RNA is instrumental to cell life in many aspects, especially gene expression regulation. Among the various known regulatory RNAs, riboswitches are particularly interesting cis‐acting molecules as they do not need cellular factor to achieve their function and are therefore highly portable from one organism to the other. These molecules usually found in the 5′ untranslated region of bacterial messenger RNAs are able to specifically sense a target ligand via an aptamer domain prior to transmitting this recognition event to an expression platform that turns on, or off, the expression of downstream genes. In addition to their obvious scientific interest, these modular molecules can also serve for the development of synthetic RNA devices with applications ranging from the control of transgene expression in gene therapy to the specific biosensing of small molecules. The engineering of such nanomachines is greatly facilitated by the proper understanding of their structure as well as the introduction of new technologies. Herein, a general overview of the current knowledge on natural riboswitches prior to explaining the main strategies used to develop new synthetic structure‐switching molecules (riboswitches or biosensors) controlled by small molecules is given.
an impressive level by riboswitches, a class of RNA motifs mostly found in the 5′ untranslated regions of some bacterial messenger RNAs (mRNAs). These sequences are able to specifically switch their structure in the presence of a target ligand, usually a small molecule, in order to control downstream gene expression (transcription or translation). [1][2][3][4] Riboswitches usually comprise two domains: a sensor aptamer that specifically recognizes a target molecule (with an affinity sometimes exceeding that of antibodies for their antigen), and a regulatory platform that remodels a region of the mRNA to modulate its transcription and/or its translation. The two domains are usually functionally connected by a transducing element, also called communication module. [1] Their modular structure makes riboswitches an excellent starting point for molecular engineering. For instance, exchanging the sensor aptamer domain for another one, natural or synthetic, already enabled the development of engineered riboswitches for application in living cells as well as in cell-free systems. [5][6][7] Thus, using such a construct to control the expression of a reporter gene (e.g., coding for a fluorescent protein, luciferase or another reporting enzyme), would yield a biosensor, that is, a biological molecule (or system) that converts the presence of a specific molecule into a measurable signal. [8] Among the wide palette of potential target ligands for this technology, small negatively charged molecules are expected to be the most difficult to target using RNA-based sensors as they offer only a low number of possible interaction sites and because strong electrostatic repulsion with polyanionic nucleic acids is anticipated. Fluoride, the smallest and the most electronegative anion, represents an extreme case in this context. However, its toxicity and often-problematic levels in drinking water make it a highly relevant target for specific detection. [9] Fluoride is also a potential degradation product of fluorinated compounds, an emerging class of persistent and toxic chemicals. [10] Specific detection of this ion would be useful in the development of high-throughput activity assays for the discovery of defluorination enzymes, of which only a few are currently known. [11] To be applicable, a corresponding sensor should be biocompatible, highly selective, easy to use, nontoxic, commercially available or easy to produce, and generate a fluorescent signal compatible with very high-throughput screening pipelines. [12] At present, fluoride is detectable by fluoride-specific electrodes [13,14] or colorimetry, [15,16] but none of these approaches fulfills all the above-mentioned requirements. Nucleic acids are not only essential actors of cell life but also extremely appealing molecular objects in the development of synthetic molecules for biotechnological application, such as biosensors to report on the presence and concentration of a target ligand by emission of a measurable signal. In this work, FluorMango, a fluorogenic ribonucleic acid...
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