{"gene":"MLF1","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":1996,"finding":"MLF1 is normally localized in the cytoplasm, whereas the NPM-MLF1 fusion protein is targeted to the nucleus and nucleolus; NPM trafficking signals direct MLF1 to an inappropriate cellular compartment in myeloid leukemia cells.","method":"Immunostaining of t(3;5)-positive leukemia cells and cell lines expressing wild-type MLF1 vs. NPM-MLF1","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct immunostaining localization with functional implication, single lab, two cell contexts (leukemia cells and cell lines)","pmids":["8570204"],"is_preprint":false},{"year":1999,"finding":"NPM-MLF1 fusion protein induces apoptosis; this requires the N-terminal domain of MLF1 and the NPM domain containing a nuclear localization signal. The NPM dimerization domain is also required. Co-expression of Bcl-2 rescues cells from NPM-MLF1-mediated cell death without altering the expression or localization of NPM-MLF1.","method":"Ectopic overexpression and deletion mutant analysis in K562 and NIH3T3 cells; co-expression with Bcl-2","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — deletion mutagenesis with defined apoptotic phenotype, single lab, multiple cell lines and constructs","pmids":["10391679"],"is_preprint":false},{"year":2004,"finding":"MLF1 physically interacts with a novel protein MLF1IP (MLF1-interacting protein); the interaction was demonstrated by yeast two-hybrid and pulldown assays, and MLF1IP colocalizes with MLF1 in both the nucleus and cytoplasm.","method":"Yeast two-hybrid, in vitro pulldown assay, co-localization by immunofluorescence","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — two orthogonal binding methods (Y2H + pulldown), co-localization confirmed, single lab","pmids":["15116101"],"is_preprint":false},{"year":2007,"finding":"MLF1 is a cytoplasmic-nuclear-shuttling protein with a functional nuclear export signal (NES). Treatment with leptomycin B induces nuclear accumulation of MLF1. Mutation of the NES enhances MLF1 antiproliferative activity. Fusion with NPM translocates MLF1 to the nucleolus and abolishes its growth-suppressing activity. Disruption of the MLF1 NES completely abolishes the growth-promoting activity of NPM-MLF1 in murine fibroblasts and hematopoietic cells.","method":"Leptomycin B treatment, NES mutagenesis, subcellular localization studies, transformation assays in murine embryonic fibroblasts and hematopoietic cells","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — NES mutagenesis with multiple orthogonal functional readouts (proliferation, transformation, p53 regulation), chemical inhibition of nuclear export, replicated across cell types","pmids":["17967869"],"is_preprint":false},{"year":2007,"finding":"MLF1 stabilizes p53 activity by suppressing its E3 ubiquitin ligase COP1 through a third component of the COP9 signalosome (CSN3); nucleolar sequestration of MLF1 by NPM prevents full induction of p53 in response to genotoxic and oncogenic stress.","method":"Genetic epistasis and molecular analysis in cell-based assays; oncogenic transformation assay in murine embryonic fibroblasts with Ras","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pathway epistasis established (MLF1–CSN3–COP1–p53), functional transformation assay, single lab","pmids":["17967869"],"is_preprint":false},{"year":2012,"finding":"MLF1 binds to 14-3-3ε adapter proteins via a phosphoserine-dependent interaction at Ser34 (motif MLF1(29-42)pSer34); crystal structure of the 14-3-3ε/MLF1(29-42)pSer34 complex resolved at high resolution (PDB: 3UAL).","method":"X-ray crystallography and isothermal titration calorimetry (ITC)","journal":"The FEBS journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with ITC quantification, two orthogonal methods, structure deposited in PDB","pmids":["22151054"],"is_preprint":false},{"year":2012,"finding":"The subcellular localization of full-length human MLF1 is independent of 14-3-3 proteins, in contrast to mouse MLF1; localization is likely regulated by other unknown proteins.","method":"Live cell imaging with GFP-fused human MLF1, mutations and deletions of 14-3-3 binding site","journal":"Cellular & molecular biology letters","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — live-cell imaging with mutant constructs, negative finding for 14-3-3 dependence, single lab, single method","pmids":["23271436"],"is_preprint":false},{"year":2015,"finding":"MLF1 interacts with MRJ (a heat shock protein/DNAJB6); MLF1 overexpression in transgenic mouse skeletal muscle results in non-pathogenic protein aggregate formation that does not impair muscle function.","method":"Co-interaction assay, transgenic mouse model with histological and RotaRod functional testing","journal":"Journal of the neurological sciences","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single interaction assay cited without detailed method, aggregate phenotype described but no pathological consequence demonstrated","pmids":["17854834"],"is_preprint":false},{"year":1997,"finding":"MNDA binds the NPM-MLF1 chimeric protein (which retains NPM residues 1-175); binding requires NPM residues 117-175 (containing a nuclear localization signal and clusters of acidic residues) that are absent in NPM-ALK (residues 1-117), which MNDA does not bind.","method":"In vitro binding assay and co-immunoprecipitation","journal":"Experimental hematology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal/orthogonal binding assays (in vitro + co-IP), domain-mapped binding, single lab","pmids":["9328447"],"is_preprint":false},{"year":2017,"finding":"MLF1 physically associates with HAX1 and HtrA2 mitochondrial proteins; increased MLF1-HAX1/HtrA2 interaction displaces HtrA2 from the HOP (HAX1/HtrA2/PARL) complex, inhibits HtrA2 cleavage/activation, and results in apoptosis. Genetic deletion of Mlf1 reverses B- and T-cell lymphopenia and neurodegeneration in Hax1-/- mice, doubling their lifespan.","method":"Co-immunoprecipitation, overexpression/knockdown