{"gene":"DMXL1","run_date":"2026-06-09T23:54:42","timeline":{"discoveries":[{"year":2000,"finding":"DMXL1 encodes a large WD repeat protein (3027 amino acids) and is the human homologue of the Drosophila DmX gene; the gene was mapped to chromosome 5q22 by radiation hybrid mapping and FISH, and its open reading frame structure was characterized by cDNA cloning.","method":"cDNA cloning, radiation hybrid mapping, fluorescence in situ hybridization (FISH)","journal":"Genomics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct molecular cloning and chromosomal mapping with two orthogonal localization methods; no functional assay performed","pmids":["10708522"],"is_preprint":false},{"year":2015,"finding":"DMXL1 physically associates with the V-ATPase complex in kidney tissue; knockdown of DMXL1 inhibits V-ATPase-mediated intracellular vesicle acidification in a kidney cell line.","method":"Proteomic co-immunoprecipitation / V-ATPase interactome mapping; siRNA knockdown with vesicle acidification assay","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal MS-based interactome plus functional siRNA knockdown phenotype in a cell line, single lab","pmids":["26442671"],"is_preprint":false},{"year":2021,"finding":"TPL-2 kinase phosphorylates DMXL1, and this phosphorylation promotes V-ATPase assembly and phagosome acidification in macrophages, thereby enabling efficient killing of phagocytosed bacteria (Staphylococcus aureus, Citrobacter rodentium).","method":"Quantitative phagosome proteomics; TPL-2 catalytic inhibitor experiments; phosphorylation detected by mass spectrometry; phagosome acidification assay; bacterial killing assay in primary mouse and human macrophages","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (proteomics, inhibitor, phosphorylation MS, functional acidification and killing assays) in primary human and mouse cells; single lab but rigorous multi-method study","pmids":["33881780"],"is_preprint":false},{"year":2024,"finding":"Kidney intercalated cell-specific Dmxl1 knockout mice show elevated urine pH and impaired proton pumping, with decreased B1 (V1) subunit levels and reduced co-localization of V1 (B1) and VO (a4) subunits at the membrane, indicating that Dmxl1 is required for V-ATPase holoenzyme assembly in vivo.","method":"Conditional knockout mouse model; western blotting; immunofluorescence co-localization; subcellular fractionation; proximity ligation assay (PLA) for B1-a4 subunit association","journal":"Function (Oxford, England)","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo conditional KO with multiple orthogonal readouts (urine pH, western blot, fractionation, PLA, immunofluorescence), single lab but rigorous multi-method study","pmids":["38984989"],"is_preprint":false},{"year":2024,"finding":"Human cytomegalovirus degrades DMXL1 via its US33A protein, which recruits the E3 ubiquitin ligase KPC (Kip1 ubiquitination-promoting complex) to ubiquitinate and degrade DMXL1; this degradation inhibits lysosome acidification, autophagic cargo degradation, and delays virion assembly compartment formation.","method":"Quantitative proteomics of HCMV-infected cells; systematic viral deletion mutant comparison; co-immunoprecipitation to identify KPC recruitment; lysosome acidification assay; autophagic cargo degradation assay; viral replication kinetics","journal":"Cell host & microbe","confidence":"High","confidence_rationale":"Tier 2 / Strong — systematic deletion mutant analysis with multiple orthogonal functional readouts and mechanistic identification of ubiquitin ligase; single lab but rigorous multi-method study","pmids":["38479395"],"is_preprint":false},{"year":2025,"finding":"DMXL1 forms a heterotrimeric metazoan RAVE complex (mRAVE) with WDR7 and ROGDI (linker); DMXL1 and DMXL2 interact with V1 subunits A and D of inactive V1, and upon dissipation of proton gradients, mRAVE binds both V1 and VO to form a supercomplex and catalyzes V1-VO assembly, enabling lysosomal acidification, neurotransmitter loading into vesicles, and ATG16L1/LC3-ATG8 conjugation onto single membranes.","method":"Cryo-EM/AlphaFold structural modeling, cross-linking mass spectrometry, co-immunoprecipitation, lysosomal acidification assay, neurotransmitter loading assay, ATG8 conjugation assay, mutagenesis of interaction interface","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — structural modeling supported by cross-linking proteomics plus mutagenesis of interaction interface plus multiple orthogonal functional assays; independent replication consistent with PMID 