Mapping RNAPII CTD Phosphorylation Reveals That the Identity and Modification of Seventh Heptad Residues Direct Tyr1 Phosphorylation
Mapping RNAPII CTD Phosphorylation Reveals That the Identity and Modification of Seventh Heptad Residues Direct Tyr1 Phosphorylation
复制标题
DOI：
10.1021/acschembio.9b00610
复制
发表时间：
2019-10-01
影响因子：
4
通讯作者：
Zhang, Yan
中科院分区：
文献类型：
--
作者：
Burkholder, Nathaniel T.;Sipe, Sarah N.;Zhang, Yan
真核 RNA 聚合酶 II 最大亚基的 C 端结构域 (CTD) 具有重复的七肽序列 Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7，负责招募转录调节因子。哺乳动物中的第七个七肽残基不太保守，并且容易发生各种翻译后修饰，但这种变异的后果尚不清楚。在这项研究中，我们使用紫外光解离质谱、动力学分析和结构分析来剖析第七个七肽位置上的不同残基或修饰如何改变 Tyr1 磷酸化。我们发现该位置带负电的残基促进相邻 Tyr1 位点的磷酸化，而带正电的残基则对其产生歧视。改变第七七肽残基上的电荷的修饰（例如精氨酸瓜氨酸化）否定了这种区别。这种特异性可以通过 ABL1 及其同源物活性位点附近的保守带正电荷的口袋来解释。我们的结果揭示了第七个七肽位置的变异或修饰的新机制，指导其他 CTD 位点的后续磷酸化，这可能有助于形成可能影响转录调控的各种修饰组合。
The C-terminal domain (CTD) of the largest subunit in eukaryotic RNA polymerase II has a repetitive heptad sequence of Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7 which is responsible for recruiting transcriptional regulatory factors. The seventh heptad residues in mammals are less conserved and subject to various post-translational modifications, but the consequences of such variations are not well understood. In this study, we use ultraviolet photodissociation mass spectrometry, kinetic assays, and structural analyses to dissect how different residues or modifications at the seventh heptad position alter Tyr1 phosphorylation. We found that negatively charged residues in this position promote phosphorylation of adjacent Tyr1 sites, whereas positively charged residues discriminate against it. Modifications that alter the charges on seventh heptad residues such as arginine citrullination negate such distinctions. Such specificity can be explained by conserved, positively charged pockets near the active sites of ABL1 and its homologues. Our results reveal a novel mechanism for variations or modifications in the seventh heptad position directing subsequent phosphorylation of other CTD sites, which can contribute to the formation of various modification combinations that likely impact transcriptional regulation.