TRMT2B is responsible for both tRNA and rRNA m⁵U-methylation in human mitochondria

RNA species play host to a plethora of post-transcriptional modifications which together make up the epitranscriptome. 5-methyluridine (m⁵U) is one of the most common modifications made to cellular RNA, where it is found almost ubiquitously in bacterial and eukaryotic cytosolic tRNAs at position 54. Here, we demonstrate that m⁵U54 in human mitochondrial tRNAs is catalysed by the nuclear-encoded enzyme TRMT2B, and that its repertoire of substrates is expanded to ribosomal RNAs, catalysing m⁵U429 in 12S rRNA. We show that TRMT2B is not essential for viability in human cells and that knocking-out the gene shows no obvious phenotype with regards to RNA stability, mitochondrial translation, or cellular growth.