40527988","pmids":["40646309"],"is_preprint":false},{"year":2025,"finding":"DMXL1 is recruited to lysosomes and Salmonella-containing vacuoles in a CASM (conjugation of ATG8 to single membranes)-dependent manner upon TRPML1 activation; it assembles with ROGDI and WDR7 and associates with V0 and V1 subunits of the lysosomal V-ATPase; TRPML1-driven V1 subunit recruitment to lysosomes requires both DMXL1 and DMXL2; an ATP6V1A binding interface in DMXL1 was identified by AlphaFold modeling and cross-linking proteomics, and its mutation disrupts interaction and function.","method":"Quantitative lysosome proteomics; CRISPR knockout cells; immunofluorescence; co-immunoprecipitation; AlphaFold structural modeling; cross-linking proteomics; lysosomal pH measurement; hydrolytic activity assay; mutagenesis of ATP6V1A interface","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — quantitative proteomics, structural modeling with mutagenesis, multiple orthogonal functional assays and genetic knockouts; multiple independent methods and consistent with independent study PMID 40646309","pmids":["40527988"],"is_preprint":false},{"year":2025,"finding":"ROGDI was identified as a novel subunit of the mammalian Rabconnectin-3 complex (which contains DMXL1/Rabconnectin-3α and DMXL2/Rabconnectin-3β); ROGDI binds to the N-terminal domains of both Rabconnectin-3α (DMXL1) and Rabconnectin-3β, structurally mimicking yeast Rav2 binding to Rav1, and co-immunoprecipitates with Rabconnectin-3 subunits from mammalian cell lysates; ROGDI partially localizes with Rabconnectin-3α in acidic perinuclear lysosomes.","method":"Yeast complementation assay; co-immunoprecipitation; immunofluorescence microscopy; structural homology modeling; immunopurified lysosome analysis","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP, yeast complementation, and localization data from a single lab; consistent with independent structural studies","pmids":["40049412"],"is_preprint":false},{"year":2025,"finding":"Loss of Dmxl1 causes very early embryonic lethality in mice (before organogenesis), demonstrating that Dmxl1 is an essential mammalian gene required for early development; missense variants orthologous to human patient variants did not show pathogenic effects in this allelic series.","method":"CRISPR genome editing allelic series in mice (small deletion and two missense alleles); embryonic lethal phenotype assessment","journal":"Differentiation; research in biological diversity","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo knockout with clear lethal phenotype; single lab; negative result for missense alleles is also informative","pmids":["41330162"],"is_preprint":false},{"year":2025,"finding":"DMXL1 was identified as a host factor required for influenza A virus replication in chicken lung epithelial cells, with functional validation showing DMXL1 contributes to multiple stages of the IAV life cycle.","method":"Genome-wide CRISPR/Cas9 knockout screen; functional validation experiments","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — preprint; single CRISPR screen with limited mechanistic detail about DMXL1 specifically; no molecular mechanism described","pmids":["bio_10.1101_2025.10.03.680283"],"is_preprint":true}],"current_model":"DMXL1 (Rabconnectin-3α) is the central subunit of the metazoan RAVE complex (mRAVE), together with WDR7 and ROGDI, which acts as a V-ATPase assembly factor: upon dissipation of proton gradients or TRPML1/CASM-dependent lysosomal signaling, DMXL1 binds inactive V1 (via subunits A and D) and bridges it to the membrane-embedded VO to catalyze holoenzyme assembly and drive organelle acidification; DMXL1 function is regulated by TPL-2 kinase-mediated phosphorylation and is exploited by human cytomegalovirus, which targets DMXL1 for ubiquitin-mediated degradation via US33A/KPC to block lysosomal acidification."},"narrative":{"mechanistic_narrative":"DMXL1 (Rabconnectin-3α) is a large WD-repeat protein that functions as a V-ATPase assembly factor controlling organelle acidification [PMID:10708522, PMID:26442671]. It is the central subunit of the heterotrimeric metazoan RAVE complex (mRAVE), associating with WDR7 and the linker subunit ROGDI, which structurally mimics yeast Rav2 binding to Rav1 [PMID:40646309, PMID:40049412]. Through an ATP6V1A (subunit A) and subunit D binding interface defined by structural modeling and cross-linking proteomics, DMXL1 (together with DMXL2) engages inactive cytosolic V1 and bridges it to membrane-embedded VO, catalyzing holoenzyme assembly upon dissipation of proton gradients or TRPML1/CASM-dependent lysosomal