Mitochondrial RNA modifications shape metabolic plasticity in metastasis
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE201993 | GEO | 2022/05/05
REPOSITORIES: GEO
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Project description:Mitochondrial gene expression uses a non-universal genetic code in mammals. Besides reading the conventional AUG codon, mitochondrial (mt-)tRNAMet mediates incorporation of methionine on AUA and AUU codons during translation initiation and on AUA codons during elongation. We show that the RNA methyltransferase NSUN3 localises to mitochondria and interacts with mt-tRNAMet to methylate cytosine 34 (C34) at the wobble position. NSUN3 specifically recognises the anticodon stem loop (ASL) of the tRNA, explaining why a mutation that compromises ASL basepairing leads to disease. We further identify ALKBH1/ABH1 as the dioxygenase responsible for oxidising m5C34 of mt-tRNAMet to generate an f5C34 modification. In vitro codon recognition studies with mitochondrial translation factors reveal preferential utilization of m5C34 mt-tRNAMet in initiation. Depletion of either NSUN3 or ABH1 strongly affects mitochondrial translation in human cells, implying that modifications generated by both enzymes are necessary for mt-tRNAMet function. Together, our data reveal how modifications in mt-tRNAMet are generated by the sequential action of NSUN3 and ABH1, allowing the single mitochondrial tRNAMet to recognise the different codons encoding methionine. HEK293 cell lines expressing His-FLAG-tagged NSUN3 or the His-FLAG tag alone were crosslinked using UV or treated with 5-azacytidine and analysed by CRAC
2016-08-09 | E-GEOD-84664 | biostudies-arrayexpress
Project description:Mitochondrial gene expression uses a non-universal genetic code in mammals. Besides reading the conventional AUG codon, mitochondrial (mt-)tRNAMet mediates incorporation of methionine on AUA and AUU codons during translation initiation and on AUA codons during elongation. We show that the RNA methyltransferase NSUN3 localises to mitochondria and interacts with mt-tRNAMet to methylate cytosine 34 (C34) at the wobble position. NSUN3 specifically recognises the anticodon stem loop (ASL) of the tRNA, explaining why a mutation that compromises ASL basepairing leads to disease. We further identify ALKBH1/ABH1 as the dioxygenase responsible for oxidising m5C34 of mt-tRNAMet to generate an f5C34 modification. In vitro codon recognition studies with mitochondrial translation factors reveal preferential utilization of m5C34 mt-tRNAMet in initiation. Depletion of either NSUN3 or ABH1 strongly affects mitochondrial translation in human cells, implying that modifications generated by both enzymes are necessary for mt-tRNAMet function. Together, our data reveal how modifications in mt-tRNAMet are generated by the sequential action of NSUN3 and ABH1, allowing the single mitochondrial tRNAMet to recognise the different codons encoding methionine.
2016-08-09 | GSE84664 | GEO
Project description:Modified nucleotides in tRNAs are important determinants of folding, structure and function. Here, we identify METTL8 as a mitochondrial matrix protein and active RNA methyltransferase responsible for installing m3C32 in the human mitochondrial (mt-)tRNAThr and mt-tRNASer(UCN). METTL8 crosslinks to the anticodon stem loop (ASL) of many mt-tRNAs in cells, raising the question of how methylation target specificity is achieved. Dissection of mt-tRNA recognition elements revealed U34G35 and t6A37/(ms2)i6A37, present concomitantly only in the ASLs of the two substrate mt-tRNAs, as key determinations for METTL8-mediated methylation of C32. Several lines of evidence demonstrate the influence of U34, G35, and the m3C32 and t6A37/(ms2)i6A37 modifications in mt-tRNAThr/Ser(UCN) on the structure of these mt-tRNAs. Although mt-tRNAThr/Ser(UCN) lacking METTL8-mediated m3C32 are efficiently aminoacylated and associate with mitochondrial ribosomes, mitochondrial translation is mildly impaired by lack of METTL8. Together these results define the cellular targets of METTL8 and shed new light on the role of m3C32 within mt-tRNAs.
