Figure 5.
CEP57 depletion leads to an increase of PCM at the acentriolar pole and facilitates spindle bipolarization in one-centrosome cells.(A) Mitotic spindle pole structures of one-centrosome cells upon CEP57 depletion. Green, red, and blue represent centrin, pericentrin, and DNA, respectively. Z-projections: 20 planes, 0.5 µm apart. Scale bar, 5 µm. Arrowheads indicate the PCM at the acentriolar spindle pole. (B) The signal intensity of pericentrin on centrosomes or PCM poles in A. Line and error bars represent the mean and SD (n ≥ 50 cells from two independent experiments). Kruskal–Wallis test was used to determine the significance of the difference. *, P < 0.01. (C) Schematic illustration of CEP57-depletion-induced pericentrin accumulation at the PCM pole. (D) Time-lapse observation of NuMA structures and microtubules upon CEP57 depletion. Centrinone-treated one-centrosome HeLa cells expressing EGFP-centrin1 and mCherry-NuMA were observed with a 40× objective. Red, green, and gray represent mCherry-NuMA, EGFP-centrin1, and SiR-tubulin, respectively. Z-projections: 10 planes, 2.2 µm apart. Scale bar, 10 µm. Time zero corresponds to NEBD. (E) The time required for the initial establishment of two poles of NuMA in D. Line and error bars represent the mean and SD (n ≥ 50 cells from two independent experiments). The Mann–Whitney U test (two tailed) was used to obtain a P value. n.s., not significantly different (P > 0.05). (F) Mitotic duration, the time required from NEBD to cytokinesis, in D. Line and error bars represent the mean and SD (n ≥ 50 cells from two independent experiments). The Mann–Whitney U test (two tailed) was used to obtain a P value. *, P < 0.0001. (G) Frequency of mitotic spindle structures upon CEP57 depletion. Values are presented as mean percentages ± SD (n = 6, triplicates, two independent experiments, at least 29 spindles in each assay).

CEP57 depletion leads to an increase of PCM at the acentriolar pole and facilitates spindle bipolarization in one-centrosome cells. (A) Mitotic spindle pole structures of one-centrosome cells upon CEP57 depletion. Green, red, and blue represent centrin, pericentrin, and DNA, respectively. Z-projections: 20 planes, 0.5 µm apart. Scale bar, 5 µm. Arrowheads indicate the PCM at the acentriolar spindle pole. (B) The signal intensity of pericentrin on centrosomes or PCM poles in A. Line and error bars represent the mean and SD (n ≥ 50 cells from two independent experiments). Kruskal–Wallis test was used to determine the significance of the difference. *, P < 0.01. (C) Schematic illustration of CEP57-depletion-induced pericentrin accumulation at the PCM pole. (D) Time-lapse observation of NuMA structures and microtubules upon CEP57 depletion. Centrinone-treated one-centrosome HeLa cells expressing EGFP-centrin1 and mCherry-NuMA were observed with a 40× objective. Red, green, and gray represent mCherry-NuMA, EGFP-centrin1, and SiR-tubulin, respectively. Z-projections: 10 planes, 2.2 µm apart. Scale bar, 10 µm. Time zero corresponds to NEBD. (E) The time required for the initial establishment of two poles of NuMA in D. Line and error bars represent the mean and SD (n ≥ 50 cells from two independent experiments). The Mann–Whitney U test (two tailed) was used to obtain a P value. n.s., not significantly different (P > 0.05). (F) Mitotic duration, the time required from NEBD to cytokinesis, in D. Line and error bars represent the mean and SD (n ≥ 50 cells from two independent experiments). The Mann–Whitney U test (two tailed) was used to obtain a P value. *, P < 0.0001. (G) Frequency of mitotic spindle structures upon CEP57 depletion. Values are presented as mean percentages ± SD (n = 6, triplicates, two independent experiments, at least 29 spindles in each assay).

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