Abstract
The explosion in genome-wide sequencing has revealed that noncoding RNAs are ubiquitous and highly conserved in biology. New molecular tools are needed for their study in live cells. Fluorescent RNA-small molecule complexes have emerged as powerful counterparts to fluorescent proteins, which are well established, universal tools in the study of proteins in cell biology. No naturally fluorescent RNAs are known; all current fluorescent RNA tags are in vitro evolved or engineered molecules that bind a conditionally fluorescent small molecule and turn on its fluorescence by up to 5000-fold. Structural analyses of several such fluorescence turn-on aptamers show that these compact (30-100 nucleotides) RNAs have diverse molecular architectures that can restrain their photoexcited fluorophores in their maximally fluorescent states, typically by stacking between planar nucleotide arrangements, such as G-quadruplexes, base triples, or base pairs. The diversity of fluorogenic RNAs as well as fluorophores that are cell permeable and bind weakly to endogenous cellular macromolecules has already produced RNA-fluorophore complexes that span the visual spectrum and are useful for tagging and visualizing RNAs in cells. Because the ligand binding sites of fluorogenic RNAs are not constrained by the need to autocatalytically generate fluorophores as are fluorescent proteins, they may offer more flexibility in molecular engineering to generate photophysical properties that are tailored to experimental needs.
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