Structural Principles of Fluorescent RNA Aptamers

Fluorescent aptamers, RNAs selected in vitro to bind small molecules and dramatically enhance their intrinsic fluorescence, are useful as in vivo sensors of metabolites and tags for monitoring the transcriptome. High-resolution structures of fluorescent aptamers have provided understanding of the inherent limitations of each of these RNA tags and, in two cases, helped improve function. Fluorescent aptamers exhibit flat binding pockets containing base quadruples (tetrads). These provide a flat surface for the conjugated aromatic fluorophores to stack on. Stacking can maximize fluorescence enhancement by allowing the excited fluorophores to adopt a planar conformation. Independently selected fluorescent aptamers have converged on structures containing G-quadruplexes, a particularly stable type of base tetrad. Several aptamer RNAs have been selected in vitro that bind to otherwise weakly fluorescent small molecules and enhance their fluorescence several thousand-fold. By genetically tagging cellular RNAs of interest with these aptamers and soaking cells in their cell-permeable cognate small-molecule fluorophores, it is possible to use them to study RNA localization and trafficking. These aptamers have also been fused to metabolite-binding RNAs to generate fluorescent biosensors. The 3D structures of three unrelated fluorogenic RNAs have been determined, and reveal a shared reliance on base quadruples (tetrads) to constrain the photo-excited chromophore. The structural diversity of fluorogenic RNAs and the chemical diversity of potential fluorophores to be activated are likely to yield a variety of future fluorogenic RNA tags that are optimized for different applications in RNA imaging and in the design of fluorescent RNA biosensors. Several aptamer RNAs have been selected in vitro that bind to otherwise weakly fluorescent small molecules and enhance their fluorescence several thousand-fold. By genetically tagging cellular RNAs of interest with these aptamers and soaking cells in their cell-permeable cognate small-molecule fluorophores, it is possible to use them to study RNA localization and trafficking. These aptamers have also been fused to metabolite-binding RNAs to generate fluorescent biosensors. The 3D structures of three unrelated fluorogenic RNAs have been determined, and reveal a shared reliance on base quadruples (tetrads) to constrain the photo-excited chromophore. The structural diversity of fluorogenic RNAs and the chemical diversity of potential fluorophores to be activated are likely to yield a variety of future fluorogenic RNA tags that are optimized for different applications in RNA imaging and in the design of fluorescent RNA biosensors.