Figure 5.

Acute removal of CPAP does not perturb centrosome PCM recruitment or organization of PCM components during mitosis. (A) Cells were treated with IAA for 6 h and immunolabeled for indicated PCM proteins. Imaging and quantification of the intensities of PCM proteins show comparable centrosomal levels in control and IAA-treated cells. Histograms show the average intensity ± SD, n = centrosome number. (B) 2DSTED imaging shows that the patterns of localization of three major components of the mitotic PCM lattice, Cep192, Cep215, and pericentrin, are not changed after 2.5 h of IAA treatment. (C) MT nucleation recovery after cold treatment. IAA treatment for 2.5 h does not change the recovery of MT nucleation or centrosomal levels of Cep192. (D) Structurally intact mother centrioles organize control-looking mitotic spindles. Cells were treated with IAA or depleted by siRNA, and centrinone was added to prevent centriole duplication, as described in Fig. 3 G. Cells were fixed at 24 h when they were undergoing mitosis, immunolabeled for γ-tubulin and CPAP, and imaged by wide-field microscopy; spindle poles were imaged by STED. Scale bars: 5 µm (A); 0.5 µm (B); 5 µm (C); 1 µm (enlarged centrosomes in C); 10 µm (wide-field in D); 1 µm (STED in D).