Duplex Structure of Double-Stranded RNA Provides Stability against Hydrolysis Relative to Single-Stranded RNA
Duplex Structure of Double-Stranded RNA Provides Stability against Hydrolysis Relative to Single-Stranded RNA
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DOI：
10.1021/acs.est.1c01255
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发表时间：
2021-05-25
影响因子：
11.4
通讯作者：
Parker，Kimberly M.
中科院分区：
文献类型：
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作者：
Zhang，Ke;Hodge，Joseph;Parker，Kimberly M.
已知双链（ds）RNA和单链（ss）RNA的主链中的磷酸二酯键经历碱性水解。因此，用于新兴RNA干扰（RNAi）产物的dsRNA试剂已被假定在溶液中表现出低化学持久性。然而，尚未评估双链RNA的双链结构对碱性水解的影响。在这项研究中，我们证明了dsRNA经历的数量级慢于ssRNA的碱性水解。此外，我们观察到dsRNA在中性pH下保持完整数月，挑战了dsRNA化学不稳定的假设。在能够使dsRNA酶促降解和碱性水解的系统中，我们发现增加pH有效地减弱了酶促降解，而不诱导对ssRNA观察到的碱性水解。总的来说，我们的研究结果首次证明了dsRNA的关键降解途径与ssRNA的降解途径显著不同。考虑到dsRNA的独特性质，将能够在新兴RNAi技术的应用过程中更好地控制dsRNA的稳定性，并更准确地评估其在环境和生物系统中的命运，以及为更广泛的应用领域提供见解，包括dsRNA分离，dsRNA病毒的检测和灭活，以及益生元分子进化。
Phosphodiester bonds in the backbones of double-stranded (ds)RNA and single-stranded (ss)RNA are known to undergo alkaline hydrolysis. Consequently, dsRNA agents used in emerging RNA interference (RNAi) products have been assumed to exhibit low chemical persistence in solutions. However, the impact of the duplex structure of dsRNA on alkaline hydrolysis has not yet been evaluated. In this study, we demonstrated that dsRNA undergoes orders-of-magnitude slower alkaline hydrolysis than ssRNA. Furthermore, we observed that dsRNA remains intact for multiple months at neutral pH, challenging the assumption that dsRNA is chemically unstable. In systems enabling both enzymatic degradation and alkaline hydrolysis of dsRNA, we found that increasing pH effectively attenuated enzymatic degradation without inducing alkaline hydrolysis that was observed for ssRNA. Overall, our findings demonstrated, for the first time, that key degradation pathways of dsRNA significantly differ from those of ssRNA. Consideration of the unique properties of dsRNA will enable greater control of dsRNA stability during the application of emerging RNAi technology and more accurate assessment of its fate in environmental and biological systems, as well as provide insights into broader application areas including dsRNA isolation, detection and inactivation of dsRNA viruses, and prebiotic molecular evolution.