A quantitative map of human Condensins provides new insights into mitotic chromosome architecture

The two Condensincomplexes in human cells are essential for mitotic chromosome structure. We used homozygous genome editingto fluorescently tag CondensinI and II subunits and mapped their absolute abundance, spacing, and dynamic localization during mitosisby fluorescence correlation spectroscopy(FSC)–calibrated live-cell imagingand superresolutionmicroscopy. Although ∼35,000 CondensinII complexes are stably bound to chromosomes throughout mitosis, ∼195,000 CondensinI complexes dynamically bind in two steps: prometaphaseand early anaphase. The two Condensinsrarely colocalize at the chromatidaxis, where CondensinII is centrally confined, but CondensinI reaches ∼50% of the chromatiddiameter from its center. Based on our comprehensive quantitative data, we propose a three-step hierarchical loop modelof mitotic chromosome compaction: CondensinII initially fixes loops of a maximum size of ∼450 kb at the chromatidaxis, whose size is then reduced by CondensinI binding to ∼90 kb in prometaphaseand ∼70 kb in anaphase, achieving maximum chromosome compaction upon sister chromatidsegregation.