Linking aptamer‐ligand binding and expression platform folding in riboswitches: prospects for mechanistic modeling and design

Abstract

The power of riboswitches in regulation of bacterial metabolism derives from coupling of two characteristics: recognition and folding. Riboswitches contain aptamers, which function as biosensors. Upon detection of the signaling molecule, the riboswitch transduces the signal into a genetic decision. The genetic decision is coupled to refolding of the expression platform, which is distinct from, although overlapping with, the aptamer. Early biophysical studies of riboswitches focused on recognition of the ligand by the aptamer‐an important consideration for drug design. A mechanistic understanding of ligand‐induced riboswitch RNA folding can further enhance riboswitch ligand design, and inform efforts to tune and engineer riboswitches with novel properties. X‐ray structures of aptamer/ligand complexes point to mechanisms through which the ligand brings together distal strand segments to form a P1 helix. Transcriptional riboswitches must detect the ligand and form this P1 helix within the timescale of transcription. Depending on the cell's metabolic state and cellular environmental conditions, the folding and genetic outcome may therefore be affected by kinetics of ligand binding, RNA folding, and transcriptional pausing, among other factors. Although some studies of isolated riboswitch aptamers found homogeneous, prefolded conformations, experimental, and theoretical studies point to functional and structural heterogeneity for nascent transcripts. Recently it has been shown that some riboswitch segments, containing the aptamer and partial expression platforms, can form binding‐competent conformers that incorporate an incomplete aptamer secondary structure. Consideration of the free energy landscape for riboswitch RNA folding suggests models for how these conformers may act as transition states—facilitating rapid, ligand‐mediated aptamer folding. WIREs RNA 2015, 6:631–650. doi: 10.1002/wrna.1300For further resources related to this article, please visit the WIREs website.

An integrated perspective on RNA aptamer ligand-recognition models: clearing muddy waters

“…17,[21][22][23][24][25][26] Others have argued that while diffusion on the FEL in configuration space is the most realistic model of protein folding, CS and IF describe not the FEL itself, but two different ways the ligand can reshape the FEL. [27][28][29] In this interpretation, CS is associated with constraining diffusion across the landscape, and IF is associated with 'forcing and tilting', adjusting the effective height of energetic barriers to local minima 30 and enabling transitions to previously energetically inaccessible subspaces. 17 Each binding mechanism, CS or IF, can be defined in one of two ways (Fig.…”

Regulatory effects of cotranscriptional RNA structure formation and transitions

“…Riboswitches act as RNA sensors that regulate the transcript in which they reside in cis via binding a small molecule ligand . Many riboswitches have been studied, including adenine, purine, lysine, thiamine pyrophosphate (TPP), S ‐adenosylmethionine (SAM), and Flavin mononucleotide (FMN) . By binding a ligand, numerous factors can interact with riboswitches as they are regulating the process of transcription or translation in cells.…”

“…By binding a ligand, numerous factors can interact with riboswitches as they are regulating the process of transcription or translation in cells. Timing and cooperation between the binding and association among these factors, the movement of transcriptional complex, and RNA folding are essential to the regulatory outcome . One well‐illustrated study is the SAM riboswitch.…”

“…This interaction can affect the cotranscriptional RNA folding pathways and the resulting RNA structures. Examples of such interactions include proteins that specifically bind to RNA to assist RNA folding , and riboswitches that interact with RNA and affect RNA structural formation . In addition, RNA chaperone and helicase can nonspecifically bind to RNA to aid RNA folding by refolding misfolded structures, and some distinct intermolecular RNA transcripts can either stabilize or destabilize existing RNA structures, which affects the cotranscriptional folding pathways and the resulting RNA conformations …”

“…Changes in many RNA structures regulate gene expression at the transcriptional level, resulting in either transcription termination or activation. Structural changes of some riboswitches upon ligand binding can regulate RNA transcription . In the adenine riboswitch cotranscriptional folding mechanism, the aptamer domain is formed prior to the expression domain by RNA polymerase, and the nascent aptamer domain is able to response to cellular cues and potentially bind to a cognate ligand before the transcription and folding of the expression domain .…”