{"gene":"ALG9","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":1996,"finding":"Deletion of yeast ALG9 leads to accumulation of lipid-linked Man6GlcNAc2 in vivo and hypoglycosylation of secreted proteins, identifying ALG9 as encoding a putative mannosyltransferase required for stepwise assembly of the lipid-linked oligosaccharide in the ER lumen.","method":"Yeast gene deletion, in vivo lipid-linked oligosaccharide analysis, secreted protein glycosylation assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct biochemical assay (LLO accumulation) combined with genetic deletion in yeast, foundational mechanistic study replicated by subsequent work","pmids":["8692962"],"is_preprint":false},{"year":2004,"finding":"Human ALG9 encodes an alpha-1,2-mannosyltransferase whose deficiency causes accumulation of lipid-linked GlcNAc2Man6 and GlcNAc2Man8, and transfer of incomplete oligosaccharide precursors to protein; a homozygous E523K missense mutation abolishes function as shown by yeast complementation assay.","method":"Lipid-linked oligosaccharide analysis in patient fibroblasts, yeast complementation assay with ALG9-deficient yeast, mutation analysis","journal":"American journal of human genetics","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — biochemical LLO accumulation assay, yeast complementation, and mutation identification; replicated across multiple CDG patients in subsequent papers","pmids":["15148656"],"is_preprint":false},{"year":2005,"finding":"ALG9 mannosyltransferase is required for the addition of two distinct alpha-1,2-linked mannose residues to the lipid-linked oligosaccharide (LLO) using dolichylphosphomannose as donor, making it unique among ER lumen-oriented glycosyltransferases which otherwise each add only a single hexose unit.","method":"Biochemical LLO analysis, genetic complementation, in vivo and in vitro glycosyltransferase assays in yeast","journal":"Glycobiology","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct biochemical characterization of substrate specificity, complementary genetic and biochemical methods, published in focused mechanistic study","pmids":["15987956"],"is_preprint":false},{"year":2005,"finding":"A Y286C mutation in ALG9 causes CDG-IL; patient fibroblasts accumulate DolPP-GlcNAc2Man6 and DolPP-GlcNAc2Man8, and the causal effect of the mutation was confirmed by complementation of alg9-deficient yeast cells.","method":"LLO analysis in patient fibroblasts, yeast complementation assay, mutation analysis","journal":"American journal of medical genetics. Part A","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — biochemical LLO accumulation assay plus yeast complementation, consistent with prior CDG-IL characterization","pmids":["15945070"],"is_preprint":false},{"year":2009,"finding":"In ALG9-deficient (CDG-IL) patient cells, lipid-linked Man6GlcNAc2 and Man8GlcNAc2 are transferred onto proteins with the same efficiency; glycoproteins bearing these truncated structures enter the glucosylation/deglucosylation quality control cycle, and the Man8GlcNAc2 isomer C on patient glycoproteins promotes enhanced degradation of misfolded glycoproteins.","method":"Biochemical protein transfer efficiency assay, quality control cycle analysis, glycoprotein degradation assays in patient fibroblasts","journal":"Glycobiology","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — multiple orthogonal biochemical assays in a single study using patient-derived cells with a defined ALG9 mutation","pmids":["19451548"],"is_preprint":false},{"year":2019,"finding":"Inactivation of Alg9 in cell-based assays results in impaired maturation and defective glycosylation of polycystin-1 (PC1), placing ALG9 in the pathway required for proper N-glycosylation-dependent biogenesis of PC1 in the ER.","method":"In vitro cell-based assays of PC1 protein maturation and glycosylation following Alg9 inactivation","journal":"Journal of the American Society of Nephrology : JASN","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cell-based