{"gene":"DDOST","run_date":"2026-06-09T23:54:41","timeline":{"discoveries":[{"year":1996,"finding":"The AGE-binding protein p60 isolated from rat liver membranes is identical to OST-48 (DDOST), a 48-kDa subunit of the oligosaccharyltransferase complex. Immunoprecipitated OST-48 from rat rough endoplasmic reticulum fractions exhibited AGE-specific binding, and anti-OST-48 IgG inhibited AGE-BSA binding to cell membranes in a dose-dependent manner. OST-48 is also surface-expressed on mononuclear, endothelial, renal, and neuronal cells.","method":"N-terminal protein sequencing, immunoprecipitation, AGE-ligand binding assay, Western blot, immunostaining, flow cytometry","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal immunoprecipitation plus AGE-binding assay and blocking antibody experiment in a single lab with multiple orthogonal methods","pmids":["8855306"],"is_preprint":false},{"year":2001,"finding":"ER localisation of OST48 (DDOST) is conferred by two cytosolic C-terminal lysine residues (Lys-3 and Lys-5 from the C-terminus). These lysines act as a retrieval/relocation signal rather than a primary retention signal. Co-expression with ribophorin I quantitatively retained an OST48 surface-escape mutant in the ER, indicating that ER localisation is primarily achieved through complex formation with ribophorin I involving their luminal domains.","method":"Site-directed mutagenesis of OST48 cytosolic lysines, immunofluorescence microscopy of COS-1 cells expressing OST48 mutants and hybrid proteins, co-expression epistasis","journal":"Biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — mutagenesis combined with immunofluorescence localization and co-expression rescue, single lab","pmids":["11530934"],"is_preprint":false},{"year":2012,"finding":"OST48 (DDOST) and DAD1 are required for the stable assembly of both STT3A- and STT3B-containing oligosaccharyltransferase complexes. Knockdown of OST48 destabilises these complexes and causes pronounced global hypoglycosylation of N-glycosylation substrates.","method":"siRNA knockdown of OST48/DAD1/KCP2 in mammalian cells, biochemical analysis of OST complex stability, glycosylation efficiency assays","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — subunit-specific knockdown with OST complex stability readout and N-glycosylation phenotype, multiple subunits tested in parallel as controls, single lab with multiple orthogonal methods","pmids":["22467853"],"is_preprint":false},{"year":2012,"finding":"Biallelic loss-of-function mutations in DDOST (a 22 bp deletion and a missense mutation) cause decreased N-glycosylation in patient fibroblasts, and complementation with wild-type DDOST cDNA restores glycosylation, establishing DDOST as a causative gene in congenital disorders of glycosylation (CDG).","method":"Whole-exome sequencing, biochemical glycosylation assays (three biomarkers), cDNA complementation in patient fibroblasts","journal":"American journal of human genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic identification combined with functional complementation in patient cells using multiple biochemical readouts","pmids":["22305527"],"is_preprint":false},{"year":2012,"finding":"DDOST (OST48) physically interacts with the testis-specific protein phosphatase isoform PPP1CC2. The interaction was identified by tandem affinity purification and confirmed by reciprocal in vitro sedimentation assay. DDOST localises to the nuclear envelope in dissociated spermatogenic cells.","method":"Tandem affinity purification from knock-in mouse embryonic stem cells, reciprocal in vitro sedimentation assay, immunolocalization","journal":"Biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — TAP identification confirmed by reciprocal in vitro sedimentation, single lab","pmids":["23140390"],"is_preprint":false},{"year":2022,"finding":"Upon DNA viral infection, MITA/STING undergoes DDOST-mediated N-glycosylation in the ER. Selective mutation of the DDOST-dependent N-glycosylation sites on MITA abolished MITA oligomerization and downstream immune signalling. Increasing Ddost expression in mouse brain enhanced local immune response to HSV-1 and prolonged survival in HSV encephalitis.","method":"N-glycosylation site mutagenesis of MITA, DDOST knockdown/overexpression in cells and in vivo mouse brain, MITA oligomerization assay, viral infection model","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 1 / Strong — mutagenesis of substrate glycosylation sites combined with functional oligomerization assay and in vivo rescue experiment","pmids":["36449507"],"is_preprint":false},{"year":2023,"finding":"OST48 (DDOST) transiently interacts with lysyl oxidase (LOX), a secreted extracellular matrix cross-linking enzyme, and is required for efficient N-glycosylation of LOX. An assay based on LOX N-glycosylation was used to functionally classify DDOST variants, identifying p.S243F and p.E286del as pathogenic loss-of-function variants.","method":"Proximity labelling to identify OST48 interaction with LOX, LOX N-glycosylation functional assay, variant complementation testing in cells","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proximity labelling interaction plus functional glycosylation assay with variant testing, single lab","pmids":["37848450"],"is_preprint":false},{"year":2024,"finding":"DDOST knockdown in pancreatic ductal adenocarcinoma (PDAC) cells decreased proliferation and