the merged image. Graphic display of Li’s correlation coefficient between PrP and LysoTracker Red in WT MEFs compared with ATG5−/− MEFs is to the right (**, student’s -test, each).
