LEDGF/p75 promotes transcriptional pausing through preventing SPT5 phosphorylation

The transcriptional progression is regulated by SPT5, which is involved in both promoter-proximal RNA polymerase II (Pol II) pausing and elongation, which serve as rate-limiting steps in metazoan. However, the mechanism for SPT5 coordinating these processes remains unclear. Here we report that the disordered sub-regions within SPT5 contain a prion-like domain (PLD) that is required for Pol II pausing, while the phosphorylation site-rich domain (PRD) is essential for transcript elongation. Mechanically, SPT5-PLD helps stabilize Pol II pausing by preventing SPT5-PRD phosphorylation mediated by the super elongation complex (SEC). SPT5-PLD also maintains Pol II pausing by recruiting a newly identified transcriptional pause regulator, PSIP1, which inhibits phosphorylation required for elongation. Additionally, SPT5-PLD prevents premature release of Pol II pausing through PSIP1's IBD domain by counteracting SEC. Our results demonstrate that SPT5 possesses intrinsic characteristics that balance the transcriptional process by preventing premature pause release and promoting progression to elongation. Overall design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for AFF4,Pol II, SPT5,SPT5-pCTR1, NELFE in HCT116 cells. Cleavage Under Targets and Tagmentation (CUT&Tag) for LEDGF in HCT116 cells.