assays, Mlf1-/-/Hax1-/- double-knockout mice","journal":"Biochimica et biophysica acta. Molecular cell research","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, in vivo genetic epistasis with double knockout mice, multiple orthogonal functional readouts (apoptosis, lymphopenia rescue, lifespan extension)","pmids":["28137643"],"is_preprint":false},{"year":2019,"finding":"NPM and NPM-MLF1 interact with subunits of chromatin remodeling complexes ISWI, NuRD, and P/BAF; NPM-MLF1 expression differentially alters gene transcription regulated by NPM and enhances recruitment of NuRD to gene regulatory regions.","method":"Proteomic analysis (mass spectrometry), chromatin immunoprecipitation, gene expression analysis in NPM knockdown and NPM-MLF1 expressing cells","journal":"PLoS genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS-based interactome plus ChIP functional validation, single lab, multiple orthogonal methods","pmids":["31675375"],"is_preprint":false},{"year":2025,"finding":"MLF1 functions as a transcriptional activator that recruits the acetyltransferase EP300 to target gene promoters, promoting H3K27ac deposition and chromatin opening at senescence effector loci (e.g., IL1B, p21) in cardiomyocytes; inhibition of EP300 (but not PRC2) reverses MLF1-dependent chromatin accessibility changes.","method":"RNA-seq, ATAC-seq, CUT&Tag, MLF1 knockdown/overexpression in human AC16 cardiomyocytes, EP300 and PRC2 inhibitor treatments","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal genome-wide methods (RNA-seq, ATAC-seq, CUT&Tag) plus pharmacological epistasis, mechanistic pathway established in a single rigorous study","pmids":["39657728"],"is_preprint":false},{"year":1999,"finding":"The murine MLF1 homologue (HLS7) enforces erythroid-to-myeloid lineage switching when overexpressed in J2E erythroleukemic cells, suppresses erythropoietin-induced erythroid differentiation, and promotes maturation of M1 monoblastoid cells and myeloid colony formation, without impeding intracellular signaling activated by erythropoietin.","method":"Enforced expression of HLS7 in murine erythroleukemic cell lines, semi-solid colony cultures, erythropoietin signaling assays","journal":"The EMBO journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — defined cellular phenotype with loss/gain of function across multiple assays, negative result for EPO signaling interference, single lab","pmids":["10523300"],"is_preprint":false}],"current_model":"MLF1 is a cytoplasmic-nuclear shuttling protein that stabilizes p53 by suppressing the COP1 E3 ubiquitin ligase through CSN3, recruits the acetyltransferase EP300 to target gene promoters to promote H3K27ac deposition and chromatin opening, physically interacts with HAX1 and HtrA2 to antagonize the pro-survival HOP mitochondrial complex and promote apoptosis, binds 14-3-3ε via phospho-Ser34, and interacts with MLF1IP and MRJ/DNAJB6; its oncogenic fusion with NPM sequesters MLF1 in the nucleolus, abrogating its growth-suppressive and p53-stabilizing activities and conferring leukemogenic properties through aberrant NuRD chromatin remodeling complex recruitment."},"narrative":{"mechanistic_narrative":"MLF1 is a cytoplasmic–nuclear shuttling protein that links subcellular compartmentalization to control of cell proliferation, differentiation, apoptosis, and chromatin-based gene regulation [PMID:17967869]. It carries a functional nuclear export signal whose disruption enhances its antiproliferative activity, indicating that cytoplasmic retention restrains its growth-suppressive function [PMID:17967869]. MLF1 stabilizes p53 by suppressing the COP1 E3 ubiquitin ligase through the COP9 signalosome subunit CSN3, thereby coupling MLF1 localization to the p53-dependent response to genotoxic and oncogenic stress [PMID:17967869]. In the nucleus, MLF1 acts as a transcriptional activator that recruits the acetyltransferase EP300 to target promoters to deposit H3K27ac and open chromatin at senescence effector loci such as IL1B and p21, a function that is EP300- but not PRC2-dependent [PMID:39657728]. At mitochondria, MLF1 physically associates with HAX1 and HtrA2, displacing HtrA2 from the pro-survival HAX1/HtrA2/PARL (HOP) complex to promote apoptosis; deletion of Mlf1 rescues lymphopenia and neurodegeneration in Hax1-deficient mice [PMID:28137643]. MLF1 also engages 14-3-3ε through a phosphoserine motif at Ser34, resolved by crystallography [PMID:22151054]. The leukemogenic NPM-MLF1 fusion redirects MLF1 to the nucleus and nucleolus, abolishing its growth-suppressing and p53-stabilizing activities and instead promoting transformation through aberrant recruitment of the NuRD chromatin remodeling complex [PMID:8570204, PMID:17967869, PMID:31675375].","teleology":[{"year":1996,"claim":"Established that the difference between normal MLF1 and its leukemic fusion is one of localization—wild-type MLF1 is cytoplasmic while NPM-MLF1 is mislocalized to the nucleus and nucleolus—framing aberrant compartmentalization as central to its oncogenic conversion.","evidence":"Immunostaining of t(3;5)-positive leukemia cells and cell lines expressing wild-type MLF1 versus NPM-MLF1","pmids":["8570204"],"confidence":"Medium","gaps":["Did not define the molecular consequence of mislocalization","No identification of MLF1's normal cytoplasmic function"]},{"year":1997,"claim":"Mapped the NPM-derived determinants of the fusion, showing MNDA binds NPM residues 117-175 retained in NPM-MLF1, distinguishing it from NPM-ALK and beginning to define fusion-specific protein interactions.","evidence":"In vitro binding assay and co-immunoprecipitation with domain-mapped NPM fragments","pmids":["9328447"],"confidence":"Medium","gaps":["Functional consequence of MNDA binding to NPM-MLF1 not established","Binding concerns NPM portion, not MLF1 sequences"]},{"year":1999,"claim":"Defined a pro-apoptotic activity of the NPM-MLF1 fusion requiring the MLF1 N-terminus and NPM NLS/dimerization domains, and showed Bcl-2 rescues without altering fusion expression, placing the death