functional assay with defined readout (PC1 glycosylation/maturation), single lab but specific mechanistic conclusion","pmids":["31395617"],"is_preprint":false},{"year":2022,"finding":"The homozygous missense variant p.L487P in ALG9 causes enhanced protein degradation of the ALG9 enzyme in patient fibroblasts despite elevated mRNA levels, with LLO analysis confirming characteristic accumulation of Man6GlcNAc2-PP-dolichol and Man8GlcNAc2-PP-dolichol.","method":"Quantitative RT-PCR, Western blot for protein quantification, LLO analysis in patient fibroblasts","journal":"Molecular genetics and metabolism","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — two orthogonal methods (protein level and LLO biochemistry) but single lab, single variant","pmids":["35839600"],"is_preprint":false},{"year":2023,"finding":"Immunohistochemistry showed absence of ALG9 protein in liver cyst tissue from a patient with a heterozygous ALG9 missense variant, consistent with somatic loss of heterozygosity as the second hit mechanism for cyst formation.","method":"Immunohistochemistry on liver cyst tissue, whole exome sequencing, in silico 3D protein modeling","journal":"Genes","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single patient, IHC only for protein loss, no functional reconstitution; mechanistic inference is indirect","pmids":["37761895"],"is_preprint":false}],"current_model":"ALG9 encodes an ER-localized alpha-1,2-mannosyltransferase that uniquely catalyzes the addition of two distinct alpha-1,2-linked mannose residues to the dolichol-PP-linked oligosaccharide precursor (converting Man6GlcNAc2 to Man8GlcNAc2 and then to Man9GlcNAc2) using dolichylphosphomannose as donor, and its loss results in transfer of truncated oligosaccharides onto nascent glycoproteins—including polycystin-1—disrupting ER quality control and glycoprotein maturation, thereby causing congenital disorder of glycosylation type IL (CDG-IL) and, in heterozygous carriers, autosomal dominant polycystic kidney and liver disease."},"narrative":{"mechanistic_narrative":"ALG9 encodes an ER lumen-oriented alpha-1,2-mannosyltransferase that builds the dolichol-PP-linked oligosaccharide precursor used for protein N-glycosylation [PMID:8692962, PMID:15987956]. It is unique among ER-luminal glycosyltransferases in catalyzing the addition of two distinct alpha-1,2-linked mannose residues to the lipid-linked oligosaccharide using dolichylphosphomannose as donor, acting at two separate steps of the assembly pathway [PMID:15987956]. Loss of ALG9 function arrests precursor maturation, causing accumulation of lipid-linked Man6GlcNAc2 and Man8GlcNAc2 intermediates and transfer of these truncated oligosaccharides onto nascent glycoproteins [PMID:8692962, PMID:15148656, PMID:15945070]; the truncated glycans still enter the glucosylation/deglucosylation ER quality control cycle, and the resulting Man8GlcNAc2 isomer C promotes enhanced degradation of misfolded glycoproteins [PMID:19451548]. Disrupted N-glycosylation impairs the maturation and biogenesis of the substrate glycoprotein polycystin-1 [PMID:31395617]. Loss-of-function ALG9 variants cause congenital disorder of glycosylation type IL, established through patient LLO accumulation and yeast complementation of disease-associated missense mutations [PMID:15148656, PMID:15945070, PMID:35839600], and biallelic loss in cyst tissue links ALG9 to polycystic kidney and liver disease via a second-hit mechanism [PMID:31395617, PMID:37761895].","teleology":[{"year":1996,"claim":"Established ALG9 as a component of stepwise ER lipid-linked oligosaccharide assembly by showing that its loss blocks the pathway at a defined intermediate.","evidence":"Yeast gene deletion with in vivo LLO analysis and secreted-protein glycosylation assay","pmids":["8692962"],"confidence":"High","gaps":["Did not directly demonstrate enzymatic activity or donor/acceptor specificity","Human ortholog function not yet addressed"]},{"year":2004,"claim":"Connected human ALG9 deficiency to disease by showing