Tyrosine phosphorylation of RNA Polymerase II CTD is associated with antisense promoter transcription and active enhancers in mammalian cells

  1. Nicolas Descostes
  2. Martin Heidemann
  3. Lionel Spinelli
  4. Roland Schüller
  5. Muhammad A Maqbool
  6. Romain Fenouil
  7. Frederic Koch
  8. Charlène Innocenti
  9. Marta Gut
  10. Ivo Gut
  11. Dirk Eick
  12. Jean-Christophe Andrau  Is a corresponding author
  1. Centre d'Immunologie de Marseille Luminy, Université Aix-Marseille, France
  2. Helmholtz Center Munich, Center of Integrated Protein Science Munich (CIPSM), Germany
  3. Centre d'Immunologie de Marseille-Luminy, Université Aix-Marseille, France
  4. Institut de Génétique Moléculaire de Montpellier (IGMM), CNRS-UMR, France
  5. Max Planck Institute for Molecular Genetics, Germany
  6. Institut de Génomique Fonctionnelle, France
  7. Centre Nacional D'Anàlisi Genòmica, Spain
  8. Insitut de Génétique Moléculaire de Montpellier (IGMM), CNRS-UMR, France

Abstract

In mammals, the carboxy-terminal domain (CTD) of RNA polymerase (Pol) II consists of 52 conserved heptapeptide repeats containing the consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. Post-translational modifications of the CTD coordinate the transcription cycle and various steps of mRNA maturation. Here we describe Tyr1 phosphorylation (Tyr1P) as a hallmark of promoter (5' associated) Pol II in mammalian cells, in contrast to what was described in yeast. Tyr1P is predominantly found in antisense orientation at promoters but is also specifically enriched at active enhancers. Mutation of Tyr1 to phenylalanine (Y1F) prevents the formation of the hyper-phosphorylated Pol IIO form, induces degradation of Pol II to the truncated Pol IIB form and results in a lethal phenotype. Our results suggest that Tyr1P has evolved specialized and essential functions in higher eukaryotes associated with antisense promoter and enhancer transcription, and Pol II stability.

Article and author information

Author details

  1. Nicolas Descostes

    Centre d'Immunologie de Marseille Luminy, Université Aix-Marseille, Marseille, France
    Competing interests
    The authors declare that no competing interests exist.
  2. Martin Heidemann

    Helmholtz Center Munich, Center of Integrated Protein Science Munich (CIPSM), Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  3. Lionel Spinelli

    Centre d'Immunologie de Marseille-Luminy, Université Aix-Marseille, Marseilles, France
    Competing interests
    The authors declare that no competing interests exist.
  4. Roland Schüller

    Helmholtz Center Munich, Center of Integrated Protein Science Munich (CIPSM), Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  5. Muhammad A Maqbool

    Institut de Génétique Moléculaire de Montpellier (IGMM), CNRS-UMR, Montpellier, France
    Competing interests
    The authors declare that no competing interests exist.
  6. Romain Fenouil

    Centre d'Immunologie de Marseille-Luminy, Université Aix-Marseille, Marseilles, France
    Competing interests
    The authors declare that no competing interests exist.
  7. Frederic Koch

    Max Planck Institute for Molecular Genetics, Berlin, Germany
    Competing interests
    The authors declare that no competing interests exist.
  8. Charlène Innocenti

    Institut de Génomique Fonctionnelle, Montpellier, France
    Competing interests
    The authors declare that no competing interests exist.
  9. Marta Gut

    Centre Nacional D'Anàlisi Genòmica, Barcelona, Spain
    Competing interests
    The authors declare that no competing interests exist.
  10. Ivo Gut

    Centre Nacional D'Anàlisi Genòmica, Barcelona, Spain
    Competing interests
    The authors declare that no competing interests exist.
  11. Dirk Eick

    Helmholtz Center Munich, Center of Integrated Protein Science Munich (CIPSM), Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  12. Jean-Christophe Andrau

    Insitut de Génétique Moléculaire de Montpellier (IGMM), CNRS-UMR, Montpellier, France
    For correspondence
    jean-christophe.andrau@igmm.cnrs.fr
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Danny Reinberg, HHMI / NYU School of Medicine, United States

Version history

  1. Received: December 17, 2013
  2. Accepted: May 8, 2014
  3. Accepted Manuscript published: May 9, 2014 (version 1)
  4. Version of Record published: June 3, 2014 (version 2)

Copyright

© 2014, Descostes 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.

