Transcription-Replication conflicts are linked to histone H3K79 methylation and R-loop dependent nucleosome eviction [WGS]

Transcription-replication conflicts (TRCs) can provoke genome instability. Progressing transcription and replication machineries profoundly impact their underlying chromatin template and thus, conflict sites are vulnerable to chromatin and epigenome alterations. Here, we engineered an inducible TRC reporter system using a genome-integrated R-loop-prone sequence. Using this system, we characterized the dynamic changes of the local chromatin structure inflicted by TRCs, leading to reduced nucleosome occupancy and replication fork blockage. Consequently, inducing a handful TRCs on the genome results in a measurable global replication stress response. We also find a TRC-specific increase in H3K79 methylation at the R-loop forming TRC-site. Accordingly, inhibition of the H3K79 methyltransferase DOT1L leads to increased cellular TRCs and exacerbated DNA damage response, suggesting that deposition of this mark is required for effective detection and resolution of TRCs. Thus, our work highlights the molecular dynamics and reveals specific epigenetic modifiers bookmarking TRC sites, relevant to cancer and other diseases. Whole Genome Sequencing of sonicated genomic DNA of a monoclonal cell line