Fig. 6: Visualising amyloidogenic APP cleavage in live cells.
a Sketch depicting the principle of the APP cleavage detection assay at the single-molecule level: before cleavage, the N-terminus of APP has a slow membrane dynamics while it acquires a fast luminal dynamics after BACE1 cleavage, created with Biorender59. b Representative COS-7 cells expressing the ER retention and marker streptavidin::StayGoldER without (top) or with (bottom) BACE1-GFP overexpression as shown by the fuzzy signal in the green channel (left column). Right four panels show trajectories of the N-terminus tagged APP-HaloTag (stained with PA-JF646) reconstructed without (middle column) or with (right column) FidlTrack. Trajectories are colour-coded depending on whether they were classified as cleaved (orange) or uncleaved (cyan) based on their average trajectory displacement length. c Barplot of the average percentage of cleaved trajectories for three WT cells (n = 3, left, grey) and three BACE1 overexpressing cells (n = 3, right, pink) reconstructed without or with FidlTrack. d Barplot of the fold increase of average percent cleaved trajectories from BACE1 overexpression to WT (from (c)) without (left) or with (right) FidlTrack reconstruction. e Example of detected cleavage events in trajectories reconstructed with FidlTrack from a BACE1-overexpressing cell. The top row presents the instantaneous displacement length along the trajectories (smoothed with a 3-points moving average) with the pink arrow showing the cleavage point. The horizontal dashed lines represent the average displacement for the cleaved (orange) and uncleaved (cyan) populations. The bottom row shows the trajectories overlaid on the local ER structure and colour-coded by displacement length. Source data are provided as a Source Data file.