Metrics

  • 4,847
    Page views
  • 596
    Downloads
  • 62
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Nicolas Descostes
  2. Martin Heidemann
  3. Lionel Spinelli
  4. Roland Schüller
  5. Muhammad A Maqbool
  6. Romain Fenouil
  7. Frederic Koch
  8. Charlène Innocenti
  9. Marta Gut
  10. Ivo Gut
  11. Dirk Eick
  12. Jean-Christophe Andrau
(2014)
Tyrosine phosphorylation of RNA Polymerase II CTD is associated with antisense promoter transcription and active enhancers in mammalian cells
eLife 3:e02105.
https://doi.org/10.7554/eLife.02105

Share this article

https://doi.org/10.7554/eLife.02105

Further reading

    1. Chromosomes and Gene Expression
    2. Genetics and Genomics
    Maria L Adelus, Jiacheng Ding ... Casey E Romanoski
    Research Article

    Heterogeneity in endothelial cell (EC) sub-phenotypes is becoming increasingly appreciated in atherosclerosis progression. Still, studies quantifying EC heterogeneity across whole transcriptomes and epigenomes in both in vitro and in vivo models are lacking. Multiomic profiling concurrently measuring transcriptomes and accessible chromatin in the same single cells was performed on six distinct primary cultures of human aortic ECs (HAECs) exposed to activating environments characteristic of the atherosclerotic microenvironment in vitro. Meta-analysis of single-cell transcriptomes across 17 human ex vivo arterial specimens was performed and two computational approaches quantitatively evaluated the similarity in molecular profiles between heterogeneous in vitro and ex vivo cell profiles. HAEC cultures were reproducibly populated by four major clusters with distinct pathway enrichment profiles and modest heterogeneous responses: EC1-angiogenic, EC2-proliferative, EC3-activated/mesenchymal-like, and EC4-mesenchymal. Quantitative comparisons between in vitro and ex vivo transcriptomes confirmed EC1 and EC2 as most canonically EC-like, and EC4 as most mesenchymal with minimal effects elicited by siERG and IL1B. Lastly, accessible chromatin regions unique to EC2 and EC4 were most enriched for coronary artery disease (CAD)-associated single-nucleotide polymorphisms from Genome Wide Association Studies (GWAS), suggesting that these cell phenotypes harbor CAD-modulating mechanisms. Primary EC cultures contain markedly heterogeneous cell subtypes defined by their molecular profiles. Surprisingly, the perturbations used here only modestly shifted cells between subpopulations, suggesting relatively stable molecular phenotypes in culture. Identifying consistently heterogeneous EC subpopulations between in vitro and ex vivo models should pave the way for improving in vitro systems while enabling the mechanisms governing heterogeneous cell state decisions.

    1. Chromosomes and Gene Expression
    Allison Coté, Aoife O'Farrell ... Arjun Raj
    Research Article

    Splicing is the stepwise molecular process by which introns are removed from pre-mRNA and exons are joined together to form mature mRNA sequences. The ordering and spatial distribution of these steps remain controversial, with opposing models suggesting splicing occurs either during or after transcription. We used single-molecule RNA FISH, expansion microscopy, and live-cell imaging to reveal the spatiotemporal distribution of nascent transcripts in mammalian cells. At super-resolution levels, we found that pre-mRNA formed clouds around the transcription site. These clouds indicate the existence of a transcription-site-proximal zone through which RNA move more slowly than in the nucleoplasm. Full-length pre-mRNA undergo continuous splicing as they move through this zone following transcription, suggesting a model in which splicing can occur post-transcriptionally but still within the proximity of the transcription site, thus seeming co-transcriptional by most assays. These results may unify conflicting reports of co-transcriptional versus post-transcriptional splicing.