Interdisciplinary Research Center on Biology and Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 100 Hai Ke Rd., Pudong, Shanghai 201210, China.
University of Chinese Academy of Sciences, Beijing 100049, China.
Nucleic Acids Res. 2023 Jan 25;51(2):e12. doi: 10.1093/nar/gkac1136.
The hub metabolite, nicotinamide adenine dinucleotide (NAD), can be used as an initiating nucleotide in RNA synthesis to result in NAD-capped RNAs (NAD-RNA). Since NAD has been heightened as one of the most essential modulators in aging and various age-related diseases, its attachment to RNA might indicate a yet-to-be discovered mechanism that impacts adult life-course. However, the unknown identity of NAD-linked RNAs in adult and aging tissues has hindered functional studies. Here, we introduce ONE-seq method to identify the RNA transcripts that contain NAD cap. ONE-seq has been optimized to use only one-step chemo-enzymatic biotinylation, followed by streptavidin capture and the nudix phosphohydrolase NudC-catalyzed elution, to specifically recover NAD-capped RNAs for epitranscriptome and gene-specific analyses. Using ONE-seq, we discover more than a thousand of previously unknown NAD-RNAs in the mouse liver and reveal epitranscriptome-wide dynamics of NAD-RNAs with age. ONE-seq empowers the identification of NAD-capped RNAs that are responsive to distinct physiological states, facilitating functional investigation into this modification.
中心代谢物烟酰胺腺嘌呤二核苷酸（NAD）可用作 RNA 合成的起始核苷酸，从而产生 NAD 帽 RNA（NAD-RNA）。由于 NAD 已被提升为衰老和各种与年龄相关疾病中最重要的调节剂之一，其与 RNA 的结合可能表明存在尚未发现的影响成年生命过程的机制。然而，成年和衰老组织中与 NAD 相关的未知 RNA 的身份阻碍了功能研究。在这里，我们介绍了 ONE-seq 方法来鉴定含有 NAD 帽的 RNA 转录本。ONE-seq 经过优化，只需一步化学-酶促生物素化，然后进行链霉亲和素捕获和 NudC 核苷酸磷酸水解酶催化的洗脱，即可特异性回收 NAD 帽 RNA，用于外转录组和基因特异性分析。使用 ONE-seq，我们在小鼠肝脏中发现了一千多种以前未知的 NAD-RNA，并揭示了 NAD-RNA 在外转录组范围内随年龄的动态变化。ONE-seq 能够识别对不同生理状态有反应的 NAD 帽 RNA，促进对这种修饰的功能研究。
