Elucidate the functions of the BET bromodomain protein BRD4 in co-transcriptional splicing

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Transcription by RNA polymerase II (Pol II) underlies fundamental cellular processes in eukaryotic cells and is strictly regulated. A major control point occurs during early elongation before Pol II is released into productive elongation. Prior research has linked the BET bromodomain protein family, with its members BRD2, BRD3 and BRD4, with transcription elongation in human cells. Our research has recently provided evidence that the BET bromodomain protein BRD4 underlies the general 5’-elongation checkpoint which helps to assemble a functional Pol II elongation complex and stimulates productive elongation. Transcription elongation is tightly coupled and temporally coordinated with RNA processing to enable RNA maturation while the RNA is being synthesized. We recently uncovered that BRD4 serves as a molecular link between transcription elongation and RNA processing. We found that BRD4 recruits 3’-RNA processing factors to transcriptionally engaged Pol II during the 5’-elongation checkpoint to enable RNA cleavage at the 3’-end of genes (Arnold et al., Mol Cell, 2021). We also found that BRD4 interacts with subunits of the spliceosome and that acute BET protein ablation impacts the splicing pattern at a large set of genes. These findings indicate that BRD4 is implicated in co-transcriptional splicing, similarly to transcription-coupled 3’-RNA processing. In this project, we aim to identify the molecular mechanisms of splicing control by BRD4 and to characterize the binding interface between BRD4 and the spliceosome in human cells. In order to accomplish the objectives, we will apply a high-resolution and quantitative multiomics approach.