Fig. 5

Functional analysis of both variants (p.T55 M and p.S308 T) in AP5Z1. (a and b) HEK293 cells were respectively transfected with WT and mutant plasmids (T55 M, S308 T). Western blot analysis revealed that both AP5Z1 variants decreased the level of AP-5 ζ protein. c ICH analysis revealed that Hela cells transfected with mutant plasmids (T55 M, S308 T) showed larger and brighter LAMP1-positive puncta as compared with cells transfected with the WT plasmid. Scale bar = 20 μm. (d and e) LAMP1 fluorescence intensity and area per cell were quantified in more than 100 cells quantified per visual field. Experiments were replicated three times. f Hela cells were respectively transfected with WT and mutant plasmids (T55 M, S308 T), then assessed for morphological changes by TEM. Ultrastructural analysis revealed that both AP5Z1 variants led to the accumulation of enlarged morphologically defined endocytic structures filled with aberrant storage material, including many intraluminal vesicles. Scale bar = 500 nm. Error bars represent SEM, *p < 0.05