{"gene":"BHMT","run_date":"2026-06-09T22:02:44","timeline":{"discoveries":[{"year":2008,"finding":"BHMT variant allozymes expressed in COS-1 cells showed no significant differences from wild type in enzyme activity or immunoreactive protein levels, but had statistically significant differences in apparent Km values, indicating altered substrate binding. BHMT2 protein could not be functionally expressed alone but was stabilized by co-transfection with BHMT and co-precipitated with BHMT, indicating a direct physical interaction between the two paralogs.","method":"COS-1 cell expression of variant allozymes, enzyme activity assays, immunoreactive protein quantification, Km determination, co-transfection and co-immunoprecipitation","journal":"Molecular genetics and metabolism","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal functional assays plus co-IP in a single lab with multiple orthogonal methods","pmids":["18457970"],"is_preprint":false},{"year":2011,"finding":"Full-length endogenous BHMT (45 kDa cytosolic enzyme) is cleaved in an autophagy-dependent (3-methyladenine-sensitive) manner in isolated rat hepatocytes, generating a novel N-terminal 10-kDa fragment (p10). The cleavage is performed by the asparaginyl proteinase legumain (inhibitable by AJN-230) and occurs primarily in amphisomes rather than lysosomes. This makes BHMT a useful probe/reporter for autophagic flux.","method":"Isolated rat hepatocytes, 3-methyladenine inhibition, specific legumain inhibitor (AJN-230), mass spectrometry identification of p10 fragment, leupeptin protection, asparagine inhibition of amphisome-lysosome fusion","journal":"Autophagy","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro hepatocyte assay with multiple specific inhibitors, MS characterization, and multiple orthogonal controls","pmids":["21610319"],"is_preprint":false},{"year":2011,"finding":"A splicing variant of BHMT exon 4 generates a frameshift with a premature termination codon in exon 5, resulting in complete loss of BHMT enzymatic activity and undetectable protein expression in HepG2 hepatocellular carcinoma cells and 5 of 6 tumor samples; this variant is absent from normal adult and fetal liver.","method":"RT-qPCR, exon junction analysis and sequencing, Western blot, immunohistochemistry, enzyme activity assay in HepG2 cells and tumor tissue","journal":"The international journal of biochemistry & cell biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal methods (sequencing, western blot, IHC, enzymatic activity) confirming loss-of-function mechanism","pmids":["22138536"],"is_preprint":false},{"year":2011,"finding":"Depletion of Bhmt in zebrafish morphants causes hypoplasia of liver, exocrine pancreas, and intestine, but paradoxically increases β-cell number. Genetic epistasis experiments showed that Bhmt depletion elevates sonic hedgehog (shh) transcript levels, and Bhmt depletion in Shh-deficient (syu) mutants failed to rescue the isletless phenotype, placing Shh downstream of Bhmt in the pathway controlling β-cell development.","method":"Antisense morpholino knockdown in zebrafish, genetic epistasis with syu (shh-null) mutant, qRT-PCR for shh transcripts, histological analysis of organ development","journal":"Endocrinology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis in an ortholog model with multiple phenotypic readouts and replicated pathway placement","pmids":["21952238"],"is_preprint":false},{"year":2015,"finding":"The GST-BHMT fusion reporter is cleaved in lysosomes in an autophagy-dependent manner. Under nutrient-rich conditions, proteasome inhibition induces GST-BHMT processing through a mechanism distinct from starvation-induced autophagy: it does not require MTOR or PRKAA/AMPK, but requires cargo receptors SQSTM1/p62 and NBR1, and depends on ER stress signaling via ERN1/IRE1 and MAPK8/JNK1 (but not XBP1), regulating BCL2-BECN1 dissociation. Additionally, the multimerization domain of GST-BHMT is required for proteasome inhibition-induced processing but is dispensable for starvation-induced processing.","method":"GST-BHMT reporter fragmentation assay, pharmacological and genetic proteasome inhibition, siRNA knockdown of MTOR, AMPK, SQSTM1, NBR1, ERN1, MAPK8, XBP1, domain deletion mutagenesis","journal":"Autophagy","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution-style reporter assay with mutagenesis and multiple genetic perturbations identifying a distinct pathway","pmids":["25984893"],"is_preprint":false},{"year":2017,"finding":"Bhmt-null mice develop elevated S-adenosylhomocysteine concentrations and preneoplastic foci in the liver (increased placental GST and CK8-18 activity starting at 12 weeks). Whole-genome methylation analysis identified differentially methylated CpGs leading to repression of Iqgap2 and F2rl2 genes, indicating that BHMT activity is required to maintain adequate DNA methylation and normal expression of these loci in liver.","method":"Bhmt-null mouse model, whole-genome bisulfite sequencing, RNA expression profiling, immunohistochemistry for preneoplastic markers, metabolite measurement (S-adenosylhomocysteine)","journal":"FASEB journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — defined KO phenotype with multiple orthogonal methods (metabolomics, epigenomics, transcriptomics, histology) in a single rigorous study","pmids":["28179424"],"is_preprint":false},{"year":2021,"finding":"BHMT is present in the nucleus of oligodendrocytes (confirmed by immunocytochemistry in human MO3.13 cell line, primary rat oligodendrocytes, and MS postmortem tissue). BHMT expression is increased ~2-fold after oxidative insult. Chromatin fractionation showed direct interaction of BHMT on chromatin, and co-IP demonstrated interaction between BHMT and DNMT3a. Betaine administration increased both histone methyltransferase and DNA methyltransferase activity, and this effect was dependent on BHMT expression (abolished by siRNA knockdown), establishing BHMT as a chromatin-associated regulator of epigenetic marks in oligodendrocytes.","method":"Immunocytochemistry, chromatin fractionation, co-immunoprecipitation, qRT-PCR, siRNA knockdown, histone and DNA methyltransferase activity assays","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus functional siRNA rescue, single lab, multiple orthogonal methods","pmids":["33975330"],"is_preprint":false},{"year":2023,"finding":"After irradiation, SDC1 transports TGM2 from the cell membrane into the cytoplasm and to lysosomes via binding to FLOT1; TGM2 then recognizes BHMT on autophagosomes to coordinate the encounter between autophagosomes and lysosomes. This SDC1-TGM2-FLOT1-BHMT complex maintains autophagic flux in irradiated GBM cells and enhances radioresistance.","method":"Co-IP, immunofluorescence, mRFP-GFP-LC3 reporter, transmission electron microscopy, flow cytometry, colony formation, Western blot, qPCR in radioresistant GBM cells","journal":"Theranostics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP identifying complex plus functional autophagy reporter assay, single lab with multiple orthogonal methods","pmids":["37441590"],"is_preprint":false},{"year":2023,"finding":"In a monkey hydroxynonenal-injection model of hepatocyte injury, BHMT protein was carbonylated (~2-fold increase by proteomics) and showed increased cleavage without upregulation. Impaired rough ER caused deficient BHMT synthesis, contributing to hepatic steatosis consistent with BHMT's role in phosphatidylcholine metabolism via choline pathway.","method":"Proteomics (DIA), Western blot, immunohistochemistry, electron microscopy in monkey liver model","journal":"Nutrients","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single animal model study, no direct mechanistic rescue experiments for BHMT specifically","pmids":["37111122"],"is_preprint":false},{"year":2024,"finding":"Betaine upregulates BHMT expression, leading to increased NADPH production. Elevated NADPH in turn upregulates FTO expression. FTO then reduces m6A methylation in the CDS of Ppargc1α (PGC1α) mRNA, increasing PGC1α expression and inhibiting hepatic lipid accumulation, placing BHMT upstream in a betaine→BHMT→NADPH→FTO→m6A-PGC1α axis.","method":"Cell-based assays, siRNA knockdown, Western blot, m6A methylation quantification, gene expression analysis in NAFLD models","journal":"The Journal of nutritional biochemistry","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, pathway placement inferred from knockdown and expression data without direct biochemical reconstitution","pmids":["39154792"],"is_preprint":false},{"year":2026,"finding":"BHMT overexpression in renal cells reduces ROS production and apoptosis under hypoxia/reoxygenation by enhancing SAM synthesis, which increases DNMT activity, leading to methylation of the NOX4 promoter and suppression of NOX4 transcription and expression. Rescue assays confirmed BHMT-mediated protection is dependent on NOX4 downregulation.","method":"RT-qPCR, Western blot, MTT assay, flow cytometry, ROS assays, promoter methylation analysis, BHMT overexpression and rescue experiments in vitro and in vivo IRI model","journal":"Archives of biochemistry and biophysics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods including rescue experiments establishing BHMT→SAM→DNMT→NOX4 methylation axis, single lab","pmids":["41643883"],"is_preprint":false},{"year":2026,"finding":"In murine hepatocytes, BRCA1 deficiency suppresses Bhmt expression. Concomitant knockdown of BRCA1 and BHMT in human hepatoma cells produced additive accumulation of DNA double-strand breaks and increased cell death susceptibility. BHMT repression under BRCA1-deficiency was associated with downregulation of choline metabolism (confirmed by metabolomics), identifying BHMT as a mediator of BRCA1's effects on choline metabolism and DNA damage response.","method":"Single nuclear RNA sequencing, siRNA knockdown in human hepatoma cells, DNA damage assays (DSBs), metabolomics in Brca1-deficient mice","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — combinatorial knockdown with defined phenotypic readout plus metabolomics, single lab with multiple orthogonal methods","pmids":["41781465"],"is_preprint":false},{"year":2016,"finding":"Betaine supplementation in MCD diet-fed rats reversed the reduction of methionine and SAM and elevation of homocysteine by inducing BHMT and MAT expression. In H4IIE cells (which express BHMT), betaine prevented homocysteine-induced reduction of pAMPK/pACC/pSREBP-1c/pLKB1, but not in HepG2 cells (which lack BHMT), demonstrating that BHMT expression is required for the betaine-mediated activation of the AMPK pathway and anti-lipogenic effect.","method":"Rat dietary model, cell-based assays in H4IIE and HepG2 cells, Western blot for phosphorylated signaling proteins, BHMT and MAT protein expression, homocysteine and SAM measurement","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cell comparison exploiting natural BHMT expression difference as a functional control, multiple signaling readouts, single lab","pmids":["27320863"],"is_preprint":false},{"year":2023,"finding":"In Bhmt-KO mice, deletion of BHMT in the liver leads to homocysteine-induced ER stress, activation of hepatic transcription factor CREBH, and increased hepatic and plasma FGF21, which in turn promotes adipose browning and atrophy. This establishes a liver-to-adipose signaling axis downstream of BHMT loss.","method":"Bhmt-KO mouse model (C57Bl6/J background), histological analysis, gene expression profiling, metabolite/hormone measurements (Hcy, FGF21), ER stress marker analysis","journal":"Heliyon","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — defined KO model with molecular pathway tracing and multiple readouts, single lab","pmids":["36755585"],"is_preprint":false}],"current_model":"BHMT is a cytosolic hepatic enzyme that catalyzes betaine-dependent remethylation of homocysteine to methionine (producing dimethylglycine and SAM); it is processed in amphisomes by the asparaginyl proteinase legumain during autophagy; it forms a complex with SDC1-TGM2-FLOT1 on autophagosomes to facilitate autophagosome-lysosome fusion; it localizes to the nucleus in oligodendrocytes where it associates with chromatin and DNMT3a to regulate histone and DNA methylation; it promotes SAM synthesis to suppress NOX4 via promoter methylation; its loss in liver elevates S-adenosylhomocysteine, alters DNA methylation at specific loci (including Iqgap2 and F2rl2), and triggers FGF21-mediated adipose browning via homocysteine-induced ER stress; and in zebrafish, Bhmt controls β-cell development by acting upstream of Sonic Hedgehog signaling."