Homo sapiens
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA380795 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5382420.fastq.gz | Fastqsanger.gz | |||
| SRR5382430.fastq.gz | Fastqsanger.gz | |||
| SRR5382421.fastq.gz | Fastqsanger.gz | |||
| SRR5382422.fastq.gz | Fastqsanger.gz | |||
| SRR5382423.fastq.gz | Fastqsanger.gz |
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Project description:Eukaryotic elongation factor 1 alpha (eEF1A) delivers aminoacyl-tRNA to the ribosome and thereby plays a key role in protein synthesis. Human eEF1A is post-translationally methylated on several lysine residues as well as on the N-terminus and until recently the responsible methyltransferases were largely elusive. Using mass spectrometry based screens, gene targeted cells and in vitro enzymology we identify the human methyltransferase like proteins 13 (METTL13), which contains two distinct methyltransferase domains, as an enzyme catalyzing both dimethylation of Lys55 and trimethylation of the N-terminus of eEF1A. Moreover, through ribosome profiling we demonstrate that loss of METTL13 function alters translation dynamics and results in changed translation rate of specific codons. In line with the established descriptive nomenclature for this type of enzymes we suggest that METTL13 is redubbed eEF1A dual methyltransferase (eEF1A-DMT / EEF1ADMT).
2018-07-04 | GSE104033 | GEO
Project description:Eukaryotic elongation factor 1A (eEF1A) is an essential, highly methylated protein that facilitates translational elongation by delivering aminoacyl-tRNAs to ribosomes. Here we report a new eukaryotic protein N-terminal methyltransferase, Saccharomyces cerevisiae YLR285W, which methylates eEF1A at a previously undescribed high-stoichiometry N-terminal site and the adjacent lysine. Deletion of YLR285W resulted in the loss of N-terminal and lysine methylation in vivo, whereas overexpression of YLR285W resulted in an increase of methylation at these sites. This was confirmed by in vitro methylation of eEF1A by recombinant YLR285W. Accordingly, we name YLR285W as elongation factor methyltransferase 7 (Efm7). This enzyme is a new type of eukaryotic N-terminal methyltransferase as, unlike the three other known eukaryotic N-terminal methyltransferases, its substrate does not have an N-terminal [A/P/S]-P-K motif. We show that the N-terminal methylation of eEF1A is also present in human; this conservation over a large evolutionary distance suggests it to be of functional importance. This study also reports that the trimethylation of K79 in eEF1A is conserved from yeast to human. The methyltransferase responsible for K79 methylation of human eEF1A is shown to be N6AMT2, previously documented as a putative N(6)-adenine-specific DNA methyltransferase. It is the direct ortholog of the recently described yeast Efm5 and we show that Efm5 and N6AMT2 can methylate eEF1A from either species in vitro. We therefore rename N6AMT2 as eEF1A-KMT1. Including the present work, yeast eEF1A is now documented to be methylated by five different methyltransferases, making it one of the few eukaryotic proteins to be extensively methylated by independent enzymes. This implies more extensive regulation of eEF1A by this post-translational modification than previously appreciated.
2015-11-10 | PXD002941 | Pride
Project description:Many cellular proteins are post-translationally modified by methylation of lysine residues. This has been most intensively studied in the case of histone proteins, where lysine methylations in the flexible tails are determinants of chromatin state and gene expression. 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. While the majority of identified human KMTs belong to the SET (Su(var)3-9, Enhancer-of-zeste, Trithorax)-domain class of methyltransferases (MTases), several recently discovered KMTs belong to a different MTase class, the seven-beta-strand (7BS) MTases. Here, we have investigated an uncharacterized human 7BS MTase currently annotated as being part of the endothelin converting enzyme 2 (ECE2). However, transcriptomics data indicate that this MTase is not part of ECE2, and should be considered a separate protein. Combining in vitro enzymology and analyses 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 demonstrate that eEF1A-KMT4 is part of a novel family of human KMTs, defined by a shared sequence motif in the active site, and we show the importance of this motif for catalytic activity.
2017-05-23 | GSE97140 | GEO
Project description:Lysine methylation is abundant on histone proteins, representing a dynamic regulator of chromatin state and gene activity, but is also frequent on many nonhistone proteins, including eukaryotic elongation factor 1 alpha (eEF1A). However, the functional significance of eEF1Amethylation 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 ribosome 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.
