Affinage

METTL9

Protein-L-histidine N-pros-methyltransferase · UniProt Q9H1A3

Length
318 aa
Mass
36.5 kDa
Annotated
2026-06-10
15 papers in source corpus 11 papers cited in narrative 11 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

METTL9 is a broad-specificity, SAM-dependent protein methyltransferase that generates the bulk of N1 (Nπ)-methylhistidine found in mammalian proteomes by modifying the second histidine of His-x-His (HxH) motifs in diverse substrates (PMID:33563959, PMID:34562450). Structural analysis shows the first histidine of the motif serves as a recognition anchor and the small 'x' residue is held within the substrate pocket, while an active-site aspartate stabilizes the imidazole N3 to present N1 to the SAM methyl donor, and tandem HxH repeats are methylated processively in a C-to-N direction (PMID:37398635); the catalytic SAM pocket has been confirmed pharmacologically by a selective inhibitor that abolishes cellular 1-methylhistidine (PMID:41870122). A recurrent functional consequence of this modification is attenuation of zinc binding at target sites, demonstrated for the metal-binding protein S100A9 at His-107 and for HxH-containing peptides (PMID:33563959, PMID:34562450). Characterized substrates and their physiological outputs include NDUFB3, where methylation enhances mitochondrial Complex I-dependent respiration (PMID:33563959), and the zinc transporter SLC39A7 at His45/His49, where methylation suppresses ferroptosis through PERK/ATF4 and SLC7A11 signaling (PMID:40414869). Through effects on Complex I activity and on SLC7A11-dependent ferroptosis, METTL9 promotes gastric cancer migration/invasion and hepatocellular carcinoma progression (PMID:35402738, PMID:38017014, PMID:40523929). Distinct from its catalytic activity, METTL9 supports vertebrate neural development by interacting with regulators of cellular transport, endocytosis, and Golgi integrity, with its loss causing Golgi fragmentation (PMID:40745158).

Mechanistic history

Synthesis pass · year-by-year structured walk · 11 steps
  1. 2021 High

    Established METTL9 as the principal enzyme generating proteome-wide 1-methylhistidine and defined its HxH motif specificity, linking the modification to Complex I respiration and zinc binding.

    Evidence In vitro methyltransferase assays, proteome-wide LC-MS/MS of 1MH, knockout mice, Complex I activity and zinc-binding measurements

    PMID:33563959

    Open questions at the time
    • Full substrate repertoire beyond S100A9 and NDUFB3 not enumerated
    • Physiological role of zinc-binding attenuation not mapped to whole-organism phenotypes
  2. 2021 High

    Pinpointed a specific physiological methylation site (S100A9 His-107) and showed the mark overlaps the zinc-binding site without disrupting S100A8 heterodimerization, providing a concrete substrate-level mechanism.

    Evidence siRNA screening with methylhistidine LC-MS/MS, in vitro methyltransferase assay, zinc-binding biochemistry

    PMID:34562450

    Open questions at the time
    • Downstream functional consequence of reduced S100A9 zinc affinity not resolved
    • Whether site occupancy is regulated in vivo unknown
  3. 2022 Medium

    Tied METTL9 to a disease context by showing mitochondrial localization and dependence of Complex I activity on METTL9 in metastatic gastric cancer cells.

    Evidence shRNA knockdown, subcellular fractionation, Complex I assay, migration/invasion assays

    PMID:35402738

    Open questions at the time
    • Does not confirm methylation of a specific mitochondrial substrate in this system
    • Single lab, single cancer cell context
  4. 2023 High

    Resolved the structural basis of HxH recognition, explaining why the second histidine is methylated and how the aspartate and substrate pocket enforce specificity and directional processing.

    Evidence X-ray crystallography of METTL9-substrate complex with active-site mutagenesis

    PMID:37398635

    Open questions at the time
    • Conformational dynamics during processive methylation not captured
    • Determinants of full-length protein substrate selection beyond the peptide motif unclear
  5. 2023 Medium

    Connected METTL9 to ferroptosis control via SLC7A11, identifying a tumor-promoting axis in hepatocellular carcinoma.

