Determining the defining lengths between mature microRNAs/small interfering RNAs and tinyRNAs.
Determining the defining lengths between mature microRNAs/small interfering RNAs and tinyRNAs.
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DOI：
10.1038/s41598-023-46562-6
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发表时间：
2023-11-13
影响因子：
4.6
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微小RNAs(MiRNAs)和小干扰RNAs(SiRNAs)被装载到ArgAerte(AGO)蛋白中，形成RNA诱导的沉默复合体(RISCs)。组装过程在加载的指南上建立种子区域、中央区域、3‘补充区域和尾部区域，使RISC能够识别用于沉默的目标RNA。这种引导片段是通过将3‘端锚定在AGO PAZ结构域上而引起的，但构象所需的最小引导长度仍有待研究，因为由迪格尔处理定义的当前miRNA大小是模糊的。使用3‘ → 5’核酸外切酶ISG20，我们确定了AGO相关的miR-20a和let-7a的长度，其中3‘端不再到达PAZ结构域。出乎意料的是，miR-20a和let-7a需要不同的长度，分别为19和20个核苷酸来维持它们的RISC构象。这种差异可以用PAZ结构域对let-7a的G19的腺苷的低亲和力来解释，这表明尾区序列略有改变最小引导长度。我们还提出了17-nT导向器足够短，可以作为tinyRNAs(TyRNAs)发挥作用，其3‘端不锚定在PAZ结构域。由于tyRNA不具有标准化指南分割的先决条件，它们识别靶标的方式与miRNAs和siRNAs不同。
MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) are loaded into Argonaute (AGO) proteins, forming RNA-induced silencing complexes (RISCs). The assembly process establishes the seed, central, 3′ supplementary, and tail regions across the loaded guide, enabling the RISC to recognize target RNAs for silencing. This guide segmentation is caused by anchoring the 3′ end at the AGO PAZ domain, but the minimum guide length required for the conformation remains to be studied because the current miRNA size defined by Dicer processing is ambiguous. Using a 3′ → 5′ exonuclease ISG20, we determined the lengths of AGO-associated miR-20a and let-7a with 3′ ends that no longer reach the PAZ domain. Unexpectedly, miR-20a and let-7a needed different lengths, 19 and 20 nt, respectively, to maintain their RISC conformation. This difference can be explained by the low affinity of the PAZ domain for the adenosine at g19 of let-7a, suggesting that the tail-region sequence slightly alters the minimum guide length. We also present that 17-nt guides are sufficiently short enough to function as tinyRNAs (tyRNAs) whose 3′ ends are not anchored at the PAZ domain. Since tyRNAs do not have the prerequisite anchoring for the standardized guide segmentation, they would recognize targets differently from miRNAs and siRNAs.