Structure of transcribing mammalian RNA polymerase II
Structure of transcribing mammalian RNA polymerase II
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DOI：
10.1038/nature16482
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发表时间：
2016-01-28
期刊：
影响因子：
64.8
通讯作者：
Cramer, Patrick
中科院分区：
文献类型：
--
作者：
Bernecky, Carrie;Herzog, Franz;Cramer, Patrick
RNA 聚合酶 (Pol) II 在所有真核细胞中蛋白质编码基因的转录过程中产生信使 RNA。通过 X 射线晶体学，可以清楚地了解两种酵母 (1-3) 的 Pol II 结构。然而，哺乳动物 Pol II 的结构研究仍然仅限于对人类 Pol II 及其与各种蛋白质的复合物进行低分辨率电子显微镜分析 (4-10)。在这里，我们报道了哺乳动物 Pol II 的 3.4 埃分辨率冷冻电子显微镜结构，其形式为包含 DNA 模板和 RNA 转录物的转录复合物。我们使用牛 Pol II，除了七个氨基酸残基外，它与人类酶相同。获得的原子模型与酵母模型非常相似，但也揭示了未知的特征。核酸与聚合酶的结合涉及移动 Pol II 夹和活性中心区域的“诱导配合”。转录泡下游的 DNA 与 Pol II 亚基 RPB5 下颌结构域中的保守“TPSA 基序”接触，这种相互作用显然在转录起始过程中已经建立(7)。上游 DNA 从活性中心裂口发出，与下游 DNA 成约 105 度角。上游 DNA 的这个位置允许结合通用转录延伸因子 DSIF (SPT4-SPT5)，我们将其定位在 Pol II 钳结构域上的保守位置的活性中心裂口上。我们的结果定义了哺乳动物 Pol II 在其功能状态下的结构，表明之前对酵母 Pol II 的晶体学分析与理解所有真核生物中的基因转录相关，并为人类转录的机制分析提供了起点。
RNA polymerase (Pol) II produces messenger RNA during transcription of protein-coding genes in all eukaryotic cells. The Pol II structure is known at high resolution from X-ray crystallography for two yeast species(1-3). Structural studies of mammalian Pol II, however, remain limited to low-resolution electron microscopy analysis of human Pol II and its complexes with various proteins(4-10). Here we report the 3.4 angstrom resolution cryo-electron microscopy structure of mammalian Pol II in the form of a transcribing complex comprising DNA template and RNA transcript. We use bovine Pol II, which is identical to the human enzyme except for seven amino-acid residues. The obtained atomic model closely resembles its yeast counterpart, but also reveals unknown features. Binding of nucleic acids to the polymerase involves 'induced fit' of the mobile Pol II clamp and active centre region. DNA downstream of the transcription bubble contacts a conserved 'TPSA motif' in the jaw domain of the Pol II subunit RPB5, an interaction that is apparently already established during transcription initiation(7). Upstream DNA emanates from the active centre cleft at an angle of approximately 105 degrees with respect to downstream DNA. This position of upstream DNA allows for binding of the general transcription elongation factor DSIF (SPT4-SPT5) that we localize over the active centre cleft in a conserved position on the clamp domain of Pol II. Our results define the structure of mammalian Pol II in its functional state, indicate that previous crystallographic analysis of yeast Pol II is relevant for understanding gene transcription in all eukaryotes, and provide a starting point for a mechanistic analysis of human transcription.