    Evidence shRNA knockdown/overexpression, western blotting, viability/invasion assays, PDX tumor models

    PMID:38017014

    Open questions at the time
    • Whether the SLC7A11 effect requires catalytic methylation not established here
    • Direct substrate mediating SLC7A11 regulation not identified
  6. 2024 Low

    Extended METTL9 substrates to a signaling regulator by linking it to histidine methylation of NF-κB RELA and suppression of NLRP3-driven neuronal pyroptosis.

    Evidence MPTP Parkinson's model, METTL9 deficiency, luciferase/ChIP for NF-κB-NLRP3 promoter binding, TUNEL/flow cytometry

    PMID:39072406

    Open questions at the time
    • Direct biochemical validation of RELA histidine methylation is limited
    • Methylation site on RELA not mapped
    • Single lab, indirect evidence
  7. 2025 Medium

    Identified SLC39A7 (His45/His49) as a methylation substrate and placed METTL9 upstream of PERK/ATF4-SLC7A11 in ferroptosis and redox control during adipogenesis.

    Evidence In vitro/in vivo methyltransferase assays, site-specific mutagenesis, ROS measurement, adipogenic differentiation, OVX mouse model

    PMID:40414869

    Open questions at the time
    • Mechanistic link from SLC39A7 methylation to PERK/ATF4 activation not fully dissected
    • Single lab
  8. 2025 High

    Revealed a catalysis-independent function: METTL9 maintains Golgi/secretory pathway integrity and is required for vertebrate neural development, separating its developmental role from methyltransferase activity.

    Evidence ESC KO, degron, and catalytically inactive knock-in lines, neural differentiation, multi-omics, Xenopus knockdown, Co-IP with transport/Golgi regulators, Golgi imaging

    PMID:40745158

    Open questions at the time
    • Identity of the functionally critical transport/Golgi partners not specified
    • Molecular mechanism by which METTL9 preserves Golgi integrity unknown
  9. 2025 Medium

    Provided reciprocal physical evidence that METTL9 binds and stabilizes SLC7A11, defining a GPX4-independent route to ferroptosis modulation in HCC.

    Evidence Reciprocal Co-IP, RNA-seq, CUT&Tag, knockdown/overexpression, in vivo tumor models

    PMID:40523929

    Open questions at the time
    • Whether stabilization depends on histidine methylation of SLC7A11 or a non-catalytic interaction is unresolved
    • Single lab
  10. 2025 High

    Showed evolutionary conservation of HxH methyltransferase activity across eukaryotic orthologues with divergent substrate specificities, mapping structural determinants of flanking-residue tolerance.

    Evidence In vitro methyltransferase assays, peptide array profiling, X-ray structure of Ostreococcus tauri METTL9

    PMID:40451431

    Open questions at the time
    • Physiological substrates of non-human orthologues not defined
    • Functional significance of specificity divergence unknown
  11. 2026 High

    Delivered a first-in-class selective METTL9 inhibitor binding the SAM pocket, confirming the catalytic site and enabling chemical interrogation of histidine methylation in cells.

    Evidence Enzymatic IC50, X-ray of inhibitor-enzyme complex, selectivity panel, cellular target engagement, global 1-MH quantification by MS

    PMID:41870122

    Open questions at the time
    • In vivo efficacy and selectivity not addressed
    • Effect on individual substrate methylation events not parsed

Open questions

Synthesis pass · forward-looking unresolved questions
  • How METTL9's catalytic (HxH histidine methylation) and non-catalytic (Golgi/secretory) functions are partitioned across tissues and disease contexts, and whether its ferroptosis and signaling roles require methylation, remains unresolved.
  • No unified model linking enzymatic activity to the Golgi-integrity phenotype
  • Whether SLC7A11 stabilization and RELA effects are methylation-dependent unclear
  • Comprehensive in vivo substrate map lacking

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0016740 transferase activity 6 GO:0140096 catalytic activity, acting on a protein 4
Localization
GO:0005739 mitochondrion 1 GO:0005794 Golgi apparatus 1
Pathway
R-HSA-392499 Metabolism of proteins 3 R-HSA-5357801 Programmed Cell Death 3