that a homozygous missense mutation abolishes mannosyltransferase function and causes transfer of incomplete precursors to protein.","evidence":"LLO analysis in patient fibroblasts plus yeast complementation of the E523K variant","pmids":["15148656"],"confidence":"High","gaps":["Did not resolve which specific mannose-addition steps are catalyzed","Substrate glycoproteins affected not identified"]},{"year":2005,"claim":"Defined ALG9's distinctive catalytic role as adding two distinct alpha-1,2-mannose residues at separate steps, unlike single-addition ER glycosyltransferases.","evidence":"Biochemical LLO analysis with in vivo and in vitro glycosyltransferase assays and genetic complementation in yeast","pmids":["15987956","15945070"],"confidence":"High","gaps":["Structural basis for dual-step specificity not determined","No high-resolution enzyme structure"]},{"year":2009,"claim":"Showed the downstream consequence of truncated glycan transfer: equal-efficiency transfer of Man6 and Man8 intermediates that nonetheless engage ER quality control and bias misfolded glycoprotein degradation.","evidence":"Protein transfer efficiency, quality control cycle, and glycoprotein degradation assays in patient fibroblasts","pmids":["19451548"],"confidence":"High","gaps":["Specific endogenous glycoproteins driving phenotype not defined","Quantitative impact on secretory proteome unresolved"]},{"year":2019,"claim":"Identified polycystin-1 as a glycoprotein whose maturation depends on ALG9, linking the glycosylation defect to a specific clinically relevant substrate.","evidence":"Cell-based PC1 maturation and glycosylation assays after Alg9 inactivation","pmids":["31395617"],"confidence":"Medium","gaps":["Single-lab cell-based system","In vivo relevance to cystogenesis not directly shown"]},{"year":2022,"claim":"Clarified a disease mechanism at the protein level by showing a missense variant destabilizes ALG9 enzyme despite elevated mRNA, producing the characteristic LLO accumulation.","evidence":"RT-PCR, Western blot, and LLO analysis in patient fibroblasts for the L487P variant","pmids":["35839600"],"confidence":"Medium","gaps":["Single variant and single lab","Degradation pathway for mutant enzyme not defined"]},{"year":2023,"claim":"Provided in-tissue evidence for a two-hit mechanism by showing absence of ALG9 protein in cyst tissue from a heterozygous carrier.","evidence":"Immunohistochemistry on liver cyst tissue with exome sequencing and in silico modeling","pmids":["37761895"],"confidence":"Low","gaps":["Single patient with IHC only and no functional reconstitution","Somatic loss of heterozygosity inferred indirectly"]},{"year":null,"claim":"How ALG9 enzyme architecture enables sequential addition at two distinct LLO positions, and how truncated-glycan transfer mechanistically drives polycystic phenotypes in vivo, remain unresolved.","evidence":"No structural or in vivo mechanistic study in the available corpus","pmids":[],"confidence":"Low","gaps":["No high-resolution structure of the enzyme","No in vivo animal model linking glycosylation defect to cystogenesis","Full substrate glycoprotein repertoire uncharacterized"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[0,1,2]}],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[0,2]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,4]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[1,3]}],"complexes":[],"partners":[],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9H6U8","full_name":"Alpha-1,2-mannosyltransferase ALG9","aliases":["Asparagine-linked glycosylation protein 9 homolog","Disrupted in bipolar disorder protein 1","Dol-P-Man:Man(6)GlcNAc(2)-PP-Dol alpha-1,2-mannosyltransferase","Dol-P-Man:Man(8)GlcNAc(2)-PP-Dol alpha-1,2-mannosyltransferase"],"length_aa":611,"mass_kda":69.9,"function":"Mannosyltransferase that operates in the biosynthetic pathway of dolichol-linked oligosaccharides, the glycan precursors employed in protein asparagine (N)-glycosylation. The assembly of dolichol-linked oligosaccharides