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Project description:Modified nucleotides in tRNAs are important determinants of folding, structure and function. Here, we identify METTL8 as a mitochondrial matrix protein and active RNA methyltransferase responsible for installing m3C32 in the human mitochondrial (mt-)tRNAThr and mt-tRNASer(UCN). METTL8 crosslinks to the anticodon stem loop (ASL) of many mt-tRNAs in cells, raising the question of how methylation target specificity is achieved. Dissection of mt-tRNA recognition elements revealed U34G35 and t6A37/(ms2)i6A37, present concomitantly only in the ASLs of the two substrate mt-tRNAs, as key determinations for METTL8-mediated methylation of C32. Several lines of evidence demonstrate the influence of U34, G35, and the m3C32 and t6A37/(ms2)i6A37 modifications in mt-tRNAThr/Ser(UCN) on the structure of these mt-tRNAs. Although mt-tRNAThr/Ser(UCN) lacking METTL8-mediated m3C32 are efficiently aminoacylated and associate with mitochondrial ribosomes, mitochondrial translation is mildly impaired by lack of METTL8. Together these results define the cellular targets of METTL8 and shed new light on the role of m3C32 within mt-tRNAs.
2021-12-04 | GSE174448 | GEO
Project description:Mitochondria are organelles that generate most of the energy in eukaryotic cells in the form of ATP via oxidative phosphorylation in eukaryote. Twenty-two species of mitochondrial (mt-)tRNAs encoded in mtDNA are required to translate essential subunits of the respiratory chain complexes involved in oxidative phosphorylation. mt-tRNAs contain post-transcriptional modifications introduced by nuclear-encoded tRNA-modifying enzymes. These modifications are required for deciphering genetic code accurately, as well as stabilizing tRNA. Loss of tRNA modifications frequently results in severe pathological consequences. We performed a comprehensive analysis of post-transcriptional modifications of all human mt-tRNAs, including 14 previously-uncharacterized species, and revised the modification status of some of the previously studied species. In total, we found 17 kinds of RNA modifications at 137 positions (8.7% in 1,575 nucleobases) in 22 species of human mt-tRNAs. An up-to-date list of 34 genes responsible for human mt-tRNA modifications are provided. We here demonstrated that both QTRT1 and QTRT2 are required for biogenesis of queuosine (Q) at position 34 of four mt-tRNAs. Our results provide insight into the molecular mechanisms underlying the mitochondrial decoding system, and could help to elucidate the molecular pathogenesis of human mitochondrial diseases caused by aberrant tRNA modifications.
2020-08-28 | GSE150439 | GEO
Project description:Mitochondria contain a specific translation machinery for the synthesis of respiratory chain components encoded on the mitochondrial genome. Mitochondrial tRNAs (mt-tRNAs) are also generated from the mitochondrial genome and, similar to their cytoplasmic counterparts, are modified at various positions. Here, we find that the RNA methyltransferase METTL8, is a mitochondrial protein that facilitates m3C methylation at position C32 of mt-tRNASer(UCN) and mt-tRNAThr. METTL8 knock out cells show reduced and over expressing cells enhanced respiratory chain activity. In pancreatic cancer, METTL8 levels are high, which correlates with patient survival. Indeed, METTL8 up regulation stimulates respiratory chain activity in these cells. Ribosome occupancy analysis using ribosome profiling revealed ribosome stalling on mt-tRNASer(UCN) and mt-tRNAThr codons and mass spectrometry analysis of native ribosomal subcomplexes unraveled reduced respiratory chain incorporation of the mitochondria encoded proteins ND6 and ND1. A well-balanced translation of mt-tRNASer(UCN) and mt-tRNAThr codons through METTL8-mediated C32 methylation might therefore provide optimal respiratory chain compositions and function.