phenotype downstream of mitochondrial apoptotic control.","evidence":"Ectopic overexpression and deletion-mutant analysis in K562 and NIH3T3 cells with Bcl-2 co-expression","pmids":["10391679"],"confidence":"Medium","gaps":["Apoptotic mechanism not molecularly resolved","Relationship of fusion-induced death to native MLF1 function unclear"]},{"year":1999,"claim":"Showed the murine homologue HLS7 enforces erythroid-to-myeloid lineage switching without disrupting erythropoietin signaling, establishing MLF1 as a regulator of hematopoietic differentiation rather than a signaling effector.","evidence":"Enforced expression in murine erythroleukemic lines, colony cultures, and EPO signaling assays","pmids":["10523300"],"confidence":"Medium","gaps":["Molecular targets of the lineage-switch activity not identified","Mechanism linking MLF1 to transcriptional/differentiation programs unknown at this stage"]},{"year":2004,"claim":"Identified MLF1IP as a direct binding partner colocalizing in both nucleus and cytoplasm, expanding the MLF1 interactome and reinforcing its dual-compartment biology.","evidence":"Yeast two-hybrid, in vitro pulldown, and co-localization by immunofluorescence","pmids":["15116101"],"confidence":"Medium","gaps":["Functional role of the MLF1-MLF1IP interaction undefined","No pathway placed for MLF1IP"]},{"year":2007,"claim":"Demonstrated MLF1 is an active nuclear-export-driven shuttling protein and connected its localization to growth control and p53 stabilization via CSN3-mediated suppression of COP1, providing the first mechanistic explanation for both its tumor-suppressive role and the loss of that role upon nucleolar sequestration by NPM.","evidence":"Leptomycin B treatment, NES mutagenesis, subcellular localization, genetic epistasis, and Ras transformation assays in MEFs and hematopoietic cells","pmids":["17967869"],"confidence":"High","gaps":["Direct biochemical demonstration of MLF1-CSN3 and CSN3-COP1 contacts not fully resolved","Identity of factors gating MLF1 nuclear import not defined"]},{"year":2012,"claim":"Resolved the structural basis of the MLF1–14-3-3ε interaction, defining a phospho-Ser34 motif as the recognition element and providing an atomic-resolution view of a regulatory contact.","evidence":"X-ray crystallography (PDB 3UAL) and isothermal titration calorimetry","pmids":["22151054"],"confidence":"High","gaps":["Cellular consequence of 14-3-3ε binding to human MLF1 not established","Kinase responsible for Ser34 phosphorylation unknown"]},{"year":2012,"claim":"Showed that, unlike mouse MLF1, human MLF1 localization is independent of 14-3-3 binding, indicating species-specific regulation and pointing to additional unidentified localization determinants.","evidence":"Live-cell imaging of GFP-fused human MLF1 with 14-3-3 binding-site mutants and deletions","pmids":["23271436"],"confidence":"Medium","gaps":["Identity of the proteins controlling human MLF1 localization unknown","Negative result from a single method and lab"]},{"year":2015,"claim":"Linked MLF1 to the chaperone MRJ/DNAJB6 and showed that muscle overexpression produces benign aggregates, addressing whether MLF1 aggregation is intrinsically pathogenic.","evidence":"Co-interaction assay and transgenic mouse muscle with histology and RotaRod testing","pmids":["17854834"],"confidence":"Low","gaps":["Interaction reported without detailed method and not independently validated","No demonstrated pathological consequence of aggregation","Functional significance of the MRJ interaction unclear"]},{"year":2017,"claim":"Established a mitochondrial pro-apoptotic mechanism whereby MLF1 binds HAX1 and HtrA2 to displace HtrA2 from the pro-survival HOP complex, validated in vivo by genetic epistasis showing Mlf1 deletion rescues Hax1-/- pathology and doubles lifespan.","evidence":"Reciprocal co-immunoprecipitation, overexpression/knockdown, and Mlf1-/-/Hax1-/- double-knockout mice","pmids":["28137643"],"confidence":"High","gaps":["Signals that drive MLF1 to mitochondria not defined","Integration with nuclear/p53 functions of MLF1 unresolved"]},{"year":2019,"claim":"Connected NPM-MLF1 oncogenesis to chromatin regulation by showing the fusion interacts with ISWI, NuRD, and P/BAF subunits and enhances NuRD recruitment to gene regulatory regions, providing a transcriptional mechanism for leukemogenesis.","evidence":"Mass spectrometry interactome, ChIP, and gene expression analysis in NPM knockdown and NPM-MLF1 expressing cells","pmids":["31675375"],"confidence":"Medium","gaps":["Direct transcriptional targets driving transformation not pinpointed","Whether native MLF1 engages these complexes not addressed"]},{"year":2025,"claim":"Defined native MLF1 as a chromatin-acting transcriptional activator that recruits EP300 to deposit H3K27ac and open chromatin at senescence loci, establishing a direct, EP300-dependent epigenetic mechanism for its nuclear function.","evidence":"RNA-seq, ATAC-seq, CUT&Tag, knockdown/overexpression in AC16 cardiomyocytes, and EP300/PRC2 inhibitor epistasis","pmids":["39657728"],"confidence":"High","gaps":["Mechanism of MLF1 promoter targeting (sequence specificity or partner-mediated) not defined","Relationship between this EP300 activity and the cytoplasmic/mitochondrial MLF1 functions unresolved"]},{"year":null,"claim":"How MLF1's distinct cytoplasmic, mitochondrial, and chromatin-associated activities are coordinated by its shuttling and what signals partition it among compartments remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking p53 stabilization, EP300-driven transcription, and HOP-complex apoptosis","Determinants of human MLF1 nuclear import unidentified","Direct DNA/promoter-recognition mechanism unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[11]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[4,9]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[11,9]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[0,3]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,3,11]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[0,3]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[9]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[11,10]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[11,10]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[9,1]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[0,10]}],"complexes":["HOP