Modular and genetically encodable RNA-based bioluminescence resonance energy transfer (BRET) sensors for cellular target detection.

Abstract

Genetically encodable bioluminescent sensors have been broadly used for highly sensitive, low-background, low-phototoxicity bioanalysis and imaging in living biological systems. To achieve real-time quantitative measurement and high-throughput screening, powerful bioluminescence resonance energy transfer (BRET) sensors have been designed, especially based on interactions between a luciferase/luciferin donor and a fluorescent protein acceptor. While fluorescent protein-based BRET sensors have been widely applied in cellular and in vivo studies, more modular and easy-to-engineer RNA-based BRET sensors have never been reported. In this project, we report, for the first time, genetically encoded fluorogenic RNA-based BRET sensors for cellular imaging and quantification of different target analytes and RNA-protein interactions. Fluorogenic RNAs are single-stranded RNA aptamers that can bind specific small-molecule fluorophores and activate the fluorescence signals. By coupling a NanoLuc luciferase donor with different types of fluorogenic RNA acceptor, without reliance on external light, sensitive BRET signals are generated in response to target metabolite, signaling molecule, or RNA levels inside living cells. We envision that this novel genetically encoded RNA-based BRET system can be widely used in bioanalysis and nanomedicine.