Avidity-based Bright and Photostable Light-up Aptamers for Single-molecule MRNA Imaging

Overview

Journal Nat Chem Biol

Specialties Biochemistry
Biology
Chemistry

Date 2023 Jan 19

PMID 36658339

Authors

Bastian Buhler

Janin Schokolowski

Anja Benderoth

Daniel Englert

Franziska Grun

Andres Jaschke

Murat Sunbul

Affiliations

Soon will be listed here.

Abstract

Fluorescent light-up aptamers (FLAPs) have emerged as valuable tools to visualize RNAs, but are mostly limited by their poor brightness, low photostability, and high fluorescence background in live cells. Exploiting the avidity concept, here we present two of the brightest FLAPs with the strongest aptamer-dye interaction, high fluorogenicity, and remarkable photostability. They consist of dimeric fluorophore-binding aptamers (biRhoBAST and biSiRA) embedded in an RNA scaffold and their bivalent fluorophore ligands (bivalent tetramethylrhodamine TMR and silicon rhodamine SiR). Red fluorescent biRhoBAST-TMR and near-infrared fluorescent biSiRA-SiR are orthogonal to each other, facilitating simultaneous visualization of two different RNA species in live cells. One copy of biRhoBAST allows for simple and robust mRNA imaging with strikingly higher signal-to-background ratios than other FLAPs. Moreover, eight biRhoBAST repeats enable single-molecule mRNA imaging and tracking with minimal perturbation of their localization, translation, and degradation, demonstrating the potential of avidity-enhanced FLAPs for imaging RNA dynamics.

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