FIGURE 6.

Synj1 overexpression affects the homeostasis of early endosomes, but not of late endosomes, in SH-SY5Y cells. (A) SH-SY5Y cells stably transfected with cDNA encoding for wild-type Synj1 were tested for the expression of Synj1 by Western blotting. Representative immunoblotting of Synj1 in a single clone (cl5) and in a pool of clones (pool) is shown. Tubulin was used as the loading control. The molecular weight of protein markers is indicated. Densitometric analysis of three different experiments is shown. Error bars, mean ± SD. (B) Cells were stained with EEA1 antibody detected with Alexa-546-conjugated secondary antibodies. Serial confocal sections were collected from the top to the bottom of the cells; scale bars, 8 μm. For each condition, the 3D reconstruction and corresponding intensity map are shown; scale bars, 5 μm. The mean fluorescence intensity (arbitrary unit, a.u.) of EEA1-positive structures is higher in Synj1-overexpressing than in wild-type cells. The bars show the relative mean value ± SD of three independent experiments, n ≥ 50 cells. (C) Immunostaining of Rab7 in wild-type and Synj1-overexpressing cells is shown. For each condition, the corresponding intensity map is shown. Scale bars, 8 μm. Mean fluorescence intensity (arbitrary unit, a.u.) is shown. The bars show the relative mean value ± SD of three independent experiments, n ≥ 50 cells. ***p < 0.001, Student’s t -test.