Abstract

DNA oxidation by ten-eleven translocation (TET) family enzymes is essential for epigenetic reprogramming. The conversion of 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC) initiates developmental and cell-type-specific transcriptional programs through mechanisms that include changes in the chromatin structure. Here, we show that the presence of 5hmC in the transcribed gene promotes the annealing of the nascent RNA to the template DNA strand, leading to the formation of an R-loop. Depletion of TET enzymes reduced global R-loops in absence of gene expression changes, whereas CRISPR-mediated tethering of TET to an active gene promoted the formation of R-loops. The genome-wide distribution of 5hmC and R-loops show a positive correlation in mouse and human stem cells and overlap in half of all active genes. Moreover, R-loop resolution leads to differential expression of a subset of genes that are involved in crucial events during stem cell proliferation. Altogether, our data reveal that epigenetic reprogramming via TET activity promotes co-transcriptional R-loop formation, disclosing new mechanisms of gene expression regulation.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files.

The following previously published data sets were used

Article and author information

Author details

  1. João C Sabino

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7991-4291
  2. Madalena R de Almeida

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9539-3289
  3. Patrícia L Abreu

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6387-8537
  4. Ana M Ferreira

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
  5. Paulo Caldas

    Institute for Health and Bioeconomy, Universidade Nova de Lisboa, Lisboa, Portugal
    Competing interests
    The authors declare that no competing interests exist.
  6. Marco M Domingues

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
  7. Nuno C Santos

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
  8. Claus M Azzalin

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9396-1980
  9. Ana Rita Grosso

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6974-4209
  10. Sérgio Fernandes de Almeida

    Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal
    For correspondence
    sergioalmeida@fm.ul.pt
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7774-1355

Funding

Fundação para a Ciência e Tecnologia, Portugal (PTDC/BIA-MOL/30438/2017)

  • Sérgio Fernandes de Almeida

Fundação para a Ciência e Tecnologia, Portugal (PTDC/MED-OUT/4301/2020)

  • Sérgio Fernandes de Almeida

EU Horizon 2020 Research and Innovation Programme (RiboMed 857119)

  • Sérgio Fernandes de Almeida

Fundação para a Ciência e Tecnologia, Portugal (PD/BD/128292/2017)

  • João C Sabino

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

Reviewing Editor

  1. Andrés Aguilera, CABIMER, Universidad de Sevilla, Spain

Version history

  1. Received: April 16, 2021
  2. Preprint posted: April 27, 2021 (view preprint)
  3. Accepted: February 21, 2022
  4. Accepted Manuscript published: February 22, 2022 (version 1)
  5. Version of Record published: March 4, 2022 (version 2)

Copyright

© 2022, Sabino 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. João C Sabino
  2. Madalena R de Almeida
  3. Patrícia L Abreu
  4. Ana M Ferreira
  5. Paulo Caldas
  6. Marco M Domingues
  7. Nuno C Santos
  8. Claus M Azzalin
  9. Ana Rita Grosso
  10. Sérgio Fernandes de Almeida
(2022)
Epigenetic reprogramming by TET enzymes impacts co-transcriptional R-loops
eLife 11:e69476.
https://doi.org/10.7554/eLife.69476

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https://doi.org/10.7554/eLife.69476