Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Structure of transcribing mammalian RNA polymerase II

Nature volume 529, pages 551–554 (2016)Cite this article

Subjects

Abstract

RNA polymerase (Pol) II produces messenger RNA during transcription of protein-coding genes in all eukaryotic cells. The Pol II structure is known at high resolution from X-ray crystallography for two yeast species1,2,3. Structural studies of mammalian Pol II, however, remain limited to low-resolution electron microscopy analysis of human Pol II and its complexes with various proteins4,5,6,7,8,9,10. Here we report the 3.4 Å resolution cryo-electron microscopy structure of mammalian Pol II in the form of a transcribing complex comprising DNA template and RNA transcript. We use bovine Pol II, which is identical to the human enzyme except for seven amino-acid residues. The obtained atomic model closely resembles its yeast counterpart, but also reveals unknown features. Binding of nucleic acids to the polymerase involves ‘induced fit’ of the mobile Pol II clamp and active centre region. DNA downstream of the transcription bubble contacts a conserved ‘TPSA motif’ in the jaw domain of the Pol II subunit RPB5, an interaction that is apparently already established during transcription initiation7. Upstream DNA emanates from the active centre cleft at an angle of approximately 105° with respect to downstream DNA. This position of upstream DNA allows for binding of the general transcription elongation factor DSIF (SPT4–SPT5) that we localize over the active centre cleft in a conserved position on the clamp domain of Pol II. Our results define the structure of mammalian Pol II in its functional state, indicate that previous crystallographic analysis of yeast Pol II is relevant for understanding gene transcription in all eukaryotes, and provide a starting point for a mechanistic analysis of human transcription.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Cryo-EM structure of mammalian Pol II EC at 3.4 Å resolution.
Figure 2: Selected mammalian-specific features that differ from yeast Pol II (PDB accession number 1Y1W).
Figure 3: Pol II–nucleic-acid interactions.
Figure 4: Location of DSIF on the Pol II EC.

Similar content being viewed by others

Accession codes

Primary accessions

Electron Microscopy Data Bank

Protein Data Bank

Data deposits

Cryo-electron microscopy densities have been deposited in the Electron Microscopy Data Bank under accession numbers EMD-3218 (EC1), EMD-3219 (EC2), and EMD-3220 (EC3). Model coordinates have been deposited in the Protein Data Bank (PDB) under accession number 5FLM.

References

  1. Cramer, P. et al. Architecture of RNA polymerase II and implications for the transcription mechanism. Science 288, 640–649 (2000)

    Article  ADS  CAS  PubMed  Google Scholar 

  2. Cramer, P., Bushnell, D. A. & Kornberg, R. D. Structural basis of transcription: RNA polymerase II at 2.8 angstrom resolution. Science 292, 1863–1876 (2001)

    Article  ADS  CAS  PubMed  Google Scholar 

  3. Spåhr, H., Calero, G., Bushnell, D. A. & Kornberg, R. D. Schizosacharomyces pombe RNA polymerase II at 3.6-Å resolution. Proc. Natl Acad. Sci. USA 106, 9185–9190 (2009)

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  4. Kostek, S. A. et al. Molecular architecture and conformational flexibility of human RNA polymerase II. Structure 14, 1691–1700 (2006)

    Article  CAS  PubMed  Google Scholar 

  5. Kassube, S. A., Jinek, M., Fang, J., Tsutakawa, S. & Nogales, E. Structural mimicry in transcription regulation of human RNA polymerase II by the DNA helicase RECQL5. Nature Struct. Mol. Biol. 20, 892–899 (2013)

    Article  CAS  Google Scholar 

  6. Kassube, S. A. et al. Structural insights into transcriptional repression by noncoding RNAs that bind to human Pol II. J. Mol. Biol. 425, 3639–3648 (2013)

    Article  CAS  PubMed  Google Scholar 

  7. He, Y., Fang, J., Taatjes, D. J. & Nogales, E. Structural visualization of key steps in human transcription initiation. Nature 495, 481–486 (2013)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  8. Bernecky, C., Grob, P., Ebmeier, C. C., Nogales, E. & Taatjes, D. J. Molecular architecture of the human Mediator–RNA polymerase II–TFIIF assembly. PLoS Biol. 9, e1000603 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Bernecky, C. & Taatjes, D. J. Activator–mediator binding stabilizes RNA polymerase II orientation within the human mediator–RNA polymerase II–TFIIF assembly. J. Mol. Biol. 417, 387–394 (2012)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Wu, Y. M. et al. Regulation of mammalian transcription by Gdown1 through a novel steric crosstalk revealed by cryo-EM. EMBO J. 31, 3575–3587 (2012)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Hu, X. et al. A Mediator-responsive form of metazoan RNA polymerase II. Proc. Natl Acad. Sci. USA 103, 9506–9511 (2006)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  12. Gnatt, A. L., Cramer, P., Fu, J., Bushnell, D. A. & Kornberg, R. D. Structural basis of transcription: an RNA polymerase II elongation complex at 3.3 A resolution. Science 292, 1876–1882 (2001)