complex (HAX1/HtrA2/PARL)"],"partners":["HAX1","HTRA2","EP300","YWHAE","CSN3","MLF1IP","DNAJB6","NPM1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P58340","full_name":"Myeloid leukemia factor 1","aliases":["Myelodysplasia-myeloid leukemia factor 1"],"length_aa":268,"mass_kda":30.6,"function":"Involved in lineage commitment of primary hemopoietic progenitors by restricting erythroid formation and enhancing myeloid formation. Interferes with erythropoietin-induced erythroid terminal differentiation by preventing cells from exiting the cell cycle through suppression of CDKN1B/p27Kip1 levels. Suppresses COP1 activity via CSN3 which activates p53 and induces cell cycle arrest. Binds DNA and affects the expression of a number of genes so may function as a transcription factor in the nucleus","subcellular_location":"Cytoplasm; Nucleus; Cell projection, cilium; Cytoplasm, cytoskeleton, cilium basal body","url":"https://www.uniprot.org/uniprotkb/P58340/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/MLF1","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/MLF1","total_profiled":1310},"omim":[{"mim_id":"618402","title":"INTELLECTUAL DEVELOPMENTAL DISORDER, AUTOSOMAL RECESSIVE 70; MRT70","url":"https://www.omim.org/entry/618402"},{"mim_id":"613352","title":"ARGININE/SERINE-RICH COILED-COIL PROTEIN 1; RSRC1","url":"https://www.omim.org/entry/613352"},{"mim_id":"611511","title":"MLF1-INTERACTING PROTEIN; MLF1IP","url":"https://www.omim.org/entry/611511"},{"mim_id":"611332","title":"DNAJ/HSP40 HOMOLOG, SUBFAMILY B, MEMBER 6; DNAJB6","url":"https://www.omim.org/entry/611332"},{"mim_id":"603511","title":"MUSCULAR DYSTROPHY, LIMB-GIRDLE, AUTOSOMAL DOMINANT 1; LGMDD1","url":"https://www.omim.org/entry/603511"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Centrosome","reliability":"Approved"},{"location":"Basal body","reliability":"Approved"},{"location":"Mid piece","reliability":"Approved"},{"location":"Principal piece","reliability":"Approved"},{"location":"End piece","reliability":"Approved"},{"location":"Vesicles","reliability":"Additional"},{"location":"Plasma membrane","reliability":"Additional"},{"location":"Focal adhesion sites","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"skeletal muscle","ntpm":170.9},{"tissue":"testis","ntpm":340.3}],"url":"https://www.proteinatlas.org/search/MLF1"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"P58340","domains":[{"cath_id":"-","chopping":"12-33_76-205","consensus_level":"high","plddt":83.2319,"start":12,"end":205}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P58340","model_url":"https://alphafold.ebi.ac.uk/files/AF-P58340-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P58340-F1-predicted_aligned_error_v6.png","plddt_mean":67.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MLF1","jax_strain_url":"https://www.jax.org/strain/search?query=MLF1"},"sequence":{"accession":"P58340","fasta_url":"https://rest.uniprot.org/uniprotkb/P58340.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P58340/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P58340"}},"corpus_meta":[{"pmid":"8570204","id":"PMC_8570204","title":"The t(3;5)(q25.1;q34) of myelodysplastic syndrome and acute myeloid leukemia produces a novel fusion gene, NPM-MLF1.","date":"1996","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/8570204","citation_count":245,"is_preprint":false},{"pmid":"28545128","id":"PMC_28545128","title":"Common variants upstream of MLF1 at 3q25 and within CPZ at 4p16 associated with neuroblastoma.","date":"2017","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/28545128","citation_count":71,"is_preprint":false},{"pmid":"15116101","id":"PMC_15116101","title":"cDNA cloning and characterization of a novel gene encoding the MLF1-interacting protein MLF1IP.","date":"2004","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/15116101","citation_count":57,"is_preprint":false},{"pmid":"11021751","id":"PMC_11021751","title":"Elevated MLF1 expression correlates with malignant progression from myelodysplastic syndrome.","date":"2000","source":"Leukemia","url":"https://pubmed.ncbi.nlm.nih.gov/11021751","citation_count":46,"is_preprint":false},{"pmid":"10523300","id":"PMC_10523300","title":"HLS7, a hemopoietic lineage switch gene homologous to the leukemia-inducing gene MLF1.","date":"1999","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/10523300","citation_count":40,"is_preprint":false},{"pmid":"17967869","id":"PMC_17967869","title":"Shuttling imbalance of MLF1 results in p53 instability and increases susceptibility to oncogenic transformation.","date":"2007","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/17967869","citation_count":35,"is_preprint":false},{"pmid":"22151054","id":"PMC_22151054","title":"Structural insights of the MLF1/14-3-3 interaction.","date":"2012","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/22151054","citation_count":34,"is_preprint":false},{"pmid":"20149264","id":"PMC_20149264","title":"Madm (Mlf1 adapter molecule) cooperates with Bunched A to promote growth in Drosophila.","date":"2010","source":"Journal of biology","url":"https://pubmed.ncbi.nlm.nih.gov/20149264","citation_count":31,"is_preprint":false},{"pmid":"14506644","id":"PMC_14506644","title":"Detection of NPM/MLF1 fusion in t(3;5)-positive acute myeloid leukemia and myelodysplasia.","date":"2003","source":"Human pathology","url":"https://pubmed.ncbi.nlm.nih.gov/14506644","citation_count":26,"is_preprint":false},{"pmid":"10391679","id":"PMC_10391679","title":"Apoptosis induced by the myelodysplastic syndrome-associated NPM-MLF1 chimeric protein.","date":"1999","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/10391679","citation_count":25,"is_preprint":false},{"pmid":"37486965","id":"PMC_37486965","title":"Epigenetic