cell viability and increased ER stress, ROS formation, and apoptosis. Quantitative mass spectrometry identified 30 differentially expressed proteins/phosphopeptides after DDOST knockdown.","method":"siRNA knockdown of DDOST in two PDAC cell lines, quantitative mass spectrometry proteomics, cell viability and apoptosis assays, ROS measurement","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knockdown with defined cellular phenotype readout (apoptosis, ER stress, ROS) validated in two cell lines, single lab","pmids":["39223141"],"is_preprint":false},{"year":2025,"finding":"DDOST depletion in HCC cells impaired EGFR N-glycosylation, suppressing downstream AKT, ERK5 and ERK1/2 signalling and sensitising cells to lenvatinib. DDOST depletion also reduced PD-L1 glycosylation. The OST inhibitor NGI-1 phenocopied these effects in vitro and in vivo.","method":"DDOST knockdown in HCC cell lines and xenograft models, EGFR and PD-L1 N-glycosylation assays, Western blot for downstream signalling, pharmacological inhibition with NGI-1","journal":"Experimental & molecular medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knockdown with glycosylation and downstream signalling readouts confirmed in vivo, single lab","pmids":["41413687"],"is_preprint":false},{"year":2025,"finding":"Macrophage-derived exosome miR-146a-5p suppresses PNKP expression, reducing PNKP–DDOST interaction and thereby enhancing DDOST phosphorylation, which activates JAGN1-dependent neutrophil extracellular trap (NET) formation in atherosclerosis.","method":"Co-immunoprecipitation, RIP assay, dual-luciferase reporter assay, Western blot, qRT-PCR, ApoE-/- mouse AS model","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-immunoprecipitation of PNKP-DDOST complex with miR-146a-5p modulation and in vivo mouse model, single lab","pmids":["41109654"],"is_preprint":false}],"current_model":"DDOST encodes OST48, a non-catalytic but structurally essential subunit of the ER oligosaccharyltransferase complex that is required for stable assembly of both STT3A- and STT3B-containing OST complexes and hence for global protein N-glycosylation; it catalyses N-glycosylation of specific substrates including MITA/STING (enabling MITA oligomerization and innate immune signalling), EGFR, and PD-L1; it is retained in the ER via its luminal interaction with ribophorin I and a cytosolic di-lysine retrieval motif; it binds AGEs at the cell surface; it physically interacts with PPP1CC2 at the nuclear envelope in spermatogenic cells; and its phosphorylation state is regulated by the PNKP kinase, linking it to NET formation in neutrophils."},"narrative":{"mechanistic_narrative":"DDOST encodes OST48, a structurally essential subunit of the endoplasmic reticulum oligosaccharyltransferase (OST) complex that is required for the stable assembly of both STT3A- and STT3B-containing OST complexes; its loss destabilizes these complexes and causes pronounced global hypoglycosylation of N-glycosylation substrates [PMID:22467853]. Biallelic loss-of-function mutations in DDOST reduce N-glycosylation in patient fibroblasts and are rescued by wild-type cDNA, establishing DDOST as a causative gene in congenital disorders of glycosylation [PMID:22305527]. ER localization of OST48 is achieved primarily through luminal complex formation with ribophorin I, supported by a cytosolic C-terminal di-lysine retrieval signal [PMID:11530934]. Beyond its global role, OST48 mediates N-glycosylation of specific substrates with defined functional consequences: it glycosylates MITA/STING to enable MITA oligomerization and antiviral innate immune signaling [PMID:36449507], the secreted matrix cross-linker lysyl oxidase [PMID:37848450], and in cancer cells EGFR and PD-L1, where DDOST loss suppresses downstream AKT/ERK signaling and sensitizes hepatocellular carcinoma to lenvatinib [PMID:41413687]. DDOST was originally identified as the AGE-binding protein p60 with cell-surface AGE-binding activity [PMID:8855306], and additional context-specific roles include a physical interaction with the testis-specific phosphatase PPP1CC2 at the nuclear envelope in spermatogenic cells [PMID:23140390] and regulation of its phosphorylation by PNKP linking it to neutrophil extracellular trap formation [PMID:41109654].","teleology":[{"year":1996,"claim":"Established a surprising dual identity for DDOST, showing the AGE-binding membrane protein p60 is the same protein as the 48-kDa OST subunit and is also displayed at the cell surface.","evidence":"N-terminal sequencing, immunoprecipitation, AGE-binding and blocking-antibody assays in rat liver membranes","pmids":["8855306"],"confidence":"Medium","gaps":["Functional significance of surface AGE binding not established","Single-lab characterization without independent confirmation","Relationship between AGE-binding and OST function unaddressed"]},{"year":2001,"claim":"Resolved how OST48 is retained in the ER, showing localization depends primarily on luminal complex formation with ribophorin I rather than its cytosolic di-lysine motif alone.","evidence":"Site-directed mutagenesis and immunofluorescence co-expression epistasis in COS-1 cells","pmids":["11530934"],"confidence":"Medium","gaps":["Structural basis of luminal ribophorin I interaction not defined","Mechanism of di-lysine retrieval not detailed"]},{"year":2012,"claim":"Defined OST48 as structurally essential for OST complex