Site-specific pseudouridylation of human ribosomal and spliceosomal RNAs is directed by H/ACA guide RNAs composed of two hairpins carrying internal pseudouridylation guide loops. The distal “antisense” sequences of the pseudouridylation loop base-pair with the target RNA to position two unpaired target nucleotides 5′-UN-3′, including the 5′ substrate U, under the base of the distal stem topping the guide loop. Therefore, each pseudouridylation loop is expected to direct synthesis of a single pseudouridine (Ψ) in the target sequence. However, in this study, genetic depletion and restoration and RNA mutational analyses demonstrate that at least four human H/ACA RNAs (SNORA53, SNORA57, SCARNA8, and SCARNA1) carry pseudouridylation loops supporting efficient and specific synthesis of two consecutive pseudouridines (ΨΨ or ΨNΨ) in the 28S (Ψ3747/Ψ3749), 18S (Ψ1045/Ψ1046), and U2 (Ψ43/Ψ44 and Ψ89/Ψ91) RNAs, respectively. In order to position two substrate Us for pseudouridylation, the dual guide loops form alternative base-pairing interactions with their target RNAs. This remarkable structural flexibility of dual pseudouridylation loops provides an unexpected versatility for RNA-directed pseudouridylation without compromising its efficiency and accuracy. Besides supporting synthesis of at least 6% of human ribosomal and spliceosomal Ψs, evidence indicates that dual pseudouridylation loops also participate in pseudouridylation of yeast and archaeal rRNAs.

中文翻译：

---

引导 RNA 杂技：通过具有双引导能力的 H/ACA 假尿苷化环定位连续尿苷以进行假尿苷化

人类核糖体和剪接体 RNA 的位点特异性假尿苷化由 H/ACA 引导 RNA 指导，该 RNA 由两个携带内部假尿苷酸化引导环的发夹组成。假尿苷酸化环的远端“反义”序列与靶 RNA 碱基对，将两个未配对的靶核苷酸 5'-UN-3'，包括 5' 底物 U，定位在远端茎的底部下方，位于导向环顶部。因此，每个假尿苷化环都有望指导靶序列中单个假尿苷 (Ψ) 的合成。然而，在这项研究中，遗传耗竭和恢复以及 RNA 突变分析表明，至少四种人类 H/ACA RNA（SNORA53、SNORA57、SCARNA8 和 SCARNA1）携带假尿苷化环，支持有效和特异性合成两个连续假尿苷（ΨΨ 或 ΨNΨ ) 在 28S (Ψ3747/Ψ3749) 中，分别为 18S (Ψ1045/Ψ1046) 和 U2 (Ψ43/Ψ44 和 Ψ89/Ψ91) RNA。为了定位两个底物 Us 进行假尿苷化，双导向环与它们的目标 RNA 形成替代碱基配对相互作用。双假尿酰化环的这种显着的结构灵活性为 RNA 指导的假尿酰化提供了意想不到的多功能性，而不会影响其效率和准确性。除了支持至少 6% 的人类核糖体和剪接体 Ψs 的合成外，有证据表明双假尿酰化环也参与酵母和古细菌 rRNA 的假尿酰化。双假尿苷化环的这种显着的结构灵活性为 RNA 指导的假尿苷化提供了意想不到的多功能性，而不会影响其效率和准确性。除了支持至少 6% 的人类核糖体和剪接体 Ψs 的合成外，有证据表明双假尿酰化环也参与酵母和古细菌 rRNA 的假尿酰化。双假尿苷化环的这种显着的结构灵活性为 RNA 指导的假尿苷化提供了意想不到的多功能性，而不会影响其效率和准确性。除了支持至少 6% 的人类核糖体和剪接体 Ψs 的合成外，有证据表明双假尿酰化环也参与酵母和古细菌 rRNA 的假尿酰化。