},"narrative":{"mechanistic_narrative":"BHMT is a cytosolic hepatic enzyme that catalyzes betaine-dependent remethylation of homocysteine to methionine, thereby controlling the supply of S-adenosylmethionine and the cell's transmethylation capacity [PMID:28179424, PMID:27320863]. Loss of BHMT in liver elevates S-adenosylhomocysteine, alters CpG methylation to repress loci including Iqgap2 and F2rl2, and drives preneoplastic foci, establishing that BHMT activity is required to maintain normal DNA methylation in hepatocytes [PMID:28179424]. Beyond its housekeeping methyl-metabolism role, BHMT acts directly in epigenetic regulation: in oligodendrocytes it localizes to the nucleus, associates with chromatin and DNMT3a, and is required for betaine-induced histone and DNA methyltransferase activity [PMID:33975330], and in renal cells it raises SAM to enhance DNMT-mediated methylation of the NOX4 promoter, suppressing NOX4 and limiting ROS-driven apoptosis [PMID:41643883]. BHMT loss links methyl metabolism to broader physiology, triggering homocysteine-induced ER stress, CREBH activation, and FGF21-mediated adipose browning [PMID:36755585], while its expression is required for betaine-dependent AMPK pathway activation and anti-lipogenic effects [PMID:27320863]. Independently of catalysis, full-length BHMT is cleaved by the asparaginyl proteinase legumain in amphisomes in an autophagy-dependent manner, making it a reporter of autophagic flux [PMID:21610319], and in irradiated glioblastoma cells it is recognized on autophagosomes within an SDC1-TGM2-FLOT1 complex to promote autophagosome-lysosome fusion [PMID:37441590]. In zebrafish, Bhmt acts upstream of Sonic Hedgehog signaling to control endoderm-derived organ and β-cell development [PMID:21952238].","teleology":[{"year":2008,"claim":"Establishing that BHMT sequence variants alter substrate binding and that BHMT physically interacts with its paralog BHMT2 clarified the enzyme's functional determinants and quaternary context.","evidence":"COS-1 expression of variant allozymes with activity/Km assays and co-IP with BHMT2","pmids":["18457970"],"confidence":"Medium","gaps":["No structural model of the BHMT-BHMT2 complex","Physiological consequence of altered Km not assessed in vivo"]},{"year":2011,"claim":"The discovery that BHMT is cleaved by legumain in amphisomes during autophagy revealed an unexpected, catalysis-independent role for BHMT as a proteolytic substrate and a tool for monitoring autophagic flux.","evidence":"Isolated rat hepatocytes with 3-MA, legumain inhibitor AJN-230, and MS identification of the p10 fragment","pmids":["21610319"],"confidence":"High","gaps":["Functional consequence of cleavage for methionine metabolism unknown","Whether p10 fragment has activity not established"]},{"year":2011,"claim":"A frameshifting splice variant abolishing BHMT expression in hepatocellular carcinoma defined a loss-of-function event tied to liver tumor tissue.","evidence":"Exon junction sequencing, Western blot, IHC, and activity assays in HepG2 cells and tumors","pmids":["22138536"],"confidence":"High","gaps":["Causal role of BHMT loss in tumorigenesis not tested","Mechanism driving aberrant splicing unknown"]},{"year":2011,"claim":"Genetic epistasis in zebrafish placed Bhmt upstream of Sonic Hedgehog in a developmental pathway controlling endodermal organ and β-cell formation, extending BHMT's relevance beyond metabolism.","evidence":"Morpholino knockdown with syu (shh-null) epistasis and qRT-PCR for shh","pmids":["21952238"],"confidence":"High","gaps":["Molecular link between BHMT metabolism and shh transcription unresolved","Relevance to mammalian development not shown"]},{"year":2015,"claim":"Dissecting GST-BHMT reporter processing distinguished proteasome-inhibition-induced autophagy (requiring SQSTM1/NBR1, ERN1/IRE1, JNK, and the multimerization domain) from starvation-induced autophagy, refining BHMT's use as a pathway-specific flux reporter.","evidence":"GST-BHMT fragmentation assay with siRNA knockdowns and domain-deletion mutagenesis","pmids":["25984893"],"confidence":"High","gaps":["Whether endogenous BHMT follows the same route not confirmed","Significance of multimerization for native BHMT function unclear"]},{"year":2016,"claim":"Comparing BHMT-expressing and BHMT-deficient hepatocyte lines showed BHMT is required for betaine-driven AMPK pathway activation, linking its remethylation activity to anti-lipogenic signaling.","evidence":"MCD-diet rat model plus H4IIE vs HepG2 cell comparison with phospho-signaling Western blots","pmids":["27320863"],"confidence":"Medium","gaps":["Direct mechanism connecting SAM/homocysteine to LKB1/AMPK not defined","Confound from other cell-line differences besides BHMT"]},{"year":2017,"claim":"The Bhmt-null mouse demonstrated that BHMT activity is required to sustain hepatic DNA methylation, with its loss raising S-adenosylhomocysteine and causing locus-specific hypomethylation and preneoplastic foci.","evidence":"Bhmt-null mice with whole-genome bisulfite sequencing, transcriptomics, metabolomics, and histology","pmids":["28179424"],"confidence":"High","gaps":["Causal chain from Iqgap2/F2rl2 repression to neoplasia not established","Which methyltransferases are limited by SAH not pinpointed"]},{"year":2021,"claim":"Detection of nuclear, chromatin-associated BHMT interacting with DNMT3a in oligodendrocytes proposed a direct role for BHMT in epigenetic regulation beyond cytosolic methyl supply.","evidence":"Immunocytochemistry, chromatin fractionation, co-IP with DNMT3a, and siRNA-dependent methyltransferase activity assays","pmids":["33975330"],"confidence":"Medium","gaps":["Co-IP lacks reciprocal and structural validation","Mechanism of BHMT nuclear import unknown"]},{"year":2023,"claim":"Identifying BHMT as the autophagosomal partner recognized by TGM2 within an SDC1-TGM2-FLOT1 complex assigned BHMT a structural role in autophagosome-lysosome fusion underlying glioblastoma radioresistance.","evidence":"Co-IP, immunofluorescence, mRFP-GFP-LC3 reporter, and functional