assembly, demonstrating its knockdown destabilizes both STT3A and STT3B complexes and globally impairs N-glycosylation.","evidence":"Subunit-specific siRNA knockdown with OST complex stability and glycosylation readouts in mammalian cells","pmids":["22467853"],"confidence":"High","gaps":["Catalytic versus purely structural contribution not separated","No structural model of OST48 within the assembled complex"]},{"year":2012,"claim":"Established DDOST as a human disease gene by linking biallelic loss-of-function mutations to a congenital disorder of glycosylation with functional rescue.","evidence":"Whole-exome sequencing with biochemical glycosylation assays and cDNA complementation in patient fibroblasts","pmids":["22305527"],"confidence":"High","gaps":["Genotype-phenotype spectrum limited to few patients","Tissue-specific disease mechanism not dissected"]},{"year":2012,"claim":"Identified a context-specific interaction of DDOST with the testis-specific phosphatase PPP1CC2 and nuclear-envelope localization in spermatogenic cells, hinting at roles beyond ER glycosylation.","evidence":"Tandem affinity purification with reciprocal in vitro sedimentation and immunolocalization in spermatogenic cells","pmids":["23140390"],"confidence":"Medium","gaps":["Functional consequence of the PPP1CC2 interaction unknown","Mechanism of nuclear-envelope localization unexplained","Single-lab finding"]},{"year":2022,"claim":"Showed DDOST acts on a specific signaling substrate, glycosylating MITA/STING to enable oligomerization and antiviral immune signaling, with in vivo protective effect against HSV encephalitis.","evidence":"Substrate glycosylation-site mutagenesis, DDOST knockdown/overexpression, oligomerization assay and in vivo mouse brain viral infection model","pmids":["36449507"],"confidence":"High","gaps":["Substrate selectivity determinants not defined","Whether effect requires full OST complex or DDOST specifically not separated"]},{"year":2023,"claim":"Extended DDOST substrate range to the secreted matrix enzyme lysyl oxidase and leveraged this to functionally classify pathogenic DDOST variants.","evidence":"Proximity labelling, LOX N-glycosylation functional assay and variant complementation in cells","pmids":["37848450"],"confidence":"Medium","gaps":["Transient nature of the LOX interaction not structurally characterized","Single-lab assay"]},{"year":2024,"claim":"Linked DDOST to cancer cell survival, showing its knockdown in PDAC cells triggers ER stress, ROS, and apoptosis.","evidence":"siRNA knockdown in two PDAC cell lines with quantitative proteomics, viability, apoptosis and ROS assays","pmids":["39223141"],"confidence":"Medium","gaps":["Specific glycoprotein substrates driving the phenotype not identified","In vivo relevance untested"]},{"year":2025,"claim":"Connected DDOST-dependent glycosylation to oncogenic receptor signaling and therapy response, showing depletion impairs EGFR and PD-L1 glycosylation and sensitizes HCC to lenvatinib.","evidence":"DDOST knockdown and NGI-1 inhibition in HCC cell lines and xenografts with glycosylation and signaling readouts","pmids":["41413687"],"confidence":"Medium","gaps":["Direct versus complex-mediated glycosylation of EGFR/PD-L1 not distinguished","Clinical translatability untested"]},{"year":2025,"claim":"Implicated DDOST phosphorylation, controlled by PNKP, in neutrophil extracellular trap formation during atherosclerosis.","evidence":"Co-IP, RIP, dual-luciferase reporter assays and ApoE-/- mouse atherosclerosis model","pmids":["41109654"],"confidence":"Medium","gaps":["Phosphorylation site(s) on DDOST not mapped","Mechanistic link between DDOST phosphorylation and NET machinery indirect","Single-lab finding"]},{"year":null,"claim":"How DDOST achieves substrate selectivity and whether its non-glycosylation roles (AGE binding, PPP1CC2 interaction, phosphorylation-dependent NET regulation) are mechanistically separable from OST complex function remains open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of DDOST substrate engagement","Moonlighting functions not reconciled with ER glycosylation role"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[2,5,6,8]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[2,3]}],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[1,2,5]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0]},{"term_id":"GO:0005635","term_label":"nuclear envelope","supporting_discovery_ids":[4]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[2,3]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[5]}],"complexes":["oligosaccharyltransferase (OST) complex"],"partners":["RPN1","PPP1CC2","LOX","PNKP","STING1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P39656","full_name":"Dolichyl-diphosphooligosaccharide--protein glycosyltransferase 48 kDa subunit","aliases":[],"length_aa":456,"mass_kda":50.8,"function":"Subunit of the oligosaccharyl transferase (OST) complex that catalyzes the initial transfer of a defined glycan (Glc(3)Man(9)GlcNAc(2) in eukaryotes) from the lipid carrier dolichol-pyrophosphate to an asparagine residue within an Asn-X-Ser/Thr consensus motif in nascent polypeptide chains, the first step in protein N-glycosylation (PubMed:31831667). N-glycosylation occurs cotranslationally and the complex associates with the Sec61 complex at the channel-forming translocon complex that mediates protein translocation across the endoplasmic reticulum (ER). All subunits are required for a maximal enzyme activity (By similarity). Required for the assembly of both SST3A- and SS3B-containing OST complexes (PubMed:22467853)","subcellular_location":"Endoplasmic reticulum membrane","url":"https://www.uniprot.org/uniprotkb/P39656/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/DDOST","classification":"Common Essential","n_dependent_lines":1160,"n_total_lines":1208,"dependency_fraction":0.9602649006622517},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000244038","cell_line_id":"CID000180","localizations":[{"compartment":"er","grade":3}],"interactors":[{"gene":"DAD1","stoichiometry":10.0},{"gene":"KRTCAP2","stoichiometry":10.0},{"gene":"MLEC","stoichiometry":10.0},{"gene":"EMD","stoichiometry":10.0},{"gene":"STT3A","stoichiometry":10.0},{"gene":"RPN1","stoichiometry":10.0},{"gene":"RPN2","stoichiometry":10.0},{"gene":"PICALM","stoichiometry":4.0},{"gene":"CCDC47","stoichiometry":4.0},{"gene":"RAB2A","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/target/CID000180","total_profiled":1310},"omim":[{"mim_id":"614507","title":"CONGENITAL DISORDER OF GLYCOSYLATION, TYPE Ir; CDG1R","url":"https://www.omim.org/entry/614507"},{"mim_id":"602202","title":"DOLICHYL-DIPHOSPHOOLIGOSACCHARIDE-PROTEIN GLYCOSYLTRANSFERASE; DDOST","url":"https://www.omim.org/entry/602202"},{"mim_id":"600243","title":"DEFENDER AGAINST CELL DEATH 1; DAD1","url":"https://www.omim.org/entry/600243"},{"mim_id":"176914","title":"PROTEIN PHOSPHATASE 1, CATALYTIC SUBUNIT, GAMMA ISOFORM; PPP1CC","url":"https://www.omim.org/entry/176914"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Endoplasmic reticulum","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/DDOST"},"hgnc":{"alias_symbol":["OST","KIAA0115","OST48","WBP1"],"prev_symbol":[]},"alphafold":{"accession":"P39656","domains":[{"cath_id":"3.40.50,3.40.50","chopping":"47-301","consensus_level":"high","plddt":94.2656,"start":47,"end":301},{"cath_id":"2.60.40.1930","chopping":"306-412","consensus_level":"high","plddt":95.6723,"start":306,"end":412}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P39656","model_url":"https://alphafold.ebi.ac.uk/files/AF-P39656-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P39656-F1-predicted_aligned_error_v6.png","plddt_mean":89.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=DDOST","jax_strain_url":"https://www.jax.org/strain/search?query=DDOST"},"sequence":{"accession":"P39656","fasta_url":"https://rest.uniprot.org/uniprotkb/P39656.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P39656/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P39656"}},"corpus_meta":[{"pmid":"8855306","id":"PMC_8855306","title":"Molecular identity and cellular distribution of advanced glycation endproduct receptors: relationship of p60 to OST-48 and p90 to 80K-H membrane proteins.","date":"1996","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/8855306","citation_count":299,"is_preprint":false},{"pmid":"22467853","id":"PMC_22467853","title":"The oligosaccharyltransferase subunits OST48, DAD1 and KCP2 function as ubiquitous and selective modulators of mammalian N-glycosylation.","date":"2012","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/22467853","citation_count":78,"is_preprint":false},{"pmid":"22305527","id":"PMC_22305527","title":"DDOST mutations identified by whole-exome sequencing are implicated in congenital disorders of glycosylation.","date":"2012","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/22305527","citation_count":58,"is_preprint":false},{"pmid":"9367678","id":"PMC_9367678","title":"Genome organization of human 48-kDa oligosaccharyltransferase (DDOST).","date":"1997","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/9367678","citation_count":27,"is_preprint":false},{"pmid":"28947796","id":"PMC_28947796","title":"Increased liver AGEs induce hepatic injury mediated through an OST48 pathway.","date":"2017","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/28947796","citation_count":25,"is_preprint":false},{"pmid":"35812432","id":"PMC_35812432","title":"DDOST Correlated with Malignancies and Immune Microenvironment in Gliomas.","date":"2022","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/35812432","citation_count":18,"is_preprint":false},{"pmid":"29027826","id":"PMC_29027826","title":"Dietary advanced glycated end-products and medicines influence the expression of SIRT1 and DDOST in peripheral mononuclear cells from long-term type 1 diabetes patients.","date":"2017","source":"Diabetes & vascular disease research","url":"https://pubmed.ncbi.nlm.nih.gov/29027826","citation_count":15,"is_preprint":false},{"pmid":"23140390","id":"PMC_23140390","title":"Tandem affinity purification in transgenic mouse embryonic stem cells identifies DDOST as a novel PPP1CC2 interacting protein.","date":"2012","source":"Biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/23140390","citation_count":13,"is_preprint":false},{"pmid":"36449507","id":"PMC_36449507","title":"MITA oligomerization upon viral infection is dependent on its N-glycosylation mediated by DDOST.","date":"2022","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/36449507","citation_count":11,"is_preprint":false},{"pmid":"36214423","id":"PMC_36214423","title":"DDOST-CDG: Clinical and molecular characterization of a third patient with a milder and a predominantly movement disorder phenotype.","date":"2022","source":"Journal