signaling [PMID:40646309, PMID:40527988]. This assembly activity drives lysosomal acidification, autophagic and ATG8/CASM membrane conjugation, and neurotransmitter loading into vesicles [PMID:40646309, PMID:40527988]. In vivo, intercalated-cell-specific Dmxl1 deletion raises urine pH and impairs V1–VO co-assembly at the membrane, establishing DMXL1 as required for V-ATPase holoenzyme formation [PMID:38984989], and TPL-2 kinase phosphorylates DMXL1 to promote phagosome acidification and bacterial killing in macrophages [PMID:33881780]. DMXL1 is essential for early mouse development, with knockout causing pre-organogenesis embryonic lethality [PMID:41330162]. Human cytomegalovirus exploits this axis: its US33A protein recruits the KPC E3 ubiquitin ligase to ubiquitinate and degrade DMXL1, blocking lysosomal acidification and autophagic degradation [PMID:38479395].","teleology":[{"year":2000,"claim":"Established the molecular identity of DMXL1 as a large WD-repeat protein and human homologue of Drosophila DmX, providing the gene structure on which all later functional work was built.","evidence":"cDNA cloning, radiation hybrid mapping, and FISH","pmids":["10708522"],"confidence":"Medium","gaps":["No functional or biochemical role assigned","No interaction partners identified","Cellular localization not determined"]},{"year":2015,"claim":"Connected DMXL1 to the V-ATPase by showing physical association and a requirement for vesicle acidification, defining its functional pathway for the first time.","evidence":"MS-based V-ATPase interactome and siRNA knockdown acidification assay in a kidney cell line","pmids":["26442671"],"confidence":"Medium","gaps":["Did not define which V-ATPase subunits DMXL1 contacts","Mechanism of action on V-ATPase not resolved","Single cell-line context"]},{"year":2021,"claim":"Identified a regulatory input on DMXL1, showing that TPL-2 phosphorylation promotes V-ATPase assembly and links DMXL1 to innate-immune bacterial killing.","evidence":"Phagosome proteomics, TPL-2 inhibitor, phosphosite MS, and bacterial killing assays in primary mouse and human macrophages","pmids":["33881780"],"confidence":"High","gaps":["Phosphosite-to-assembly mechanistic link not structurally resolved","Other kinases/phosphatases acting on DMXL1 unknown"]},{"year":2024,"claim":"Provided in vivo proof that DMXL1 is required for V-ATPase holoenzyme assembly in a physiological epithelium, by showing impaired V1–VO co-assembly and acidification on knockout.","evidence":"Intercalated-cell conditional KO mice with urine pH, western blot, fractionation, PLA, and immunofluorescence","pmids":["38984989"],"confidence":"High","gaps":["Molecular interface mediating assembly not yet defined here","Tissue-restricted readout"]},{"year":2024,"claim":"Revealed DMXL1 as a viral target, defining a US33A/KPC ubiquitin-degradation mechanism by which HCMV disables lysosomal acidification and autophagy.","evidence":"HCMV proteomics, deletion mutants, co-IP, lysosome acidification and autophagy assays","pmids":["38479395"],"confidence":"High","gaps":["Ubiquitination site on DMXL1 not mapped","Consequences for V-ATPase complex stoichiometry not detailed"]},{"year":2025,"claim":"Resolved the molecular architecture, defining DMXL1 as the core of the heterotrimeric mRAVE complex with WDR7 and ROGDI and mapping its V1 subunit A/D binding interfaces that catalyze V1–VO supercomplex assembly.","evidence":"Cryo-EM/AlphaFold modeling, cross-linking MS, co-IP, mutagenesis, and acidification/neurotransmitter/ATG8 conjugation assays; ROGDI subunit and TRPML1/CASM-dependent recruitment confirmed across independent studies","pmids":["40646309","40527988","40049412"],"confidence":"High","gaps":["Conformational steps of catalyzed assembly not fully visualized","Division of labor between DMXL1 and DMXL2 not fully defined"]},{"year":2025,"claim":"Established DMXL1 as an essential mammalian gene, showing knockout causes pre-organogenesis embryonic lethality while patient-orthologous missense alleles were not pathogenic in mice.","evidence":"CRISPR allelic series in mice with embryonic lethality assessment","pmids":["41330162"],"confidence":"Medium","gaps":["Developmental process requiring DMXL1 not identified","Discrepancy with human variant pathogenicity unexplained"]},{"year":null,"claim":"How DMXL1-driven V-ATPase assembly is selectively deployed across tissues, signaling states, and cellular compartments — and the basis of its essential developmental role — remains open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["Functional distinction between