autophagy assays in radioresistant GBM cells","pmids":["37441590"],"confidence":"Medium","gaps":["Direct BHMT-TGM2 binding interface not mapped","Whether catalytic activity is required for fusion role unknown"]},{"year":2023,"claim":"Tracing the Bhmt-KO phenotype to homocysteine-induced ER stress, CREBH activation, and FGF21 release defined a liver-to-adipose signaling axis arising from BHMT loss.","evidence":"Bhmt-KO mice with histology, expression profiling, and hormone/metabolite measurements","pmids":["36755585"],"confidence":"Medium","gaps":["Direct trigger of CREBH by homocysteine not biochemically resolved","Contribution of other one-carbon perturbations not excluded"]},{"year":2026,"claim":"Rescue experiments established a BHMT→SAM→DNMT→NOX4-promoter-methylation axis by which BHMT overexpression suppresses NOX4 and protects renal cells from oxidative apoptosis.","evidence":"BHMT overexpression and NOX4 rescue assays with promoter methylation analysis in vitro and an IRI model","pmids":["41643883"],"confidence":"Medium","gaps":["Which DNMT methylates the NOX4 promoter not specified","Generality beyond renal context untested"]},{"year":2026,"claim":"Combinatorial knockdown linked BHMT to BRCA1's control of choline metabolism and DNA double-strand-break susceptibility, positioning BHMT as a mediator of BRCA1-dependent genome maintenance.","evidence":"snRNA-seq, dual BRCA1/BHMT knockdown with DSB assays, and metabolomics in Brca1-deficient mice","pmids":["41781465"],"confidence":"Medium","gaps":["Mechanism by which BRCA1 suppresses Bhmt expression unknown","Direct contribution of choline metabolism to DSB repair not demonstrated"]},{"year":null,"claim":"It remains unresolved how BHMT's cytosolic remethylation activity is mechanistically coupled to its proposed nuclear chromatin and autophagosomal-fusion roles, and whether these moonlighting functions require its catalytic activity.","evidence":"No timeline discovery reconciles the catalytic, nuclear, and autophagic functions in a single system","pmids":[],"confidence":"Low","gaps":["No structural basis for nuclear or autophagosomal localization","Catalysis-dependence of moonlighting functions untested","Trafficking signals directing BHMT to different compartments unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[5,12]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[1]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[6]},{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[6]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[5,12]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[1,4,7]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[5,6,10]}],"complexes":["SDC1-TGM2-FLOT1-BHMT autophagosomal complex"],"partners":["BHMT2","DNMT3A","TGM2","SDC1","FLOT1","LGMN"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q93088","full_name":"Betaine--homocysteine S-methyltransferase 1","aliases":[],"length_aa":406,"mass_kda":45.0,"function":"Involved in the regulation of homocysteine metabolism. Converts betaine and homocysteine to dimethylglycine and methionine, respectively. This reaction is also required for the irreversible oxidation of choline","subcellular_location":"Cytoplasm, cytosol; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q93088/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/BHMT","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/BHMT","total_profiled":1310},"omim":[{"mim_id":"605932","title":"BETAINE-HOMOCYSTEINE METHYLTRANSFERASE 2; BHMT2","url":"https://www.omim.org/entry/605932"},{"mim_id":"602888","title":"BETAINE-HOMOCYSTEINE METHYLTRANSFERASE; BHMT","url":"https://www.omim.org/entry/602888"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Cytosol","reliability":"Approved"}],"tissue_specificity":"Group enriched","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"kidney","ntpm":727.4},{"tissue":"liver","ntpm":1067.7}],"url":"https://www.proteinatlas.org/search/BHMT"},"hgnc":{"alias_symbol":["BHMT1"],"prev_symbol":[]},"alphafold":{"accession":"Q93088","domains":[{"cath_id":"3.20.20.330","chopping":"12-356_364-400","consensus_level":"high","plddt":94.2477,"start":12,"end":400}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q93088","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q93088-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q93088-F1-predicted_aligned_error_v6.png","plddt_mean":92.62},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=BHMT","jax_strain_url":"https://www.jax.org/strain/search?query=BHMT"},"sequence":{"accession":"Q93088","fasta_url":"https://rest.uniprot.org/uniprotkb/Q93088.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q93088/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q93088"}},"corpus_meta":[{"pmid":"12818402","id":"PMC_12818402","title":"Investigations of a common genetic variant in betaine-homocysteine methyltransferase (BHMT) in coronary artery disease.","date":"2003","source":"Atherosclerosis","url":"https://pubmed.ncbi.nlm.nih.gov/12818402","citation_count":67,"is_preprint":false},{"pmid":"11073719","id":"PMC_11073719","title":"Betaine-homocysteine methyltransferase (BHMT): genomic sequencing and relevance to hyperhomocysteinemia and vascular disease in humans.","date":"2000","source":"Molecular genetics and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/11073719","citation_count":60,"is_preprint":false},{"pmid":"18457970","id":"PMC_18457970","title":"Human betaine-homocysteine methyltransferase (BHMT) and BHMT2: common gene sequence variation and functional characterization.","date":"2008","source":"Molecular genetics and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/18457970","citation_count":59,"is_preprint":false},{"pmid":"12749058","id":"PMC_12749058","title":"Common variant in betaine-homocysteine methyltransferase (BHMT) and risk for spina bifida.","date":"2003","source":"American journal of medical genetics. 