deregulation of MLF1 drives intrahepatic cholangiocarcinoma progression through EGFR/AKT and Wnt/β-catenin signaling.","date":"2023","source":"Hepatology 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leukemia.","date":"1997","source":"Experimental hematology","url":"https://pubmed.ncbi.nlm.nih.gov/9328447","citation_count":18,"is_preprint":false},{"pmid":"17854834","id":"PMC_17854834","title":"Non-pathogenic protein aggregates in skeletal muscle in MLF1 transgenic mice.","date":"2007","source":"Journal of the neurological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/17854834","citation_count":17,"is_preprint":false},{"pmid":"25572810","id":"PMC_25572810","title":"MLF1 interacting protein: a potential gene therapy target for human prostate cancer?","date":"2015","source":"Medical oncology (Northwood, London, England)","url":"https://pubmed.ncbi.nlm.nih.gov/25572810","citation_count":14,"is_preprint":false},{"pmid":"31675375","id":"PMC_31675375","title":"NPM and NPM-MLF1 interact with chromatin remodeling complexes and influence their recruitment to specific genes.","date":"2019","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/31675375","citation_count":12,"is_preprint":false},{"pmid":"39657728","id":"PMC_39657728","title":"Downregulation of MLF1 safeguards cardiomyocytes against senescence-associated chromatin opening.","date":"2025","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/39657728","citation_count":11,"is_preprint":false},{"pmid":"17595757","id":"PMC_17595757","title":"MLF1-interacting protein is mainly localized in nucleolus through N-terminal bipartite nuclear localization signal.","date":"2007","source":"Anticancer research","url":"https://pubmed.ncbi.nlm.nih.gov/17595757","citation_count":8,"is_preprint":false},{"pmid":"28137643","id":"PMC_28137643","title":"MLF1 is a proapoptotic antagonist of HOP complex-mediated survival.","date":"2017","source":"Biochimica et biophysica acta. Molecular cell research","url":"https://pubmed.ncbi.nlm.nih.gov/28137643","citation_count":7,"is_preprint":false},{"pmid":"31949731","id":"PMC_31949731","title":"MLF1 protein is a potential therapy target for lung adenocarcinoma.","date":"2018","source":"International journal of clinical and experimental pathology","url":"https://pubmed.ncbi.nlm.nih.gov/31949731","citation_count":7,"is_preprint":false},{"pmid":"25027285","id":"PMC_25027285","title":"Development of an NPM1/MLF1 D-FISH probe set for the detection of t(3;5)(q25;q35) identified in patients with acute myeloid leukemia.","date":"2014","source":"The Journal of molecular diagnostics : JMD","url":"https://pubmed.ncbi.nlm.nih.gov/25027285","citation_count":7,"is_preprint":false},{"pmid":"23724000","id":"PMC_23724000","title":"Investigation of tissue-specific expression and functions of MLF1-IP during development and in the immune system.","date":"2013","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/23724000","citation_count":6,"is_preprint":false},{"pmid":"23271436","id":"PMC_23271436","title":"Subcellular localization of full-length human myeloid leukemia factor 1 (MLF1) is independent of 14-3-3 proteins.","date":"2012","source":"Cellular & molecular biology letters","url":"https://pubmed.ncbi.nlm.nih.gov/23271436","citation_count":4,"is_preprint":false},{"pmid":"40497606","id":"PMC_40497606","title":"A Rare Case of Juvenile Myelomonocytic Leukemia (JMML) with t(3;5)(q25;q34)/NPM::MLF1 Fusion Gene in a Pediatric Patient.","date":"2025","source":"Clinical laboratory","url":"https://pubmed.ncbi.nlm.nih.gov/40497606","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2024.09.29.615655","title":"Integrated data from R405W desmin knock-in mice highlight alterations of mitochondrial function, protein quality control, and myofibrillar structure in the initial stages of myofibrillar myopathy","date":"2024-10-01","source":"bioRxiv","url":"https://doi.org/10.1101/2024.09.29.615655","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":15730,"output_tokens":3208,"usd":0.047655,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10670,"output_tokens":4361,"usd":0.081187,"stage2_stop_reason":"end_turn"},"total_usd":0.128842,"stage1_batch_id":"msgbatch_01BbTCMyrRPyhtEq6b1MdwCN","stage2_batch_id":"msgbatch_011dEed5bbxYvTAaQWKJevxu","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1996,\n      \"finding\": \"MLF1 is normally localized in the cytoplasm, whereas the NPM-MLF1 fusion protein is targeted to the nucleus and nucleolus; NPM trafficking signals direct MLF1 to an inappropriate cellular compartment in myeloid leukemia cells.\",\n      \"method\": \"Immunostaining of t(3;5)-positive leukemia cells and cell lines expressing wild-type MLF1 vs. NPM-MLF1\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct immunostaining localization with functional implication, single lab, two cell contexts (leukemia cells and cell lines)\",\n      \"pmids\": [\"8570204\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"NPM-MLF1 fusion protein induces apoptosis; this requires the N-terminal domain of MLF1 and the NPM domain containing a nuclear localization signal. The NPM dimerization domain is also required. Co-expression of Bcl-2 rescues cells from NPM-MLF1-mediated cell death without altering the expression or localization of NPM-MLF1.\",\n      \"method\": \"Ectopic overexpression and deletion mutant analysis in K562 and NIH3T3 cells; co-expression with Bcl-2\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — deletion mutagenesis with defined apoptotic phenotype, single lab, multiple cell lines and constructs\",\n      \"pmids\": [\"10391679\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"MLF1 physically interacts with a novel protein MLF1IP (MLF1-interacting protein); the interaction was demonstrated by yeast two-hybrid and pulldown assays, and MLF1IP colocalizes with MLF1 in both the nucleus and cytoplasm.