DMXL1 and DMXL2 not resolved","Developmental substrate of essentiality unknown","Full catalytic cycle of assembly not structurally defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[3,5,6]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[5,6]}],"localization":[{"term_id":"GO:0005764","term_label":"lysosome","supporting_discovery_ids":[5,6,7]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[5]}],"pathway":[{"term_id":"R-HSA-382551","term_label":"Transport of small molecules","supporting_discovery_ids":[1,3]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[5,6,4]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[3,5]}],"complexes":["metazoan RAVE complex (mRAVE)","Rabconnectin-3 complex","V-ATPase"],"partners":["WDR7","ROGDI","DMXL2","ATP6V1A","TPL-2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y485","full_name":"DmX-like protein 1","aliases":[],"length_aa":3027,"mass_kda":337.8,"function":"","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q9Y485/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/DMXL1","classification":"Not Classified","n_dependent_lines":39,"n_total_lines":1208,"dependency_fraction":0.03228476821192053},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"ATP6V1B2","stoichiometry":0.2},{"gene":"ATP6V1E1","stoichiometry":0.2},{"gene":"ATP6V1F","stoichiometry":0.2},{"gene":"ATP6V1G1","stoichiometry":0.2},{"gene":"ATP6V1H","stoichiometry":0.2},{"gene":"DDOST","stoichiometry":0.2},{"gene":"OST4","stoichiometry":0.2},{"gene":"RPN1","stoichiometry":0.2},{"gene":"RPN2","stoichiometry":0.2},{"gene":"STK4","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/DMXL1","total_profiled":1310},"omim":[{"mim_id":"612186","title":"DMX-LIKE 2; DMXL2","url":"https://www.omim.org/entry/612186"},{"mim_id":"605671","title":"DMX-LIKE 1; DMXL1","url":"https://www.omim.org/entry/605671"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoli","reliability":"Approved"},{"location":"Nucleoli fibrillar center","reliability":"Approved"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/DMXL1"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"Q9Y485","domains":[],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y485","model_url":"","pae_url":"","plddt_mean":null},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=DMXL1","jax_strain_url":"https://www.jax.org/strain/search?query=DMXL1"},"sequence":{"accession":"Q9Y485","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y485.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y485/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y485"}},"corpus_meta":[{"pmid":"26442671","id":"PMC_26442671","title":"Mapping the H(+) (V)-ATPase interactome: identification of proteins involved in trafficking, folding, assembly and phosphorylation.","date":"2015","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/26442671","citation_count":110,"is_preprint":false},{"pmid":"16865689","id":"PMC_16865689","title":"Identification of novel genes associated with astrocytoma progression using suppression subtractive hybridization and real-time reverse transcription-polymerase chain reaction.","date":"2006","source":"International journal of cancer","url":"https://pubmed.ncbi.nlm.nih.gov/16865689","citation_count":55,"is_preprint":false},{"pmid":"21310917","id":"PMC_21310917","title":"Copy number variations and primary open-angle glaucoma.","date":"2011","source":"Investigative ophthalmology & visual science","url":"https://pubmed.ncbi.nlm.nih.gov/21310917","citation_count":33,"is_preprint":false},{"pmid":"36776048","id":"PMC_36776048","title":"Machine Learning Selection of Most Predictive Brain Proteins Suggests Role of Sugar Metabolism in Alzheimer's Disease.","date":"2023","source":"Journal of Alzheimer's disease : JAD","url":"https://pubmed.ncbi.nlm.nih.gov/36776048","citation_count":30,"is_preprint":false},{"pmid":"33881780","id":"PMC_33881780","title":"TPL-2 kinase induces phagosome acidification to promote macrophage killing of bacteria.","date":"2021","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/33881780","citation_count":23,"is_preprint":false},{"pmid":"15742475","id":"PMC_15742475","title":"Interstitial deletion of the long arm of chromosome 5 in a boy with multiple congenital anomalies and mental retardation: Molecular characterization of the deleted region to 5q22.3q23.3.","date":"2005","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/15742475","citation_count":23,"is_preprint":false},{"pmid":"10708522","id":"PMC_10708522","title":"Mapping and structure of DMXL1, a human homologue of the DmX gene from Drosophila