A Programmable mRNA Platform for miRNA Detection via miRNA–mRNA 2 Triplex-Mediated Ribosomal Frameshifting

作者：Yanyu Chen, Wanqi Zhao, Huizhen Chen, Hanting Zhou, Shijian Fan, Liping Xing, Yun Lian, Hongyu Duan, Yuze Dai, Zheng Zhao, Rongguang Lu, Chenzhong Li, Cheng Jiang, Ho Yin Edwin Chan, Gang Chen · 发表于：ACS Synthetic Biology · 年份：2026 · DOI：10.1021/acssynbio.5c00883 · 研究领域：RNA and protein synthesis mechanisms、RNA Interference and Gene Delivery、MicroRNA in disease regulation

Programmed −1 ribosomal frameshifting (−1 PRF) is a recoding mechanism utilized by viruses to expand their coding capacity and modulate the stoichiometric ratio of −1 frame and 0 frame translation products. The stability of mRNA secondary structure at the ribosomal entry site within the frameshifting stimulating elements (FSEs) determines the frameshifting efficiency. Here, we report the development of a programmable mRNA-based platform that detects specific mature microRNA (miRNA or miR) by converting their presence into a quantifiable protein output through miRNA-triggered −1 PRF. We designed a triplex-forming mRNA (TF-mRNA) platform to selectively trap target miRNAs through the formation of major-groove mRNA–miRNA–mRNA (miR–mRNA 2 ) triplexes. Biolayer interferometry and fluorescence binding studies confirmed that TF-mRNA forms stable complexes with cognate miRNAs with low nanomolar affinity and prolonged dissociation rate. Critically, the formation of miR–mRNA 2 triplex robustly stimulated ribosomal frameshifting in a cell-free dual-luciferase translation system, acting as a miRNA-dependent molecular switch. The generality of this TF-mRNA platform has been verified for several disease-associated purine-rich miRNAs, and it is suitable for targeting a wide range of other purine-enriched miRNAs. This programmable TF-mRNA platform establishes a foundation for developing novel diagnostic tools and synthetic biology circuits that convert the presence of miRNA into a quantifiabl...