Monitoring RNA dynamics in native transcriptional complexes
Adrien Chauvier, Patrick St-Pierre, Jean‐François Nadon, Elsa D. M. Hien, Cibran Pérez‐González, Sébastien H. Eschbach, Anne-Marie Lamontagne, J. Carlos Penedo, Daniel A. Lafontaine
Abstract
Significance Transcription of DNA into RNA is crucial to life, and understanding RNA polymerase (RNAP) function has received considerable attention. In contrast, how the nascent RNA folds into structures that impact transcription itself and regulate gene expression remains poorly understood. Here, we combine single-molecule Förster resonance energy transfer and site-specific fluorescent labelling of transcripts within native complexes to enable real-time cotranscriptional folding studies of a metabolite-sensing riboswitch from Escherichia coli . By monitoring the folding of riboswitches stalled at RNAP pausing sites and during active elongation, we reveal a crucial role for RNAP, which directs RNA folding to allow thiamin pyrophosphate sensing within a precise, transcriptional hotspot. Our approach offers a unique opportunity to unveil cotranscriptional processes in eukaryotic and bacterial systems.