Department of Molecular Cell Biology and Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Jangangu, Suwon, Republic of Korea.
Cell Cycle. 2010 May 15;9(10):2027-36. doi: 10.4161/cc.9.10.11753.
During mitosis, regulation of protein structures and functions by phosphorylation plays critical roles in orchestrating a series of complex events essential for the cell division process. Tumor-associated microtubule-associated protein (TMAP), also known as cytoskeleton-associated protein 2 (CKAP2), is a novel player in spindle assembly and chromosome segregation. We have previously reported that TMAP is phosphorylated at multiple residues specifically during mitosis. However, the mechanisms and functional importance of phosphorylation at most of the sites identified are currently unknown. Here, we report that TMAP is a novel substrate of the Aurora B kinase. Ser627 of TMAP was specifically phosphorylated by Aurora B both in vitro and in vivo. Ser627 and neighboring conserved residues were strictly required for efficient phosphorylation of TMAP by Aurora B, as even minor amino acid substitutions of the phosphorylation motif significantly diminished the efficiency of the substrate phosphorylation. Nearly all mutations at the phosphorylation motif had dramatic effects on the subcellular localization of TMAP. Instead of being localized to the chromosome region during late mitosis, the mutants remained associated with microtubules and centrosomes throughout mitosis. However, the changes in the subcellular localization of these mutants could not be completely explained by the phosphorylation status on Ser627. Our findings suggest that the motif surrounding Ser627 ((625) RRSRRL (630)) is a critical part of a functionally important sequence motif which not only governs the kinase-substrate recognition, but also regulates the subcellular localization of TMAP during mitosis.
在有丝分裂过程中，蛋白质结构和功能的磷酸化调控对于协调一系列复杂事件至关重要，这些事件对于细胞分裂过程是必不可少的。肿瘤相关微管相关蛋白（TMAP），也称为细胞骨架相关蛋白 2（CKAP2），是纺锤体组装和染色体分离的新成员。我们之前报道过 TMAP 在有丝分裂过程中多个残基被特异性磷酸化。然而，目前大多数鉴定出的磷酸化位点的磷酸化机制和功能重要性尚不清楚。在这里，我们报告 TMAP 是 Aurora B 激酶的一个新底物。TMAP 的 Ser627 在体外和体内均被 Aurora B 特异性磷酸化。Ser627 和相邻的保守残基对于 Aurora B 对 TMAP 的有效磷酸化是严格必需的，因为即使磷酸化模体中的微小氨基酸取代也会显著降低底物磷酸化的效率。磷酸化模体上的几乎所有突变都对 TMAP 的亚细胞定位产生了巨大影响。与晚期有丝分裂时定位于染色体区域不同，这些突变体在整个有丝分裂过程中仍然与微管和中心体结合。然而，这些突变体的亚细胞定位的变化并不能完全用 Ser627 的磷酸化状态来解释。我们的发现表明，Ser627 周围的模体 ((625) RRSRRL (630)) 是一个功能重要的序列模体的关键部分，它不仅控制激酶-底物识别，而且在有丝分裂过程中调节 TMAP 的亚细胞定位。
