HITS-CLIP analysis of human ALKBH8 reveals interactions with fully processed substrate tRNAs and with specific noncoding RNAs
- Ivana Cavallin1,
- Marek Bartosovic1,5,
- Tomas Skalicky1,2,
- Praveenkumar Rengaraj1,
- Martin Demko1,
- Martina Christina Schmidt-Dengler3,
- Aleksej Drino4,
- Mark Helm3 and
- Stepanka Vanacova1
- 1Central European Institute of Technology, Masaryk University, 625 00 Brno, Czech Republic
- 2Institute of Parasitology, Biology Centre, Czech Academy of Sciences, 370 05 České Budějovice, Czech Republic
- 3Johannes Gutenberg-Universität Mainz, Institute of Pharmaceutical and Biomedical Science (IPBS), D-55128 Mainz, Germany
- 4Medical University of Vienna, Center for Anatomy and Cell Biology, 1090 Vienna, Austria
- Corresponding author: stepanka.vanacova{at}ceitec.muni.cz
Abstract
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 (mcm5U34) or 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U34), 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.
Keywords
- Received August 16, 2022.
- Accepted September 9, 2022.
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