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

Nature volume 493, pages 437–440 (2013)Cite this article

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Abstract

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 complex1,2 indicated how TFIIB functions in DNA recruitment, but they lacked nucleic acids and half of the B-reader, leaving other TFIIB functions3,4 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 protein–nucleic 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 factors5, and the bacterial initiation factor sigma has TFIIB-like topology1,2 and contains the loop region 3.2 that resembles the B-reader loop in location, charge and function6,7,8. 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.

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Figure 1: Structure of the Pol II–TFIIB complex at 3.4 Å resolution.
Figure 2: TFIIB changes Pol II domains and stimulates catalysis.
Figure 3: Structure of initially transcribing Pol II–TFIIB complex (ITC) elucidates RNA initiation and separation, and TFIIB release.

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Protein Data Bank

Data deposits

Coordinates and structure factors of the Pol II–TFIIB complex and the ITC with 6-nucleotide RNA have been deposited in the Protein Data Bank under accession numbers 4BBR and 4BBS, respectively. Reprints and permissions information is available at www.nature.com/reprints.

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Acknowledgements

We thank C. Bäjen, A. Cheung, S. Etzold, F. Hög, D. Kostrewa, N. Pirkl and other members of the Cramer laboratory. Part of this work was performed at the Swiss Light Source (SLS) at the Paul Scherrer Institut, Villigen, Switzerland. S.S. was supported by a postdoctoral fellowship from the Alexander-von-Humboldt Foundation. P.C. was supported by the Deutsche Forschungsgemeinschaft, SFB646, TR5, GraKo1721, SFB960, CIPSM, NIM, an Advanced Grant of the European Research Council, the LMUinnovativ project Bioimaging Network, the Jung-Stiftung, and the Vallee Foundation.

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  1. Gene Center and Department of Biochemistry, Center for Integrated Protein Science CIPSM, Ludwig-Maximilians-Universität München, Feodor-Lynen-Str. 25, 81377 Munich, Germany,

    Sarah Sainsbury, Jürgen Niesser & Patrick Cramer

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  1. Sarah Sainsbury
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Contributions

S.S. carried out experiments except for functional analysis presented in Supplementary Fig. 5, which was carried out by J.N. P.C. initiated and supervised the project. S.S. and P.C. prepared the manuscript.

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Correspondence to Patrick Cramer.

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The authors declare no competing financial interests.

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This file contains Supplementary Figures 1-5, Supplementary Table 1 and additional references. (PDF 5748 kb)

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Sainsbury, S., Niesser, J. & Cramer, P. Structure and function of the initially transcribing RNA polymerase II–TFIIB complex. Nature 493, 437–440 (2013). https://doi.org/10.1038/nature11715

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