To maintain genomic stability, chromosome architecture needs to be tightly regulated as chromosomes undergo condensation during prophase and separation during anaphase, but the mechanisms remain poorly understood. Here, we show that the Plk1-binding protein PICH and Plk1 kinase coordinately maintain chromosome architecture during prometaphase. PICH knockdown results in a loss of Plk1 from the chromosome arm and an increase in highly disorganized “wavy” chromosomes that exhibit an “open” or “X-shaped” configuration, consistent with a loss of chromosome arm cohesion. Such chromosome disorganization occurs with essentially no change in the localization of condensin or cohesin on chromosomes. Interestingly, the chromosome disorganization could be prevented by treatment with a topoisomerase II inhibitor ICRF-193, suggesting that the PICH–Plk1 complex normally maintains chromosome architecture in a manner that involves topoisomerase II activity. PICH knockdown does not affect initial chromosome compaction at prophase but causes anaphase DNA bridge formation and failed abscission. Our studies suggest that the PICH–Plk1 complex plays a critical role in maintaining prometaphase chromosome architecture.