Visualising the dynamic nature of RNA polymerase II promoter proximal pausing
RNA polymerase (Pol) II is regulated during all stages of transcription to ensure appropriate gene expression. In metazoans, Pol II frequently pauses in the region proximal to the promoter through its association with two protein complexes, DRB-sensitivity inducing factor (DSIF) and negative elongation factor (NELF). Release from pausing into active elongation is mediated by the kinase, P-TEFb and transcription elongation factors including Polymerase Associated Factor 1 Complex and SPT6. To understand how pausing and pause release are precisely regulated during early transcription elongation, we have developed a fully reconstituted, massively parallel transcription assay. This assay allows us to define how sequence and factors influence Pol II pausing and elongation. We have also obtained cryo-EM structures of actively transcribing early elongation complexes, which clarify how pausing is induced. Together, our results provide the foundation for understanding the molecular basis of promoter proximal gene regulation.