Evidence

Reading pass · 11 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2021 METTL9 is a broad-specificity methyltransferase that mediates formation of the majority of 1-methylhistidine (1MH) present in mouse and human proteomes, requiring a His-x-His (HxH) motif where 'x' is preferably a small amino acid. Substrates include S100A9 and NDUFB3 (subunit of mitochondrial respiratory Complex I). METTL9-mediated methylation enhances respiration via Complex I, and 1MH in an HxH-containing peptide reduces zinc binding affinity. In vitro methyltransferase assays, LC-MS/MS detection of 1MH, knockout mouse models, genetic deletion with Complex I activity assay, zinc-binding affinity measurements Nature communications High 33563959
2021 METTL9 catalyzes Nπ-methylhistidine (N1-methylhistidine) formation on S100A9 at His-107 in vivo and in vitro. Methylation at His-107 overlaps with S100A9's zinc-binding site, attenuating its zinc affinity. METTL9 does not affect heterodimer formation of S100A9 with S100A8. siRNA screening coupled with methylhistidine analysis using LC-tandem MS; in vitro methyltransferase assay; zinc-binding affinity measurements The Journal of biological chemistry High 34562450
2023 Crystal/cryo structure of METTL9 in complex with substrate revealed it specifically methylates the second histidine of the HxH motif while the first histidine serves as a recognition signature. The small 'x' residue is confined within the substrate pocket. An aspartate residue stabilizes the N3 atom of the histidine imidazole ring, presenting the N1 atom to SAM for methylation. METTL9 exhibits preferred consecutive 'C-to-N' directional methylation of tandem HxH repeats. X-ray crystallography of METTL9-substrate complex; biochemical methyltransferase assays; mutagenesis of active-site residues Cell insight High 37398635
2022 In metastatic gastric cancer cells, METTL9 protein is predominantly localized in mitochondria, and METTL9 knockdown significantly reduces mitochondrial Complex I activity, inhibiting cell migration and invasion. shRNA knockdown; subcellular fractionation/localization; Complex I activity assay; migration and invasion assays in vitro Biochemistry and biophysics reports Medium 35402738
2023 METTL9 knockdown in hepatocellular carcinoma cells reduces SLC7A11 expression (a key suppressor of ferroptosis), promoting ferroptosis and inhibiting HCC progression in vitro and in vivo. shRNA knockdown; overexpression; western blotting; cell viability/migration/invasion assays; PDX tumor growth experiments Cell death discovery Medium 38017014
2025 METTL9 mediates N1-histidine methylation of the zinc transporter SLC39A7 at His45 and His49. This methylation suppresses ferroptosis through the PERK/ATF4 signaling pathway and downstream SLC7A11, which transports cystine for glutathione synthesis, thereby reducing intracellular ROS and inhibiting adipogenic differentiation of mesenchymal stem cells. In vitro and in vivo methyltransferase assays; site-specific mutagenesis (His45, His49); western blotting; ROS measurement; adipogenic differentiation assays; OVX mouse model Molecular medicine (Cambridge, Mass.) Medium 40414869
2025 METTL9 sustains vertebrate neural development primarily via non-catalytic functions. METTL9 interacts with key regulators of cellular transport, endocytosis, and Golgi integrity; in Mettl9 KO cells, the Golgi becomes fragmented. This developmental role occurs through modulation of the secretory pathway and is largely independent of its histidine methyltransferase catalytic activity. Mouse embryonic stem cell KO, inducible degron, and catalytically inactive knock-in lines; neural differentiation assays; multi-omics; Xenopus laevis mettl9 knockdown; co-immunoprecipitation with transport/Golgi regulators; Golgi morphology imaging Nature communications High 40745158
2024 METTL9 promotes histidine methylation of NF-κB RELA, resulting in inhibition of NLRP3 epigenetic transcription and suppression of neuronal pyroptosis in a Parkinson's disease mouse model. MPTP mouse model; electroacupuncture treatment; METTL9 deficiency; luciferase and ChIP assays for NF-κB binding to NLRP3 promoter; western blotting; TUNEL/flow cytometry Critical reviews in eukaryotic gene expression Low 39072406
2025 Co-immunoprecipitation confirmed that METTL9 binds to SLC7A11, enhancing its stability and reducing its degradation, thus modulating ferroptosis in hepatocellular carcinoma independently of the classical GPX4 pathway. Co-immunoprecipitation (Co-IP); RNA sequencing; CUT&Tag; overexpression and knockdown experiments; in vivo mouse tumor models NPJ precision oncology Medium 40523929
2025 METTL9 orthologues across eukaryotes retain in vitro methyltransferase activity on HxH-containing substrates (e.g., ARMC6 and DNAJB12), but show distinct substrate specificities. The X-ray structure of Ostreococcus tauri METTL9 revealed structural differences from human METTL9 that may explain its distinct substrate specificity. Tolerance for substitutions at the HxH flanking positions varies among orthologues (Hs > Dm > Ot). In vitro methyltransferase assays; peptide array substrate specificity profiling; X-ray crystallography of OtMETTL9 The Journal of biological chemistry High 40451431
2026 A first-in-class inhibitor (METTL9i) binds within the SAM binding pocket of METTL9 with IC50 of 0.067 µM and inhibits METTL9 with selectivity over other methyltransferases. In cells, METTL9i engages METTL9 and reduces global 1-methylhistidine levels, confirming the SAM pocket as the catalytic site. Enzymatic inhibition assay (IC50 measurement); X-ray crystallography of METTL9i-METTL9 complex; cellular target engagement assay; global 1-MH quantification by MS Angewandte Chemie (International ed. in English) High 41870122

