Human seven-ß-strand (METTL) methyltransferases - conquering the universe of protein lysine methylation.

Lysine methylation is an abundant posttranslational modification, which has been most intensively studied in the context of histone proteins, where it represents an important epigenetic mark. Lysine methylation of histone proteins is primarily catalyzed by SET-domain methyltransferases (MTases). However, it has recently become evident that also another MTase family, the so-called seven-ß-strand (7BS) MTases, often denoted METTLs (methyltransferase-like), contains several lysine (K)-specific MTases (KMTs). These enzymes catalyze the attachment of up to three methyl groups to lysine residues in specific substrate proteins, using S-adenosylmethionine (AdoMet) as methyl donor. About a decade ago, only a single human 7BS KMT was known, namely the histone-specific DOT1L, but 15 additional 7BS KMTs have now been discovered and characterized. These KMTs typically target a single nonhistone substrate that, in most cases, belongs to one of the following three protein groups: components of the cellular protein synthesis machinery, mitochondrial proteins, and molecular chaperones. This article provides an extensive overview and discussion of the human 7BS KMTs and their biochemical and biological roles.

Protein methylation in mitochondria.

Many proteins are modified by posttranslational methylation, introduced by a number of methyltransferases (MTases). Protein methylation plays important roles in modulating protein function and thus in optimizing and regulating cellular and physiological processes. Research has mainly focused on nuclear and cytosolic protein methylation, but it has been known for many years that also mitochondrial proteins are methylated. During the last decade, significant progress has been made on identifying the MTases responsible for mitochondrial protein methylation and addressing its functional significance. In particular, several novel human MTases have been uncovered that methylate lysine, arginine, histidine, and glutamine residues in various mitochondrial substrates. Several of these substrates are key components of the bioenergetics machinery, e.g., respiratory Complex I, citrate synthase, and the ATP synthase. In the present review, we report the status of the field of mitochondrial protein methylation, with a particular emphasis on recently discovered human MTases. We also discuss evolutionary aspects and functional significance of mitochondrial protein methylation and present an outlook for this emergent research field.

Human METTL18 is a histidine-specific methyltransferase that targets RPL3 and affects ribosome biogenesis and function.

Protein methylation occurs primarily on lysine and arginine, but also on some other residues, such as histidine. METTL18 is the last uncharacterized member of a group of human methyltransferases (MTases) that mainly exert lysine methylation, and here we set out to elucidate its function. We found METTL18 to be a nuclear protein that contains a functional nuclear localization signal and accumulates in nucleoli. Recombinant METTL18 methylated a single protein in nuclear extracts and in isolated ribosomes from METTL18 knockout (KO) cells, identified as 60S ribosomal protein L3 (RPL3). We also performed an RPL3 interactomics screen and identified METTL18 as the most significantly enriched MTase. We found that His-245 in RPL3 carries a 3-methylhistidine (3MH; τ-methylhistidine) modification, which was absent in METTL18 KO cells. In addition, both recombinant and endogenous METTL18 were found to be automethylated at His-154, thus further corroborating METTL18 as a histidine-specific MTase. Finally, METTL18 KO cells displayed altered pre-rRNA processing, decreased polysome formation and codon-specific changes in mRNA translation, indicating that METTL18-mediated methylation of RPL3 is important for optimal ribosome biogenesis and function. In conclusion, we have here established METTL18 as the second human histidine-specific protein MTase, and demonstrated its functional relevance.

The human methyltransferase ZCCHC4 catalyses N6-methyladenosine modification of 28S ribosomal RNA.

RNA methylations are essential both for RNA structure and function, and are introduced by a number of distinct methyltransferases (MTases). In recent years, N6-methyladenosine (m6A) modification of eukaryotic mRNA has been subject to intense studies, and it has been demonstrated that m6A is a reversible modification that regulates several aspects of mRNA function. However, m6A is also found in other RNAs, such as mammalian 18S and 28S ribosomal RNAs (rRNAs), but the responsible MTases have remained elusive. 28S rRNA carries a single m6A modification, found at position A4220 (alternatively referred to as A4190) within a stem-loop structure, and here we show that the MTase ZCCHC4 is the enzyme responsible for introducing this modification. Accordingly, we found that ZCCHC4 localises to nucleoli, the site of ribosome assembly, and that proteins involved in RNA metabolism are overrepresented in the ZCCHC4 interactome. Interestingly, the absence of m6A4220 perturbs codon-specific translation dynamics and shifts gene expression at the translational level. In summary, we establish ZCCHC4 as the enzyme responsible for m6A modification of human 28S rRNA, and demonstrate its functional significance in mRNA translation.

