Natural Products Research Center, Chengdu Institute of Biology, Chinese Academy of Science, Chengdu 610041, P. R. China.
University of Chinese Academy of Sciences, Beijing 100049, P. R. China.
Anal Chem. 2023 Sep 19;95(37):13762-13768. doi: 10.1021/acs.analchem.2c05724. Epub 2023 Sep 3.
Imaging and tracking tools for natural cellular RNA with improved biocompatibility, specificity, and sensitivity are critical to understanding RNA function and providing insights into disease therapeutics. We developed a new genetically encoded sensor using fluorogenic allosteric aptamer (FaApt) for the sensitive imaging of the localization and dynamics of RNA targets in live cells. Target RNAs can be specifically recognized with our sensor by forming perfectly complementary duplexes, which in turn can induce allosteric structural changes of the sensor to refold the native conformation of fluorogenic RNA aptamers. We demonstrated the ability of the sensor to monitor the effect of tumor necrosis factor and small-molecule inhibitor on the expression abundance of CXCL1 and survivin mRNA in human cancer cells, respectively. The asymmetrical distribution of endogenous Squint mRNA was confirmed in developing zebrafish embryos through microinjection of FaApt probes. This study provides an effective molecular tool for sensitive imaging and tracking endogenous RNA in living cells. Due to the high specificity and small size of our sensor system, it is expected to be applied to early diagnosis of RNA marker-related diseases and real-time evaluation of the treatment process.
具有改善的生物相容性、特异性和灵敏度的天然细胞 RNA 的成像和跟踪工具对于理解 RNA 功能并深入了解疾病治疗至关重要。我们开发了一种新的遗传编码传感器,该传感器使用荧光变构适体 (FaApt) 来灵敏地成像活细胞中 RNA 靶标在的定位和动态。我们的传感器可以通过形成完全互补的双链体特异性识别靶 RNA,这反过来又可以诱导传感器的变构结构变化,从而重新折叠荧光 RNA 适体的天然构象。我们证明了该传感器能够分别监测肿瘤坏死因子和小分子抑制剂对人癌细胞中 CXCL1 和 survivin mRNA 表达丰度的影响。通过微注射 FaApt 探针,在发育中的斑马鱼胚胎中证实了内源性 Squint mRNA 的不对称分布。这项研究为在活细胞中灵敏地成像和跟踪内源性 RNA 提供了有效的分子工具。由于我们的传感器系统具有高特异性和小尺寸,预计它将应用于 RNA 标记物相关疾病的早期诊断和治疗过程的实时评估。
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