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meta-analysis.","date":"2012","source":"Genetic testing and molecular biomarkers","url":"https://pubmed.ncbi.nlm.nih.gov/22339686","citation_count":8,"is_preprint":false},{"pmid":"31826386","id":"PMC_31826386","title":"Genetic and epigenetic regulation of BHMT is associated with folate therapy efficacy in hyperhomocysteinaemia.","date":"2019","source":"Asia Pacific journal of clinical nutrition","url":"https://pubmed.ncbi.nlm.nih.gov/31826386","citation_count":8,"is_preprint":false},{"pmid":"27735840","id":"PMC_27735840","title":"Maternal Folate Status and the BHMT c.716G>A Polymorphism Affect the Betaine Dimethylglycine Pathway during Pregnancy.","date":"2016","source":"Nutrients","url":"https://pubmed.ncbi.nlm.nih.gov/27735840","citation_count":8,"is_preprint":false},{"pmid":"35956270","id":"PMC_35956270","title":"Association and Interaction Effect of BHMT Gene Polymorphisms and Maternal Dietary Habits with Ventricular Septal Defect in 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sciences","url":"https://pubmed.ncbi.nlm.nih.gov/37203835","citation_count":0,"is_preprint":false},{"pmid":"41895659","id":"PMC_41895659","title":"Targeting metabolic dysregulation in HCC: Development and validation of a dual-biomarker diagnostic model integrating ADH1C and BHMT.","date":"2026","source":"Clinics and research in hepatology and gastroenterology","url":"https://pubmed.ncbi.nlm.nih.gov/41895659","citation_count":0,"is_preprint":false},{"pmid":"41643883","id":"PMC_41643883","title":"BHMT Prevents renal ischemia/reperfusion injury via suppressing ROS-induced apoptosis by targeting NOX4.","date":"2026","source":"Archives of biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/41643883","citation_count":0,"is_preprint":false},{"pmid":"41781465","id":"PMC_41781465","title":"Paradoxical oncogenic effects of hepatic Brca1 through modulating Bhmt.","date":"2026","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/41781465","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.05.07.652674","title":"Marine community metabolomes in the eastern tropical North Pacific Oxygen Deficient Zone reveal glycine betaine as a metabolic link between Prochlorococcus and SAR11","date":"2025-05-12","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.07.652674","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":22941,"output_tokens":4062,"usd":0.064877,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":11991,"output_tokens":4134,"usd":0.081653,"stage2_stop_reason":"end_turn"},"total_usd":0.14653,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2008,\n      \"finding\": \"BHMT variant allozymes expressed in COS-1 cells showed no significant differences from wild type in enzyme activity or immunoreactive protein levels, but had statistically significant differences in apparent Km values, indicating altered substrate binding. BHMT2 protein could not be functionally expressed alone but was stabilized by co-transfection with BHMT and co-precipitated with BHMT, indicating a direct physical interaction between the two paralogs.\",\n      \"method\": \"COS-1 cell expression of variant allozymes, enzyme activity assays, immunoreactive protein quantification, Km determination, co-transfection and co-immunoprecipitation\",\n      \"journal\": \"Molecular genetics and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal functional assays plus co-IP in a single lab with multiple orthogonal methods\",\n      \"pmids\": [\"18457970\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Full-length endogenous BHMT (45 kDa cytosolic enzyme) is cleaved in an autophagy-dependent (3-methyladenine-sensitive) manner in isolated rat hepatocytes, generating a novel N-terminal 10-kDa fragment (p10). The cleavage is performed by the asparaginyl proteinase legumain (inhibitable by AJN-230) and occurs primarily in amphisomes rather than lysosomes. This makes BHMT a useful probe/reporter for autophagic flux.\",\n      \"method\": \"Isolated rat hepatocytes, 3-methyladenine inhibition, specific legumain inhibitor (AJN-230), mass spectrometry identification of p10 fragment, leupeptin protection, asparagine inhibition of amphisome-lysosome fusion\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro hepatocyte assay with multiple specific inhibitors, MS characterization, and multiple orthogonal controls\",\n      \"pmids\": [\"21610319\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"A splicing variant of BHMT exon 4 generates a frameshift with a premature termination codon in exon 5, resulting in complete loss of BHMT enzymatic activity and undetectable protein expression in HepG2 hepatocellular carcinoma cells and 5 of 6 tumor samples; this variant is absent from normal adult and fetal liver.\",\n      \"method\": \"RT-qPCR, exon junction analysis and sequencing, Western blot, immunohistochemistry, enzyme activity assay in HepG2 cells and tumor tissue\",\n      \"journal\": \"The international journal of biochemistry & cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal methods (sequencing, western blot, IHC, enzymatic activity) confirming loss-of-function mechanism\",\n      \"pmids\": [\"22138536\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Depletion of Bhmt in zebrafish morphants causes hypoplasia of liver, exocrine pancreas, and intestine, but paradoxically increases β-cell number. Genetic epistasis experiments showed that Bhmt depletion elevates sonic hedgehog (shh) transcript levels, and Bhmt depletion in Shh-deficient (syu) mutants failed to rescue the isletless phenotype, placing Shh downstream of Bhmt in the pathway controlling β-cell development.\",\n      \"method\": \"Antisense morpholino knockdown in zebrafish, genetic epistasis with syu (shh-null) mutant, qRT-PCR for shh transcripts, histological analysis of organ development\",\n      \"journal\": \"Endocrinology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis in an ortholog model with multiple phenotypic readouts and replicated pathway placement\",\n      \"pmids\": [\"21952238\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The GST-BHMT fusion reporter is cleaved in lysosomes in an autophagy-dependent manner. Under nutrient-rich conditions, proteasome inhibition induces GST-BHMT processing through a mechanism distinct from starvation-induced autophagy: it does not require MTOR or PRKAA/AMPK, but requires cargo receptors SQSTM1/p62 and NBR1, and depends on ER stress signaling via ERN1/IRE1 and MAPK8/JNK1 (but not XBP1), regulating BCL2-BECN1 dissociation. Additionally, the multimerization domain of GST-BHMT is required for proteasome inhibition-induced processing but is dispensable for starvation-induced processing.