\",\n      \"method\": \"Yeast two-hybrid, in vitro pulldown assay, co-localization by immunofluorescence\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — two orthogonal binding methods (Y2H + pulldown), co-localization confirmed, single lab\",\n      \"pmids\": [\"15116101\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"MLF1 is a cytoplasmic-nuclear-shuttling protein with a functional nuclear export signal (NES). Treatment with leptomycin B induces nuclear accumulation of MLF1. Mutation of the NES enhances MLF1 antiproliferative activity. Fusion with NPM translocates MLF1 to the nucleolus and abolishes its growth-suppressing activity. Disruption of the MLF1 NES completely abolishes the growth-promoting activity of NPM-MLF1 in murine fibroblasts and hematopoietic cells.\",\n      \"method\": \"Leptomycin B treatment, NES mutagenesis, subcellular localization studies, transformation assays in murine embryonic fibroblasts and hematopoietic cells\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — NES mutagenesis with multiple orthogonal functional readouts (proliferation, transformation, p53 regulation), chemical inhibition of nuclear export, replicated across cell types\",\n      \"pmids\": [\"17967869\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"MLF1 stabilizes p53 activity by suppressing its E3 ubiquitin ligase COP1 through a third component of the COP9 signalosome (CSN3); nucleolar sequestration of MLF1 by NPM prevents full induction of p53 in response to genotoxic and oncogenic stress.\",\n      \"method\": \"Genetic epistasis and molecular analysis in cell-based assays; oncogenic transformation assay in murine embryonic fibroblasts with Ras\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pathway epistasis established (MLF1–CSN3–COP1–p53), functional transformation assay, single lab\",\n      \"pmids\": [\"17967869\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"MLF1 binds to 14-3-3ε adapter proteins via a phosphoserine-dependent interaction at Ser34 (motif MLF1(29-42)pSer34); crystal structure of the 14-3-3ε/MLF1(29-42)pSer34 complex resolved at high resolution (PDB: 3UAL).\",\n      \"method\": \"X-ray crystallography and isothermal titration calorimetry (ITC)\",\n      \"journal\": \"The FEBS journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with ITC quantification, two orthogonal methods, structure deposited in PDB\",\n      \"pmids\": [\"22151054\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The subcellular localization of full-length human MLF1 is independent of 14-3-3 proteins, in contrast to mouse MLF1; localization is likely regulated by other unknown proteins.\",\n      \"method\": \"Live cell imaging with GFP-fused human MLF1, mutations and deletions of 14-3-3 binding site\",\n      \"journal\": \"Cellular & molecular biology letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — live-cell imaging with mutant constructs, negative finding for 14-3-3 dependence, single lab, single method\",\n      \"pmids\": [\"23271436\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"MLF1 interacts with MRJ (a heat shock protein/DNAJB6); MLF1 overexpression in transgenic mouse skeletal muscle results in non-pathogenic protein aggregate formation that does not impair muscle function.\",\n      \"method\": \"Co-interaction assay, transgenic mouse model with histological and RotaRod functional testing\",\n      \"journal\": \"Journal of the neurological sciences\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single interaction assay cited without detailed method, aggregate phenotype described but no pathological consequence demonstrated\",\n      \"pmids\": [\"17854834\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"MNDA binds the NPM-MLF1 chimeric protein (which retains NPM residues 1-175); binding requires NPM residues 117-175 (containing a nuclear localization signal and clusters of acidic residues) that are absent in NPM-ALK (residues 1-117), which MNDA does not bind.\",\n      \"method\": \"In vitro binding assay and co-immunoprecipitation\",\n      \"journal\": \"Experimental hematology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal/orthogonal binding assays (in vitro + co-IP), domain-mapped binding, single lab\",\n      \"pmids\": [\"9328447\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"MLF1 physically associates with HAX1 and HtrA2 mitochondrial proteins; increased MLF1-HAX1/HtrA2 interaction displaces HtrA2 from the HOP (HAX1/HtrA2/PARL) complex, inhibits HtrA2 cleavage/activation, and results in apoptosis. Genetic deletion of Mlf1 reverses B- and T-cell lymphopenia and neurodegeneration in Hax1-/- mice, doubling their lifespan.\",\n      \"method\": \"Co-immunoprecipitation, overexpression/knockdown assays, Mlf1-/-/Hax1-/- double-knockout mice\",\n      \"journal\": \"Biochimica et biophysica acta. Molecular cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, in vivo genetic epistasis with double knockout mice, multiple orthogonal functional readouts (apoptosis, lymphopenia rescue, lifespan extension)\",\n      \"pmids\": [\"28137643\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NPM and NPM-MLF1 interact with subunits of chromatin remodeling complexes ISWI, NuRD, and P/BAF; NPM-MLF1 expression differentially alters gene transcription regulated by NPM and enhances recruitment of NuRD to gene regulatory regions.\",\n      \"method\": \"Proteomic analysis (mass spectrometry), chromatin immunoprecipitation, gene expression analysis in NPM knockdown and NPM-MLF1 expressing cells\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-based interactome plus ChIP functional validation, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"31675375\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MLF1 functions as a transcriptional activator that recruits the acetyltransferase EP300 to target gene promoters, promoting H3K27ac deposition and chromatin opening at senescence effector loci (e.g., IL1B, p21) in cardiomyocytes; inhibition of EP300 (but not PRC2) reverses MLF1-dependent chromatin accessibility changes.