Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, China.
Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, China.
J Biol Chem. 2011 Aug 19;286(33):29218-29226. doi: 10.1074/jbc.M111.252130. Epub 2011 Jun 15.
Spt6 is a highly conserved transcription elongation factor and histone chaperone. It binds directly to the RNA polymerase II C-terminal domain (RNAPII CTD) through its C-terminal region that recognizes RNAPII CTD phosphorylation. In this study, we determined the solution structure of the C-terminal region of Saccharomyces cerevisiae Spt6, and we discovered that Spt6 has two SH2 domains in tandem. Structural and phylogenetic analysis revealed that the second SH2 domain was evolutionarily distant from canonical SH2 domains and represented a novel SH2 subfamily with a novel binding site for phosphoserine. In addition, NMR chemical shift perturbation experiments demonstrated that the tandem SH2 domains recognized Tyr(1), Ser(2), Ser(5), and Ser(7) phosphorylation of RNAPII CTD with millimolar binding affinities. The structural basis for the binding of the tandem SH2 domains to different forms of phosphorylated RNAPII CTD and its physiological relevance are discussed. Our results also suggest that Spt6 may use the tandem SH2 domain module to sense the phosphorylation level of RNAPII CTD.
Spt6 是一种高度保守的转录延伸因子和组蛋白伴侣。它通过其识别 RNA 聚合酶 II C 端结构域（RNAPII CTD）磷酸化的 C 端区域直接与 RNA 聚合酶 II CTD 结合。在这项研究中，我们确定了酿酒酵母 Spt6 C 端区域的溶液结构，发现 Spt6 具有串联的两个 SH2 结构域。结构和系统发育分析表明，第二个 SH2 结构域与典型的 SH2 结构域在进化上相距甚远，代表了一个新的 SH2 亚家族，具有与磷酸丝氨酸结合的新结合位点。此外，NMR 化学位移扰动实验表明，串联 SH2 结构域以毫摩尔亲和力识别 RNA 聚合酶 II CTD 的 Tyr(1)、Ser(2)、Ser(5)和 Ser(7)磷酸化。讨论了串联 SH2 结构域与不同形式磷酸化 RNA 聚合酶 II CTD 结合的结构基础及其生理相关性。我们的结果还表明，Spt6 可能使用串联 SH2 结构域模块来感知 RNA 聚合酶 II CTD 的磷酸化水平。
