Jiang Li, Zuo Fangting, Pan Yuanyuan, Li Ruilong, Shi Yajie, Huang Xinyi, Zhang Dasheng, Zhuang Yingping, Zhao Yuzheng, Lin Qiuning, Yang Yi, Zhu Linyong, Chen Xianjun
Optogenetics & Synthetic Biology Interdisciplinary Research Center, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Mei Long Road, Shanghai, 200237, China.
School of Biomedical Engineering, Shanghai Jiao Tong University, 800 Dong Chuan Road, Shanghai, 200240, China.
Small. 2025 Mar;21(9):e2405165. doi: 10.1002/smll.202405165. Epub 2024 Oct 28.
Fluorescent RNAs (FRs), which are RNA aptamers that bind and activate their cognate small fluorogenic dyes, have provided a particularly useful approach for imaging RNAs in live cells. Although the color palette of FRs is greatly expanded, a bright and stable cyan FR with good biocompatibility and biorthogonality with currently available FRs remains desirable but is not yet developed. Herein, the development of Myosotis is described, an RNA aptamer that emits bright cyan fluorescence upon binding a novel GFP chromophore-like fluorophore called DBT. Myosotis has a nanomolar affinity for DBT and shows a weak dependence on magnesium for folding. Further studies reveal that the Myosotis-DBT complex has a long fluorescence lifetime, good photostability, and enhance cellular brightness. It is further shown that Myosotis-DBT is biorthogonal to Pepper and Clivia FRs, allowing multiplex fluorescence imaging of RNA in live bacteria. Myosotis can also use to image mRNA in live bacteria, revealing potential coupling between mRNA translation and stability. It is believed that this cyan FR will be a useful tool for studying the functionality and mechanism of RNA underlying diverse biological processes.
荧光RNA（FRs）是一类能结合并激活其同源小分子荧光染料的RNA适配体，为活细胞中的RNA成像提供了一种特别有用的方法。尽管FRs的颜色种类已大大扩展，但仍需要一种明亮且稳定的青色FR，它具有良好的生物相容性且与目前可用的FRs具有生物正交性，但尚未开发出来。在此，描述了勿忘草素（Myosotis）的开发过程，它是一种RNA适配体，在结合一种名为DBT的新型类绿色荧光蛋白发色团荧光团时会发出明亮的青色荧光。勿忘草素对DBT具有纳摩尔亲和力，并且折叠对镁的依赖性较弱。进一步的研究表明，勿忘草素-DBT复合物具有较长的荧光寿命、良好的光稳定性，并能增强细胞亮度。还进一步表明，勿忘草素-DBT与辣椒素（Pepper）和君子兰素（Clivia）FRs具有生物正交性，从而能够对活细菌中的RNA进行多重荧光成像。勿忘草素还可用于对活细菌中的mRNA进行成像，揭示mRNA翻译与稳定性之间的潜在耦合关系。据信，这种青色FR将成为研究多种生物过程中RNA功能和机制的有用工具。
