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TFIID Enables RNA Polymerase II Promoter-Proximal Pausing.
Molecular Cell ( IF 14.5 ) Pub Date : 2020-03-30 , DOI: 10.1016/j.molcel.2020.03.008 Charli B Fant 1 , Cecilia B Levandowski 1 , Kapil Gupta 2 , Zachary L Maas 1 , John Moir 1 , Jonathan D Rubin 1 , Andrew Sawyer 3 , Meagan N Esbin 1 , Jenna K Rimel 1 , Olivia Luyties 1 , Michael T Marr 3 , Imre Berger 2 , Robin D Dowell 4 , Dylan J Taatjes 1
Molecular Cell ( IF 14.5 ) Pub Date : 2020-03-30 , DOI: 10.1016/j.molcel.2020.03.008 Charli B Fant 1 , Cecilia B Levandowski 1 , Kapil Gupta 2 , Zachary L Maas 1 , John Moir 1 , Jonathan D Rubin 1 , Andrew Sawyer 3 , Meagan N Esbin 1 , Jenna K Rimel 1 , Olivia Luyties 1 , Michael T Marr 3 , Imre Berger 2 , Robin D Dowell 4 , Dylan J Taatjes 1
Affiliation
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RNA polymerase II (RNAPII) transcription is governed by the pre-initiation complex (PIC), which contains TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, RNAPII, and Mediator. After initiation, RNAPII enzymes pause after transcribing less than 100 bases; precisely how RNAPII pausing is enforced and regulated remains unclear. To address specific mechanistic questions, we reconstituted human RNAPII promoter-proximal pausing in vitro, entirely with purified factors (no extracts). As expected, NELF and DSIF increased pausing, and P-TEFb promoted pause release. Unexpectedly, the PIC alone was sufficient to reconstitute pausing, suggesting RNAPII pausing is an inherent PIC function. In agreement, pausing was lost upon replacement of the TFIID complex with TATA-binding protein (TBP), and PRO-seq experiments revealed widespread disruption of RNAPII pausing upon acute depletion (t = 60 min) of TFIID subunits in human or Drosophila cells. These results establish a TFIID requirement for RNAPII pausing and suggest pause regulatory factors may function directly or indirectly through TFIID.
中文翻译：
TFIID启用RNA聚合酶II启动子近端暂停。
RNA聚合酶II（RNAPII）的转录受预起始复合物（PIC）的控制，该复合物包含TFIIA，TFIIB，TFIID，TFIIE，TFIIF，TFIIH，RNAPII和介体。起始后，RNAPII酶在转录少于100个碱基后会暂停；究竟如何实施和调控RNAPII暂停尚不清楚。为了解决特定的机械问题，我们在体外完全重建了纯化因子（无提取物）重建了人类RNAPII启动子近端暂停。不出所料，NELF和DSIF增加了暂停，而P-TEFb促进了暂停释放。出乎意料的是，仅PIC足以重构暂停，这表明RNAPII暂停是PIC固有的功能。一致的是，用TATA结合蛋白（TBP）代替TFIID复合物后，暂停就消失了，和PRO-seq实验表明，人或果蝇细胞中TFIID亚基的急性耗竭（t = 60分钟）后，RNAPII暂停被广泛破坏。这些结果确立了TFIID对RNAPII暂停的要求，并表明暂停调节因子可能通过TFIID直接或间接起作用。
更新日期：2020-03-30
中文翻译：
TFIID启用RNA聚合酶II启动子近端暂停。
RNA聚合酶II（RNAPII）的转录受预起始复合物（PIC）的控制，该复合物包含TFIIA，TFIIB，TFIID，TFIIE，TFIIF，TFIIH，RNAPII和介体。起始后，RNAPII酶在转录少于100个碱基后会暂停；究竟如何实施和调控RNAPII暂停尚不清楚。为了解决特定的机械问题，我们在体外完全重建了纯化因子（无提取物）重建了人类RNAPII启动子近端暂停。不出所料，NELF和DSIF增加了暂停，而P-TEFb促进了暂停释放。出乎意料的是，仅PIC足以重构暂停，这表明RNAPII暂停是PIC固有的功能。一致的是，用TATA结合蛋白（TBP）代替TFIID复合物后，暂停就消失了，和PRO-seq实验表明，人或果蝇细胞中TFIID亚基的急性耗竭（t = 60分钟）后，RNAPII暂停被广泛破坏。这些结果确立了TFIID对RNAPII暂停的要求，并表明暂停调节因子可能通过TFIID直接或间接起作用。
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