\",\n      \"method\": \"GST-BHMT reporter fragmentation assay, pharmacological and genetic proteasome inhibition, siRNA knockdown of MTOR, AMPK, SQSTM1, NBR1, ERN1, MAPK8, XBP1, domain deletion mutagenesis\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution-style reporter assay with mutagenesis and multiple genetic perturbations identifying a distinct pathway\",\n      \"pmids\": [\"25984893\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Bhmt-null mice develop elevated S-adenosylhomocysteine concentrations and preneoplastic foci in the liver (increased placental GST and CK8-18 activity starting at 12 weeks). Whole-genome methylation analysis identified differentially methylated CpGs leading to repression of Iqgap2 and F2rl2 genes, indicating that BHMT activity is required to maintain adequate DNA methylation and normal expression of these loci in liver.\",\n      \"method\": \"Bhmt-null mouse model, whole-genome bisulfite sequencing, RNA expression profiling, immunohistochemistry for preneoplastic markers, metabolite measurement (S-adenosylhomocysteine)\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — defined KO phenotype with multiple orthogonal methods (metabolomics, epigenomics, transcriptomics, histology) in a single rigorous study\",\n      \"pmids\": [\"28179424\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"BHMT is present in the nucleus of oligodendrocytes (confirmed by immunocytochemistry in human MO3.13 cell line, primary rat oligodendrocytes, and MS postmortem tissue). BHMT expression is increased ~2-fold after oxidative insult. Chromatin fractionation showed direct interaction of BHMT on chromatin, and co-IP demonstrated interaction between BHMT and DNMT3a. Betaine administration increased both histone methyltransferase and DNA methyltransferase activity, and this effect was dependent on BHMT expression (abolished by siRNA knockdown), establishing BHMT as a chromatin-associated regulator of epigenetic marks in oligodendrocytes.\",\n      \"method\": \"Immunocytochemistry, chromatin fractionation, co-immunoprecipitation, qRT-PCR, siRNA knockdown, histone and DNA methyltransferase activity assays\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus functional siRNA rescue, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"33975330\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"After irradiation, SDC1 transports TGM2 from the cell membrane into the cytoplasm and to lysosomes via binding to FLOT1; TGM2 then recognizes BHMT on autophagosomes to coordinate the encounter between autophagosomes and lysosomes. This SDC1-TGM2-FLOT1-BHMT complex maintains autophagic flux in irradiated GBM cells and enhances radioresistance.\",\n      \"method\": \"Co-IP, immunofluorescence, mRFP-GFP-LC3 reporter, transmission electron microscopy, flow cytometry, colony formation, Western blot, qPCR in radioresistant GBM cells\",\n      \"journal\": \"Theranostics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP identifying complex plus functional autophagy reporter assay, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"37441590\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In a monkey hydroxynonenal-injection model of hepatocyte injury, BHMT protein was carbonylated (~2-fold increase by proteomics) and showed increased cleavage without upregulation. Impaired rough ER caused deficient BHMT synthesis, contributing to hepatic steatosis consistent with BHMT's role in phosphatidylcholine metabolism via choline pathway.\",\n      \"method\": \"Proteomics (DIA), Western blot, immunohistochemistry, electron microscopy in monkey liver model\",\n      \"journal\": \"Nutrients\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single animal model study, no direct mechanistic rescue experiments for BHMT specifically\",\n      \"pmids\": [\"37111122\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Betaine upregulates BHMT expression, leading to increased NADPH production. Elevated NADPH in turn upregulates FTO expression. FTO then reduces m6A methylation in the CDS of Ppargc1α (PGC1α) mRNA, increasing PGC1α expression and inhibiting hepatic lipid accumulation, placing BHMT upstream in a betaine→BHMT→NADPH→FTO→m6A-PGC1α axis.\",\n      \"method\": \"Cell-based assays, siRNA knockdown, Western blot, m6A methylation quantification, gene expression analysis in NAFLD models\",\n      \"journal\": \"The Journal of nutritional biochemistry\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, pathway placement inferred from knockdown and expression data without direct biochemical reconstitution\",\n      \"pmids\": [\"39154792\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"BHMT overexpression in renal cells reduces ROS production and apoptosis under hypoxia/reoxygenation by enhancing SAM synthesis, which increases DNMT activity, leading to methylation of the NOX4 promoter and suppression of NOX4 transcription and expression. Rescue assays confirmed BHMT-mediated protection is dependent on NOX4 downregulation.\",\n      \"method\": \"RT-qPCR, Western blot, MTT assay, flow cytometry, ROS assays, promoter methylation analysis, BHMT overexpression and rescue experiments in vitro and in vivo IRI model\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods including rescue experiments establishing BHMT→SAM→DNMT→NOX4 methylation axis, single lab\",\n      \"pmids\": [\"41643883\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"In murine hepatocytes, BRCA1 deficiency suppresses Bhmt expression. Concomitant knockdown of BRCA1 and BHMT in human hepatoma cells produced additive accumulation of DNA double-strand breaks and increased cell death susceptibility. BHMT repression under BRCA1-deficiency was associated with downregulation of choline metabolism (confirmed by metabolomics), identifying BHMT as a mediator of BRCA1's effects on choline metabolism and DNA damage response.