RNA polymerase II stalling at pre-mRNA splice sites is enforced by ubiquitination of the catalytic subunit

  1. Laura Milligan
  2. Camille Sayou
  3. Alex Tuck  Is a corresponding author
  4. Tatsiana Auchynnikava
  5. Jane E A Reid
  6. Ross Alexander
  7. Flavia de Lima Alves
  8. Robin Allshire
  9. Christos Spanos
  10. Juri Rappsilber
  11. Jean D Beggs
  12. Grzegorz Kudla
  13. David Tollervey  Is a corresponding author
  1. University of Edinburgh, United Kingdom
  2. Friedrich Miescher Institute for Biomedical Research, Switzerland

Abstract

Numerous links exist between co-transcriptional RNA processing and the transcribing RNAPII. In particular, pre-mRNA splicing was reported to be associated with slowed RNAPII elongation. Here we identify a site of ubiquitination (K1246) in the catalytic subunit of RNAPII close to the DNA entry path. Ubiquitination was increased in the absence of the Bre5-Ubp3 ubiquitin protease complex. Bre5 binds RNA in vivo, with a preference for exon 2 regions of intron-containing pre-mRNAs and poly(A) proximal sites. Ubiquitinated RNAPII showed similar enrichment. The absence of Bre5 led to impaired splicing and defects in RNAPII elongation in vivo on a splicing reporter construct. Strains expressing RNAPII with a K1246R mutation showed reduced cotranscriptional splicing. We propose that ubiquinitation of RNAPII is induced by RNA processing events and linked to transcriptional pausing, which is released by Bre5-Ubp3 associated with the nascent transcript.

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Article and author information

Author details

  1. Laura Milligan

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. Camille Sayou

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8226-7272
  3. Alex Tuck

    Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland
    For correspondence
    alex.tuck@fmi.ch
    Competing interests
    The authors declare that no competing interests exist.
  4. Tatsiana Auchynnikava

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Jane E A Reid

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  6. Ross Alexander

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  7. Flavia de Lima Alves

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  8. Robin Allshire

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  9. Christos Spanos

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  10. Juri Rappsilber

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  11. Jean D Beggs

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  12. Grzegorz Kudla

    MRC Human Genetics Unit, IGMM, University of Edinburgh, Edinburgh, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  13. David Tollervey

    Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, United Kingdom
    For correspondence
    d.tollervey@ed.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2894-2772

Funding

Wellcome (77248)

  • David Tollervey

Medical Research Council

  • Grzegorz Kudla

Wellcome (104648)

  • Jean D Beggs

Wellcome (108504)

  • Juri Rappsilber

Wellcome (93853)

  • Jane E A Reid

Wellcome (97383)

  • Grzegorz Kudla

Wellcome (92076)

  • David Tollervey

Wellcome (200885)

  • Robin Allshire

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Nick J Proudfoot, University of Oxford, United Kingdom

Publication history

  1. Received: March 22, 2017
  2. Accepted: October 12, 2017
  3. Accepted Manuscript published: October 13, 2017 (version 1)
  4. Version of Record published: November 6, 2017 (version 2)

Copyright

© 2017, Milligan et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Laura Milligan
  2. Camille Sayou
  3. Alex Tuck
  4. Tatsiana Auchynnikava
  5. Jane E A Reid
  6. Ross Alexander
  7. Flavia de Lima Alves
  8. Robin Allshire
  9. Christos Spanos
  10. Juri Rappsilber
  11. Jean D Beggs
  12. Grzegorz Kudla
  13. David Tollervey
(2017)
RNA polymerase II stalling at pre-mRNA splice sites is enforced by ubiquitination of the catalytic subunit
eLife 6:e27082.
https://doi.org/10.7554/eLife.27082

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