    Article  ADS  CAS  PubMed  Google Scholar 

  13. Armache, K. J., Kettenberger, H. & Cramer, P. Architecture of initiation-competent 12-subunit RNA polymerase II. Proc. Natl Acad. Sci. USA 100, 6964–6968 (2003)

    Article  ADS  CAS  Google Scholar 

  14. Bushnell, D. A. & Kornberg, R. D. Complete, 12-subunit RNA polymerase II at 4.1-A resolution: implications for the initiation of transcription. Proc. Natl Acad. Sci. USA 100, 6969–6973 (2003)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  15. Plaschka, C. et al. Architecture of the RNA polymerase II–Mediator core initiation complex. Nature 518, 376–380 (2015)

    Article  ADS  CAS  PubMed  Google Scholar 

  16. Chang, L., Zhang, Z., Yang, J., McLaughlin, S. H. & Barford, D. Atomic structure of the APC/C and its mechanism of protein ubiquitination. Nature 522, 450–454 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  17. Liang, B. et al. Structure of the L protein of vesicular stomatitis virus from electron cryomicroscopy. Cell 162, 314–327 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Urnavicius, L. et al. The structure of the dynactin complex and its interaction with dynein. Science 347, 1441–1446 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  19. Bai, X. C. et al. An atomic structure of human γ-secretase. Nature 525, 212–217 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  20. Kettenberger, H., Armache, K. J. & Cramer, P. Complete RNA polymerase II elongation complex structure and its interactions with NTP and TFIIS. Mol. Cell 16, 955–965 (2004)

    Article  CAS  PubMed  Google Scholar 

  21. Martinez-Rucobo, F. W. & Cramer, P. Structural basis of transcription elongation. Biochim. Biophys. Acta 1829, 9–19 (2013)

    Google Scholar 

  22. Barnes, C. O. et al. Crystal structure of a transcribing RNA polymerase II complex reveals a complete transcription bubble. Mol. Cell 59, 258–269 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Fishburn, J., Tomko, E., Galburt, E. & Hahn, S. Double-stranded DNA translocase activity of transcription factor TFIIH and the mechanism of RNA polymerase II open complex formation. Proc. Natl Acad. Sci. USA 112, 3961–3966 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  24. Wang, D., Bushnell, D. A., Westover, K. D., Kaplan, C. D. & Kornberg, R. D. Structural basis of transcription: role of the trigger loop in substrate specificity and catalysis. Cell 127, 941–954 (2006)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Andrecka, J. et al. Nano positioning system reveals the course of upstream and nontemplate DNA within the RNA polymerase II elongation complex. Nucleic Acids Res. 37, 5803–5809 (2009)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Martinez-Rucobo, F. W., Sainsbury, S., Cheung, A. C. & Cramer, P. Architecture of the RNA polymerase-Spt4/5 complex and basis of universal transcription processivity. EMBO J. 30, 1302–1310 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Li, W., Giles, C. & Li, S. Insights into how Spt5 functions in transcription elongation and repressing transcription coupled DNA repair. Nucleic Acids Res. 42, 7069–7083 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Hirtreiter, A. et al. Spt4/5 stimulates transcription elongation through the RNA polymerase clamp coiled-coil motif. Nucleic Acids Res. 38, 4040–4051 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Belogurov, G. A., Sevostyanova, A., Svetlov, V. & Artsimovitch, I. Functional regions of the N-terminal domain of the antiterminator RfaH. Mol. Microbiol. 76, 286–301 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Sevostyanova, A. & Artsimovitch, I. Functional analysis of Thermus thermophilus transcription factor NusG. Nucleic Acids Res. 38, 7432–7445 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Wenzel, S., Martins, B. M., Rösch, P. & Wöhrl, B. M. Crystal structure of the human transcription elongation factor DSIF hSpt4 subunit in complex with the hSpt5 dimerization interface. Biochem. J. 425, 373–380 (2010)

    Article  CAS  Google Scholar 

  32. Korinek, A., Beck, F., Baumeister, W., Nickell, S. & Plitzko, J. M. Computer controlled cryo-electron microscopy—TOM2 a software package for high-throughput applications. J. Struct. Biol. 175, 394–405 (2011)

