RNA editing leveraging ADARs (adenosine deaminases acting on RNA) shows promising potential for in vivo biosensing beyond gene therapy. However, current ADAR sensors sense only a single target of RNA transcripts, thus limiting their use in different biosensing scenarios. Here, we report a hairpin RNA sensor that exploits new mechanisms to generate intramolecular duplex substrates for efficient ADAR recruitment and editing and apply it to detection of various intracellular molecules, including messenger RNA, small molecules and proteins. We utilize the base pairing interactions between neighbouring bases for enhanced stability, as well as the reverse effects to sense RNA transcripts and single-nucleotide variants with high sensitivity and specificity, irrespective of sequence requirement for complementarity to an UAG stop codon. In addition, we integrate RNA aptamers into the hairpin RNA sensor to realize the detection of the primary energy-supplying molecule, ATP, and a transcription factor, nuclear factor-kappa B (NF-κB), in live cells via a simple conformational change for programming the activation of hairpin RNA. This sensor not only broadens the detection of applicable molecules, but also offers potential for diverse cell manipulation.

中文翻译：

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编程 ADAR 募集发夹 RNA 传感器以检测内源性分子


利用 ADARs（作用于 RNA 的腺苷脱氨酶）进行 RNA 编辑显示出超越基因治疗之外的体内生物传感的潜力。然而，目前的 ADAR 传感器只能感应 RNA 转录本的单个靶标，从而限制了它们在不同生物传感场景中的使用。在这里，我们报道了一种发夹 RNA 传感器，它利用新机制产生分子内双链底物以实现高效的 ADAR 募集和编辑，并将其应用于检测各种细胞内分子，包括信使 RNA、小分子和蛋白质。我们利用相邻碱基之间的碱基配对相互作用来增强稳定性，并利用反向效应以高灵敏度和特异性感知 RNA 转录本和单核苷酸变体，而不管与 UAG 终止密码子互补的序列要求如何。此外，我们将 RNA 适配体集成到发夹 RNA 传感器中，通过简单的构象变化实现对发夹 RNA 激活进行编程，从而实现活细胞中主要能量供应分子 ATP 和转录因子核因子-κB （NF-κB） 的检测。该传感器不仅拓宽了适用分子的检测范围，还为多种细胞操作提供了潜力。