Fig. 6
Bub1 regulates CENP-AS7 phosphorylation and Aurora-A targeting at centromeres. a, b HeLa cells depleted of Bub1 by siRNA were immunostained with anti-p-CENP-AS7 (a) or anti-Aurora-A (b) and CREST antibodies. Magnifications (× 9) of single chromosomes are shown. Means ± SEM are shown (on the right of the panel) for the quantification of signal intensity relative to CREST obtained with antibodies against p-CENP-AS7 or Aurora A antibodies in a representative experiment (for p-CENP-AS7 intensity, n = 20 cells in each condition; for Aurora A intensity, n = 25 cells in control and 20 cells in Bub1 RNAi conditions; ****P < 0.0001; two-tailed unpaired nonparametric Mann–Whitney U-test). c Metaphase chromosome spreads from CENP-A-HA-WT, CENP-A-HA-S7A, and CENP-A-HA-S7D cell lines depleted of Bub1 were immunostained with antibodies against Bub1 and Sgo1. Scale bar = 5 µm. Inter-kinetochore distance means ± SEM of five to six representative cells with joined SC are shown (n = 10–50 chromosomes for each cell). Statistical analysis were performed on the mean of the distance measured for each cell (n = 5–6; ****P < 0.0001; ***P < 0.001; NS, not significant; two-way ANOVA with Tukey’s post hoc analysis)