    Article  PubMed  Google Scholar 

  33. Li, X. et al. Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM. Nature Methods 10, 584–590 (2013)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Scheres, S. H. RELION: implementation of a Bayesian approach to cryo-EM structure determination. J. Struct. Biol. 180, 519–530 (2012)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Rohou, A. & Grigorieff, N. CTFFIND4: fast and accurate defocus estimation from electron micrographs. J. Struct. Biol. 192, 216–221 (2015)

    Article  PubMed  PubMed Central  Google Scholar 

  36. Tang, G. et al. EMAN2: an extensible image processing suite for electron microscopy. J. Struct. Biol. 157, 38–46 (2007)

    Article  CAS  PubMed  Google Scholar 

  37. Scheres, S. H. Semi-automated selection of cryo-EM particles in RELION-1.3. J. Struct. Biol. 189, 114–122 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  38. Scheres, S. H. Beam-induced motion correction for sub-megadalton cryo-EM particles. eLife 3, e03665 (2014)

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  39. Rosenthal, P. B. & Henderson, R. Optimal determination of particle orientation, absolute hand, and contrast loss in single-particle electron cryomicroscopy. J. Mol. Biol. 333, 721–745 (2003)

    Article  CAS  PubMed  Google Scholar 

  40. Scheres, S. H. & Chen, S. Prevention of overfitting in cryo-EM structure determination. Nature Methods 9, 853–854 (2012)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Chen, S. et al. High-resolution noise substitution to measure overfitting and validate resolution in 3D structure determination by single particle electron cryomicroscopy. Ultramicroscopy 135, 24–35 (2013)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Cardone, G., Heymann, J. B. & Steven, A. C. One number does not fit all: mapping local variations in resolution in cryo-EM reconstructions. J. Struct. Biol. 184, 226–236 (2013)

    Article  PubMed  Google Scholar 

  43. Pettersen, E. F. et al. UCSF Chimera—a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004)

    Article  CAS  PubMed  Google Scholar 

  44. Sievers, F. et al. Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega. Mol. Syst. Biol. 7, 539 (2011)

    Article  PubMed  PubMed Central  Google Scholar 

  45. Sainsbury, S., Niesser, J. & Cramer, P. Structure and function of the initially transcribing RNA polymerase II–TFIIB complex. Nature 493, 437–440 (2013)

    Article  ADS  CAS  PubMed  Google Scholar 

  46. Winn, M. D. et al. Overview of the CCP4 suite and current developments. Acta Crystallogr. D 67, 235–242 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Stein, N. CHAINSAW: a program for mutating pdb files used as templates in molecular replacement. J. Appl. Crystallogr. 41, 641–643 (2008)

    Article  CAS  Google Scholar 

  48. Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D 66, 486–501 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Meka, H., Werner, F., Cordell, S. C., Onesti, S. & Brick, P. Crystal structure and RNA binding of the Rpb4/Rpb7 subunits of human RNA polymerase II. Nucleic Acids Res. 33, 6435–6444 (2005)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D 66, 213–221 (2010)

    Google Scholar 

  51. Chen, V. B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D 66, 12–21 (2010)

    Google Scholar 

  52. Barad, B. A. et al. EMRinger: side chain-directed model and map validation for 3D cryo-electron microscopy. Nature Methods 12, 943–946 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Cheng, B. & Price, D. H. Analysis of factor interactions with RNA polymerase II elongation complexes using a new electrophoretic mobility shift assay. Nucleic Acids Res. 36, e135 (2008)

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  54. Martinez-Rucobo, F. W. et al. Molecular basis of transcription-coupled pre-mRNA capping. Mol. Cell 58, 1079–1089 (2015)

    Article  CAS  PubMed  Google Scholar 

  55. Herzog, F. et al. Structural probing of a protein phosphatase 2A network by chemical cross-linking and mass spectrometry. Science 337, 1348–1352 (2012)

    Article  ADS  CAS  PubMed  Google Scholar 

  56. Missra, A. & Gilmour, D. S. Interactions between DSIF (DRB sensitivity inducing factor), NELF (negative elongation factor), and the Drosophila RNA polymerase II transcription elongation complex. Proc. Natl Acad. Sci. USA 107, 11301–11306 (2010)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  57. Brueckner, F., Hennecke, U., Carell, T. & Cramer, P. CPD damage recognition by transcribing RNA polymerase II. Science 315, 859–862 (2007)

    Article  ADS  CAS  PubMed  Google Scholar 

  58. Brueckner, F. & Cramer, P. Structural basis of transcription inhibition by α-amanitin and implications for RNA polymerase II translocation. Nature Struct. Mol. Biol. 15, 811–818 (2008)

