The Plk1-dependent Phosphoproteome of the Early Mitotic Spindle
The Plk1-dependent Phosphoproteome of the Early Mitotic Spindle
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DOI：
10.1074/mcp.m110.004457
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发表时间：
2011-01-01
影响因子：
7
通讯作者：
Nigg, Erich A.
中科院分区：
文献类型：
--
作者：
Santamaria, Anna;Wang, Bin;Nigg, Erich A.
Polo样激酶调节从酵母到人类的有丝分裂和减数分裂进程的许多方面。在有丝分裂早期，哺乳动物Polo样激酶1（Plk 1）控制中心体成熟、纺锤体组装和微管附着到动粒。然而，尽管Plk 1的基本和多样的功能，全系列的Plk 1基板仍有待探索。为了研究人类有丝分裂纺锤体的Plk 1依赖性磷酸化蛋白质组，我们结合了稳定同位素标记的氨基酸在细胞培养与Plk 1失活或耗尽，然后纺锤体分离和质谱。我们的研究确定了358个独特的Plk 1依赖的纺锤体蛋白磷酸化位点，包括新的底物，说明了Plk 1依赖的信号网络的复杂性。通过体外磷酸化肽阵列验证了超过100个位点，从而扩大了Plk 1共有基序。总的来说，我们的数据提供了丰富的信息来源Plk 1依赖的磷酸化，Plk 1对接基板，磷酸化对蛋白定位的影响，以及Plk 1和极光A之间的功能相互作用的早期有丝分裂纺锤体。Molecular & Cellular Proteomics 10：10.1074/mcp. M110.004457，1-18，2011.
Polo-like kinases regulate many aspects of mitotic and meiotic progression from yeast to man. In early mitosis, mammalian Polo-like kinase 1 (Plk1) controls centrosome maturation, spindle assembly, and microtubule attachment to kinetochores. However, despite the essential and diverse functions of Plk1, the full range of Plk1 substrates remains to be explored. To investigate the Plk1-dependent phosphoproteome of the human mitotic spindle, we combined stable isotope labeling by amino acids in cell culture with Plk1 inactivation or depletion followed by spindle isolation and mass spectrometry. Our study identified 358 unique Plk1-dependent phosphorylation sites on spindle proteins, including novel substrates, illustrating the complexity of the Plk1-dependent signaling network. Over 100 sites were validated by in vitro phosphorylation of peptide arrays, resulting in a broadening of the Plk1 consensus motif. Collectively, our data provide a rich source of information on Plk1-dependent phosphorylation, Plk1 docking to substrates, the influence of phosphorylation on protein localization, and the functional interaction between Plk1 and Aurora A on the early mitotic spindle. Molecular & Cellular Proteomics 10: 10.1074/mcp.M110.004457, 1-18, 2011.