State Key Laboratory of Molecular Oncology, Cancer Institute, Chinese Academy of Medical Sciences and Peking Union Medical College, China.
Cancer Biol Ther. 2010 Nov 1;10(9):945-52. doi: 10.4161/cbt.10.9.13368.
The formation of proper mitotic spindles is required for appropriate chromosome segregation during cell division. Aberrant spindle formation often causes aneuploidy and results in tumorigenesis. However, the underlying mechanism of regulating spindle formation and chromosome separation remains to be further defined. Centrosomal Nlp (ninein-like protein) is a recently characterized BRCA1-regulated centrosomal protein and plays an important role in centrosome maturation and spindle formation. In this study, we show that Nlp can be phosphorylated by cell cycle protein kinase Cdc2/cyclin B1. The phosphorylation sites of Nlp are mapped at Ser185 and Ser589. Interestingly, the Cdc2/cyclin B1 phosphorylation site Ser185 of Nlp is required for its recognition by PLK1, which enable Nlp depart from centrosomes to allow the establishment of a mitotic scaffold at the onset of mitosis . PLK1 fails to dissociate the Nlp mutant lacking Ser185 from centrosome, suggesting that Cdc2/cyclin B1 might serve as a primary kinase of PLK1 in regulating Nlp subcellular localization. However, the phosphorylation at the site Ser589 by Cdc2/cyclin B1 plays an important role in Nlp protein stability probably due to its effect on protein degradation. Furthermore, we show that deregulated expression or subcellular localization of Nlp lead to multinuclei in cells, indicating that scheduled levels of Nlp and proper subcellular localization of Nlp are critical for successful completion of normal cell mitosis, These findings demonstrate that Cdc2/cyclin B1 is a key regulator in maintaining appropriate degradation and subcellular localization of Nlp, providing novel insights into understanding on the role of Cdc2/cyclin B1 in mitotic progression.
有丝分裂纺锤体的形成是细胞分裂过程中染色体正确分离所必需的。纺锤体的异常形成常常导致非整倍体，并导致肿瘤发生。然而，调节纺锤体形成和染色体分离的潜在机制仍有待进一步确定。中心体 Nlp（类九肽蛋白）是一种最近被描述的 BRCA1 调节的中心体蛋白，在中心体成熟和纺锤体形成中发挥重要作用。在这项研究中，我们表明细胞周期蛋白激酶 Cdc2/周期蛋白 B1 可以使 Nlp 磷酸化。Nlp 的磷酸化位点定位于 Ser185 和 Ser589。有趣的是，Nlp 的 Cdc2/周期蛋白 B1 磷酸化位点 Ser185 对于其被 PLK1 识别是必需的，这使得 Nlp 离开中心体，从而在有丝分裂开始时在中心体建立有丝分裂支架。PLK1 未能使缺乏 Ser185 的 Nlp 突变体从中心体解离，这表明 Cdc2/周期蛋白 B1 可能作为调节 Nlp 亚细胞定位的 PLK1 的主要激酶。然而，Cdc2/周期蛋白 B1 在 Ser589 位点的磷酸化对 Nlp 蛋白稳定性起着重要作用，可能是因为它对蛋白降解的影响。此外，我们表明 Nlp 的失调表达或亚细胞定位导致细胞多核化，表明 Nlp 的预定水平和适当的亚细胞定位对于正常细胞有丝分裂的成功完成至关重要。这些发现表明 Cdc2/周期蛋白 B1 是维持 Nlp 适当降解和亚细胞定位的关键调节剂，为理解 Cdc2/周期蛋白 B1 在有丝分裂进程中的作用提供了新的见解。
