Abstract
METTL20 is a seven-β-strand methyltransferase that is localised to the mitochondria and tri-methylates the electron transfer flavoprotein (ETF) β subunit (ETFB) at lysines 200 and 203. It has been shown that METTL20 decreases the ability of ETF to extract electrons from medium-chain acyl-coenzyme A (CoA) dehydrogenase (MCAD) and glutaryl-CoA dehydrogenase in vitro. METTL20-mediated methylation of ETFB influences the oxygen consumption rate in permeabilised mitochondria, suggesting that METTL20-mediated ETFB methylation may also play a regulatory role in mitochondrial metabolism. In this study, we generated Mettl20 knockout (KO) mice to uncover the in vivo functions of METTL20. The KO mice were viable, and a loss of ETFB methylation was confirmed. In vitro enzymatic assays revealed that mitochondrial ETF activity was higher in the KO mice than in wild-type mice, suggesting that the KO mice had higher β-oxidation capacity. Calorimetric analysis showed that the KO mice fed a ketogenic diet had higher oxygen consumption and heat production. A subsequent cold tolerance test conducted after 24 h of fasting indicated that the KO mice had a better ability to maintain their body temperature in cold environments. Thus, METTL20 regulates ETF activity and heat production through lysine methylation when β-oxidation is highly activated.
Highlights
Protein post-translational modifications (PTMs) are involved in diverse cellular processes such as gene expression, signal transduction, and intracellular interactions
The two METTL20 methylation sites in ETFB, i.e. K200 and K203, are located in the proximity of the recognition loop responsible for the interaction between electron transfer flavoprotein (ETF) and medium-chain acyl-coenzyme A (CoA) dehydrogenase (MCAD)[7]; it is believed to be involved in interactions with other ETF-dependent dehydrogenases
We found that ETFB is a major substrate; this result is consistent with other findings[5,6]
Summary
Protein post-translational modifications (PTMs) are involved in diverse cellular processes such as gene expression, signal transduction, and intracellular interactions. There are approximately 40 MTase genes that may participate in mitochondrial functioning because the encoded proteins contain mitochondria-targeting signal sequences[4]. Several approaches have been developed to probe lysine-methylated proteins . It has been reported that an engineered version of the triple malignant-brain-tumour domain region, i.e. L3MBTL1, can serve as a probe for the detection and enrichment of proteins containing mono- and di-methylated lysine residue(s)[10,11]. Another approach is to use cofactor analogues for labelling, detection, and pull-down via copper-catalysed azide-alkyne cycloaddition, known as click chemistry, to enrich PTM targets. To explore the biological function of METTL20-mediated ETFB methylation, we created Mettl[20] knockout (KO) mice and characterised phenotypes related to their metabolism
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