melanogaster coding for a WD repeat protein.","date":"2000","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/10708522","citation_count":16,"is_preprint":false},{"pmid":"38984989","id":"PMC_38984989","title":"Dmxl1 Is an Essential Mammalian Gene that Is Required for V-ATPase Assembly and Function In Vivo.","date":"2024","source":"Function (Oxford, England)","url":"https://pubmed.ncbi.nlm.nih.gov/38984989","citation_count":13,"is_preprint":false},{"pmid":"40527988","id":"PMC_40527988","title":"DMXL1 promotes recruitment of V1-ATPase to lysosomes upon TRPML1 activation.","date":"2025","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/40527988","citation_count":11,"is_preprint":false},{"pmid":"18224412","id":"PMC_18224412","title":"Gene profiling of growth factor independence 1B gene (Gfi-1B) in leukemic cells.","date":"2007","source":"International journal of hematology","url":"https://pubmed.ncbi.nlm.nih.gov/18224412","citation_count":11,"is_preprint":false},{"pmid":"38479395","id":"PMC_38479395","title":"Human cytomegalovirus degrades DMXL1 to inhibit autophagy, lysosomal acidification, and viral assembly.","date":"2024","source":"Cell host & microbe","url":"https://pubmed.ncbi.nlm.nih.gov/38479395","citation_count":9,"is_preprint":false},{"pmid":"35287291","id":"PMC_35287291","title":"Swelling-induced upregulation of miR-141-3p inhibits hepatocyte proliferation.","date":"2022","source":"JHEP reports : innovation in hepatology","url":"https://pubmed.ncbi.nlm.nih.gov/35287291","citation_count":9,"is_preprint":false},{"pmid":"35702937","id":"PMC_35702937","title":"Genom-wide analysis identifies single nucleotide polymorphism variations and altered pathways associated with poor semen quality in breeding bulls.","date":"2022","source":"Reproduction in domestic animals = Zuchthygiene","url":"https://pubmed.ncbi.nlm.nih.gov/35702937","citation_count":6,"is_preprint":false},{"pmid":"40646309","id":"PMC_40646309","title":"A heterotrimeric protein complex assembles the metazoan V-ATPase upon dissipation of proton gradients.","date":"2025","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/40646309","citation_count":4,"is_preprint":false},{"pmid":"40049412","id":"PMC_40049412","title":"The ROGDI protein mutated in Kohlschutter-Tonz syndrome is a novel subunit of the Rabconnectin-3 complex implicated in V-ATPase assembly.","date":"2025","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/40049412","citation_count":4,"is_preprint":false},{"pmid":"37559881","id":"PMC_37559881","title":"Secondary leiomyosarcoma of the nasal cavity in a treated patient with possible hereditary retinoblastoma with germline reciprocal translocation of RB1 and DMXL1 and somatic TP53 mutation: A case report.","date":"2023","source":"Molecular and clinical oncology","url":"https://pubmed.ncbi.nlm.nih.gov/37559881","citation_count":2,"is_preprint":false},{"pmid":"39401425","id":"PMC_39401425","title":"IQGAP-2: a novel interacting partner with the human colonic thiamin pyrophosphate transporter.","date":"2024","source":"American journal of physiology. 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the gene was mapped to chromosome 5q22 by radiation hybrid mapping and FISH, and its open reading frame structure was characterized by cDNA cloning.\",\n      \"method\": \"cDNA cloning, radiation hybrid mapping, fluorescence in situ hybridization (FISH)\",\n      \"journal\": \"Genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct molecular cloning and chromosomal mapping with two orthogonal localization methods; no functional assay performed\",\n      \"pmids\": [\"10708522\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"DMXL1 physically associates with the V-ATPase complex in kidney tissue; knockdown of DMXL1 inhibits V-ATPase-mediated intracellular vesicle acidification in a kidney cell line.\",\n      \"method\": \"Proteomic co-immunoprecipitation / V-ATPase interactome mapping; siRNA knockdown with vesicle acidification assay\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal MS-based interactome plus functional siRNA knockdown phenotype in a cell line, single lab\",\n      \"pmids\": [\"26442671\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TPL-2 kinase phosphorylates DMXL1, and this phosphorylation promotes V-ATPase assembly and phagosome acidification in macrophages, thereby enabling efficient killing of phagocytosed bacteria (Staphylococcus aureus, Citrobacter rodentium).