Chromosomes as physical barrier can perturb centrosome clustering in diploid cells with centrosome amplification. (A–D) Stills of time-lapse videos of mitotic SakOE, mad2^mut NBs expressing tubulin-GFP (green) and histone-RFP (red and gray in the bottom insets). Orange and white dotted circles surround cells and nuclei, respectively. Time of mitosis is indicated in minutes:seconds, and time 00:00 corresponds to NEBD. (A) SakOE, mad2^mut NB with several MT-nucleating sites builds up a bipolar spindle as centrosomes (arrows) are not blocked by a DNA barrier. (B) SakOE, mad2^mut NB presents several MT-nucleating sites and assembles a tripolar spindle containing DNA (arrowhead) between two poles (arrows). As mitosis proceeds, the DNA is pushed toward the metaphase plate, and the spindle becomes bipolar. (C) SakOE, mad2^mut NB with several MT-nucleating sites assembles first a tripolar spindle. A DNA mass acts as a barrier blocking the clustering of two poles (arrows). At anaphase onset, a pseudo-bipolar spindle, resulting from partial centrosome clustering, generates two nuclei. (D) SakOE, mad2^mut NB with several MT-nucleating sites displays multipolar division, as unresolved DNA barrier inhibits pole coalescence. (E) Table showing the number and the percentage of cells in each indicated category in SakOE, mad2^mut NBs (n = 59 NBs).