Medicinal Chemistry and Bioinformatics Center, Shanghai Jiao Tong University, School of Medicine,Shanghai 200025, China.
Department of Anesthesiology, Changhai Hospital, Naval Medical University, Shanghai 200433, China.
Biomolecules. 2022 Nov 3;12(11):1631. doi: 10.3390/biom12111631.
While host miRNA usually plays an antiviral role, the relentless tides of viral evolution have carved out a mechanism to recruit host miRNA as a viral protector. By complementing miR-122 at the 5' end of the genome, the hepatitis C virus (HCV) gene can form a complex with Argonaute 2 (Ago2) protein to protect the 5' end of HCV RNA from exonucleolytic attacks. Experiments showed that the disruption of the stem-loop 1(SL1) structure and the 9th nucleotide (T9) of HCV site 1 RNA could enhance the affinity of the Ago2 protein to the HCV site 1 RNA (target RNA). However, the underlying mechanism of how the conformation and dynamics of the Ago2: miRNA: target RNA complex is affected by the SL1 and T9 remains unclear. To address this, we performed large-scale molecular dynamics simulations on the AGO2-miRNA complex binding with the WT target, T9-abasic target and SL1-disruption target, respectively. The results revealed that the T9 and SL1 structures could induce the departing motion of the PAZ, PIWI and N domains, propping up the mouth of the central groove which accommodates the target RNA, causing the instability of the target RNA and disrupting the Ago2 binding. The coordinated motion among the PAZ, PIWI and N domains were also weakened by the T9 and SL1 structures. Moreover, we proposed a new model wherein the Ago2 protein could adopt a more constraint conformation with the proximity and more correlated motions of the PAZ, N and PIWI domains to protect the target RNA from dissociation. These findings reveal the mechanism of the Ago2-miRNA complex's protective effect on the HCV genome at the atomic level, which will offer guidance for the design of drugs to confront the protection effect and engineering of Ago2 as a gene-regulation tool.
虽然宿主 miRNA 通常发挥抗病毒作用，但病毒不断进化的浪潮已经开辟了一种机制，将宿主 miRNA 招募为病毒保护剂。丙型肝炎病毒 (HCV) 基因通过在基因组的 5'端互补 miR-122，可以与 Argonaute 2 (Ago2) 蛋白形成复合物，保护 HCV RNA 的 5'端免受外切核酸酶的攻击。实验表明，破坏 HCV 位点 1 RNA 的茎环 1 (SL1) 结构和第 9 个核苷酸 (T9) 可以增强 Ago2 蛋白与 HCV 位点 1 RNA (靶 RNA) 的亲和力。然而，Ago2:miRNA:靶 RNA 复合物的构象和动力学如何受到 SL1 和 T9 影响的机制尚不清楚。为了解决这个问题，我们分别对 AGO2-miRNA 复合物与 WT 靶标、T9-脱碱基靶标和 SL1 破坏靶标进行了大规模的分子动力学模拟。结果表明，T9 和 SL1 结构可以诱导 PAZ、PIWI 和 N 结构域的离去运动，支撑中央槽的口，容纳靶 RNA，导致靶 RNA 的不稳定并破坏 Ago2 结合。PAZ、PIWI 和 N 结构域之间的协调运动也被 T9 和 SL1 结构削弱。此外，我们提出了一个新模型，其中 Ago2 蛋白可以采用更受限的构象，PAZ、N 和 PIWI 结构域更接近且运动更相关，以保护靶 RNA 免受解离。这些发现揭示了 Ago2-miRNA 复合物在原子水平上保护 HCV 基因组的机制，为设计对抗保护作用的药物和工程化 Ago2 作为基因调控工具提供了指导。
