Year: 2026
Type: article
Abstract: Multiple lines of evidence suggest that RNase H1-sensitive RNA:DNA hybrids could disrupt the stability and restart of stalled replication forks. The prolonged over-expression of RNase H1 is the most popular method to manipulate RNA:DNA hybrids in situ but it cannot discriminate between direct and indirect effects resulting from their loss. To overcome this limitation, we developed iCoRD (in Cellulo RNase H Delivery) to rapidly deliver comparable amounts of either Escherichia coli RNase H1 (RnhA) or control proteins in live cells, with a negligible impact on the transcriptome. iCoRD-delivered RnhA recognises RNA:DNA hybrids in situ, is enriched in the vicinity of active replication forks and rescues replication fork progression in different stress conditions. While iCoRD-delivered RnhA had a clear impact on stress-induced, fork-proximal RNA:DNA hybrids and R-ChIP signals, it had no effect on DRIP or S9.6 Cut&Tag signals, even when these were induced by stress. Our results strongly suggest the existence of distinct populations of RNA:DNA hybrids that are best mapped by different methods and are differently sensitive to RNase H1 activity in situ.
Source: The EMBO Journal
Institutions: Lyon 1 Université, École Normale Supérieure de Lyon, Centre National de la Recherche Scientifique, Inserm, Université de Lyon, Laboratoire de Biologie et Modélisation de la Cellule, Institut de Génétique Humaine, University of Exeter, MRC Centre for Medical Mycology, Helmholtz Munich, Institute of Epigenetics and Stem Cells
Language: English
Topic: DNA Repair Mechanisms
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $7,990
Funders: IDEXLYON, European Commission, Agence Nationale de la Recherche, Institut National de la Santé et de la Recherche Médicale, Fondation pour la Recherche Médicale, Fondation ARC pour la Recherche sur le Cancer, Ligue Contre le Cancer, Centre National de la Recherche Scientifique, Université Claude Bernard Lyon 1, Université de Lyon, École Normale Supérieure de Lyon, Laboratoire d'Excellence Ecofect, HORIZON EUROPE European Research Council
Awards: G1682526469, G169068394, Replication Stress caused by Post-replicative RNA:DNA hybrids, G2125209779, PROJET AVENIR LYON SAINT-ETIENNE, G3008589337, G3785575267, Dynamiques eco-évolutives des maladies infectieuses, G5649854608, Unveiling the role of deregulated transcription termination and R-loop metabolism in motor neuron disease, G6215407190, G657559870, G6888366887, G7080048889, RNA:DNA hybrids and replication fork instability
OpenAlex ID: W7223099450