Figure 4: Cholesterol is redistributed to late endosomes/lysosomes in VPS35 KD HeLa cells. | Nature Communications

Figure 4: Cholesterol is redistributed to late endosomes/lysosomes in VPS35 KD HeLa cells.

From: Arf6 controls retromer traffic and intracellular cholesterol distribution via a phosphoinositide-based mechanism

Figure 4

(a) Western blot analysis of VPS35 levels in HeLa cells transfected with scramble or VPS35 siRNA. Tubulin was used as an equal loading marker. (b) Representative confocal images of HeLa cells transfected with scramble or VPS35 siRNA, immunostained for LAMP1 (magenta) and VPS35 (green) and labelled with filipin (blue). Scale bar, 5 μm. (c) Normalized integrated densities of filipin staining in scramble (100±5%,±indicates s.e.m., n=29 cells, three experiments) and VPS35 siRNA cells (124±9%, n=28 cells, three experiments). *P<0.05 in t-test with Welch’s correction. (d) Quantification (left) and frequency distribution (right) of the number of filipin puncta in scramble (22±4 puncta per cell, ± indicates s.e.m., n=33 cells, three experiments) and VPS35 siRNA cells (39±5 puncta per cell, n=32 cells, three experiments). *P<0.05 in Student’s t-test and ***P<0.001 in χ2-test. (e) Scatter dot blot of the size of filipin puncta in scramble (0.71±0.03 μm2, ± indicates s.e.m., 741 puncta from 33 cells) and VPS35 siRNA cells (0.73±0.02 μm2, 1,241 puncta from 32 cells). **P<0.01 in Mann–Whitney test.

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