    Article  CAS  Google Scholar 

  59. Cheung, A. C. & Cramer, P. Structural basis of RNA polymerase II backtracking, arrest and reactivation. Nature 471, 249–253 (2011)

    Article  ADS  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank E. Wolf for providing bovine thymus. We thank D. Pöllmann for preparation of human DSIF, and M. Hemann for generation of peptides for mass spectrometry. We thank J. Schuller and C. Plaschka for assistance with EM data collection. We thank D. Tegunov and C. Plaschka for discussions about EM data processing. We thank D. Haselbach and W. Liu for assistance with cryo-EM grid preparation. We thank O. Berninghausen for providing access to a Tecnai Spirit microscope. C.B. was supported by the Alexander von Humboldt Foundation. P.C. was supported by the Deutsche Forschungsgemeinschaft (SFB860), the Advanced Grant ‘TRANSIT’ of the European Research Council, and the Volkswagen Foundation.

Author information

Authors and Affiliations

  1. Department of Molecular Biology, Max Planck Institute for Biophysical Chemistry, Am Faßberg 11, Göttingen, 37077, Germany

    Carrie Bernecky & Patrick Cramer

  2. Gene Center Munich, Ludwig-Maximilians-Universität München, Feodor-Lynen-Straße 25, Munich, 81377, Germany

    Franz Herzog

  3. Department of Molecular Structural Biology, Max Planck Institute for Biochemistry, Am Klopferspitz 18, Martinsried, 82152, Germany

    Wolfgang Baumeister & Jürgen M. Plitzko

Authors
  1. Carrie Bernecky
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Franz Herzog
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Wolfgang Baumeister
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jürgen M. Plitzko
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Patrick Cramer
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

C.B. designed and performed all experiments and data analysis. F.H. supervised mass spectrometry. W.B. and J.P. provided access to a high-end EM facility. J.P. supervised EM data collection. P.C. designed and supervised research. C.B. and P.C. prepared the manuscript.

Corresponding author

Correspondence to Patrick Cramer.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Extended data figures and tables

Extended Data Figure 1 Bovine Pol II purification and transcription activity.

a, SDS–polyacrylamide gel electrophoresis analysis of size-exclusion-purified 12-subunit bovine Pol II, recombinant Gdown1, and reconstituted complete Pol II with all 13 subunits (right). b, Extension of RNA by bovine Pol II. Activity assay was performed on Pol II ECs assembled on fully complementary DNA. Complexes were assembled with 12-subunit Pol II (Pol II) or 12-subunit Pol II in complex with Gdown1 (Pol II (G)). Transcription was allowed to proceed for 10 min at 30 °C in the presence of the indicated concentration of NTPs. Purified endogenous bovine Pol II is active in transcription. Approximately 40% of the RNA appears to be extended in the presence of Gdown1. c, Extension of RNA by bovine Pol II–human Gdown1 on the bubble scaffold used for structural studies. Uncrosslinked Pol II EC was incubated with the indicated amount of UTP at 30 °C for 10 min to allow the incorporation of two additional nucleotides. Approximately 30% of RNA is extended. d, Observation of individual Pol II ECs of the expected size under cryo-EM conditions with a K2 direct electron detection device. e, Fifty representative 2D class averages resulting from classification of the full Pol II EC data set.

Extended Data Figure 2 Computational sorting of cryo-EM particle images.

Particle images were sorted by 3D classification in RELION to reveal additional features of the Pol II EC. To reduce computational costs, all classifications were performed without image alignment. For global classifications, a mask encompassing the entire Pol II EC was used, whereas for local classifications masks encompassed only the area of interest as indicated in the diagram. The percentage of total data included in each class is indicated above each panel. Density was coloured to assist in visualization. For global classifications (upper left), density was coloured according to observed mobile regions. Green: clamp, anchor, part of the RPB1 cleft, part of the hybrid-binding region, and part of the active site. Orange: RPB9, RPB1 upper jaw, and part of the RPB1 cleft. Purple: RPB4–RPB7 stalk. Blue: nucleic acids. For focused classifications, density for the region of interest was coloured. This corresponded to the region within the classification mask, except for the classification of upstream DNA, in which case the density in the region of the entire nucleic-acid scaffold was coloured. The three reconstructions used for modelling the Pol II EC are shown in yellow, red, and blue boxes. The EC class displaying a mobile jaw-lobe region is highlighted with an orange box, and the EC class displaying a mobile clamp-active site region is highlighted with a green box.

