METTL15 introduces N4-methylcytidine into human mitochondrial 12S rRNA and is required for mitoribosome biogenesis.

Lindsey Van Haute,Shujing Ding,Michal Minczuk,Pedro Rebelo-Guiomar,Aaron R D’Souza,Ian M Fearnley,Alan G Hendrick,Michael E Harbour,Byron Andrews,Christopher A Powell

doi:10.1093/nar/gkz735

Abstract

Post-transcriptional RNA modifications, the epitranscriptome, play important roles in modulating the functions of RNA species. Modifications of rRNA are key for ribosome production and function. Identification and characterization of enzymes involved in epitranscriptome shaping is instrumental for the elucidation of the functional roles of specific RNA modifications. Ten modified sites have been thus far identified in the mammalian mitochondrial rRNA. Enzymes responsible for two of these modifications have not been characterized. Here, we identify METTL15, show that it is the main N4-methylcytidine (m4C) methyltransferase in human cells and demonstrate that it is responsible for the methylation of position C839 in mitochondrial 12S rRNA. We show that the lack of METTL15 results in a reduction of the mitochondrial de novo protein synthesis and decreased steady-state levels of protein components of the oxidative phosphorylation system. Without functional METTL15, the assembly of the mitochondrial ribosome is decreased, with the late assembly components being unable to be incorporated efficiently into the small subunit. We speculate that m4C839 is involved in the stabilization of 12S rRNA folding, therefore facilitating the assembly of the mitochondrial small ribosomal subunits. Taken together our data show that METTL15 is a novel protein necessary for efficient translation in human mitochondria.

Highlights

Mammalian mitochondrial DNA encodes 13 polypeptides that are essential subunits of the oxidative phosphorylation (OxPhos) system
METTL15 has no predicted mitochondrial targeting sequence, our results clearly indicate that METTL15 is an intra-mitochondrial protein in human cells
Our results are consistent with a large-scale proteomic study based on proximity labelling that suggested mitochondrial matrix localization of METTL15 [36], as well as with a recent BioID study suggesting that METTL15 interacts with TRUB2, a mitochondrial matrix pseudouridine synthase [13]

Summary

Introduction

Mammalian mitochondrial DNA (mtDNA) encodes 13 polypeptides that are essential subunits of the oxidative phosphorylation (OxPhos) system. MtDNA contains the genes for all 22 mitochondrial (mt-) tRNAs and two mtrRNAs (small 12S and large 16S). The maintenance and expression of the mitochondrial genome is entirely dependent on proteins encoded in the nucleus, which upon translation on cytoplasmic ribosomes are imported into mitochondria. Expression of mitochondrial genes shows very distinct differences as compared to their nuclear-encoded counterparts, with respect to protein synthesis. The 5S rRNA conventionally present as a third rRNA component in bacterial and cytoplasmic ribosomes has been replaced by mt-tRNAVal (e.g. human, rat) or mt-tRNAPhe (e.g. pig, cow) in the central protuberance of the mitochondrial large ribosomal subunit (mt-LSU) [1,4,5,6].

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Conclusion