Fig. 4

Inhibition of VMP1 palmitoylation influences endosome-MVB-lysosome system. A Representative immunofluorescence images for the colocalization between ectopically expressed WT-VMP1/ MT-VMP1-Flag and LAMP1, the marker for lysosome. The white arrows in the magnified photos indicate colocalized VMP1 and LAMP1 in SCs. Right panel: Statistical analysis of the co-localization rate between Flag and LAMP1 in two groups. B Representative immunofluorescence showing the colocalization between WT-VMP1/ MT-VMP1-Flag and Rab11b, the specific marker for recycling endosome. Right panel is the co-localization analysis of Flag and Rab11b in two groups. C Representative immunofluorescence shows CD63 expression (marker of MVBs) in SCs transfected with WT-VMP1 or MT-VMP1-Flag. Right graph: quantification of relative CD63 fluorescence intensity of MT group normalized to WT group (control). D Electron microscopy displays the morphology and size of MVBs in SC transfected with MT-VMP1 or WT-VMP1-Flag (The red arrow represents the intraluminal vesicles in MVBs). E The statistical analysis of MVB size and number and intraluminal vesicles per MVB between two groups in image D. F Immunofluorescence of CD63 expression (MVB marker) in SCs overexpressing WT-VMP1 or MT-VMP1 treated with Baf A1 (20 nM) or DMSO. Right graph: quantification of the relative CD63 fluorescence intensity in the four groups. Data were presented as mean ± SEM, *P < 0.05. ns: not significant. G Western blot of CD63 expression (MVB marker) in SCs transfected with WT-VMP1 or MT-VMP1 treated with Baf1 or DMSO. Representative results from three independent experiments are shown