Quantitative Phosphoproteomics Reveals Widespread Full Phosphorylation Site Occupancy During Mitosis
Quantitative Phosphoproteomics Reveals Widespread Full Phosphorylation Site Occupancy During Mitosis
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DOI：
10.1126/scisignal.2000475
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发表时间：
2010-01-12
影响因子：
7.3
通讯作者：
Mann, Matthias
中科院分区：
文献类型：
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作者：
Olsen, Jesper V.;Vermeulen, Michiel;Mann, Matthias
真核细胞通过一系列复杂的进化上保守的事件进行复制，这些事件在细胞分裂周期的确定阶段受到严格调控。通过该循环的进展涉及大量专用蛋白质复合物和信号传导途径，并且该过程的失调与肿瘤发生有关。我们应用基于高分辨率质谱的蛋白质组学在全球范围内研究了人类细胞周期的蛋白质组和磷酸化蛋白质组，并量化了6027种蛋白质和20，443个独特的磷酸化位点及其动力学。共调节蛋白和磷酸化位点根据其细胞周期动力学进行分组，并与公开可用的信使RNA微阵列数据进行比较。大多数检测到的磷酸化位点和超过20%的所有定量蛋白质显示出实质性的调节，主要是在有丝分裂细胞。激酶基序分析显示，在S期的DNA损伤反应网络，这是由ATM或ATR或DNA依赖性蛋白激酶的磷酸化介导的全球激活。我们确定了超过5000个位点的位点特异性化学计量，发现大多数上调的细胞周期蛋白依赖性激酶1（CDK1）或CDK2磷酸化的位点在有丝分裂细胞中几乎完全磷酸化。特别是，核蛋白和参与调节代谢过程的蛋白质在有丝分裂中具有高磷酸化位点占有率。这表明，这些蛋白质可能被有丝分裂细胞中的磷酸化失活。
Eukaryotic cells replicate by a complex series of evolutionarily conserved events that are tightly regulated at defined stages of the cell division cycle. Progression through this cycle involves a large number of dedicated protein complexes and signaling pathways, and deregulation of this process is implicated in tumorigenesis. We applied high-resolution mass spectrometry-based proteomics to investigate the proteome and phosphoproteome of the human cell cycle on a global scale and quantified 6027 proteins and 20,443 unique phosphorylation sites and their dynamics. Co-regulated proteins and phosphorylation sites were grouped according to their cell cycle kinetics and compared to publicly available messenger RNA microarray data. Most detected phosphorylation sites and more than 20% of all quantified proteins showed substantial regulation, mainly in mitotic cells. Kinase-motif analysis revealed global activation during S phase of the DNA damage response network, which was mediated by phosphorylation by ATM or ATR or DNA-dependent protein kinases. We determined site-specific stoichiometry of more than 5000 sites and found that most of the up-regulated sites phosphorylated by cyclin-dependent kinase 1 (CDK1) or CDK2 were almost fully phosphorylated in mitotic cells. In particular, nuclear proteins and proteins involved in regulating metabolic processes have high phosphorylation site occupancy in mitosis. This suggests that these proteins may be inactivated by phosphorylation in mitotic cells.