Supra-second tracking and live-cell karyotyping reveal principles of mitotic chromosome dynamics.

Mitotic chromosome dynamics are essential for the three-dimensional organization of the genome during the cell cycle, but the spatiotemporal characteristics of this process remain unclear due to methodological challenges. While Hi-C methods capture interchromosomal contacts, they lack single-cell temporal dynamics, whereas microscopy struggles with bleaching and phototoxicity. Here, to overcome these limitations, we introduce Facilitated Segmentation and Tracking of Chromosomes in Mitosis Pipeline (FAST CHIMP), pairing time-lapse super-resolution microscopy with deep learning. FAST CHIMP tracked all human chromosomes with 8-s resolution from prophase to telophase, identified 15 out of 23 homologue pairs in single cells and compared chromosomal positioning between mother and daughter cells. It revealed a centrosome-motion-dependent flow that governs the mapping between chromosome locations at prophase and their metaphase plate position. In addition, FAST CHIMP measured supra-second dynamics of intra- and interchromosomal contacts. This tool adds a dynamic dimension to the study of chromatin behaviour in live cells, promising advances beyond the scope of existing methods.