Co-transcriptional folding of the glmS ribozyme enables a rapid response to metabolite

• Published in: Nucleic acids research 2024-01, Vol.52 (2), p.872-884
• Main Authors: Lou, Yuan, Woodson, Sarah A
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - metabolism; Base Sequence; Catalytic Domain; Nucleic Acid Conformation; Riboswitch; RNA and RNA-protein complexes; RNA, Catalytic - chemistry
• ISSN: 0305-1048; 1362-4962; 1362-4962
• DOI: 10.1093/nar/gkad1120

Abstract The glmS ribozyme riboswitch, located in the 5′ untranslated region of the Bacillus subtilis glmS messenger RNA (mRNA), regulates cell wall biosynthesis through ligand-induced self-cleavage and decay of the glmS mRNA. Although self-cleavage of the refolded glmS ribozyme has been studied extensively, it is not known how early the ribozyme folds and self-cleaves during transcription. Here, we combine single-molecule fluorescence with kinetic modeling to show that self-cleavage can occur during transcription before the ribozyme is fully synthesized. Moreover, co-transcriptional folding of the RNA at a physiological elongation rate allows the ribozyme catalytic core to react without the downstream peripheral stability domain. Dimethyl sulfate footprinting further revealed how slow sequential folding favors formation of the native core structure through fraying of misfolded helices and nucleation of a native pseudoknot. Ribozyme self-cleavage at an early stage of transcription may benefit glmS regulation in B. subtilis, as it exposes the mRNA to exoribonuclease before translation of the open reading frame can begin. Our results emphasize the importance of co-transcriptional folding of RNA tertiary structure for cis-regulation of mRNA stability. Graphical Abstract Graphical Abstract