Figure 6
CP110 depletion suppresses p27K effects on centrosome reduplication and micronucleation. (a) Parental (Par) and p27K (clones 2 and 8) cells were transfected with 40 nM non-targeting (NT) or CP110 (CP) siRNA using Dharmafect 1. Cells received Dox for 32–36 h after transfection. Cell extracts were immunoblotted with antibody to CP110, HA or actin (loading control). (b, c) Cells on coverslips were transfected with siRNAs and treated with Dox as in panel (a) and stained with DAPI and immunostained with antibody to γ-tubulin. Number of cells with more than two centrosomes (b) or with a phenotype indicative of mitotic catastrophe (c) was determined. Three independent experiments were performed, and at least 100 cells were examined per sample with an Axio Imager Z1, Carl Zeiss upright fluorescent microscope at 63 × magnification. Error bars indicate standard deviation. Statistical significance was evaluated with a two-tailed, unpaired Student's t test (assuming unequal variances) using Microsoft's Excel 2003 software. P values <0.05 (denoted by double asterisks) are considered significant; P values between 0.05 and 0.1 (denoted by single asterisk) are considered suggestive. P values for the data in panel (b) are: 0.845 (parental), 0.02 (clone 2) and 0.046 (clone 8). P values for the data in panel (c) are: 0.378 (parental), 0.033 (clone 2) and 0.032 (clone 8).
