Architecture of the RNA polymerase II preinitiation complex and mechanism of ATP-dependent promoter opening

Grünberg, Sebastian; Warfield, Linda; Hahn, Steven

doi:10.1038/nsmb.2334

Cited by 125 publications

(169 citation statements)

References 60 publications

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“…This result implies that the tight interaction network between RNAP, TBP, TFB, and DNA overrides the effect of the stalk and controls the clamp conformation. TFE in archaea and TFIIE in the eukaryotic pol II system stabilizes the PIC and stimulates transcription by promoting DNA melting (13,15,16,18,39,41,42). TFE is composed of winged helix (WH) and zinc ribbon domains that are anchored to the RNAP clamp and stalk domains: these binding sites are ideally suited to manipulate clamp movements.…”

Section: Resultsmentioning

confidence: 99%

“…The RNAP clamp provides binding sites for the transcription initiation factor TFE (homologous to TFIIEα) and the elongation factor Spt4/5, both of which bind competitively to the clamp coiled coil motif (15)(16)(17) of archaeal-eukaryotic RNAPs. TFE facilitates open complex formation by interacting with the RNAP clamp and stalk and the nontemplate strand (NTS) (15,18).…”

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confidence: 99%

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TFE and Spt4/5 open and close the RNA polymerase clamp during the transcription cycle

Schulz

Gietl

Smollett

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Transcription is an intrinsically dynamic process and requires the coordinated interplay of RNA polymerases (RNAPs) with nucleic acids and transcription factors. Classical structural biology techniques have revealed detailed snapshots of a subset of conformational states of the RNAP as they exist in crystals. A detailed view of the conformational space sampled by the RNAP and the molecular mechanisms of the basal transcription factors E (TFE) and Spt4/5 through conformational constraints has remained elusive. We monitored the conformational changes of the flexible clamp of the RNAP by combining a fluorescently labeled recombinant 12-subunit RNAP system with single-molecule FRET measurements. We measured and compared the distances across the DNA binding channel of the archaeal RNAP. Our results show that the transition of the closed to the open initiation complex, which occurs concomitant with DNA melting, is coordinated with an opening of the RNAP clamp that is stimulated by TFE. We show that the clamp in elongation complexes is modulated by the nontemplate strand and by the processivity factor Spt4/5, both of which stimulate transcription processivity. Taken together, our results reveal an intricate network of interactions within transcription complexes between RNAP, transcription factors, and nucleic acids that allosterically modulate the RNAP during the transcription cycle.

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

TFE and Spt4/5 open and close the RNA polymerase clamp during the transcription cycle

Schulz

Gietl

Smollett

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Our results lead to a unified, highly conserved architecture of the core transcription initiation complex. The location of the remaining general transcription factors TFIIE and TFIIH differs to some extent in three published studies [6][7][8] and may be analysed in the future.…”

Section: Discussionmentioning

confidence: 99%

“…Architectural models of the yeast Pol II PIC were obtained by site-specific protein cleavage mapping [4][5][6] . The architecture of the human PIC was obtained by electron microscopy (EM) 7 , and generally resembled that of the yeast PIC.…”

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confidence: 99%

Conserved architecture of the core RNA polymerase II initiation complex

et al. 2014

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During transcription initiation at promoters of protein-coding genes, RNA polymerase (Pol) II assembles with TBP, TFIIB and TFIIF into a conserved core initiation complex that recruits additional factors. The core complex stabilizes open DNA and initiates RNA synthesis, and it is conserved in the Pol I and Pol III transcription systems. Here, we derive the domain architecture of the yeast core pol II initiation complex during transcription initiation. The yeast complex resembles the human initiation complex and reveals that the TFIIF Tfg2 winged helix domain swings over promoter DNA. An 'arm' and a 'charged helix' in TFIIF function in transcription start site selection and initial RNA synthesis, respectively, and apparently extend into the active centre cleft. Our model provides the basis for further structure-function analysis of the entire transcription initiation complex.

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“…The previously mobile 10 tip of the protrusion forms a b-sheet and an ordered loop (a11-a12) that is required for initiation complex stability 11 (Fig. 1d).…”

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confidence: 99%

Structure and function of the initially transcribing RNA polymerase II–TFIIB complex

Sainsbury

Niesser

Cramer

2012

Nature

176

195

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The general transcription factor (TF) IIB is required for RNA polymerase (Pol) II initiation and extends with its B-reader element into the Pol II active centre cleft. Low-resolution structures of the Pol II-TFIIB complex 1,2 indicated how TFIIB functions in DNA recruitment, but they lacked nucleic acids and half of the B-reader, leaving other TFIIB functions 3,4 enigmatic. Here we report crystal structures of the Pol II-TFIIB complex from the yeast Saccharomyces cerevisiae at 3.4 Å resolution and of an initially transcribing complex that additionally contains the DNA template and a 6-nucleotide RNA product. The structures reveal the entire B-reader and proteinnucleic acid interactions, and together with functional data lead to a more complete understanding of transcription initiation. TFIIB partially closes the polymerase cleft to position DNA and assist in its opening. The B-reader does not reach the active site but binds the DNA template strand upstream to assist in the recognition of the initiator sequence and in positioning the transcription start site. TFIIB rearranges active-site residues, induces binding of the catalytic metal ion B, and stimulates initial RNA synthesis allosterically. TFIIB then prevents the emerging DNA-RNA hybrid duplex from tilting, which would impair RNA synthesis. When the RNA grows beyond 6 nucleotides, it is separated from DNA and is directed to its exit tunnel by the B-reader loop. Once the RNA grows to 12-13 nucleotides, it clashes with TFIIB, triggering TFIIB displacement and elongation complex formation. Similar mechanisms may underlie all cellular transcription because all eukaryotic and archaeal RNA polymerases use TFIIB-like factors 5 , and the bacterial initiation factor sigma has TFIIB-like topology 1,2 and contains the loop region 3.2 that resembles the B-reader loop in location, charge and function [6][7][8] . TFIIB and its counterparts may thus account for the two fundamental properties that distinguish RNA from DNA polymerases: primer-independent chain initiation and product separation from the template.Our previous X-ray analysis of the Pol II-TFIIB complex at 4.3 Å resolution provided a partial backbone model of TFIIB 1 . To obtain a complete and atomic structure, we co-crystallized Pol II with a TFIIB variant lacking the mobile amino-terminal tail and carboxy-terminal cyclin fold (Methods, Supplementary Table and Supplementary Fig. 1). The resulting Pol II-TFIIB structure at 3.4 Å resolution provides details of the interactions of the four TFIIB domains with Pol II: the B-ribbon with the dock, the B-core N-terminal cyclin fold with the wall, the B-reader helix with the RNA exit tunnel, and the B-linker helix with the coiled-coil of the clamp (Fig. 1).The structure reveals the entire course of the TFIIB polypeptide chain through the Pol II cleft, including the previously lacking 1 B-reader loop (residues 67-79) and a new 'B-reader strand' (residues 80-83). The B-reader loop does not reach the active site, but instead interacts with the Pol II rudder and fork loop ...

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Architecture of the RNA polymerase II preinitiation complex and mechanism of ATP-dependent promoter opening

Cited by 125 publications

References 60 publications

TFE and Spt4/5 open and close the RNA polymerase clamp during the transcription cycle

TFE and Spt4/5 open and close the RNA polymerase clamp during the transcription cycle

Conserved architecture of the core RNA polymerase II initiation complex

Structure and function of the initially transcribing RNA polymerase II–TFIIB complex

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