Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15219, USA.
Université de Lorraine, CNRS, INSERM, UAR2008/US40 IBSLor, F-54000 Nancy, France.
RNA. 2022 Nov;28(11):1542-1552. doi: 10.1261/rna.079219.122. Epub 2022 Sep 13.
Epstein-Barr virus (EBV) expresses two highly abundant noncoding RNAs called EBV-encoded RNA 1 (EBER1) and EBER2, which are preserved in all clinical isolates of EBV, thus underscoring their essential function in the viral life cycle. Recent epitranscriptomics studies have uncovered a vast array of distinct RNA modifications within cellular as well as viral noncoding RNAs that are instrumental in executing their function. Here we show that EBER2 is marked by pseudouridylation, and by using HydraPsiSeq the modification site was mapped to a single nucleotide within the 3' region of EBER2. The writer enzyme was identified to be the snoRNA-dependent pseudouridine synthase Dyskerin, which is the catalytic subunit of H/ACA small nucleolar ribonucleoprotein complexes, and is guided to EBER2 by SNORA22. Similar to other noncoding RNAs for which pseudouridylation has a positive effect on RNA stability, loss of EBER2 pseudouridylation results in a decrease in RNA levels. Furthermore, pseudouridylation of EBER2 is required for the prolific accumulation of progeny viral genomes, suggesting that this single modification in EBER2 is important for efficient viral lytic replication. Taken together, our findings add to the list of RNA modifications that are essential for noncoding RNAs to implement their physiological roles.
Epstein-Barr 病毒（EBV）表达两种高度丰富的非编码 RNA，称为 EBV 编码 RNA1（EBER1）和 EBER2，它们存在于 EBV 的所有临床分离株中，因此强调了它们在病毒生命周期中的重要功能。最近的转录组学研究揭示了细胞内以及病毒非编码 RNA 中存在大量不同的 RNA 修饰，这些修饰对于执行其功能至关重要。在这里，我们表明 EBER2 被假尿嘧啶化标记，并且通过使用 HydraPsiSeq 将修饰位点映射到 EBER2 的 3' 区域内的单个核苷酸上。该写入酶被鉴定为 snoRNA 依赖性假尿嘧啶合酶 Dyskerin，它是 H/ACA 小核仁核糖核蛋白复合物的催化亚基，并由 SNORA22 引导至 EBER2。与其他假尿嘧啶化对 RNA 稳定性有积极影响的非编码 RNA 类似，EBER2 假尿嘧啶化的缺失导致 RNA 水平降低。此外，EBER2 的假尿嘧啶化对于产生大量病毒基因组的积累是必需的，这表明 EBER2 中的这种单一修饰对于有效的病毒裂解复制很重要。总之，我们的发现增加了 RNA 修饰对于非编码 RNA 执行其生理功能的重要性的列表。