Identification of FAM173B as a protein methyltransferase promoting chronic pain.

Chronic pain is a debilitating problem, and insights in the neurobiology of chronic pain are needed for the development of novel pain therapies. A genome-wide association study implicated the 5p15.2 region in chronic widespread pain. This region includes the coding region for FAM173B, a functionally uncharacterized protein. We demonstrate here that FAM173B is a mitochondrial lysine methyltransferase that promotes chronic pain. Knockdown and sensory neuron overexpression strategies showed that FAM173B is involved in persistent inflammatory and neuropathic pain via a pathway dependent on its methyltransferase activity. FAM173B methyltransferase activity in sensory neurons hyperpolarized mitochondria and promoted macrophage/microglia activation through a reactive oxygen species-dependent pathway. In summary, we uncover a role for methyltransferase activity of FAM173B in the neurobiology of pain. These results also highlight FAM173B methyltransferase activity as a potential therapeutic target to treat debilitating chronic pain conditions.

Human FAM173A is a mitochondrial lysine-specific methyltransferase that targets adenine nucleotide translocase and affects mitochondrial respiration.

Lysine methylation is a common posttranslational modification of nuclear and cytoplasmic proteins but is also present in mitochondria. The human protein denoted "family with sequence similarity 173 member B" (FAM173B) was recently uncovered as a mitochondrial lysine (K)-specific methyltransferase (KMT) targeting the c-subunit of mitochondrial ATP synthase (ATPSc), and was therefore renamed ATPSc-KMT. We here set out to investigate the biochemical function of its yet uncharacterized paralogue FAM173A. We demonstrate that FAM173A localizes to mitochondria, mediated by a noncanonical targeting sequence that is partially retained in the mature protein. Immunoblotting analysis using methyllysine-specific antibodies revealed that FAM173A knock-out (KO) abrogates lysine methylation of a single mitochondrial protein in human cells. Mass spectrometry analysis identified this protein as adenine nucleotide translocase (ANT), represented by two highly similar isoforms ANT2 and ANT3. We found that methylation occurs at Lys-52 of ANT, which was previously reported to be trimethylated. Complementation of KO cells with WT or enzyme-dead FAM173A indicated that the enzymatic activity of FAM173A is required for ANT methylation at Lys-52 to occur. Both in human cells and in rat organs, Lys-52 was exclusively trimethylated, indicating that this modification is constitutive, rather than regulatory and dynamic. Moreover, FAM173A-deficient cells displayed increased mitochondrial respiration compared with FAM173A-proficient cells. In summary, we demonstrate that FAM173A is the long-sought KMT responsible for ANT methylation at Lys-52, and point out the functional significance of Lys-52 methylation in ANT. Based on the established naming nomenclature for KMTs, we propose to rename FAM173A to ANT-KMT (gene name ANTKMT).

Lysine methylation by the mitochondrial methyltransferase FAM173B optimizes the function of mitochondrial ATP synthase.

Lysine methylation is an important post-translational modification that is also present on mitochondrial proteins, but the mitochondrial lysine-specific methyltransferases (KMTs) responsible for modification are in most cases unknown. Here, we set out to determine the function of human family with sequence similarity 173 member B (FAM173B), a mitochondrial methyltransferase (MTase) reported to promote chronic pain. Using bioinformatics analyses and biochemical assays, we found that FAM173B contains an atypical, noncleavable mitochondrial targeting sequence responsible for its localization to mitochondria. Interestingly, CRISPR/Cas9-mediated KO of FAM173B in mammalian cells abrogated trimethylation of Lys-43 in ATP synthase c-subunit (ATPSc), a modification previously reported as ubiquitous among metazoans. ATPSc methylation was restored by complementing the KO cells with enzymatically active human FAM173B or with a putative FAM173B orthologue from the nematode Caenorhabditis elegans Interestingly, lack of Lys-43 methylation caused aberrant incorporation of ATPSc into the ATP synthase complex and resulted in decreased ATP-generating ability of the complex, as well as decreased mitochondrial respiration. In summary, we have identified FAM173B as the long-sought KMT responsible for methylation of ATPSc, a key protein in cellular ATP production, and have demonstrated functional significance of ATPSc methylation. We suggest renaming FAM173B to ATPSc-KMT (gene name ATPSCKMT).

