Figure 3.

PrP^∗ Traffics to the Plasma Membrane during RESET

(A) Time-lapse images of YFP-PrP^∗-expressing cells treated with TG + methyl-β-cyclodextrin (MβCD) (top) or TG alone (bottom) taken at two different focal planes. Mid-cell indicates a focal plane at the widest point of the nucleus and coverslip indicates a focal plane close to the coverslip where the largest portion of the plasma membrane is in focus.

(B) YFP-PrP^∗ cells were untreated or treated with TG and MβCD and then fixed after 90 min and stained with anti-GFP antibody without permeabilization to detect cell surface YFP-PrP^∗.

(C) Antibody uptake assay for internalization of YFP-PrP^∗ from the plasma membrane. YFP-PrP^∗-expressing or untransfected (Untf’d) cells were treated for 1 hr with TG in the presence of leupeptin and either anti-GFP or anti-Myc. The cells were then washed, fixed, permeabilized, and stained with Cy5-conjugated secondary antibody to detect internalized antibody.

Scale bars, 10 μm.