of inherited metabolic disease","url":"https://pubmed.ncbi.nlm.nih.gov/36214423","citation_count":7,"is_preprint":false},{"pmid":"37155292","id":"PMC_37155292","title":"LncRNA TSIX aggravates spinal cord injury by regulating the PI3K/AKT pathway via the miR-532-3p/DDOST axis.","date":"2023","source":"Journal of biochemical and molecular toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/37155292","citation_count":7,"is_preprint":false},{"pmid":"34277994","id":"PMC_34277994","title":"The AGE receptor, OST48 drives podocyte foot process effacement and basement membrane expansion (alters structural composition).","date":"2021","source":"Endocrinology, diabetes & metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/34277994","citation_count":7,"is_preprint":false},{"pmid":"11530934","id":"PMC_11530934","title":"Analysis of structural signals conferring localisation of pig OST48 to the endoplasmic reticulum.","date":"2001","source":"Biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/11530934","citation_count":6,"is_preprint":false},{"pmid":"7607543","id":"PMC_7607543","title":"PCR-mediated cloning and sequencing of the DmOST50 gene, a WBP1/AvOST50/OST48 homologue, from Drosophila melanogaster.","date":"1995","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/7607543","citation_count":6,"is_preprint":false},{"pmid":"37848450","id":"PMC_37848450","title":"Functional classification of DDOST variants of uncertain clinical significance in congenital disorders of glycosylation.","date":"2023","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/37848450","citation_count":5,"is_preprint":false},{"pmid":"34462534","id":"PMC_34462534","title":"The second DDOST-CDG patient with lactose intolerance, developmental delay, and situs inversus totalis.","date":"2021","source":"Journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/34462534","citation_count":5,"is_preprint":false},{"pmid":"39223141","id":"PMC_39223141","title":"Induction of oxidative- and endoplasmic-reticulum-stress dependent apoptosis in pancreatic cancer cell lines by DDOST knockdown.","date":"2024","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/39223141","citation_count":4,"is_preprint":false},{"pmid":"31541173","id":"PMC_31541173","title":"Globally elevating the AGE clearance receptor, OST48, does not protect against the development of diabetic kidney disease, despite improving insulin secretion.","date":"2019","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/31541173","citation_count":4,"is_preprint":false},{"pmid":"41109654","id":"PMC_41109654","title":"Macrophage-derived exosome miR-146a-5p modulates PNKP/DDOST/JAGN1 complex to regulate NETs formation in atherosclerosis.","date":"2025","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/41109654","citation_count":2,"is_preprint":false},{"pmid":"38460058","id":"PMC_38460058","title":"DDOST is associated with tumor immunosuppressive microenvironment in cervical cancer.","date":"2024","source":"Discover oncology","url":"https://pubmed.ncbi.nlm.nih.gov/38460058","citation_count":1,"is_preprint":false},{"pmid":"41413687","id":"PMC_41413687","title":"Targeting DDOST improves the efficacy of lenvatinib and immunotherapy in hepatocellular carcinoma.","date":"2025","source":"Experimental & molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/41413687","citation_count":0,"is_preprint":false},{"pmid":"40601285","id":"PMC_40601285","title":"Progress in research on DDOST dysregulation in related diseases.","date":"2025","source":"Glycoconjugate journal","url":"https://pubmed.ncbi.nlm.nih.gov/40601285","citation_count":0,"is_preprint":false},{"pmid":"41392699","id":"PMC_41392699","title":"DDOST-Congenital Disorder of Glycosylation: Defining the Clinical Spectrum and First Report of a Structural Variant.","date":"2025","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/41392699","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12890,"output_tokens":2697,"usd":0.039563,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9880,"output_tokens":3529,"usd":0.068812,"stage2_stop_reason":"end_turn"},"total_usd":0.108375,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1996,\n      \"finding\": \"The AGE-binding protein p60 isolated from rat liver membranes is identical to OST-48 (DDOST), a 48-kDa subunit of the oligosaccharyltransferase complex. Immunoprecipitated OST-48 from rat rough endoplasmic reticulum fractions exhibited AGE-specific binding, and anti-OST-48 IgG inhibited AGE-BSA binding to cell membranes in a dose-dependent manner. OST-48 is also surface-expressed on mononuclear, endothelial, renal, and neuronal cells.\",\n      \"method\": \"N-terminal protein sequencing, immunoprecipitation, AGE-ligand binding assay, Western blot, immunostaining, flow cytometry\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal immunoprecipitation plus AGE-binding assay and blocking antibody experiment in a single lab with multiple orthogonal methods\",\n      \"pmids\": [\"8855306\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"ER localisation of OST48 (DDOST) is conferred by two cytosolic C-terminal lysine residues (Lys-3 and Lys-5 from the C-terminus). These lysines act as a retrieval/relocation signal rather than a primary retention signal. Co-expression with ribophorin I quantitatively retained an OST48 surface-escape mutant in the ER, indicating that ER localisation is primarily achieved through complex formation with ribophorin I involving their luminal domains.