Extended Data Figure 3 Quality of the cryo-EM reconstructions.

a, Angular distribution of particle images for the Pol II EC1, EC2, and EC3 reconstructions. Black shading indicates the number of particles assigned to a given view, while red dots indicate represented views (at least one particle was assigned within 1° of the point). b, FSC plots for the reconstructions described in a, as well as the FSC plot for the Pol II EC model versus the unsharpened Pol II EC1 map. The first data point after phase randomization is omitted. c, Top–front view and middle slice of the B-factor sharpened Pol II EC1 reconstruction coloured by local resolution. The top–front view (also shown in Extended Data Figure 2) is a rotation of either the top view or front view by 45°. d, Top–front view and middle slice of the unsharpened Pol II EC2 reconstruction coloured by local resolution. e, Top–front view and middle slice of the unsharpened Pol II EC3 reconstruction coloured by local resolution. f, RPB4–RPB7 stalk density for the unsharpened Pol II EC1, Pol II EC2, and stalk focused refinements shown at the same threshold level and filtered to the nominal resolutions (3.4 Å, 3.6 Å, and 4.2 Å). Continuous density for the stalk is visible in the lower resolution focused refinement. g, Density for the protrusion in the unsharpened Pol II EC1 and protrusion-focused refinements shown at the same threshold level and filtered to the nominal resolutions (3.4 Å and 3.8 Å). Density for the protrusion tip is more visible in the focused refinement map.

Extended Data Figure 4 Structural comparison of bovine and yeast Pol II.

a, Structures of the bovine and yeast EC (PDB 1Y1W) were superimposed by alignment of the active site. The protein regions of the bovine EC are shown in two views, coloured by r.m.s.d. of the yeast and bovine models. Insertions and uniquely structured regions within the bovine EC are coloured purple. b, Surface conservation between yeast and bovine Pol II. Strong and weak conservation groups were assigned according to Clustal conventions. c, Structural rearrangements within the RPB5 jaw domain. Axes were drawn through the four longest helices of the bovine and yeast RPB5 jaw using UCSF Chimera. d, Comparison of the backbone path of the trigger loop to previously known trigger loop conformations. The trigger loop backbone of the bovine Pol II EC is shown in thick, dark grey; the open conformation of the yeast EC (PDB 1Y1W)20 is in blue; the closed conformation of the yeast EC in the presence of GTP (PDB 2E2H)24 is in yellow; the wedged conformation of the yeast EC in the presence of α-amanitin (PDB 2VUM)58 is in dark green; the trapped conformation of the arrested and backtracked yeast EC (PDB 3PO2)59 is in purple; the locked conformation of the yeast EC reactivation intermediate in the presence of TFIIS (PDB 3PO3)59 is in orange; and the new open position observed in the ten-subunit yeast transcription complex crystallized in the presence of TFIIF is in light green. The binding site for α-amanitin is denoted in dashed green lines. The binding site for the incoming NTP is outlined in dashed yellow lines. e, Comparison of the path of DNA in the closed initiation complex (yellow7) and the elongation complex (blue, this study). Downstream DNA was extended.

Extended Data Figure 5 Modelled position of the human SPT5 NGN domain on the bovine Pol II EC.

a, Zoom-in view of Pol II EC3 nucleic-acid density superimposed on the Pol II–SPT4–SPT5-NGN EC model reveals proximity of non-template DNA to the SPT5 NGN domain. The human SPT4–SPT5-NGN crystal structure (PDB 3H7H)31 was positioned on the Pol II EC model by alignment with the archaeal RNA polymerase clamp–SPT4–SPT5 crystal structure (PDB 3QQC)26. b, Colouring a surface representation of the X-ray structure of the SPT4–SPT5-NGN by Coulombic surface charge reveals positively charged patches in close proximity to non-template and upstream DNA. Positive charge is in blue; negative charge is in red.

Extended Data Table 1 Model refinement statistics
Full size table
Extended Data Table 2 Summary of differences between the yeast EC (X-ray, PDB 1Y1W) and bovine EC (cryo-EM)
Full size table

Supplementary information

Supplementary Table 1

This table contains a list of observed Pol II EC-DSIF protein-protein crosslinks. Inter-protein crosslinks are listed first, followed by intra-protein crosslinks. Within these categories, crosslinks are listed in order of decreasing Id-Score. Crosslink precursor ions with unique M/z values were treated as unique crosslinks in the analysis. (PDF 46 kb)

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bernecky, C., Herzog, F., Baumeister, W. et al. Structure of transcribing mammalian RNA polymerase II. Nature 529, 551–554 (2016). https://doi.org/10.1038/nature16482

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature16482

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing