Epigenetic reprogramming by TET enzymes impacts co-transcriptional R-loops
- Universidade de Lisboa, Portugal
- Universidade Nova de Lisboa, Portugal
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.
Article and author information
Author details
Madalena R de Almeida
Sérgio Fernandes de Almeida
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
- Andrés Aguilera, CABIMER, Universidad de Sevilla, Spain
Version history
- Received: April 16, 2021
- Preprint posted: April 27, 2021 (view preprint)
- Accepted: February 21, 2022
- Accepted Manuscript published: February 22, 2022 (version 1)
- 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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Epigenetic reprogramming by TET enzymes impacts co-transcriptional R-loops
eLife 11:e69476.
https://doi.org/10.7554/eLife.69476
Further reading
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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.
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