\",\n      \"method\": \"Site-directed mutagenesis of OST48 cytosolic lysines, immunofluorescence microscopy of COS-1 cells expressing OST48 mutants and hybrid proteins, co-expression epistasis\",\n      \"journal\": \"Biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — mutagenesis combined with immunofluorescence localization and co-expression rescue, single lab\",\n      \"pmids\": [\"11530934\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"OST48 (DDOST) and DAD1 are required for the stable assembly of both STT3A- and STT3B-containing oligosaccharyltransferase complexes. Knockdown of OST48 destabilises these complexes and causes pronounced global hypoglycosylation of N-glycosylation substrates.\",\n      \"method\": \"siRNA knockdown of OST48/DAD1/KCP2 in mammalian cells, biochemical analysis of OST complex stability, glycosylation efficiency assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — subunit-specific knockdown with OST complex stability readout and N-glycosylation phenotype, multiple subunits tested in parallel as controls, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"22467853\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Biallelic loss-of-function mutations in DDOST (a 22 bp deletion and a missense mutation) cause decreased N-glycosylation in patient fibroblasts, and complementation with wild-type DDOST cDNA restores glycosylation, establishing DDOST as a causative gene in congenital disorders of glycosylation (CDG).\",\n      \"method\": \"Whole-exome sequencing, biochemical glycosylation assays (three biomarkers), cDNA complementation in patient fibroblasts\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic identification combined with functional complementation in patient cells using multiple biochemical readouts\",\n      \"pmids\": [\"22305527\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"DDOST (OST48) physically interacts with the testis-specific protein phosphatase isoform PPP1CC2. The interaction was identified by tandem affinity purification and confirmed by reciprocal in vitro sedimentation assay. DDOST localises to the nuclear envelope in dissociated spermatogenic cells.\",\n      \"method\": \"Tandem affinity purification from knock-in mouse embryonic stem cells, reciprocal in vitro sedimentation assay, immunolocalization\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — TAP identification confirmed by reciprocal in vitro sedimentation, single lab\",\n      \"pmids\": [\"23140390\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Upon DNA viral infection, MITA/STING undergoes DDOST-mediated N-glycosylation in the ER. Selective mutation of the DDOST-dependent N-glycosylation sites on MITA abolished MITA oligomerization and downstream immune signalling. Increasing Ddost expression in mouse brain enhanced local immune response to HSV-1 and prolonged survival in HSV encephalitis.\",\n      \"method\": \"N-glycosylation site mutagenesis of MITA, DDOST knockdown/overexpression in cells and in vivo mouse brain, MITA oligomerization assay, viral infection model\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — mutagenesis of substrate glycosylation sites combined with functional oligomerization assay and in vivo rescue experiment\",\n      \"pmids\": [\"36449507\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"OST48 (DDOST) transiently interacts with lysyl oxidase (LOX), a secreted extracellular matrix cross-linking enzyme, and is required for efficient N-glycosylation of LOX. An assay based on LOX N-glycosylation was used to functionally classify DDOST variants, identifying p.S243F and p.E286del as pathogenic loss-of-function variants.\",\n      \"method\": \"Proximity labelling to identify OST48 interaction with LOX, LOX N-glycosylation functional assay, variant complementation testing in cells\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proximity labelling interaction plus functional glycosylation assay with variant testing, single lab\",\n      \"pmids\": [\"37848450\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"DDOST knockdown in pancreatic ductal adenocarcinoma (PDAC) cells decreased proliferation and cell viability and increased ER stress, ROS formation, and apoptosis. Quantitative mass spectrometry identified 30 differentially expressed proteins/phosphopeptides after DDOST knockdown.\",\n      \"method\": \"siRNA knockdown of DDOST in two PDAC cell lines, quantitative mass spectrometry proteomics, cell viability and apoptosis assays, ROS measurement\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockdown with defined cellular phenotype readout (apoptosis, ER stress, ROS) validated in two cell lines, single lab\",\n      \"pmids\": [\"39223141\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"DDOST depletion in HCC cells impaired EGFR N-glycosylation, suppressing downstream AKT, ERK5 and ERK1/2 signalling and sensitising cells to lenvatinib. DDOST depletion also reduced PD-L1 glycosylation. The OST inhibitor NGI-1 phenocopied these effects in vitro and in vivo.