Vesicle capture and mitochondrial aggregation using rerouting of TPD52-like proteins. (A) Representative confocal micrographs showing the corerouting of GFP-TPD52 after rerouting of mCherry-FKBP-TPD54 (top) or mCherry-FKBP-TPD54(L53P,L67P; PP; bottom) to dark MitoTrap by addition of 200 nM rapamycin. Inset: 5× zoom. Scale bar, 10 µm. (B) CLEM experiment to test for vesicle capture using TPD52. Cells expressing GFP-FKBP-TPD52 and mCherry-MitoTrap with rapamycin 200 nM for 3 min. Insets: 4× zoom. Scale bars, 500 nm or 50 nm (inset). (C) Representative confocal micrographs showing rerouting and mitochondrial aggregation. GFP-FKBP (control) or GFP-FKBP-TPD52-like (POI, green) proteins were coexpressed with mCherry-MitoTrap (red) in cells treated with control siRNA (siControl) or TPD54 siRNA as indicated and treated with 200 nM rapamycin for 30 min. Scale bar, 10 µm. (D) Quantification of mitochondrial aggregation. Dots show average mitochondrial shape per cell. Box plots show IQR, and bar represents the median and whiskers show 9th and 91st percentiles. Right: P values from Dunnett’s post hoc tests using respective GFP-FKBP as control.