Olsen Jesper V, Blagoev Blagoy, Gnad Florian, Macek Boris, Kumar Chanchal, Mortensen Peter, Mann Matthias
Center for Experimental BioInformatics, Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense, Denmark.
Cell. 2006 Nov 3;127(3):635-48. doi: 10.1016/j.cell.2006.09.026.
Cell signaling mechanisms often transmit information via posttranslational protein modifications, most importantly reversible protein phosphorylation. Here we develop and apply a general mass spectrometric technology for identification and quantitation of phosphorylation sites as a function of stimulus, time, and subcellular location. We have detected 6,600 phosphorylation sites on 2,244 proteins and have determined their temporal dynamics after stimulating HeLa cells with epidermal growth factor (EGF) and recorded them in the Phosida database. Fourteen percent of phosphorylation sites are modulated at least 2-fold by EGF, and these were classified by their temporal profiles. Surprisingly, a majority of proteins contain multiple phosphorylation sites showing different kinetics, suggesting that they serve as platforms for integrating signals. In addition to protein kinase cascades, the targets of reversible phosphorylation include ubiquitin ligases, guanine nucleotide exchange factors, and at least 46 different transcriptional regulators. The dynamic phosphoproteome provides a missing link in a global, integrative view of cellular regulation.
细胞信号传导机制通常通过翻译后蛋白质修饰来传递信息，其中最重要的是可逆性蛋白质磷酸化。在此，我们开发并应用了一种通用的质谱技术，用于鉴定和定量磷酸化位点，并将其作为刺激、时间和亚细胞定位的函数。我们在2244种蛋白质上检测到6600个磷酸化位点，并在用人表皮生长因子（EGF）刺激HeLa细胞后确定了它们的时间动态，并将其记录在Phosida数据库中。14%的磷酸化位点受到EGF至少2倍的调节，并根据它们的时间谱进行分类。令人惊讶的是，大多数蛋白质含有多个显示不同动力学的磷酸化位点，这表明它们作为整合信号的平台。除了蛋白激酶级联反应外，可逆磷酸化的靶点还包括泛素连接酶、鸟嘌呤核苷酸交换因子以及至少46种不同的转录调节因子。动态磷酸化蛋白质组在细胞调节的全局、综合视图中提供了缺失的环节。