\",\n      \"method\": \"DDOST knockdown in HCC cell lines and xenograft models, EGFR and PD-L1 N-glycosylation assays, Western blot for downstream signalling, pharmacological inhibition with NGI-1\",\n      \"journal\": \"Experimental & molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockdown with glycosylation and downstream signalling readouts confirmed in vivo, single lab\",\n      \"pmids\": [\"41413687\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Macrophage-derived exosome miR-146a-5p suppresses PNKP expression, reducing PNKP–DDOST interaction and thereby enhancing DDOST phosphorylation, which activates JAGN1-dependent neutrophil extracellular trap (NET) formation in atherosclerosis.\",\n      \"method\": \"Co-immunoprecipitation, RIP assay, dual-luciferase reporter assay, Western blot, qRT-PCR, ApoE-/- mouse AS model\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-immunoprecipitation of PNKP-DDOST complex with miR-146a-5p modulation and in vivo mouse model, single lab\",\n      \"pmids\": [\"41109654\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"DDOST encodes OST48, a non-catalytic but structurally essential subunit of the ER oligosaccharyltransferase complex that is required for stable assembly of both STT3A- and STT3B-containing OST complexes and hence for global protein N-glycosylation; it catalyses N-glycosylation of specific substrates including MITA/STING (enabling MITA oligomerization and innate immune signalling), EGFR, and PD-L1; it is retained in the ER via its luminal interaction with ribophorin I and a cytosolic di-lysine retrieval motif; it binds AGEs at the cell surface; it physically interacts with PPP1CC2 at the nuclear envelope in spermatogenic cells; and its phosphorylation state is regulated by the PNKP kinase, linking it to NET formation in neutrophils.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"DDOST encodes OST48, a structurally essential subunit of the endoplasmic reticulum oligosaccharyltransferase (OST) complex that is required for the stable assembly of both STT3A- and STT3B-containing OST complexes; its loss destabilizes these complexes and causes pronounced global hypoglycosylation of N-glycosylation substrates [#2]. Biallelic loss-of-function mutations in DDOST reduce N-glycosylation in patient fibroblasts and are rescued by wild-type cDNA, establishing DDOST as a causative gene in congenital disorders of glycosylation [#3]. ER localization of OST48 is achieved primarily through luminal complex formation with ribophorin I, supported by a cytosolic C-terminal di-lysine retrieval signal [#1]. Beyond its global role, OST48 mediates N-glycosylation of specific substrates with defined functional consequences: it glycosylates MITA/STING to enable MITA oligomerization and antiviral innate immune signaling [#5], the secreted matrix cross-linker lysyl oxidase [#6], and in cancer cells EGFR and PD-L1, where DDOST loss suppresses downstream AKT/ERK signaling and sensitizes hepatocellular carcinoma to lenvatinib [#8]. DDOST was originally identified as the AGE-binding protein p60 with cell-surface AGE-binding activity [#0], and additional context-specific roles include a physical interaction with the testis-specific phosphatase PPP1CC2 at the nuclear envelope in spermatogenic cells [#4] and regulation of its phosphorylation by PNKP linking it to neutrophil extracellular trap formation [#9].\",\n  \"teleology\": [\n    {\n      \"year\": 1996,\n      \"claim\": \"Established a surprising dual identity for DDOST, showing the AGE-binding membrane protein p60 is the same protein as the 48-kDa OST subunit and is also displayed at the cell surface.\",\n      \"evidence\": \"N-terminal sequencing, immunoprecipitation, AGE-binding and blocking-antibody assays in rat liver membranes\",\n      \"pmids\": [\n        \"8855306\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Functional significance of surface AGE binding not established\",\n        \"Single-lab characterization without independent confirmation\",\n        \"Relationship between AGE-binding and OST function unaddressed\"\n      ]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Resolved how OST48 is retained in the ER, showing localization depends primarily on luminal complex formation with ribophorin I rather than its cytosolic di-lysine motif alone.\",\n      \"evidence\": \"Site-directed mutagenesis and immunofluorescence co-expression epistasis in COS-1 cells\",\n      \"pmids\": [\n        \"11530934\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Structural basis of luminal ribophorin I interaction not defined\",\n        \"Mechanism of di-lysine retrieval not detailed\"\n      ]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Defined OST48 as structurally essential for OST complex assembly, demonstrating its knockdown destabilizes both STT3A and STT3B complexes and globally impairs N-glycosylation.\",\n      \"evidence\": \"Subunit-specific siRNA knockdown with OST complex stability and glycosylation readouts in mammalian cells\",\n      \"pmids\": [\n        \"22467853\"\n      ],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Catalytic versus purely structural contribution not separated\",\n        \"No structural model of OST48 within the assembled complex\"\n      ]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Established DDOST as a human disease gene by linking biallelic loss-of-function mutations to a congenital disorder of glycosylation with functional rescue.