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Fig. 5 | Cell Division

Fig. 5

From: Nucleoporin Nup58 localizes to centrosomes and mid-bodies during mitosis

Fig. 5

Nup58 depletion induced centrosomal abnormalities and delayed cytokinesis. a Brief agenda of collecting cells after Nup58 knockdown via siRNA transfection. b HeLa cells were transfected with control or Nup58 siRNA, then cell lysates were collected 72 h after transfection and analyzed by immunoblot of Nup58, Nup62, SAS-6, γ-tubulin, and β-actin expression. c qRTPCR analysis of Nup58, Nup62, and SAS-6 mRNA in HeLa cells after transfection with siRNA Nup58 for 72 h. Data show mean ± SD of three separate experiments. Significance difference was assessed with a Student’s t-test, **p < 0.01. d Quantification (relative %) of abnormal centrosome phenotypes in control or Nup58 siRNA-transfected HeLa cells. Values are based on three independent experiments (n = 500 cells). Mean values ± SD (error bars) are shown. Significance difference was assessed with a Student’s t-test, ****p < 0.0001. e Deconvoluted confocal images of synchronous HeLa cells, transfected with control or Nup58 siRNA, 72-h post-transfection. Green, anti-Nup58; red, anti-α-tubulin; blue, chromatin (DAPI). Scale bars, 5 µm. White arrows indicate bipolar and monopolar spindles as centrosome abnormalities when Nup58 was completely depleted. f Deconvoluted confocal images of HeLa cells 72-h post-transfection with control or Nup58 siRNA. Green, anti-Nup58; red, anti-acetylated-γ-tubulin; blue, chromatin (DAPI). Scale bars, 5 µm. Insets are magnifications of the midbody areas in observed cells. g Midbody maturation to abscission time in cells transfected with Nup58 siRNA (n = 30 cells) compared with control (n = 30 cells). Mean values ± SD (error bars) are shown, and significant differences were assessed with a Mann–Whitney U test, ****p < 0.0001. h Confocal live-cell images of stable GFP-α-tubulin expression in HeLa cells transfected with Nup58 siRNA shows delayed cytokinesis expression compared with the control. Insets are magnifications of microtubules areas in observed cells (Additional file 6: Video 4). Arrowheads indicate microtubules abscission. Scale bars, 5 µm

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