\",\n      \"method\": \"Single nuclear RNA sequencing, siRNA knockdown in human hepatoma cells, DNA damage assays (DSBs), metabolomics in Brca1-deficient mice\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — combinatorial knockdown with defined phenotypic readout plus metabolomics, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"41781465\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Betaine supplementation in MCD diet-fed rats reversed the reduction of methionine and SAM and elevation of homocysteine by inducing BHMT and MAT expression. In H4IIE cells (which express BHMT), betaine prevented homocysteine-induced reduction of pAMPK/pACC/pSREBP-1c/pLKB1, but not in HepG2 cells (which lack BHMT), demonstrating that BHMT expression is required for the betaine-mediated activation of the AMPK pathway and anti-lipogenic effect.\",\n      \"method\": \"Rat dietary model, cell-based assays in H4IIE and HepG2 cells, Western blot for phosphorylated signaling proteins, BHMT and MAT protein expression, homocysteine and SAM measurement\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cell comparison exploiting natural BHMT expression difference as a functional control, multiple signaling readouts, single lab\",\n      \"pmids\": [\"27320863\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In Bhmt-KO mice, deletion of BHMT in the liver leads to homocysteine-induced ER stress, activation of hepatic transcription factor CREBH, and increased hepatic and plasma FGF21, which in turn promotes adipose browning and atrophy. This establishes a liver-to-adipose signaling axis downstream of BHMT loss.\",\n      \"method\": \"Bhmt-KO mouse model (C57Bl6/J background), histological analysis, gene expression profiling, metabolite/hormone measurements (Hcy, FGF21), ER stress marker analysis\",\n      \"journal\": \"Heliyon\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — defined KO model with molecular pathway tracing and multiple readouts, single lab\",\n      \"pmids\": [\"36755585\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"BHMT is a cytosolic hepatic enzyme that catalyzes betaine-dependent remethylation of homocysteine to methionine (producing dimethylglycine and SAM); it is processed in amphisomes by the asparaginyl proteinase legumain during autophagy; it forms a complex with SDC1-TGM2-FLOT1 on autophagosomes to facilitate autophagosome-lysosome fusion; it localizes to the nucleus in oligodendrocytes where it associates with chromatin and DNMT3a to regulate histone and DNA methylation; it promotes SAM synthesis to suppress NOX4 via promoter methylation; its loss in liver elevates S-adenosylhomocysteine, alters DNA methylation at specific loci (including Iqgap2 and F2rl2), and triggers FGF21-mediated adipose browning via homocysteine-induced ER stress; and in zebrafish, Bhmt controls β-cell development by acting upstream of Sonic Hedgehog signaling.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"BHMT is a cytosolic hepatic enzyme that catalyzes betaine-dependent remethylation of homocysteine to methionine, thereby controlling the supply of S-adenosylmethionine and the cell's transmethylation capacity [#5, #12]. Loss of BHMT in liver elevates S-adenosylhomocysteine, alters CpG methylation to repress loci including Iqgap2 and F2rl2, and drives preneoplastic foci, establishing that BHMT activity is required to maintain normal DNA methylation in hepatocytes [#5]. Beyond its housekeeping methyl-metabolism role, BHMT acts directly in epigenetic regulation: in oligodendrocytes it localizes to the nucleus, associates with chromatin and DNMT3a, and is required for betaine-induced histone and DNA methyltransferase activity [#6], and in renal cells it raises SAM to enhance DNMT-mediated methylation of the NOX4 promoter, suppressing NOX4 and limiting ROS-driven apoptosis [#10]. BHMT loss links methyl metabolism to broader physiology, triggering homocysteine-induced ER stress, CREBH activation, and FGF21-mediated adipose browning [#13], while its expression is required for betaine-dependent AMPK pathway activation and anti-lipogenic effects [#12]. Independently of catalysis, full-length BHMT is cleaved by the asparaginyl proteinase legumain in amphisomes in an autophagy-dependent manner, making it a reporter of autophagic flux [#1], and in irradiated glioblastoma cells it is recognized on autophagosomes within an SDC1-TGM2-FLOT1 complex to promote autophagosome-lysosome fusion [#7]. In zebrafish, Bhmt acts upstream of Sonic Hedgehog signaling to control endoderm-derived organ and \\u03b2-cell development [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Establishing that BHMT sequence variants alter substrate binding and that BHMT physically interacts with its paralog BHMT2 clarified the enzyme's functional determinants and quaternary context.\",\n      \"evidence\": \"COS-1 expression of variant allozymes with activity/Km assays and co-IP with BHMT2\",\n      \"pmids\": [\"18457970\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model of the BHMT-BHMT2 complex\", \"Physiological consequence of altered Km not assessed in vivo\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"The discovery that BHMT is cleaved by legumain in amphisomes during autophagy revealed an unexpected, catalysis-independent role for BHMT as a proteolytic substrate and a tool for monitoring autophagic flux.\",\n      \"evidence\": \"Isolated rat hepatocytes with 3-MA, legumain inhibitor AJN-230, and MS identification of the p10 fragment\",\n      \"pmids\": [\"21610319\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of cleavage for methionine metabolism unknown\", \"Whether p10 fragment has activity not established\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"A frameshifting splice variant abolishing BHMT expression in hepatocellular carcinoma defined a loss-of-function event tied to liver tumor tissue.\",\n      \"evidence\": \"Exon junction sequencing, Western blot, IHC, and activity assays in HepG2 cells and tumors\",\n      \"pmids\": [\"22138536\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Causal role of BHMT loss in tumorigenesis not tested\", \"Mechanism driving aberrant splicing unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Genetic epistasis in zebrafish placed Bhmt upstream of Sonic Hedgehog in a developmental pathway controlling endodermal organ and \\u03b2-cell formation, extending BHMT's relevance beyond metabolism.