\",\n      \"method\": \"RNA-seq, ATAC-seq, CUT&Tag, MLF1 knockdown/overexpression in human AC16 cardiomyocytes, EP300 and PRC2 inhibitor treatments\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal genome-wide methods (RNA-seq, ATAC-seq, CUT&Tag) plus pharmacological epistasis, mechanistic pathway established in a single rigorous study\",\n      \"pmids\": [\"39657728\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"The murine MLF1 homologue (HLS7) enforces erythroid-to-myeloid lineage switching when overexpressed in J2E erythroleukemic cells, suppresses erythropoietin-induced erythroid differentiation, and promotes maturation of M1 monoblastoid cells and myeloid colony formation, without impeding intracellular signaling activated by erythropoietin.\",\n      \"method\": \"Enforced expression of HLS7 in murine erythroleukemic cell lines, semi-solid colony cultures, erythropoietin signaling assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — defined cellular phenotype with loss/gain of function across multiple assays, negative result for EPO signaling interference, single lab\",\n      \"pmids\": [\"10523300\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MLF1 is a cytoplasmic-nuclear shuttling protein that stabilizes p53 by suppressing the COP1 E3 ubiquitin ligase through CSN3, recruits the acetyltransferase EP300 to target gene promoters to promote H3K27ac deposition and chromatin opening, physically interacts with HAX1 and HtrA2 to antagonize the pro-survival HOP mitochondrial complex and promote apoptosis, binds 14-3-3ε via phospho-Ser34, and interacts with MLF1IP and MRJ/DNAJB6; its oncogenic fusion with NPM sequesters MLF1 in the nucleolus, abrogating its growth-suppressive and p53-stabilizing activities and conferring leukemogenic properties through aberrant NuRD chromatin remodeling complex recruitment.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MLF1 is a cytoplasmic–nuclear shuttling protein that links subcellular compartmentalization to control of cell proliferation, differentiation, apoptosis, and chromatin-based gene regulation [#3, #4]. It carries a functional nuclear export signal whose disruption enhances its antiproliferative activity, indicating that cytoplasmic retention restrains its growth-suppressive function [#3]. MLF1 stabilizes p53 by suppressing the COP1 E3 ubiquitin ligase through the COP9 signalosome subunit CSN3, thereby coupling MLF1 localization to the p53-dependent response to genotoxic and oncogenic stress [#4]. In the nucleus, MLF1 acts as a transcriptional activator that recruits the acetyltransferase EP300 to target promoters to deposit H3K27ac and open chromatin at senescence effector loci such as IL1B and p21, a function that is EP300- but not PRC2-dependent [#11]. At mitochondria, MLF1 physically associates with HAX1 and HtrA2, displacing HtrA2 from the pro-survival HAX1/HtrA2/PARL (HOP) complex to promote apoptosis; deletion of Mlf1 rescues lymphopenia and neurodegeneration in Hax1-deficient mice [#9]. MLF1 also engages 14-3-3ε through a phosphoserine motif at Ser34, resolved by crystallography [#5]. The leukemogenic NPM-MLF1 fusion redirects MLF1 to the nucleus and nucleolus, abolishing its growth-suppressing and p53-stabilizing activities and instead promoting transformation through aberrant recruitment of the NuRD chromatin remodeling complex [#0, #3, #10].\",\n  \"teleology\": [\n    {\n      \"year\": 1996,\n      \"claim\": \"Established that the difference between normal MLF1 and its leukemic fusion is one of localization—wild-type MLF1 is cytoplasmic while NPM-MLF1 is mislocalized to the nucleus and nucleolus—framing aberrant compartmentalization as central to its oncogenic conversion.\",\n      \"evidence\": \"Immunostaining of t(3;5)-positive leukemia cells and cell lines expressing wild-type MLF1 versus NPM-MLF1\",\n      \"pmids\": [\"8570204\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define the molecular consequence of mislocalization\", \"No identification of MLF1's normal cytoplasmic function\"]\n    },\n    {\n      \"year\": 1997,\n      \"claim\": \"Mapped the NPM-derived determinants of the fusion, showing MNDA binds NPM residues 117-175 retained in NPM-MLF1, distinguishing it from NPM-ALK and beginning to define fusion-specific protein interactions.\",\n      \"evidence\": \"In vitro binding assay and co-immunoprecipitation with domain-mapped NPM fragments\",\n      \"pmids\": [\"9328447\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of MNDA binding to NPM-MLF1 not established\", \"Binding concerns NPM portion, not MLF1 sequences\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Defined a pro-apoptotic activity of the NPM-MLF1 fusion requiring the MLF1 N-terminus and NPM NLS/dimerization domains, and showed Bcl-2 rescues without altering fusion expression, placing the death phenotype downstream of mitochondrial apoptotic control.\",\n      \"evidence\": \"Ectopic overexpression and deletion-mutant analysis in K562 and NIH3T3 cells with Bcl-2 co-expression\",\n      \"pmids\": [\"10391679\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Apoptotic mechanism not molecularly resolved\", \"Relationship of fusion-induced death to native MLF1 function unclear\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Showed the murine homologue HLS7 enforces erythroid-to-myeloid lineage switching without disrupting erythropoietin signaling, establishing MLF1 as a regulator of hematopoietic differentiation rather than a signaling effector.\",\n      \"evidence\": \"Enforced expression in murine erythroleukemic lines, colony cultures, and EPO signaling assays\",\n      \"pmids\": [\"10523300\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular targets of the lineage-switch activity not identified\", \"Mechanism linking MLF1 to transcriptional/differentiation programs unknown at this stage\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Identified MLF1IP as a direct binding partner colocalizing in both nucleus and cytoplasm, expanding the MLF1 interactome and reinforcing its dual-compartment biology.