Institute of Genetics, Rheinische Friedrich-Wilhelms-Universität Bonn, Karlrobert-Kreiten-Str. 13, 53115 Bonn, Germany.
Cells. 2022 Apr 1;11(7):1189. doi: 10.3390/cells11071189.
Centrosomes represent main microtubule organizing centers (MTOCs) in animal cells. Their duplication in S-phase enables the establishment of two MTOCs in M-phase that define the poles of the spindle and ensure equal distribution of chromosomes and centrosomes to the two daughter cells. While key functions of many centrosomal proteins have been addressed in RNAi experiments and chronic knockdown, knockout experiments with complete loss of function in all cells enable quantitative analysis of cellular phenotypes at all cell-cycle stages. Here, we show that the centriolar satellite proteins SSX2IP and WDR8 and the centriolar protein CEP135 form a complex before centrosome assembly in vertebrate oocytes and further functionally interact in somatic cells with established centrosomes. We present stable knockouts of , , and in human cells. While loss of SSX2IP and WDR8 are compensated for, knockout cells display compromised PCM recruitment, reduced MTOC function, and premature centrosome splitting with imbalanced PCMs. Defective knockout centrosomes, however, manage to establish balanced spindle poles, allowing unperturbed mitosis and regular cell proliferation. Our data show essential functions of CEP135 in interphase MTOCs and demonstrate that loss of individual functions of SSX2IP, WDR8, and CEP135 are fully compensated for in mitosis.
中心体是动物细胞中主要的微管组织中心（MTOC）。它们在 S 期的复制使 M 期能够建立两个 MTOC，从而确定纺锤体的两极，并确保染色体和中心体均等分配到两个子细胞中。虽然许多中心体蛋白的关键功能已在 RNAi 实验和慢性敲低实验中得到解决，但在所有细胞中完全丧失功能的 knockout 实验可在所有细胞周期阶段对细胞表型进行定量分析。在这里，我们表明在脊椎动物卵母细胞中，中心粒卫星蛋白 SSX2IP 和 WDR8 以及中心粒蛋白 CEP135 在中心体组装之前形成复合物，并在具有已建立的中心体的体细胞中进一步发挥功能相互作用。我们在人类细胞中展示了 、 和 的稳定 knockout。虽然 SSX2IP 和 WDR8 的缺失得到了补偿，但 knockout 细胞显示出 PCM 募集受损、MTOC 功能降低以及中心体过早分裂导致 PCM 失衡。然而，功能缺陷的 knockout 中心体设法建立了平衡的纺锤体极，允许不受干扰的有丝分裂和正常的细胞增殖。我们的数据显示 CEP135 在间期中 MTOC 中的重要功能，并证明 SSX2IP、WDR8 和 CEP135 的单个功能缺失在有丝分裂中得到了充分补偿。
