Figure 5.

Dispersal and mobility of centriolar satellites is impaired by sorbitol. (A) U2OS cells were left untreated (mock) or incubated in the presence of sorbitol (1 h). Cells were immunostained with antibodies against CEP131 and γ-tubulin, counterstained for nuclear content with DAPI, and subjected to confocal microscopy. Scale bars, 10 μm. (B) Cells from (A) were immunostained with antibodies against PCM1 and pericentrin, counterstained for nuclear content with DAPI, and subjected to 3D-SIM microscopy. Pictures represent reconstructed images shown as 3D projections. (C) U2OS cells were pre-treated with sorbitol (1 h) and incubated in the presence of nocodazole (3 h) as indicated. Cells were immunostained with antibodies against CEP131 and α-tubulin and counterstained for nuclear content with DAPI. Scale bars, 10 μm. (D) U2OS cells stably expressing low levels of GFP-CEP131 were incubated in the presence or absence of sorbitol and subjected to time-lapse microscopy. Note the high mobility of satellites in mock-treated cells compared to their near-complete immobility in sorbitol-treated cells (see also Supplementary movies 1 and 2). Scale bars, 5 μm. (E) Quantification of satellite velocities from (D). Trajectories of individual satellites were identified using the MOSAIC ImageJ/Fiji plug-in. Trajectories from three independent movie streams were used to calculate velocities of individual satellites. The two groups were compared using Wilcoxon rank sum test with continuity correction. p < 2.2 × 10⁻¹⁶. (see also Supplementary movies 3 and 4).