The RPB2 flap loop of human RNA polymerase II is dispensable for transcription initiation and elongation.
The RPB2 flap loop of human RNA polymerase II is dispensable for transcription initiation and elongation.
复制标题
人 RNA 聚合酶 II 的 RPB2 瓣环对于转录起始和延伸是可有可无的。
DOI：
10.1128/mcb.05318-11
复制
发表时间：
2011
影响因子：
5.3
通讯作者：
Landick,Robert
中科院分区：
文献类型：
--
作者：
Palangat,Murali;Grass,JeffreyA;Langelier,Marie-France;Coulombe,Benoit;Landick,Robert
多亚基RNA聚合酶（RNAP）的瓣状结构域，也称为壁，形成RNA出口通道的一侧。在细菌RNAP中，皮瓣的移动的部分被称为皮瓣尖端，并与起始和延伸因子进行必要的接触。共晶体结构表明真核RNAPII的直系同源部分（称为瓣环）与转录因子IIB（TFIIB）接触，但瓣环的功能尚未评估。我们构建并测试了人RNAPII（亚基RPB 2 Δ873-884）中瓣环的缺失，其去除了瓣环与TFIIB的相互作用界面。在人胚肾细胞系（HEK 293）中表达的具有瓣环缺失的RNAPII的分布的全基因组分析揭示瓣环对整体转录起始、RNAPII在基因内的占据或启动子逃逸和生产性延伸的效率没有影响。TFIIS刺激的转录物切割，或负延伸因子（NELF）和5，6-二氯-1-β-d-呋喃核糖基苯并咪唑（DRB）敏感性诱导因子（DSIF）复合物抑制转录物延伸。TFIIF抑制了对转录延长和暂停的适度影响。虽然类似于细菌RNAP的瓣尖，但RNAPII瓣环并不是同等重要的。
The flap domain of multisubunit RNA polymerases (RNAPs), also called the wall, forms one side of the RNA exit channel. In bacterial RNAP, the mobile part of the flap is called the flap tip and makes essential contacts with initiation and elongation factors. Cocrystal structures suggest that the orthologous part of eukaryotic RNAPII, called the flap loop, contacts transcription factor IIB (TFIIB), but the function of the flap loop has not been assessed. We constructed and tested a deletion of the flap loop in human RNAPII (subunit RPB2 Δ873-884) that removes the flap loop interaction interface with TFIIB. Genome-wide analysis of the distribution of the RNAPII with the flap loop deletion expressed in a human embryonic kidney cell line (HEK 293) revealed no effect of the flap loop on global transcription initiation, RNAPII occupancy within genes, or the efficiency of promoter escape and productive elongation.In vitro, the flap loop deletion had no effect on promoter binding, abortive initiation or promoter escape, TFIIS-stimulated transcript cleavage, or inhibition of transcript elongation by the complex of negative elongation factor (NELF) and 5,6-dichloro-1-β-d-ribofuranosylbenzimidazole (DRB) sensitivity-inducing factor (DSIF). A modest effect on transcript elongation and pausing was suppressed by TFIIF. Although similar to the flap tip of bacterial RNAP, the RNAPII flap loop is not equivalently essential.