RNA-Driven Control of Transcriptional Pausing and Termination

Abstract

The diverse mechanisms by which bacterial riboswitches regulate gene expression through communication with the RNA polymerase (RNAP) are incompletely understood. A key aspect of transcriptional regulation is transcriptional pausing, believed to be initiated and stabilized by a consensus DNA pause sequence. Remarkably, we find that the fluoride-sensing riboswitch stabilizes a pause and favors termination in the absence of its ligand despite lacking this consensus sequence, suggesting a purely RNA-driven mechanism. We combine biochemical assays, cryo-electron microscopy, and molecular dynamics simulations to investigate the coupling between a prototypical fluoride-sensing riboswitch and termination of bacterial transcription. We present structural evidence that ligand binding initiates an event cascade, leading to global conformational changes of RNAP that promote recommitment to transcription. Strikingly, upon ligand binding the riboswitch rotates within the RNA exit channel, preventing zipping of the nascent terminator hairpin. Our work captures a snapshot of a riboswitch toggling between transcriptional readthrough and termination.

Keywords: riboswitches, transcription, RNAP, cryo-EM, RNA-dependent pausing