Pervasive transcription fine-tunes replication origin activity
- Université Paris-Diderot, France
Abstract
RNA polymerase (RNAPII) transcription occurs pervasively, raising the important question of its functional impact on other DNA-associated processes, including replication. In budding yeast, replication originates from Autonomously Replicating Sequences (ARSs), generally located in intergenic regions. The influence of transcription on ARSs function has been studied for decades, but these earlier studies have neglected the role of non-annotated transcription. We studied the relationships between pervasive transcription and replication origin activity using high-resolution transcription maps. We show that ARSs alter the pervasive transcription landscape by pausing and terminating neighboring RNAPII transcription, thus limiting the occurrence of pervasive transcription within origins. We propose that quasi-symmetrical binding of the ORC complex to ARS borders and/or pre-RC formation are responsible for pausing and termination. We show that low, physiological levels of pervasive transcription impact the function of replication origins. Overall, our results have important implications for understanding the impact of genomic location on origin function.
Data availability
All data analyzed in this manuscript have been previously published and appropriate GEO accession codes and references have been provided.
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Genome-wide mapping of yeast RNA polymerase II termination.NCBI Gene Expression Omnibus, GSE56435.
Article and author information
Author details
Funding
Centre National de la Recherche Scientifique
- Domenico Libri
Fondation pour la Recherche Médicale (F.R.M. programme équipes 2013)
- Domenico Libri
Agence Nationale de la Recherche (Grant ANR-16-CE12-0022-01)
- Domenico Libri
Labex Who Am I? (ANR-11-LABX-0071)
- Domenico Libri
French Ministry of Research (Fellowship)
- Tito Candelli
Ligue Nationale Contre le Cancer (GB/MA/CD/IQ - 12031)
- Julien Gros
Labex Who Am I? (ANR-11-IDEX-0005-02)
- Domenico Libri
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2018, Candelli 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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Pervasive transcription fine-tunes replication origin activity
eLife 7:e40802.
https://doi.org/10.7554/eLife.40802
Further reading
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- Chromosomes and Gene Expression
The association between late replication timing and low transcription rates in eukaryotic heterochromatin is well known, yet the specific mechanisms underlying this link remain uncertain. In Saccharomyces cerevisiae, the histone deacetylase Sir2 is required for both transcriptional silencing and late replication at the repetitive ribosomal DNA (rDNA) arrays. We have previously reported that in the absence of SIR2, a de-repressed RNA PolII repositions MCM replicative helicases from their loading site at the ribosomal origin, where they abut well-positioned, high-occupancy nucleosomes, to an adjacent region with lower nucleosome occupancy. By developing a method that can distinguish activation of closely spaced MCM complexes, here we show that the displaced MCMs at rDNA origins have increased firing propensity compared to the nondisplaced MCMs. Furthermore, we found that both activation of the repositioned MCMs and low occupancy of the adjacent nucleosomes critically depend on the chromatin remodeling activity of FUN30. Our study elucidates the mechanism by which Sir2 delays replication timing, and it demonstrates, for the first time, that activation of a specific replication origin in vivo relies on the nucleosome context shaped by a single chromatin remodeler.
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- Chromosomes and Gene Expression
- Structural Biology and Molecular Biophysics
Type II nuclear receptors (T2NRs) require heterodimerization with a common partner, the retinoid X receptor (RXR), to bind cognate DNA recognition sites in chromatin. Based on previous biochemical and overexpression studies, binding of T2NRs to chromatin is proposed to be regulated by competition for a limiting pool of the core RXR subunit. However, this mechanism has not yet been tested for endogenous proteins in live cells. Using single-molecule tracking (SMT) and proximity-assisted photoactivation (PAPA), we monitored interactions between endogenously tagged RXR and retinoic acid receptor (RAR) in live cells. Unexpectedly, we find that higher expression of RAR, but not RXR, increases heterodimerization and chromatin binding in U2OS cells. This surprising finding indicates the limiting factor is not RXR but likely its cadre of obligate dimer binding partners. SMT and PAPA thus provide a direct way to probe which components are functionally limiting within a complex TF interaction network providing new insights into mechanisms of gene regulation in vivo with implications for drug development targeting nuclear receptors.
