Central European Institute of Technology, Masaryk University, 625 00 Brno, Czech Republic.
Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 370 05 České Budějovice, Czech Republic.
RNA. 2022 Dec;28(12):1568-1581. doi: 10.1261/rna.079421.122. Epub 2022 Oct 3.
Transfer RNAs acquire a large plethora of chemical modifications. Among those, modifications of the anticodon loop play important roles in translational fidelity and tRNA stability. Four human wobble U-containing tRNAs obtain 5-methoxycarbonylmethyluridine (mcmU) or 5-methoxycarbonylmethyl-2-thiouridine (mcmsU), which play a role in decoding. This mark involves a cascade of enzymatic activities. The last step is mediated by alkylation repair homolog 8 (ALKBH8). In this study, we performed a transcriptome-wide analysis of the repertoire of ALKBH8 RNA targets. Using a combination of HITS-CLIP and RIP-seq analyses, we uncover ALKBH8-bound RNAs. We show that ALKBH8 targets fully processed and CCA modified tRNAs. Our analyses uncovered the previously known set of wobble U-containing tRNAs. In addition, both our approaches revealed ALKBH8 binding to several other types of noncoding RNAs, in particular C/D box snoRNAs.
转移 RNA 获得了大量的化学修饰。其中，反密码环的修饰在翻译保真度和 tRNA 稳定性方面起着重要作用。四种含有 wobble U 的人类 tRNA 获得 5-甲氧基羰基甲基尿嘧啶 (mcmU) 或 5-甲氧基羰基甲基-2-硫代尿嘧啶 (mcmsU)，它们在解码中发挥作用。这个标记涉及一系列酶促活性。最后一步由烷基化修复同源物 8 (ALKBH8) 介导。在这项研究中，我们对 ALKBH8 RNA 靶标的转录组范围进行了分析。我们使用 HITS-CLIP 和 RIP-seq 分析的组合，揭示了 ALKBH8 结合的 RNA。我们表明，ALKBH8 靶向完全加工和 CCA 修饰的 tRNA。我们的分析揭示了以前已知的 wobble U 含有 tRNA 组。此外，我们的两种方法都揭示了 ALKBH8 与其他几种非编码 RNA（特别是 C/D 盒 snoRNA）的结合。
