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Structural mimicry in transcription regulation of human RNA polymerase II by the DNA helicase RECQL5

Nature Structural & Molecular Biology volume 20, pages 892–899 (2013)Cite this article

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Abstract

RECQL5 is a member of the highly conserved RecQ family of DNA helicases involved in DNA repair. RECQL5 interacts with RNA polymerase II (Pol II) and inhibits transcription of protein-encoding genes by an unknown mechanism. We show that RECQL5 contacts the Rpb1 jaw domain of Pol II at a site that overlaps with the binding site for the transcription elongation factor TFIIS. Our cryo-EM structure of elongating Pol II arrested in complex with RECQL5 shows that the RECQL5 helicase domain is positioned to sterically block elongation. The crystal structure of the RECQL5 KIX domain reveals similarities with TFIIS, and binding of RECQL5 to Pol II interferes with the ability of TFIIS to promote transcriptional read-through in vitro. Together, our findings reveal a dual mode of transcriptional repression by RECQL5 that includes structural mimicry of the Pol II–TFIIS interaction.

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Figure 1: An extended helicase domain fragment of RECQL5 interacts with the upper jaw domain of Pol II.
Figure 2: The RECQL5 KIX domain resembles domain II of TFIIS.
Figure 3: The RECQL5 IRI domain binds to a loop element in the Rpb1 jaw of Pol II.
Figure 4: Cryo-EM reconstruction of RECQL5-stalled Pol II elongation complex.
Figure 5: Both RECQL5 helicase and IRI domains are required for repression of transcription.
Figure 6: RECQL5 IRI domain competes with TFIIS for binding to Pol II and inhibits TFIIS-mediated read-through of intrinsic elongation blocks.
Figure 7: Dual mechanism of RECQL5-mediated transcriptional repression in DNA repair.

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Acknowledgements

We thank G. Lander and P. Grob for advice on EM data collection and processing, and F. Bleichert for critical reading of the manuscript. We thank C. Kane (University of California, Berkeley) for providing the pGEMTerm plasmid, advice on the transcriptional read-through experiment and critical reading of the manuscript. We are grateful to J. Holton (beamline 8.3.1, Advanced Light Source, Lawrence Berkeley National Laboratory) for assistance with synchrotron data collection. We thank D. King (Howard Hughes Medical Institute, University of California, Berkeley) for synthesis of the CTD peptide used for purification of Pol II. S.A.K. was supported by a fellowship from the Boehringer Ingelheim Fonds. The work was supported by National Institute of General Medical Sciences grant GM63072 (E.N.). S.T. and the SIBYLS beamline (12.3.1) at the Advanced Light Source are supported by NIH grant P01 CA092584 and the US Department of Energy Integrated Diffraction Analysis Technologies (IDAT) under contract number DE-AC02-05CH11231. E.N. is a Howard Hughes Medical Institute Investigator.

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  1. Martin Jinek

    Present address: Present address: Department of Biochemistry, University of Zurich, Zurich, Switzerland.,

Authors and Affiliations

  1. Biophysics Graduate Group, University of California, Berkeley, Berkeley, California, USA

    Susanne A Kassube

  2. Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, California, USA

    Martin Jinek, Jie Fang & Eva Nogales

  3. Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA

    Martin Jinek & Eva Nogales

  4. Life Science Division, Lawrence Berkeley National Lab, Berkeley, California, USA

    Susan Tsutakawa & Eva Nogales

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Contributions

S.A.K. conceived the study, designed and performed experiments and analyzed the data. M.J. and S.A.K. collected and processed X-ray diffraction data. J.F. and S.A.K. purified human Pol II. S.T. collected and processed SAXS data. E.N. oversaw the project and S.A.K., M.J. and E.N. prepared the manuscript.

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Correspondence to Susanne A Kassube or Eva Nogales.

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Supplementary Figures 1–6 (PDF 3635 kb)

Supplementary Movie 1

Cryo-EM model of a RECQL5-stalled Pol II elongation complex. (MOV 62768 kb)

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Kassube, S., Jinek, M., Fang, J. et al. Structural mimicry in transcription regulation of human RNA polymerase II by the DNA helicase RECQL5. Nat Struct Mol Biol 20, 892–899 (2013). https://doi.org/10.1038/nsmb.2596

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