Abstract
Proper chromosome segregation during cell division is essential for genomic integrity and organismal development. This process is monitored by the spindle assembly checkpoint (SAC), which delays anaphase onset until all chromosomes are properly attached to the mitotic spindle. The kinetochore protein KNL1 plays a critical role in recruiting SAC proteins. Here, we reveal that Arabidopsis KNL1 regulates SAC silencing through the direct recruitment of type one protein phosphatase (TOPP) to kinetochores. We show that KNL1 interacts with all nine TOPPs via a conserved RVSF motif in its N terminus, and this interaction is required for the proper localization of TOPPs to kinetochores during mitosis. Disrupting KNL1-TOPP interaction leads to persistent SAC activation, resulting in a severe metaphase arrest and defects in plant growth and development. Our findings highlight the evolutionary conservation of KNL1 in coordinating kinetochore-localized phosphatase to ensure timely SAC silencing and faithful chromosome segregation in Arabidopsis.