期刊
NUCLEIC ACIDS RESEARCH
卷 50, 期 4, 页码 2051-2073出版社
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkac035
关键词
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资金
- National Institutes of Health
- NIH-IMSD-T32 [R25 GM056765]
- NIH-MCB-T32 [T32 GM007377]
- NCI-F31 [F31 CA213871]
- National Cancer Institute, NCI-K22 [K22CA188106, P30CA093373]
- National Institutes of General Medical Sciences [NIH R35 GM139549]
- National Institutes of Health Shared Instrumentation Grant [S10 OD010786-01]
- National Institute of Health [S100D018223]
Conflicts between transcription and replication machinery can lead to replication stress and genome instability. The newly developed TRIPn-Seq method allows the identification of genomic loci prone to transcription-replication interactions. Using TRIPn-Seq, the authors mapped 1009 unique transcription-replication interactions in mouse primary B cells, which were enriched at transcription start sites and early replicating regions.
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.
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