Source papers

Stage 0 corpus · 15 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2021 The methyltransferase METTL9 mediates pervasive 1-methylhistidine modification in mammalian proteomes. Nature communications 90 33563959
2023 METTL9-SLC7A11 axis promotes hepatocellular carcinoma progression through ferroptosis inhibition. Cell death discovery 24 38017014
2023 Circular RNA METTL9 contributes to neuroinflammation following traumatic brain injury by complexing with astrocytic SND1. Journal of neuroinflammation 22 36803376
2021 siRNA screening identifies METTL9 as a histidine Nπ-methyltransferase that targets the proinflammatory protein S100A9. The Journal of biological chemistry 19 34562450
2023 Molecular basis for protein histidine N1-specific methylation of the "His-x-His" motifs by METTL9. Cell insight 16 37398635
2023 METTL9 derived circular RNA circ-METTL9 sponges miR-551b-5p to accelerate colorectal cancer progression by upregulating CDK6. Carcinogenesis 12 37158456
2022 Elevated METTL9 is associated with peritoneal dissemination in human scirrhous gastric cancers. Biochemistry and biophysics reports 11 35402738
2025 METTL9 mediated N1-Histidine methylation of SLC39A7 confers ferroptosis resistance and inhibits adipogenic differentiation in mesenchymal stem cells. Molecular medicine (Cambridge, Mass.) 7 40414869
2025 METTL9 sustains vertebrate neural development primarily via non-catalytic functions. Nature communications 4 40745158
2025 Orthologues of the human protein histidine methyltransferase METTL9 display distinct substrate specificities. The Journal of biological chemistry 3 40451431
2024 Electroacupuncture Alleviates Parkinson's Disease by Promoting METTL9-Catalyzed Histidine Methylation of Nuclear Factor-κВ. Critical reviews in eukaryotic gene expression 2 39072406
2025 Targeting SIX2 as a novel sensitization strategy of sorafenib treatment on advanced hepatocellular carcinoma through modulating METTL9-SLC7A11 axis. NPJ precision oncology 1 40523929
2026 METTL9 tests Candida's mettle by limiting metal acquisition. Cell host & microbe 0 41539299
2026 Discovery of a Potent and Selective Cell-Active Inhibitor of Histidine-N1 Methyltransferase METTL9. Angewandte Chemie (International ed. in English) 0 41870122
2026 The Protein Histidine Methyltransferase METTL9-From Mechanism to Biological Function. Life (Basel, Switzerland) 0 41900964

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