A CDC20 allelic series reveals stage-specific control of metaphase-to-anaphase transitions during mitosis and meiosis in rice.

The metaphase-to-anaphase transition is a critically controlled checkpoint essential for precise chromosome segregation. In yeast and animals, cell division cycle 20 (CDC20) serves as an essential coactivator of the anaphase-promoting complex/cyclosome (APC/C), triggering the degradation of key substrates to initiate anaphase. Plants, however, often possess multiple CDC20 homologs, whose functional specificity remains poorly characterized. Employing CRISPR/Cas9 technology, we generated a series of cdc20 triple mutants in rice (Oryza sativa) and performed a systematic cytological analysis to investigate the functions of all three CDC20 genes. Complete loss of CDC20s' function resulted in gametophytic mitotic defects, ultimately causing bilateral gametophyte lethality. We obtained hypomorphic alleles specifically displaying disrupted meiosis without mitotic defects. The cdc20-triple-1 and cdc20-triple-2 mutants were arrested at metaphase I with normal chromosome alignment, but failed to remove sister chromatid cohesin 3 (SCC3) from chromosome arms. In contrast, the cdc20-triple-3 and cdc20-triple-4 alleles progressed through meiosis I but arrested at metaphase II, exhibiting aberrant spindles and chromosome missegregation. We further demonstrated that CDC20s regulated shugoshin 1 (SGO1) distributive pattern: SGO1 loaded normally in cdc20-triple-1 but persisted at centromeres and failed to detach, even when sister chromatids separated aberrantly. However, cdc20-triple-3 mutants showed normal SGO1 unloading, suggesting that timely SGO1 removal from centromeres depends on a CDC20-mediated metaphase I-to-anaphase I transition. Our findings reveal that rice CDC20 homologs play a critical role in controlling the metaphase-to-anaphase transition throughout both mitosis and meiosis, thereby ensuring tight regulation and enhancing genomic stability during sexual reproduction.