Department Biochemistry and Molecular Genetics, RNA Bioscience Initiative, University of Colorado School of Medicine, Aurora, CO 80045, USA.
Department Biochemistry and Molecular Genetics, RNA Bioscience Initiative, University of Colorado School of Medicine, Aurora, CO 80045, USA.
Mol Cell. 2019 Jan 3;73(1):107-118.e4. doi: 10.1016/j.molcel.2018.10.031. Epub 2018 Nov 29.
In addition to phosphodiester bond formation, RNA polymerase II has an RNA endonuclease activity, stimulated by TFIIS, which rescues complexes that have arrested and backtracked. How TFIIS affects transcription under normal conditions is poorly understood. We identified backtracking sites in human cells using a dominant-negative TFIIS (TFIIS) that inhibits RNA cleavage and stabilizes backtracked complexes. Backtracking is most frequent within 2 kb of start sites, consistent with slow elongation early in transcription, and in 3' flanking regions where termination is enhanced by TFIIS, suggesting that backtracked pol II is a favorable substrate for termination. Rescue from backtracking by RNA cleavage also promotes escape from 5' pause sites, prevents premature termination of long transcripts, and enhances activation of stress-inducible genes. TFIIS slowed elongation rates genome-wide by half, suggesting that rescue of backtracked pol II by TFIIS is a major stimulus of elongation under normal conditions.
除了磷酸二酯键形成，RNA 聚合酶 II 还具有 RNA 内切酶活性，受 TFIIS 刺激，可挽救停滞和回溯的复合物。TFIIS 在正常条件下如何影响转录尚不清楚。我们使用一种显性负 TFIIS（TFIIS）来鉴定人细胞中的回溯位点，该蛋白可抑制 RNA 切割并稳定回溯复合物。回溯在启动子附近 2kb 内最频繁，这与转录早期伸长缓慢一致，并且在终止受 TFIIS 增强的 3'侧翼区域，表明回溯的 pol II 是终止的有利底物。RNA 切割的回溯挽救也促进了从 5'暂停位点的逃逸，防止长转录物的过早终止，并增强应激诱导基因的激活。TFIIS 使全基因组的伸长速度减慢了一半，这表明 TFIIS 对回溯 pol II 的挽救是正常条件下伸长的主要刺激因素。