\",\n      \"evidence\": \"Morpholino knockdown with syu (shh-null) epistasis and qRT-PCR for shh\",\n      \"pmids\": [\"21952238\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular link between BHMT metabolism and shh transcription unresolved\", \"Relevance to mammalian development not shown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Dissecting GST-BHMT reporter processing distinguished proteasome-inhibition-induced autophagy (requiring SQSTM1/NBR1, ERN1/IRE1, JNK, and the multimerization domain) from starvation-induced autophagy, refining BHMT's use as a pathway-specific flux reporter.\",\n      \"evidence\": \"GST-BHMT fragmentation assay with siRNA knockdowns and domain-deletion mutagenesis\",\n      \"pmids\": [\"25984893\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether endogenous BHMT follows the same route not confirmed\", \"Significance of multimerization for native BHMT function unclear\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Comparing BHMT-expressing and BHMT-deficient hepatocyte lines showed BHMT is required for betaine-driven AMPK pathway activation, linking its remethylation activity to anti-lipogenic signaling.\",\n      \"evidence\": \"MCD-diet rat model plus H4IIE vs HepG2 cell comparison with phospho-signaling Western blots\",\n      \"pmids\": [\"27320863\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct mechanism connecting SAM/homocysteine to LKB1/AMPK not defined\", \"Confound from other cell-line differences besides BHMT\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"The Bhmt-null mouse demonstrated that BHMT activity is required to sustain hepatic DNA methylation, with its loss raising S-adenosylhomocysteine and causing locus-specific hypomethylation and preneoplastic foci.\",\n      \"evidence\": \"Bhmt-null mice with whole-genome bisulfite sequencing, transcriptomics, metabolomics, and histology\",\n      \"pmids\": [\"28179424\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Causal chain from Iqgap2/F2rl2 repression to neoplasia not established\", \"Which methyltransferases are limited by SAH not pinpointed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Detection of nuclear, chromatin-associated BHMT interacting with DNMT3a in oligodendrocytes proposed a direct role for BHMT in epigenetic regulation beyond cytosolic methyl supply.\",\n      \"evidence\": \"Immunocytochemistry, chromatin fractionation, co-IP with DNMT3a, and siRNA-dependent methyltransferase activity assays\",\n      \"pmids\": [\"33975330\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Co-IP lacks reciprocal and structural validation\", \"Mechanism of BHMT nuclear import unknown\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identifying BHMT as the autophagosomal partner recognized by TGM2 within an SDC1-TGM2-FLOT1 complex assigned BHMT a structural role in autophagosome-lysosome fusion underlying glioblastoma radioresistance.\",\n      \"evidence\": \"Co-IP, immunofluorescence, mRFP-GFP-LC3 reporter, and functional autophagy assays in radioresistant GBM cells\",\n      \"pmids\": [\"37441590\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct BHMT-TGM2 binding interface not mapped\", \"Whether catalytic activity is required for fusion role unknown\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Tracing the Bhmt-KO phenotype to homocysteine-induced ER stress, CREBH activation, and FGF21 release defined a liver-to-adipose signaling axis arising from BHMT loss.\",\n      \"evidence\": \"Bhmt-KO mice with histology, expression profiling, and hormone/metabolite measurements\",\n      \"pmids\": [\"36755585\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct trigger of CREBH by homocysteine not biochemically resolved\", \"Contribution of other one-carbon perturbations not excluded\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Rescue experiments established a BHMT\\u2192SAM\\u2192DNMT\\u2192NOX4-promoter-methylation axis by which BHMT overexpression suppresses NOX4 and protects renal cells from oxidative apoptosis.\",\n      \"evidence\": \"BHMT overexpression and NOX4 rescue assays with promoter methylation analysis in vitro and an IRI model\",\n      \"pmids\": [\"41643883\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Which DNMT methylates the NOX4 promoter not specified\", \"Generality beyond renal context untested\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Combinatorial knockdown linked BHMT to BRCA1's control of choline metabolism and DNA double-strand-break susceptibility, positioning BHMT as a mediator of BRCA1-dependent genome maintenance.\",\n      \"evidence\": \"snRNA-seq, dual BRCA1/BHMT knockdown with DSB assays, and metabolomics in Brca1-deficient mice\",\n      \"pmids\": [\"41781465\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which BRCA1 suppresses Bhmt expression unknown\", \"Direct contribution of choline metabolism to DSB repair not demonstrated\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how BHMT's cytosolic remethylation activity is mechanistically coupled to its proposed nuclear chromatin and autophagosomal-fusion roles, and whether these moonlighting functions require its catalytic activity.\",\n      \"evidence\": \"No timeline discovery reconciles the catalytic, nuclear, and autophagic functions in a single system\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structural basis for nuclear or autophagosomal localization\", \"Catalysis-dependence of moonlighting functions untested\", \"Trafficking signals directing BHMT to different compartments unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [5, 12]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [6]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [5, 12]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [1, 4, 7]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [5, 6, 10]}\n    ],\n    \"complexes\": [\n      \"SDC1-TGM2-FLOT1-BHMT autophagosomal complex\"\n    ],\n    \"partners\": [\n      \"BHMT2\",\n      \"DNMT3a\",\n      \"TGM2\",\n      \"SDC1\",\n      \"FLOT1\",\n      \"LGMN\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}