Abstract
Effective spatio-temporal control of transcription and replication during S-phase is paramount to maintaining genomic integrity and cell survival. Dysregulation of these systems can lead to conflicts between the transcription and replication machinery, causing DNA damage and cell death. BRD4 allows efficient transcriptional elongation by stimulating phosphorylation of RNA polymerase II (RNAPII). We report that bromodomain and extra-terminal domain (BET) protein loss of function (LOF) causes RNAPII pausing on the chromatin and DNA damage affecting cells in S-phase. This persistent RNAPII-dependent pausing leads to an accumulation of RNA:DNA hybrids (R-loops) at sites of BRD4 occupancy, leading to transcription-replication conflicts (TRCs), DNA damage, and cell death. Finally, our data show that the BRD4 C-terminal domain, which interacts with P-TEFb, is required to prevent R-loop formation and DNA damage caused by BET protein LOF.