Methylation of human eukaryotic elongation factor alpha (eEF1A) by a member of a novel protein lysine methyltransferase family modulates mRNA translation.

Many cellular proteins are methylated on lysine residues and this has been most intensively studied for histone proteins. Lysine methylations on non-histone proteins are also frequent, but in most cases the functional significance of the methylation event, as well as the identity of the responsible lysine (K) specific methyltransferase (KMT), remain unknown. Several recently discovered KMTs belong to the so-called seven-ß-strand (7BS) class of MTases and we have here investigated an uncharacterized human 7BS MTase currently annotated as part of the endothelin converting enzyme 2, but which should be considered a separate enzyme. Combining in vitro enzymology and analyzes of knockout cells, we demonstrate that this MTase efficiently methylates K36 in eukaryotic translation elongation factor 1 alpha (eEF1A) in vitro and in vivo. We suggest that this novel KMT is named eEF1A-KMT4 (gene name EEF1AKMT4), in agreement with the recently established nomenclature. Furthermore, by ribosome profiling we show that the absence of K36 methylation affects translation dynamics and changes translation speed of distinct codons. Finally, we show that eEF1A-KMT4 is part of a novel family of human KMTs, defined by a shared sequence motif in the active site and we demonstrate the importance of this motif for catalytic activity.

The novel lysine specific methyltransferase METTL21B affects mRNA translation through inducible and dynamic methylation of Lys-165 in human eukaryotic elongation factor 1 alpha (eEF1A).

Lysine methylation is abundant on histone proteins, representing a dynamic regulator of chromatin state and gene activity, but is also frequent on many non-histone proteins, including eukaryotic elongation factor 1 alpha (eEF1A). However, the functional significance of eEF1A methylation remains obscure and it has remained unclear whether eEF1A methylation is dynamic and subject to active regulation. We here demonstrate, using a wide range of in vitro and in vivo approaches, that the previously uncharacterized human methyltransferase METTL21B specifically targets Lys-165 in eEF1A in an aminoacyl-tRNA- and GTP-dependent manner. Interestingly, METTL21B-mediated eEF1A methylation showed strong variation across different tissues and cell lines, and was induced by altering growth conditions or by treatment with certain ER-stress-inducing drugs, concomitant with an increase in METTL21B gene expression. Moreover, genetic ablation of METTL21B function in mammalian cells caused substantial alterations in mRNA translation, as measured by ribosomal profiling. A non-canonical function for eEF1A in organization of the cellular cytoskeleton has been reported, and interestingly, METTL21B accumulated in centrosomes, in addition to the expected cytosolic localization. In summary, the present study identifies METTL21B as the enzyme responsible for methylation of eEF1A on Lys-165 and shows that this modification is dynamic, inducible and likely of regulatory importance.

Uncovering human METTL12 as a mitochondrial methyltransferase that modulates citrate synthase activity through metabolite-sensitive lysine methylation.

Lysine methylation is an important and much-studied posttranslational modification of nuclear and cytosolic proteins but is present also in mitochondria. However, the responsible mitochondrial lysine-specific methyltransferases (KMTs) remain largely elusive. Here, we investigated METTL12, a mitochondrial human S-adenosylmethionine (AdoMet)-dependent methyltransferase and found it to methylate a single protein in mitochondrial extracts, identified as citrate synthase (CS). Using several in vitro and in vivo approaches, we demonstrated that METTL12 methylates CS on Lys-395, which is localized in the CS active site. Interestingly, the METTL12-mediated methylation inhibited CS activity and was blocked by the CS substrate oxaloacetate. Moreover, METTL12 was strongly inhibited by the reaction product S-adenosylhomocysteine (AdoHcy). In summary, we have uncovered a novel human mitochondrial KMT that introduces a methyl modification into a metabolic enzyme and whose activity can be modulated by metabolic cues. Based on the established naming nomenclature for similar enzymes, we suggest that METTL12 be renamed CS-KMT (gene name CSKMT).

Regulation of eukaryotic elongation factor 1 alpha (eEF1A) by dynamic lysine methylation.