\",\n      \"method\": \"Quantitative phagosome proteomics; TPL-2 catalytic inhibitor experiments; phosphorylation detected by mass spectrometry; phagosome acidification assay; bacterial killing assay in primary mouse and human macrophages\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (proteomics, inhibitor, phosphorylation MS, functional acidification and killing assays) in primary human and mouse cells; single lab but rigorous multi-method study\",\n      \"pmids\": [\"33881780\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Kidney intercalated cell-specific Dmxl1 knockout mice show elevated urine pH and impaired proton pumping, with decreased B1 (V1) subunit levels and reduced co-localization of V1 (B1) and VO (a4) subunits at the membrane, indicating that Dmxl1 is required for V-ATPase holoenzyme assembly in vivo.\",\n      \"method\": \"Conditional knockout mouse model; western blotting; immunofluorescence co-localization; subcellular fractionation; proximity ligation assay (PLA) for B1-a4 subunit association\",\n      \"journal\": \"Function (Oxford, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo conditional KO with multiple orthogonal readouts (urine pH, western blot, fractionation, PLA, immunofluorescence), single lab but rigorous multi-method study\",\n      \"pmids\": [\"38984989\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Human cytomegalovirus degrades DMXL1 via its US33A protein, which recruits the E3 ubiquitin ligase KPC (Kip1 ubiquitination-promoting complex) to ubiquitinate and degrade DMXL1; this degradation inhibits lysosome acidification, autophagic cargo degradation, and delays virion assembly compartment formation.\",\n      \"method\": \"Quantitative proteomics of HCMV-infected cells; systematic viral deletion mutant comparison; co-immunoprecipitation to identify KPC recruitment; lysosome acidification assay; autophagic cargo degradation assay; viral replication kinetics\",\n      \"journal\": \"Cell host & microbe\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — systematic deletion mutant analysis with multiple orthogonal functional readouts and mechanistic identification of ubiquitin ligase; single lab but rigorous multi-method study\",\n      \"pmids\": [\"38479395\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"DMXL1 forms a heterotrimeric metazoan RAVE complex (mRAVE) with WDR7 and ROGDI (linker); DMXL1 and DMXL2 interact with V1 subunits A and D of inactive V1, and upon dissipation of proton gradients, mRAVE binds both V1 and VO to form a supercomplex and catalyzes V1-VO assembly, enabling lysosomal acidification, neurotransmitter loading into vesicles, and ATG16L1/LC3-ATG8 conjugation onto single membranes.\",\n      \"method\": \"Cryo-EM/AlphaFold structural modeling, cross-linking mass spectrometry, co-immunoprecipitation, lysosomal acidification assay, neurotransmitter loading assay, ATG8 conjugation assay, mutagenesis of interaction interface\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — structural modeling supported by cross-linking proteomics plus mutagenesis of interaction interface plus multiple orthogonal functional assays; independent replication consistent with PMID 40527988\",\n      \"pmids\": [\"40646309\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"DMXL1 is recruited to lysosomes and Salmonella-containing vacuoles in a CASM (conjugation of ATG8 to single membranes)-dependent manner upon TRPML1 activation; it assembles with ROGDI and WDR7 and associates with V0 and V1 subunits of the lysosomal V-ATPase; TRPML1-driven V1 subunit recruitment to lysosomes requires both DMXL1 and DMXL2; an ATP6V1A binding interface in DMXL1 was identified by AlphaFold modeling and cross-linking proteomics, and its mutation disrupts interaction and function.\",\n      \"method\": \"Quantitative lysosome proteomics; CRISPR knockout cells; immunofluorescence; co-immunoprecipitation; AlphaFold structural modeling; cross-linking proteomics; lysosomal pH measurement; hydrolytic activity assay; mutagenesis of ATP6V1A interface\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — quantitative proteomics, structural modeling with mutagenesis, multiple orthogonal functional assays and genetic knockouts; multiple independent methods and consistent with independent study PMID 40646309\",\n      \"pmids\": [\"40527988\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ROGDI was identified as a novel subunit of the mammalian Rabconnectin-3 complex (which contains DMXL1/Rabconnectin-3α and DMXL2/Rabconnectin-3β); ROGDI binds to the N-terminal domains of both Rabconnectin-3α (DMXL1) and Rabconnectin-3β, structurally mimicking yeast Rav2 binding to Rav1, and co-immunoprecipitates with Rabconnectin-3 subunits from mammalian cell lysates; ROGDI partially localizes with Rabconnectin-3α in acidic perinuclear lysosomes.