State Key Laboratory of Protein and Plant Gene Research, Beijing 100871, China.
College of Life Sciences, Peking University, Beijing 100871, China.
Plant Physiol. 2022 Feb 4;188(2):1111-1128. doi: 10.1093/plphys/kiab543.
Chromosome segregation must be under strict regulation to maintain chromosome euploidy and stability. Cell Division Cycle 20 (CDC20) is an essential cell cycle regulator that promotes the metaphase-to-anaphase transition and functions in the spindle assembly checkpoint, a surveillance pathway that ensures the fidelity of chromosome segregation. Plant CDC20 genes are present in multiple copies, and whether CDC20s have the same functions in plants as in yeast and animals is unclear, given the potential for divergence or redundancy among the multiple copies. Here, we studied all three CDC20 genes in rice (Oryza sativa) and constructed two triple mutants by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9-mediated genome editing to explore their roles in development. Knocking out all three CDC20 genes led to total sterility but did not affect vegetative development. Loss of the three CDC20 proteins did not alter mitotic division but severely disrupted meiosis as a result of asynchronous and unequal chromosome segregation, chromosome lagging, and premature separation of chromatids. Immunofluorescence of tubulin revealed malformed meiotic spindles in microsporocytes of the triple mutants. Furthermore, cytokinesis of meiosis I was absent or abnormal, and cytokinesis II was completely prevented in all mutant microsporocytes; thus, no tetrads or pollen formed in either cdc20 triple mutant. Finally, the subcellular structures and functions of the tapetum were disturbed by the lack of CDC20 proteins. These findings demonstrate that the three rice CDC20s play redundant roles but are indispensable for faithful meiotic chromosome segregation and cytokinesis, which are required for the production of fertile microspores.
染色体分离必须受到严格的调控，以维持染色体的整倍性和稳定性。细胞周期蛋白 20（CDC20）是一种必不可少的细胞周期调节剂，它促进了从中期到后期的转变，并在纺锤体组装检查点（spindle assembly checkpoint）中发挥作用，该检查点是一种确保染色体分离保真度的监测途径。植物 CDC20 基因存在多个副本，鉴于多个副本之间可能存在分歧或冗余，CDC20 在植物中的功能是否与酵母和动物中的相同尚不清楚。在这里，我们研究了水稻（Oryza sativa）中的三个 CDC20 基因，并通过成簇规律间隔短回文重复（CRISPR）/CRISPR 相关蛋白 9 介导的基因组编辑构建了两个三突变体，以探索它们在发育中的作用。敲除所有三个 CDC20 基因导致完全不育，但不影响营养生长发育。三个 CDC20 蛋白的缺失并没有改变有丝分裂分裂，但由于染色体分离的异步和不均匀、染色体滞后和染色单体的过早分离，严重破坏了减数分裂。微孢子细胞中微管的免疫荧光显示三突变体中的减数分裂纺锤体畸形。此外，减数分裂 I 的胞质分裂缺失或异常，所有突变体微孢子细胞中的胞质分裂 II 完全被阻止；因此，在 cdc20 三突变体中均未形成四分体或花粉。最后，缺乏 CDC20 蛋白扰乱了绒毡层的亚细胞结构和功能。这些发现表明，三个水稻 CDC20 基因发挥冗余作用，但对于减数分裂中染色体的正确分离和胞质分裂是必不可少的，这是产生有活力的小孢子所必需的。