Lysine methylation is a frequent post-translational protein modification, which has been intensively studied in the case of histone proteins. Lysine methylations are also found on many non-histone proteins, and one prominent example is eukaryotic elongation factor 1 alpha (eEF1A). Besides its essential role in the protein synthesis machinery, a number of non-canonical functions have also been described for eEF1A, such as regulation of the actin cytoskeleton and the promotion of viral replication. The functional significance of the extensive lysine methylations on eEF1A, as well as the identity of the responsible lysine methyltransferases (KMTs), have until recently remained largely elusive. However, recent discoveries and characterizations of human eEF1A-specific KMTs indicate that lysine methylation of eEF1A can be dynamic and inducible, and modulates mRNA translation in a codon-specific fashion. Here, we give a general overview of eEF1A lysine methylation and discuss its possible functional and regulatory significance, with particular emphasis on newly discovered human KMTs.

The METTL20 Homologue from Agrobacterium tumefaciens Is a Dual Specificity Protein-lysine Methyltransferase That Targets Ribosomal Protein L7/L12 and the ß Subunit of Electron Transfer Flavoprotein (ETFß).

Human METTL20 is a mitochondrial, lysine-specific methyltransferase that methylates the ß-subunit of electron transfer flavoprotein (ETFß). Interestingly, putative METTL20 orthologues are found in a subset of α-proteobacteria, including Agrobacterium tumefaciens Using an activity-based approach, we identified in bacterial extracts two substrates of recombinant METTL20 from A. tumefaciens (AtMETTL20), namely ETFß and the ribosomal protein RpL7/L12. We show that AtMETTL20, analogous to the human enzyme, methylates ETFß on Lys-193 and Lys-196 both in vitro and in vivo ETF plays a key role in mediating electron transfer from various dehydrogenases, and we found that its electron transferring ability was diminished by AtMETTL20-mediated methylation of ETFß. Somewhat surprisingly, AtMETTL20 also catalyzed monomethylation of RpL7/L12 on Lys-86, a common modification also found in many bacteria that lack METTL20. Thus, we here identify AtMETTL20 as the first enzyme catalyzing RpL7/L12 methylation. In summary, here we have identified and characterized a novel bacterial lysine-specific methyltransferase with unprecedented dual substrate specificity within the seven ß-strand class of lysine-specific methyltransferases, as it targets two apparently unrelated substrates, ETFß and RpL7/L12. Moreover, the present work establishes METTL20-mediated methylation of ETFß as the first lysine methylation event occurring in both bacteria and humans.

Protein lysine methylation by seven-ß-strand methyltransferases.

Methylation of biomolecules is a frequent biochemical reaction within the cell, and a plethora of highly specific methyltransferases (MTases) catalyse the transfer of a methyl group from S-adenosylmethionine (AdoMet) to various substrates. The posttranslational methylation of lysine residues, catalysed by numerous lysine (K)-specific protein MTases (KMTs), is a very common and important protein modification, which recently has been subject to intense studies, particularly in the case of histone proteins. The majority of KMTs belong to a class of MTases that share a defining 'SET domain', and these enzymes mostly target lysines in the flexible tails of histones. However, the so-called seven-ß-strand (7BS) MTases, characterized by a twisted beta-sheet structure and certain conserved sequence motifs, represent the largest MTase class, and these enzymes methylate a wide range of substrates, including small metabolites, lipids, nucleic acids and proteins. Until recently, the histone-specific Dot1/DOT1L was the only identified eukaryotic 7BS KMT. However, a number of novel 7BS KMTs have now been discovered, and, in particular, several recently characterized human and yeast members of MTase family 16 (MTF16) have been found to methylate lysines in non-histone proteins. Here, we review the status and recent progress on the 7BS KMTs, and discuss these enzymes at the levels of sequence/structure, catalytic mechanism, substrate recognition and biological significance.

Human METTL20 is a mitochondrial lysine methyltransferase that targets the ß subunit of electron transfer flavoprotein (ETFß) and modulates its activity.

