RNA folds cotranscriptionally to traverse out-of-equilibrium intermediate structures that are important for RNA function in the context of gene regulation. To investigate this process, here we study the structure and function of the Bacillus subtilis yxjA purine riboswitch, a transcriptional riboswitch that downregulates a nucleoside transporter in response to binding guanine. Although the aptamer and expression platform domain sequences of the yxjA riboswitch do not completely overlap, we hypothesized that a strand exchange process triggers its structural switching in response to ligand binding. In vivo fluorescence assays, structural chemical probing data and experimentally informed secondary structure modeling suggest the presence of a nascent intermediate central helix. The formation of this central helix in the absence of ligand appears to compete with both the aptamer's P1 helix and the expression platform's transcriptional terminator. All-atom molecular dynamics simulations support the hypothesis that ligand binding stabilizes the aptamer P1 helix against central helix strand invasion, thus allowing the terminator to form. These results present a potential model mechanism to explain how ligand binding can induce downstream conformational changes by influencing local strand displacement processes of intermediate folds that could be at play in multiple riboswitch classes.
Riboswitches have challenged our understanding of biological regulation for almost two decades. The ability of small molecules to bind to RNA and control gene expression offers another layer of regulation and the potential for direct action by compounds in the environment. While some riboswitches have been well studied, we lack a general understanding of how changes in RNA structure switch genetic expression from “On” to “Off”. In this study, the authors propose an elegant “strand displacement” model to explain how the RNA structure shifts between “On” and “Off” states as the concentration of small molecule ligand changes. These observations help us to understand how riboswitches enable genetic decision-making. The data provide a possible general mechanism for understanding how the competition between different strand displacement outcomes can influence RNA folding. Understanding RNA folding pathways could advance the successful design of drugs that target RNA.
© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.