\",\n      \"method\": \"Yeast complementation assay; co-immunoprecipitation; immunofluorescence microscopy; structural homology modeling; immunopurified lysosome analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP, yeast complementation, and localization data from a single lab; consistent with independent structural studies\",\n      \"pmids\": [\"40049412\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Loss of Dmxl1 causes very early embryonic lethality in mice (before organogenesis), demonstrating that Dmxl1 is an essential mammalian gene required for early development; missense variants orthologous to human patient variants did not show pathogenic effects in this allelic series.\",\n      \"method\": \"CRISPR genome editing allelic series in mice (small deletion and two missense alleles); embryonic lethal phenotype assessment\",\n      \"journal\": \"Differentiation; research in biological diversity\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo knockout with clear lethal phenotype; single lab; negative result for missense alleles is also informative\",\n      \"pmids\": [\"41330162\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"DMXL1 was identified as a host factor required for influenza A virus replication in chicken lung epithelial cells, with functional validation showing DMXL1 contributes to multiple stages of the IAV life cycle.\",\n      \"method\": \"Genome-wide CRISPR/Cas9 knockout screen; functional validation experiments\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — preprint; single CRISPR screen with limited mechanistic detail about DMXL1 specifically; no molecular mechanism described\",\n      \"pmids\": [\"bio_10.1101_2025.10.03.680283\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"DMXL1 (Rabconnectin-3α) is the central subunit of the metazoan RAVE complex (mRAVE), together with WDR7 and ROGDI, which acts as a V-ATPase assembly factor: upon dissipation of proton gradients or TRPML1/CASM-dependent lysosomal signaling, DMXL1 binds inactive V1 (via subunits A and D) and bridges it to the membrane-embedded VO to catalyze holoenzyme assembly and drive organelle acidification; DMXL1 function is regulated by TPL-2 kinase-mediated phosphorylation and is exploited by human cytomegalovirus, which targets DMXL1 for ubiquitin-mediated degradation via US33A/KPC to block lysosomal acidification.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"DMXL1 (Rabconnectin-3α) is a large WD-repeat protein that functions as a V-ATPase assembly factor controlling organelle acidification [#0, #1]. It is the central subunit of the heterotrimeric metazoan RAVE complex (mRAVE), associating with WDR7 and the linker subunit ROGDI, which structurally mimics yeast Rav2 binding to Rav1 [#5, #7]. Through an ATP6V1A (subunit A) and subunit D binding interface defined by structural modeling and cross-linking proteomics, DMXL1 (together with DMXL2) engages inactive cytosolic V1 and bridges it to membrane-embedded VO, catalyzing holoenzyme assembly upon dissipation of proton gradients or TRPML1/CASM-dependent lysosomal signaling [#5, #6]. This assembly activity drives lysosomal acidification, autophagic and ATG8/CASM membrane conjugation, and neurotransmitter loading into vesicles [#5, #6]. In vivo, intercalated-cell-specific Dmxl1 deletion raises urine pH and impairs V1–VO co-assembly at the membrane, establishing DMXL1 as required for V-ATPase holoenzyme formation [#3], and TPL-2 kinase phosphorylates DMXL1 to promote phagosome acidification and bacterial killing in macrophages [#2]. DMXL1 is essential for early mouse development, with knockout causing pre-organogenesis embryonic lethality [#8]. Human cytomegalovirus exploits this axis: its US33A protein recruits the KPC E3 ubiquitin ligase to ubiquitinate and degrade DMXL1, blocking lysosomal acidification and autophagic degradation [#4].\",\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Established the molecular identity of DMXL1 as a large WD-repeat protein and human homologue of Drosophila DmX, providing the gene structure on which all later functional work was built.\",\n      \"evidence\": \"cDNA cloning, radiation hybrid mapping, and FISH\",\n      \"pmids\": [\"10708522\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No functional or biochemical role assigned\", \"No interaction partners identified\", \"Cellular localization not determined\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Connected DMXL1 to the V-ATPase by showing physical association and a requirement for vesicle acidification, defining its functional pathway for the first time.