Proteins are frequently modified by post-translational methylation of lysine residues, catalyzed by S-adenosylmethionine-dependent lysine methyltransferases (KMTs). Lysine methylation of histone proteins has been extensively studied, but it has recently become evident that methylation of non-histone proteins is also abundant and important. The human methyltransferase METTL20 belongs to a group of 10 established and putative human KMTs. We here found METTL20 to be associated with mitochondria and determined that recombinant METTL20 methylated a single protein in extracts from human cells. Using an methyltransferase activity-based purification scheme, we identified the ß-subunit of the mitochondrially localized electron transfer flavoprotein (ETFß) as the substrate of METTL20. Furthermore, METTL20 was found to specifically methylate two adjacent lysine residues, Lys(200) and Lys(203), in ETFß both in vitro and in cells. Interestingly, the residues methylated by METTL20 partially overlap with the so-called "recognition loop" in ETFß, which has been shown to mediate its interaction with various dehydrogenases. Accordingly, we found that METTL20-mediated methylation of ETFß in vitro reduced its ability to receive electrons from the medium chain acyl-CoA dehydrogenase and the glutaryl-CoA dehydrogenase. In conclusion, the present study establishes METTL20 as the first human KMT localized to mitochondria and suggests that it may regulate cellular metabolism through modulating the interaction between its substrate ETFß and dehydrogenases. Based on the previous naming of similar enzymes, we suggest the renaming of human METTL20 to ETFß-KMT.

Ikbkap/Elp1 deficiency causes male infertility by disrupting meiotic progression.

Mouse Ikbkap gene encodes IKAP--one of the core subunits of Elongator--and is thought to be involved in transcription. However, the biological function of IKAP, particularly within the context of an animal model, remains poorly characterized. We used a loss-of-function approach in mice to demonstrate that Ikbkap is essential for meiosis during spermatogenesis. Absence of Ikbkap results in defects in synapsis and meiotic recombination, both of which result in increased apoptosis and complete arrest of gametogenesis. In Ikbkap-mutant testes, a few meiotic genes are down-regulated, suggesting IKAP's role in transcriptional regulation. In addition, Ikbkap-mutant testes exhibit defects in wobble uridine tRNA modification, supporting a conserved tRNA modification function from yeast to mammals. Thus, our study not only reveals a novel function of IKAP in meiosis, but also suggests that IKAP contributes to this process partly by exerting its effect on transcription and tRNA modification.

Identification and characterization of a novel evolutionarily conserved lysine-specific methyltransferase targeting eukaryotic translation elongation factor 2 (eEF2).

The components of the cellular protein translation machinery, such as ribosomal proteins and translation factors, are subject to numerous post-translational modifications. In particular, this group of proteins is frequently methylated. However, for the majority of these methylations, the responsible methyltransferases (MTases) remain unknown. The human FAM86A (family with sequence similarity 86) protein belongs to a recently identified family of protein MTases, and we here show that FAM86A catalyzes the trimethylation of eukaryotic elongation factor 2 (eEF2) on Lys-525. Moreover, we demonstrate that the Saccharomyces cerevisiae MTase Yjr129c, which displays sequence homology to FAM86A, is a functional FAM86A orthologue, modifying the corresponding residue (Lys-509) in yeast eEF2, both in vitro and in vivo. Finally, Yjr129c-deficient yeast cells displayed phenotypes related to eEF2 function (i.e. increased frameshifting during protein translation and hypersensitivity toward the eEF2-specific drug sordarin). In summary, the present study establishes the function of the previously uncharacterized MTases FAM86A and Yjr129c, demonstrating that these enzymes introduce a functionally important lysine methylation in eEF2. Based on the previous naming of similar enzymes, we have redubbed FAM86A and Yjr129c as eEF2-KMT and Efm3, respectively.

Identification and characterization of a novel human methyltransferase modulating Hsp70 protein function through lysine methylation.

Hsp70 proteins constitute an evolutionarily conserved protein family of ATP-dependent molecular chaperones involved in a wide range of biological processes. Mammalian Hsp70 proteins are subject to various post-translational modifications, including methylation, but for most of these, a functional role has not been attributed. In this study, we identified the methyltransferase METTL21A as the enzyme responsible for trimethylation of a conserved lysine residue found in several human Hsp70 (HSPA) proteins. This enzyme, denoted by us as HSPA lysine (K) methyltransferase (HSPA-KMT), was found to catalyze trimethylation of various Hsp70 family members both in vitro and in vivo, and the reaction was stimulated by ATP. Furthermore, we show that HSPA-KMT exclusively methylates 70-kDa proteins in mammalian protein extracts, demonstrating that it is a highly specific enzyme. Finally, we show that trimethylation of HSPA8 (Hsc70) has functional consequences, as it alters the affinity of the chaperone for both the monomeric and fibrillar forms of the Parkinson disease-associated protein α-synuclein.