\",\n      \"evidence\": \"Whole-exome sequencing with biochemical glycosylation assays and cDNA complementation in patient fibroblasts\",\n      \"pmids\": [\n        \"22305527\"\n      ],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Genotype-phenotype spectrum limited to few patients\",\n        \"Tissue-specific disease mechanism not dissected\"\n      ]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Identified a context-specific interaction of DDOST with the testis-specific phosphatase PPP1CC2 and nuclear-envelope localization in spermatogenic cells, hinting at roles beyond ER glycosylation.\",\n      \"evidence\": \"Tandem affinity purification with reciprocal in vitro sedimentation and immunolocalization in spermatogenic cells\",\n      \"pmids\": [\n        \"23140390\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Functional consequence of the PPP1CC2 interaction unknown\",\n        \"Mechanism of nuclear-envelope localization unexplained\",\n        \"Single-lab finding\"\n      ]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Showed DDOST acts on a specific signaling substrate, glycosylating MITA/STING to enable oligomerization and antiviral immune signaling, with in vivo protective effect against HSV encephalitis.\",\n      \"evidence\": \"Substrate glycosylation-site mutagenesis, DDOST knockdown/overexpression, oligomerization assay and in vivo mouse brain viral infection model\",\n      \"pmids\": [\n        \"36449507\"\n      ],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Substrate selectivity determinants not defined\",\n        \"Whether effect requires full OST complex or DDOST specifically not separated\"\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended DDOST substrate range to the secreted matrix enzyme lysyl oxidase and leveraged this to functionally classify pathogenic DDOST variants.\",\n      \"evidence\": \"Proximity labelling, LOX N-glycosylation functional assay and variant complementation in cells\",\n      \"pmids\": [\n        \"37848450\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Transient nature of the LOX interaction not structurally characterized\",\n        \"Single-lab assay\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Linked DDOST to cancer cell survival, showing its knockdown in PDAC cells triggers ER stress, ROS, and apoptosis.\",\n      \"evidence\": \"siRNA knockdown in two PDAC cell lines with quantitative proteomics, viability, apoptosis and ROS assays\",\n      \"pmids\": [\n        \"39223141\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Specific glycoprotein substrates driving the phenotype not identified\",\n        \"In vivo relevance untested\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Connected DDOST-dependent glycosylation to oncogenic receptor signaling and therapy response, showing depletion impairs EGFR and PD-L1 glycosylation and sensitizes HCC to lenvatinib.\",\n      \"evidence\": \"DDOST knockdown and NGI-1 inhibition in HCC cell lines and xenografts with glycosylation and signaling readouts\",\n      \"pmids\": [\n        \"41413687\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct versus complex-mediated glycosylation of EGFR/PD-L1 not distinguished\",\n        \"Clinical translatability untested\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Implicated DDOST phosphorylation, controlled by PNKP, in neutrophil extracellular trap formation during atherosclerosis.\",\n      \"evidence\": \"Co-IP, RIP, dual-luciferase reporter assays and ApoE-/- mouse atherosclerosis model\",\n      \"pmids\": [\n        \"41109654\"\n      ],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Phosphorylation site(s) on DDOST not mapped\",\n        \"Mechanistic link between DDOST phosphorylation and NET machinery indirect\",\n        \"Single-lab finding\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How DDOST achieves substrate selectivity and whether its non-glycosylation roles (AGE binding, PPP1CC2 interaction, phosphorylation-dependent NET regulation) are mechanistically separable from OST complex function remains open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No structural model of DDOST substrate engagement\",\n        \"Moonlighting functions not reconciled with ER glycosylation role\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\n        \"term_id\": \"GO:0140096\",\n        \"supporting_discovery_ids\": [\n          2,\n          5,\n          6,\n          8\n        ]\n      },\n      {\n        \"term_id\": \"GO:0016740\",\n        \"supporting_discovery_ids\": [\n          2,\n          3\n        ]\n      }\n    ],\n    \"localization\": [\n      {\n        \"term_id\": \"GO:0005783\",\n        \"supporting_discovery_ids\": [\n          1,\n          2,\n          5\n        ]\n      },\n      {\n        \"term_id\": \"GO:0005886\",\n        \"supporting_discovery_ids\": [\n          0\n        ]\n      },\n      {\n        \"term_id\": \"GO:0005635\",\n        \"supporting_discovery_ids\": [\n          4\n        ]\n      }\n    ],\n    \"pathway\": [\n      {\n        \"term_id\": \"R-HSA-392499\",\n        \"supporting_discovery_ids\": [\n          2,\n          3\n        ]\n      },\n      {\n        \"term_id\": \"R-HSA-168256\",\n        \"supporting_discovery_ids\": [\n          5\n        ]\n      }\n    ],\n    \"complexes\": [\n      \"oligosaccharyltransferase (OST) complex\"\n    ],\n    \"partners\": [\n      \"RPN1\",\n      \"PPP1CC2\",\n      \"LOX\",\n      \"PNKP\",\n      \"STING1\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}