\",\n      \"evidence\": \"Yeast two-hybrid, in vitro pulldown, and co-localization by immunofluorescence\",\n      \"pmids\": [\"15116101\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional role of the MLF1-MLF1IP interaction undefined\", \"No pathway placed for MLF1IP\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Demonstrated MLF1 is an active nuclear-export-driven shuttling protein and connected its localization to growth control and p53 stabilization via CSN3-mediated suppression of COP1, providing the first mechanistic explanation for both its tumor-suppressive role and the loss of that role upon nucleolar sequestration by NPM.\",\n      \"evidence\": \"Leptomycin B treatment, NES mutagenesis, subcellular localization, genetic epistasis, and Ras transformation assays in MEFs and hematopoietic cells\",\n      \"pmids\": [\"17967869\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct biochemical demonstration of MLF1-CSN3 and CSN3-COP1 contacts not fully resolved\", \"Identity of factors gating MLF1 nuclear import not defined\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Resolved the structural basis of the MLF1–14-3-3ε interaction, defining a phospho-Ser34 motif as the recognition element and providing an atomic-resolution view of a regulatory contact.\",\n      \"evidence\": \"X-ray crystallography (PDB 3UAL) and isothermal titration calorimetry\",\n      \"pmids\": [\"22151054\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cellular consequence of 14-3-3ε binding to human MLF1 not established\", \"Kinase responsible for Ser34 phosphorylation unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed that, unlike mouse MLF1, human MLF1 localization is independent of 14-3-3 binding, indicating species-specific regulation and pointing to additional unidentified localization determinants.\",\n      \"evidence\": \"Live-cell imaging of GFP-fused human MLF1 with 14-3-3 binding-site mutants and deletions\",\n      \"pmids\": [\"23271436\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of the proteins controlling human MLF1 localization unknown\", \"Negative result from a single method and lab\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Linked MLF1 to the chaperone MRJ/DNAJB6 and showed that muscle overexpression produces benign aggregates, addressing whether MLF1 aggregation is intrinsically pathogenic.\",\n      \"evidence\": \"Co-interaction assay and transgenic mouse muscle with histology and RotaRod testing\",\n      \"pmids\": [\"17854834\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Interaction reported without detailed method and not independently validated\", \"No demonstrated pathological consequence of aggregation\", \"Functional significance of the MRJ interaction unclear\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Established a mitochondrial pro-apoptotic mechanism whereby MLF1 binds HAX1 and HtrA2 to displace HtrA2 from the pro-survival HOP complex, validated in vivo by genetic epistasis showing Mlf1 deletion rescues Hax1-/- pathology and doubles lifespan.\",\n      \"evidence\": \"Reciprocal co-immunoprecipitation, overexpression/knockdown, and Mlf1-/-/Hax1-/- double-knockout mice\",\n      \"pmids\": [\"28137643\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signals that drive MLF1 to mitochondria not defined\", \"Integration with nuclear/p53 functions of MLF1 unresolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Connected NPM-MLF1 oncogenesis to chromatin regulation by showing the fusion interacts with ISWI, NuRD, and P/BAF subunits and enhances NuRD recruitment to gene regulatory regions, providing a transcriptional mechanism for leukemogenesis.\",\n      \"evidence\": \"Mass spectrometry interactome, ChIP, and gene expression analysis in NPM knockdown and NPM-MLF1 expressing cells\",\n      \"pmids\": [\"31675375\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct transcriptional targets driving transformation not pinpointed\", \"Whether native MLF1 engages these complexes not addressed\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined native MLF1 as a chromatin-acting transcriptional activator that recruits EP300 to deposit H3K27ac and open chromatin at senescence loci, establishing a direct, EP300-dependent epigenetic mechanism for its nuclear function.\",\n      \"evidence\": \"RNA-seq, ATAC-seq, CUT&Tag, knockdown/overexpression in AC16 cardiomyocytes, and EP300/PRC2 inhibitor epistasis\",\n      \"pmids\": [\"39657728\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of MLF1 promoter targeting (sequence specificity or partner-mediated) not defined\", \"Relationship between this EP300 activity and the cytoplasmic/mitochondrial MLF1 functions unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How MLF1's distinct cytoplasmic, mitochondrial, and chromatin-associated activities are coordinated by its shuttling and what signals partition it among compartments remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking p53 stabilization, EP300-driven transcription, and HOP-complex apoptosis\", \"Determinants of human MLF1 nuclear import unidentified\", \"Direct DNA/promoter-recognition mechanism unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [11]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [4, 9]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [11, 9]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0, 3]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 3, 11]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [0, 3]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [9]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [11, 10]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [11, 10]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [9, 1]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [0, 10]}\n    ],\n    \"complexes\": [\"HOP complex (HAX1/HtrA2/PARL)\"],\n    \"partners\": [\"HAX1\", \"HTRA2\", \"EP300\", \"YWHAE\", \"CSN3\", \"MLF1IP\", \"DNAJB6\", \"NPM1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}