Genomic patterns of transcription–replication interactions in mouse primary B cells

• Published in: Nucleic acids research Vol. 50; no. 4; pp. 2051 - 2073
• Main Authors: St Germain, Commodore P, Zhao, Hongchang, Sinha, Vrishti, Sanz, Lionel A, Chédin, Frédéric, Barlow, Jacqueline H
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 28.02.2022
• Subjects: Animals; B-Lymphocytes - metabolism; DNA Breaks, Double-Stranded; DNA Repair; DNA Replication - genetics; Genome Integrity, Repair and; Genomic Instability; Mice; Transcription, Genetic
• ISSN: 0305-1048; 1362-4962
• DOI: 10.1093/nar/gkac035

Abstract Conflicts between transcription and replication machinery are a potent source of replication stress and genome instability; however, no technique currently exists to identify endogenous genomic locations prone to transcription–replication interactions. Here, we report a novel method to identify genomic loci prone to transcription–replication interactions termed transcription–replication immunoprecipitation on nascent DNA sequencing, TRIPn-Seq. TRIPn-Seq employs the sequential immunoprecipitation of RNA polymerase 2 phosphorylated at serine 5 (RNAP2s5) followed by enrichment of nascent DNA previously labeled with bromodeoxyuridine. Using TRIPn-Seq, we mapped 1009 unique transcription–replication interactions (TRIs) in mouse primary B cells characterized by a bimodal pattern of RNAP2s5, bidirectional transcription, an enrichment of RNA:DNA hybrids, and a high probability of forming G-quadruplexes. TRIs are highly enriched at transcription start sites and map to early replicating regions. TRIs exhibit enhanced Replication Protein A association and TRI-associated genes exhibit higher replication fork termination than control transcription start sites, two marks of replication stress. TRIs colocalize with double-strand DNA breaks, are enriched for deletions, and accumulate mutations in tumors. We propose that replication stress at TRIs induces mutations potentially contributing to age-related disease, as well as tumor formation and development.