The interplay of RNA:DNA hybrid structure and G-quadruplexes determines the outcome of R-loop-replisome collisions
Abstract
R-loops are a major source of genome instability associated with transcription-induced replication stress. However, how R-loops inherently impact replication fork progression is not understood. Here, we characterize R-loop-replisome collisions using a fully reconstituted eukaryotic DNA replication system. We find that RNA:DNA hybrids and G-quadruplexes at both co-directional and head-on R-loops can impact fork progression by inducing fork stalling, uncoupling of leading strand synthesis from replisome progression, and nascent strand gaps. RNase H1 and Pif1 suppress replication defects by resolving RNA:DNA hybrids and G-quadruplexes, respectively. We also identify an intrinsic capacity of replisomes to maintain fork progression at certain R-loops by unwinding RNA:DNA hybrids, repriming leading strand synthesis downstream of G-quadruplexes, or utilizing R-loop transcripts to prime leading strand restart during co-directional R-loop-replisome collisions. Collectively, the data demonstrates that the outcome of R-loop-replisome collisions is modulated by R-loop structure, providing a mechanistic basis for the distinction of deleterious from non-deleterious R-loops.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided.
Article and author information
Author details
Funding
National Institute of General Medical Sciences (R01-GM127428)
- Dirk Remus
National Institute of General Medical Sciences (R01-GM107239)
- Dirk Remus
National Institutes of Health (P30 CA008748)
- Dirk Remus
National Institute of Environmental Health Sciences (ES031635)
- Jack D Griffith
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Bruce Stillman, Cold Spring Harbor Laboratory, United States
Version history
- Preprint posted: July 17, 2021 (view preprint)
- Received: July 18, 2021
- Accepted: September 7, 2021
- Accepted Manuscript published: September 8, 2021 (version 1)
- Version of Record published: September 28, 2021 (version 2)
Copyright
© 2021, Kumar 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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