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Mammalian mitochondrial tRNA (mt-tRNA) plays a central role in the synthesis of the 13 subunits of the oxidative phosphorylation complex system (OXPHOS). However, many aspects of the context-dependent expression of mt-tRNAs in mammals remain unknown. To investigate the tissue-specific effects of mt-tRNAs, we performed a comprehensive analysis of mitochondrial tRNA expression across five mice tissues (brain, heart, liver, skeletal muscle, and kidney) using Northern blot analysis. Striking differences in the tissue-specific expression of 22 mt-tRNAs were observed, in some cases differing by as much as tenfold from lowest to highest expression levels among these five tissues. Overall, the heart exhibited the highest levels of mt-tRNAs, while the liver displayed markedly lower levels. Variations in the levels of mt-tRNAs showed significant correlations with total mitochondrial DNA (mtDNA) contents in these tissues. However, there were no significant differences observed in the 2-thiouridylation levels of tRNA Lys , tRNA Glu , and tRNA Gln among these tissues. A wide range of aminoacylation levels for 15 mt-tRNAs occurred among these five tissues, with skeletal muscle and kidneys most notably displaying the highest and lowest tRNA aminoacylation levels, respectively. Among these tissues, there was a negative correlation between variations in mt-tRNA aminoacylation levels and corresponding variations in mitochondrial tRNA synthetases (mt-aaRS) expression levels. Furthermore, the variable levels of OXPHOS subunits, as encoded by mtDNA or nuclear genes, may reflect differences in relative functional emphasis for mitochondria in each tissue. Our findings provide new insight into the mechanism of mt-tRNA tissue-specific effects on oxidative phosphorylation., Competing Interests: Conflict of interest All authors declare that they have no conflict of interest with contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Mitochondrial DNA gene expression is coordinately regulated both pre- and post-transcriptionally, and its perturbation can lead to human pathologies. Mitochondrial rRNAs (mt-rRNAs) undergo a series of nucleotide modifications after release from polycistronic mitochondrial RNA precursors, which is essential for mitochondrial ribosomal biogenesis. Cytosine N 4 -methylation (m 4 C) at position 839 (m 4 C839) of the 12S small subunit mt-rRNA was identified decades ago; however, its biogenesis and function have not been elucidated in detail. Here, using several approaches, including immunofluorescence, RNA immunoprecipitation and methylation assays, and bisulfite mapping, we demonstrate that human methyltransferase-like 15 (METTL15), encoded by a nuclear gene, is responsible for 12S mt-rRNA methylation at m 4 C839 both in vivo and in vitro We tracked the evolutionary history of RNA m 4 C methyltransferases and identified a difference in substrate preference between METTL15 and its bacterial ortholog rsmH. Additionally, unlike the very modest impact of a loss of m 4 C methylation in bacterial small subunit rRNA on the ribosome, we found that METTL15 depletion results in impaired translation of mitochondrial protein-coding mRNAs and decreases mitochondrial respiration capacity. Our findings reveal that human METTL15 is required for mitochondrial function, delineate the evolution of methyltransferase substrate specificities and modification patterns in rRNA, and highlight a differential impact of m 4 C methylation on prokaryotic ribosomes and eukaryotic mitochondrial ribosomes., Competing Interests: Conflict of interest—Y. S. is a co-founder/equity holder of Constellation Pharmaceuticals, Inc. and Athelas Therapeutics, Inc., an equity holder of Imago Biosciences and a consultant for Active Motif., (© 2020 Chen et al.)

Perfectly accurate translation of mRNA into protein is not a prerequisite for life. Resulting from errors in protein synthesis, mistranslation occurs in all cells, including human cells. The human genome encodes >600 tRNA genes, providing both the raw material for genetic variation and a buffer to ensure that resulting translation errors occur at tolerable levels. On the basis of data from the 1000 Genomes Project, we highlight the unanticipated prevalence of mistranslating tRNA variants in the human population and review studies on synthetic and natural tRNA mutations that cause mistranslation or de-regulate protein synthesis. Although mitochondrial tRNA variants are well known to drive human diseases, including developmental disorders, few studies have revealed a role for human cytoplasmic tRNA mutants in disease. In the context of the unexpectedly large number of tRNA variants in the human population, the emerging literature suggests that human diseases may be affected by natural tRNA variants that cause mistranslation or de-regulate tRNA expression and nucleotide modification. This review highlights examples relevant to genetic disorders, cancer, and neurodegeneration in which cytoplasmic tRNA variants directly cause or exacerbate disease and disease-linked phenotypes in cells, animal models, and humans. In the near future, tRNAs may be recognized as useful genetic markers to predict the onset or severity of human disease., (© 2019 Lant et al.)

Mitochondrial tRNAs are transcribed as long polycistronic transcripts of precursor tRNAs and undergo posttranscriptional modifications such as endonucleolytic processing and methylation required for their correct structure and function. Among them, 5'-end processing and purine 9 N1-methylation of mitochondrial tRNA are catalyzed by two proteinaceous complexes with overlapping subunit composition. The Mg 2+ -dependent RNase P complex for 5'-end cleavage comprises the methyltransferase domain-containing protein tRNA methyltransferase 10C, mitochondrial RNase P subunit (TRMT10C/MRPP1), short-chain oxidoreductase hydroxysteroid 17β-dehydrogenase 10 (HSD17B10/MRPP2), and metallonuclease KIAA0391/MRPP3. An MRPP1-MRPP2 subcomplex also catalyzes the formation of 1-methyladenosine/1-methylguanosine at position 9 using S -adenosyl-l-methionine as methyl donor. However, a lack of structural information has precluded insights into how these complexes methylate and process mitochondrial tRNA. Here, we used a combination of X-ray crystallography, interaction and activity assays, and small angle X-ray scattering (SAXS) to gain structural insight into the two tRNA modification complexes and their components. The MRPP1 N terminus is involved in tRNA binding and monomer-monomer self-interaction, whereas the C-terminal SPOUT fold contains key residues for S -adenosyl-l-methionine binding and N1-methylation. The entirety of MRPP1 interacts with MRPP2 to form the N1-methylation complex, whereas the MRPP1-MRPP2-MRPP3 RNase P complex only assembles in the presence of precursor tRNA. This study proposes low-resolution models of the MRPP1-MRPP2 and MRPP1-MRPP2-MRPP3 complexes that suggest the overall architecture, stoichiometry, and orientation of subunits and tRNA substrates., (© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.)