The DNA dioxygenase ALKBH2 protects Arabidopsis thaliana against methylation damage.

The Escherichia coli AlkB protein (EcAlkB) is a DNA repair enzyme which reverses methylation damage such as 1-methyladenine (1-meA) and 3-methylcytosine (3-meC). The mammalian AlkB homologues ALKBH2 and ALKBH3 display EcAlkB-like repair activity in vitro, but their substrate specificities are different, and ALKBH2 is the main DNA repair enzyme for 1-meA in vivo. The genome of the model plant Arabidopsis thaliana encodes several AlkB homologues, including the yet uncharacterized protein AT2G22260, which displays sequence similarity to both ALKBH2 and ALKBH3. We have here characterized protein AT2G22260, by us denoted ALKBH2, as both our functional studies and bioinformatics analysis suggest it to be an orthologue of mammalian ALKBH2. The Arabidopsis ALKBH2 protein displayed in vitro repair activities towards methyl and etheno adducts in DNA, and was able to complement corresponding repair deficiencies of the E. coli alkB mutant. Interestingly, alkbh2 knock-out plants were sensitive to the methylating agent methylmethanesulphonate (MMS), and seedlings from these plants developed abnormally when grown in the presence of MMS. The present study establishes ALKBH2 as an important enzyme for protecting Arabidopsis against methylation damage in DNA, and suggests its homologues in other plants to have a similar function.

Roles of Trm9- and ALKBH8-like proteins in the formation of modified wobble uridines in Arabidopsis tRNA.

Uridine at the wobble position of tRNA is usually modified, and modification is required for accurate and efficient protein translation. In eukaryotes, wobble uridines are modified into 5-methoxycarbonylmethyluridine (mcm(5)U), 5-carbamoylmethyluridine (ncm(5)U) or derivatives thereof. Here, we demonstrate, both by in vitro and in vivo studies, that the Arabidopsis thaliana methyltransferase AT1G31600, denoted by us AtTRM9, is responsible for the final step in mcm(5)U formation, thus representing a functional homologue of the Saccharomyces cerevisiae Trm9 protein. We also show that the enzymatic activity of AtTRM9 depends on either one of two closely related proteins, AtTRM112a and AtTRM112b. Moreover, we demonstrate that AT1G36310, denoted AtALKBH8, is required for hydroxylation of mcm(5)U to (S)-mchm(5)U in tRNA(Gly)(UCC), and has a function similar to the mammalian dioxygenase ALKBH8. Interestingly, atalkbh8 mutant plants displayed strongly increased levels of mcm(5)U, and also of mcm(5)Um, its 2'-O-ribose methylated derivative. This suggests that accumulated mcm(5)U is prone to further ribose methylation by a non-specialized mechanism, and may challenge the notion that the existence of mcm(5)U- and mcm(5)Um-containing forms of the selenocysteine-specific tRNA(Sec) in mammals reflects an important regulatory process. The present study reveals a role in for several hitherto uncharacterized Arabidopsis proteins in the formation of modified wobble uridines.

Characterization of the biochemical activity and tumor-promoting role of the dual protein methyltransferase METL-13/METTL13 in Caenorhabditis elegans.

The methyltransferase-like protein 13 (METTL13) methylates the eukaryotic elongation factor 1 alpha (eEF1A) on two locations: the N-terminal amino group and lysine 55. The absence of this methylation leads to reduced protein synthesis and cell proliferation in human cancer cells. Previous studies showed that METTL13 is dispensable in non-transformed cells, making it potentially interesting for cancer therapy. However, METTL13 has not been examined yet in whole animals. Here, we used the nematode Caenorhabditis elegans as a simple model to assess the functions of METTL13. Using methyltransferase assays and mass spectrometry, we show that the C. elegans METTL13 (METL-13) methylates eEF1A (EEF-1A) in the same way as the human protein. Crucially, the cancer-promoting role of METL-13 is also conserved and depends on the methylation of EEF-1A, like in human cells. At the same time, METL-13 appears dispensable for animal growth, development, and stress responses. This makes C. elegans a convenient whole-animal model for studying METL13-dependent carcinogenesis without the complications of interfering with essential wild-type functions.