2023-01-02 | GSE180400 | GEO
Project description:A subset of eukaryotic tRNAs is methylated in the anticodon loop to form the 3-methylcytosine (m3C) modification. In mammals, the number of tRNAs containing m3C has expanded to include mitochondrial (mt) tRNA-Ser-UGA and mt-tRNA-Thr-UGU. Whereas the enzymes catalyzing m3C formation in nuclear-encoded cytoplasmic tRNAs have been identified, the proteins responsible for m3C modification in mt-tRNAs are unknown. Here, we show that m3C formation in human mt-tRNAs is dependent upon the Methyltransferase-Like 8 (METTL8) enzyme. We find that METTL8 is a mitochondria-associated protein that interacts with mitochondrial seryl-tRNA synthetase along with mt-tRNAs containing m3C. Human cells deficient in METTL8 exhibit loss of m3C modification in mt-tRNAs but not nuclear-encoded tRNAs. Consistent with the mitochondrial import of METTL8, the formation of m3C in METTL8-deficient cells can be rescued by re-expression of wildtype METTL8 but not by a METTL8 variant lacking the N-terminal mitochondrial localization signal. Notably, METTL8-deficiency in human cells causes alterations in the native migration pattern of mt-tRNA-Ser-UGA suggesting a role for m3C in tRNA folding. Altogether, these findings demonstrate that METTL8 is required for m3C formation in mitochondrial tRNAs and uncover a potential role for m3C modification in mitochondrial tRNA structure.
2022-05-19 | PXD030418 | Pride
Project description:Spermatozoa harbor a complex and environment sensitive pool of small non-coding RNAs (sncRNA)1, which influences offspring development and adult phenotypes1-7. Whether spermatozoa in the epididymis are directly susceptible to environmental cues is not fully understood8. We used two distinct paradigms of preconception acute high fat diet to dissect epididymal vs testicular contributions to the sperm sncRNA pool and offspring health. We show that epididymal spermatozoa, but not developing germ cells, are sensitive to the environment and identify mitochondrial tRNAs and their fragments (mt-tsRNA) as sperm-borne factors. In humans, mt-tsRNAs in spermatozoa correlate with BMI and paternal overweight at conception doubles offspring obesity risk and compromises metabolic health. Sperm sncRNA-seq of mice mutant for genes involved in mitochondrial function, and metabolic phenotyping of their wild-type offspring, suggest that alterations of mt-tsRNAs are downstream of mitochondrial dysfunction. Most importantly, single embryo transcriptomics of genetically hybrid two-cell embryos demonstrated sperm-to-oocyte transfer of mt-tRNAs at fertilisation and implied them in the control of early embryo transcription. Our study supports the importance of paternal health at conception for offspring metabolism, shows that mt-tRNAs are diet-induced and sperm-borne and demonstrates, for the first time in a physiological setting, father-to-offspring transfer of sperm mitochondrial RNAs at fertilization.
2024-04-03 | GSE239815 | GEO
Project description:Mitochondrial RNA modifications shape metabolic plasticity in metastasis
| PRJNA833944 | ENA
Project description:Organoids derived from small intestine epithelial cells of C57BL/6 wild-type mice were cultured in the presence or absence of IL-28 (1000 ng/ml) for 24 hours. The experimental setup included 8 samples in a pairwise design (4 unstimulated vs 4 stimulated). Profiling of whole-genome transcriptomic patterns was performed by RNA-Seq on an Illumina HiSeq 2500 platform using total RNA at the Next Generation Sequencing core unit of the University Hospital Erlangen, Germany. Demultiplexed reads were quality filtered against rRNAs, tRNAs, mt-rRNAs and mt-tRNAs. Alignment against the mus musculus reference genome (Ensemble V.84 for GRCm38) was performed with RNA-seq aligner STAR (V.2.5.1b) and statistical analysis of unique mappings (HTseq count) was computed in R using the DESeq2 1.10.1.
2017-07-01 | E-MTAB-4862 | biostudies-arrayexpress