\",\n      \"evidence\": \"MS-based V-ATPase interactome and siRNA knockdown acidification assay in a kidney cell line\",\n      \"pmids\": [\"26442671\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define which V-ATPase subunits DMXL1 contacts\", \"Mechanism of action on V-ATPase not resolved\", \"Single cell-line context\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified a regulatory input on DMXL1, showing that TPL-2 phosphorylation promotes V-ATPase assembly and links DMXL1 to innate-immune bacterial killing.\",\n      \"evidence\": \"Phagosome proteomics, TPL-2 inhibitor, phosphosite MS, and bacterial killing assays in primary mouse and human macrophages\",\n      \"pmids\": [\"33881780\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphosite-to-assembly mechanistic link not structurally resolved\", \"Other kinases/phosphatases acting on DMXL1 unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Provided in vivo proof that DMXL1 is required for V-ATPase holoenzyme assembly in a physiological epithelium, by showing impaired V1–VO co-assembly and acidification on knockout.\",\n      \"evidence\": \"Intercalated-cell conditional KO mice with urine pH, western blot, fractionation, PLA, and immunofluorescence\",\n      \"pmids\": [\"38984989\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular interface mediating assembly not yet defined here\", \"Tissue-restricted readout\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Revealed DMXL1 as a viral target, defining a US33A/KPC ubiquitin-degradation mechanism by which HCMV disables lysosomal acidification and autophagy.\",\n      \"evidence\": \"HCMV proteomics, deletion mutants, co-IP, lysosome acidification and autophagy assays\",\n      \"pmids\": [\"38479395\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Ubiquitination site on DMXL1 not mapped\", \"Consequences for V-ATPase complex stoichiometry not detailed\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Resolved the molecular architecture, defining DMXL1 as the core of the heterotrimeric mRAVE complex with WDR7 and ROGDI and mapping its V1 subunit A/D binding interfaces that catalyze V1–VO supercomplex assembly.\",\n      \"evidence\": \"Cryo-EM/AlphaFold modeling, cross-linking MS, co-IP, mutagenesis, and acidification/neurotransmitter/ATG8 conjugation assays; ROGDI subunit and TRPML1/CASM-dependent recruitment confirmed across independent studies\",\n      \"pmids\": [\"40646309\", \"40527988\", \"40049412\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conformational steps of catalyzed assembly not fully visualized\", \"Division of labor between DMXL1 and DMXL2 not fully defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Established DMXL1 as an essential mammalian gene, showing knockout causes pre-organogenesis embryonic lethality while patient-orthologous missense alleles were not pathogenic in mice.\",\n      \"evidence\": \"CRISPR allelic series in mice with embryonic lethality assessment\",\n      \"pmids\": [\"41330162\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Developmental process requiring DMXL1 not identified\", \"Discrepancy with human variant pathogenicity unexplained\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How DMXL1-driven V-ATPase assembly is selectively deployed across tissues, signaling states, and cellular compartments — and the basis of its essential developmental role — remains open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional distinction between DMXL1 and DMXL2 not resolved\", \"Developmental substrate of essentiality unknown\", \"Full catalytic cycle of assembly not structurally defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [3, 5, 6]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [5, 6]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005764\", \"supporting_discovery_ids\": [5, 6, 7]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-382551\", \"supporting_discovery_ids\": [1, 3]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [5, 6, 4]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [3, 5]}\n    ],\n    \"complexes\": [\n      \"metazoan RAVE complex (mRAVE)\",\n      \"Rabconnectin-3 complex\",\n      \"V-ATPase\"\n    ],\n    \"partners\": [\n      \"WDR7\",\n      \"ROGDI\",\n      \"DMXL2\",\n      \"ATP6V1A\",\n      \"TPL-2\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":7,"faith_total":7,"faith_pct":100.0}}