Phosphosite Scanning reveals a complex phosphorylation code underlying CDK-dependent activation of Hcm1.
Phosphosite Scanning reveals a complex phosphorylation code underlying CDK-dependent activation of Hcm1.
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DOI：
10.1038/s41467-023-36035-9
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发表时间：
2023-01-19
影响因子：
16.6
通讯作者：
Benanti, Jennifer A.
中科院分区：
文献类型：
--
作者：
Conti, Michelle M.;Li, Rui;Narvaez Ramos, Michelle A.;Zhu, Lihua Julie;Fazzio, Thomas G.;Benanti, Jennifer A.
有序的细胞周期进程由细胞周期蛋白依赖性激酶（CDK）协调。CDK通常在聚集在无序区域内的多个位点磷酸化底物。然而，对于大多数底物，不知道哪些磷酸化位点在功能上是重要的。我们开发了一种高通量的方法，磷酸化位点扫描，测试多位点磷酸化结构域中每个磷酸化位点的重要性。我们表明，磷酸盐扫描识别多种组合的磷酸盐，可以调节蛋白质的功能，并揭示了特定的磷酸化所需的磷酸化在一个域内的其他网站。我们将这种方法应用于酵母转录因子Hcm 1，一种保守的有丝分裂基因的调节因子，对准确的染色体分离至关重要。磷酸化位点扫描揭示了一个复杂的CDK调节电路，介导体内关键激活位点的Cks1依赖性磷酸化。这些结果阐明了CDK激活Hcm 1的机制，并建立了磷酸化位点扫描作为解码多位点磷酸化结构域的有力工具。许多激酶在多个位点磷酸化蛋白质，然而在大多数情况下，不知道哪些位点在功能上是重要的。在这里，作者描述了一种高通量的方法来定量评估多位点磷酸化结构域中每个磷酸化位点的贡献。
Ordered cell cycle progression is coordinated by cyclin dependent kinases (CDKs). CDKs often phosphorylate substrates at multiple sites clustered within disordered regions. However, for most substrates, it is not known which phosphosites are functionally important. We developed a high-throughput approach, Phosphosite Scanning, that tests the importance of each phosphosite within a multisite phosphorylated domain. We show that Phosphosite Scanning identifies multiple combinations of phosphosites that can regulate protein function and reveals specific phosphorylations that are required for phosphorylation at additional sites within a domain. We applied this approach to the yeast transcription factor Hcm1, a conserved regulator of mitotic genes that is critical for accurate chromosome segregation. Phosphosite Scanning revealed a complex CDK-regulatory circuit that mediates Cks1-dependent phosphorylation of key activating sites in vivo. These results illuminate the mechanism of Hcm1 activation by CDK and establish Phosphosite Scanning as a powerful tool for decoding multisite phosphorylated domains. Many kinases phosphorylate proteins on multiple sites, however in most cases it is not known which sites are functionally important. Here, the authors describe a high-throughput approach to quantitatively evaluate the contribution of each phosphosite within a multisite phosphorylated domain.
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影响因子：
3.3
作者：
Arsenault HE;Roy J;Mapa CE;Cyert MS;Benanti JA
通讯作者：
Benanti JA
必需细胞周期基因的抑制会增加细胞适应性。
影响因子：
4.5
作者：
通讯作者：
--
影响因子：
48
作者：
Fowler, Douglas M.;Fields, Stanley
通讯作者：
Fields, Stanley
影响因子：
16.8
作者：
Koivomaegi, Mardo;Oerd, Mihkel;Iofik, Anna;Valk, Ervin;Venta, Rainis;Faustova, Ilona;Kivi, Rait;Balog, Eva Rose M.;Rubin, Seth M.;Loog, Mart
通讯作者：
Loog, Mart
影响因子：
5.3
作者：
HADWIGER, JA;WITTENBERG, C;REED, SI
通讯作者：
REED, SI