title: NSUN3 and ABH1 modify the wobble position of mt-tRNAMet to expand codon recognition in mitochondrial translation.
abstract: Mitochondrial gene expression uses a non-universal genetic code in mammals. Besides reading the conventional AUG codon, mitochondrial (mt-)tRNAMetmediates 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-tRNAMetto 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-tRNAMetto generate an f5C34 modification. In vitro codon recognition studies with mitochondrial translation factors reveal preferential utilisation of m5C34 mt-tRNAMetin 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-tRNAMetfunction. Together, our data reveal how modifications in mt-tRNAMetare generated by the sequential action of NSUN3 and ABH1, allowing the single mitochondrial tRNAMetto recognise the different codons encoding methionine.
authors: Institute for Molecular Biology, University Medical Center Göttingen Georg-August-University, Göttingen, Germany.
authors: Sara
authors: S
authors: Haag
authors: Institute for Molecular Biology, University Medical Center Göttingen Georg-August-University, Göttingen, Germany.
authors: Katherine E
authors: KE
authors: Sloan
authors: Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors: Namit
authors: N
authors: Ranjan
authors: Institute for Molecular Biology, University Medical Center Göttingen Georg-August-University, Göttingen, Germany.
authors: Ahmed S
authors: AS
authors: Warda
authors: Institute for Molecular Biology, University Medical Center Göttingen Georg-August-University, Göttingen, Germany.
authors: Jens
authors: J
authors: Kretschmer
authors: Institute for Molecular Biology, University Medical Center Göttingen Georg-August-University, Göttingen, Germany.
authors: Charlotte
authors: C
authors: Blessing
authors: Institute for Molecular Biology, University Medical Center Göttingen Georg-August-University, Göttingen, Germany.
authors: Benedikt
authors: B
authors: Hübner
authors: Institute for Organic and Biomolecular Chemistry, Georg-August-University, Göttingen, Germany Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors: Jan
authors: J
authors: Seikowski
authors: Institute for Cellular Biochemistry, University Medical Center Göttingen Georg-August-University, Göttingen, Germany.
authors: Sven
authors: S
authors: Dennerlein
authors: Max Planck Institute for Biophysical Chemistry, Göttingen, Germany Institute for Cellular Biochemistry, University Medical Center Göttingen Georg-August-University, Göttingen, Germany Göttingen Centre for Molecular Biosciences, Georg-August-University, Göttingen, Germany.
authors: Peter
authors: P
authors: Rehling
authors: Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors: Marina V
authors: MV
authors: Rodnina
authors: Institute for Organic and Biomolecular Chemistry, Georg-August-University, Göttingen, Germany Markus.Bohnsack@med.uni-goettingen.de claudia.hoebartner@chemie.uni-goettingen.de.
authors: Claudia
authors: 0000-0001-7063-5456
authors: C
authors: Höbartner
authors: Institute for Molecular Biology, University Medical Center Göttingen Georg-August-University, Göttingen, Germany Göttingen Centre for Molecular Biosciences, Georg-August-University, Göttingen, Germany Markus.Bohnsack@med.uni-goettingen.de claudia.hoebartner@chemie.uni-goettingen.de.
authors: Markus T
authors: MT
authors: Bohnsack
mesh_terms: D000818:Animals; D002265:Carboxylic Ester Hydrolases / Q000378:metabolism*; D003062:Codon / Q000378:metabolism*; D006801:Humans; D008322:Mammals; D008565:Membrane Proteins / Q000378:metabolism*; D008780:Methyltransferases / Q000378:metabolism*; D008928:Mitochondria / Q000201:enzymology* / Q000378:metabolism*; D014176:Protein Biosynthesis; D012358:RNA, Transfer, Met / Q000378:metabolism*; D017422:Sequence Analysis, DNA
chemical_list: D003062:Codon; D008565:Membrane Proteins; D012358:RNA, Transfer, Met; D008780:Methyltransferases; C000620888:NSUN3 protein, human; C476001:ABHD1 protein, human; D002265:Carboxylic Ester Hydrolases
keywords: ABH1; NSUN3; RNA modification; mitochondria; translation