2017-01-05 | GSE93133 | GEO
Project description:Lysine methylation is widespread on human proteins, however the enzymes that catalyse its addition remain largely unknown. This limits our capacity to study the function and regulation of this modification. Here we report that human METTL21B is a protein methyltransferase, which methylates lysine 165 of eukaryotic translation elongation factor 1A (eEF1A). The CRISPR/Cas9 system was used to knock out putative protein methyltransferases METTL21B and METTL23 in K562 cells. The known eEF1A methyltransferase EEF1AKMT1 was also knocked out as a control. Targeted mass spectrometry revealed the loss of lysine 165 methylation upon knock out of METTL21B, and the expected loss of lysine 79 methylation on knock out of EEF1AKMT1. No loss of eEF1A methylation was seen in the METTL23 knock out. Recombinant METTL21B was then shown to catalyse methylation on lysine 165 in eEF1A1 and eEF1A2 in vitro, confirming it as the methyltransferase responsible for this methylation site. Stable isotope labelling by amino acids in cell culture (SILAC) proteomic analysis revealed dysregulation of processes related to eEF1A function in knock outs of METTL21B and EEF1AKMT1, but specific dysregulation of ribosomal proteins in the knock out of METTL21B. This indicates a specific function for lysine 165 methylation of eEF1A in human. METTL21B is specific to vertebrates, with its target lysine showing similar evolutionary conservation. We suggest METTL21B be renamed eEF1A-KMT3. This is the first study to specifically generate CRISPR/Cas9 knock outs of the genes encoding putative protein methyltransferases, for the purpose of substrate discovery and site mapping. Our approach should prove useful for the discovery of further novel methyltransferases, and more generally for the discovery of sites for other protein-modifying enzymes.
2017-07-03 | PXD005497 | Pride
Project description:Translation elongation factor 1A (eEF1A) is an essential and highly conserved protein required for protein synthesis in eukaryotes. In both Saccharomyces cerevisiae and human, five different methyltransferase enzymes methylate specific residues on eEF1A, making eEF1A the eukaryotic protein targeted by the highest number of dedicated methyltransferases. eEF1A methyltransferases are highly selective enzymes, only targeting eEF1A and each targeting just one or two specific residues in eEF1A. However, the mechanism of this selectivity remains poorly understood. Here we have used AlphaFold modelling in combination with crosslinking mass spectrometry (XL-MS) and enzyme mutagenesis to reveal how S. cerevisiae elongation factor methyltransferase 4 (Efm4) specifically methylates eEF1A at K316. We find that a unique beta-hairpin motif, which extends out from the core methyltransferase fold, is important for methylation of eEF1A K316 in vitro. An alanine mutation of a single residue on this beta-hairpin, F212, significantly reduces Efm4 activity in vitro and in yeast cells. We show that the equivalent residue in human eEF1A-KMT2 (METTL10), F220, is also important for its activity towards eEF1A in vitro. Lastly, we find that phosphorylation of eEF1A at S314 negatively crosstalks with Efm4-mediated methylation of K316. Our findings demonstrate how protein methyltransferases can be being highly selective towards a single residue on a single protein.
2024-01-03 | PXD042599 | Pride
Project description:Cotranslational protein folding depends on general chaperones that engage highly diverse nascent chains at the ribosomes. Here we find that the universal cotranslational machinery adapts to accommodate the challenging biogenesis of abundantly expressed eukaryotic translation elongation factor 1A (eEF1A). During eEF1A synthesis, chaperone Chp1 is recruited to the ribosome with the help of the nascent polypeptide-associated complex (NAC), where it safeguards eEF1A biogenesis. Aberrant eEF1A production triggers instant proteolysis, widespread protein aggregation, activation of Hsf1 stress transcription and compromises cellular fitness. The expression of pathogenic eEF1A2 variants linked to epileptic-dyskinetic encephalopathy is protected by Chp1. Thus, eEF1A is a difficult to fold protein that necessitates dedicated folding factor Chp1 at the ribosomal tunnel exit to protect the eukaryotic cell from proteostasis collapse.
2024-01-16 | GSE221651 | GEO
Project description: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)
| PRJNA360112 | ENA
Project description: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.
2023-07-03 | PXD042540 | Pride
Project description:NSP12 of SARS-CoV-2 can recruit the elongation factor eEF1A, we intend to know whether NSP12 regulate the mRNA translation level of host cell upon viral infection
2024-08-22 | GSE182745 | GEO