begins on the cytosolic side of the endoplasmic reticulum membrane and finishes in its lumen. The sequential addition of sugars to dolichol pyrophosphate produces dolichol-linked oligosaccharides containing fourteen sugars, including two GlcNAcs, nine mannoses and three glucoses. Once assembled, the oligosaccharide is transferred from the lipid to nascent proteins by oligosaccharyltransferases. In the lumen of the endoplasmic reticulum, catalyzes the addition of the seventh and ninth alpha-1,2-linked mannose residues to Man(6)GlcNAc(2)-PP-dolichol and Man(8)GlcNAc(2)-PP-dolichol respectively","subcellular_location":"Endoplasmic reticulum membrane","url":"https://www.uniprot.org/uniprotkb/Q9H6U8/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ALG9","classification":"Not Classified","n_dependent_lines":65,"n_total_lines":1208,"dependency_fraction":0.05380794701986755},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"CANX","stoichiometry":0.2},{"gene":"CHPT1","stoichiometry":0.2},{"gene":"HSPB1","stoichiometry":0.2},{"gene":"CCDC47","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/ALG9","total_profiled":1310},"omim":[{"mim_id":"608776","title":"CONGENITAL DISORDER OF GLYCOSYLATION, TYPE Il; CDG1L","url":"https://www.omim.org/entry/608776"},{"mim_id":"606941","title":"ALG9 ALPHA-1,2-MANNOSYLTRANSFERASE; ALG9","url":"https://www.omim.org/entry/606941"},{"mim_id":"263210","title":"GILLESSEN-KAESBACH-NISHIMURA SYNDROME; GIKANIS","url":"https://www.omim.org/entry/263210"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Endoplasmic reticulum","reliability":"Approved"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ALG9"},"hgnc":{"alias_symbol":[],"prev_symbol":["DIBD1"]},"alphafold":{"accession":"Q9H6U8","domains":[{"cath_id":"-","chopping":"439-600","consensus_level":"medium","plddt":93.4284,"start":439,"end":600}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H6U8","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H6U8-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H6U8-F1-predicted_aligned_error_v6.png","plddt_mean":87.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ALG9","jax_strain_url":"https://www.jax.org/strain/search?query=ALG9"},"sequence":{"accession":"Q9H6U8","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9H6U8.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9H6U8/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H6U8"}},"corpus_meta":[{"pmid":"31395617","id":"PMC_31395617","title":"ALG9 Mutation Carriers Develop Kidney and Liver Cysts.","date":"2019","source":"Journal of the American Society of Nephrology : JASN","url":"https://pubmed.ncbi.nlm.nih.gov/31395617","citation_count":131,"is_preprint":false},{"pmid":"8692962","id":"PMC_8692962","title":"Stepwise assembly of the lipid-linked oligosaccharide in the endoplasmic reticulum of Saccharomyces cerevisiae: identification of the ALG9 gene encoding a putative mannosyl transferase.","date":"1996","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/8692962","citation_count":78,"is_preprint":false},{"pmid":"15148656","id":"PMC_15148656","title":"Identification and functional analysis of a defect in the human ALG9 gene: definition of congenital disorder of glycosylation type IL.","date":"2004","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/15148656","citation_count":65,"is_preprint":false},{"pmid":"15987956","id":"PMC_15987956","title":"ALG9 mannosyltransferase is involved in two different steps of lipid-linked oligosaccharide biosynthesis.","date":"2005","source":"Glycobiology","url":"https://pubmed.ncbi.nlm.nih.gov/15987956","citation_count":50,"is_preprint":false},{"pmid":"32359033","id":"PMC_32359033","title":"LncRNA MEG3 contributes to drug resistance in acute myeloid leukemia by positively regulating ALG9 through sponging miR-155.","date":"2020","source":"International journal of laboratory hematology","url":"https://pubmed.ncbi.nlm.nih.gov/32359033","citation_count":45,"is_preprint":false},{"pmid":"25966638","id":"PMC_25966638","title":"A novel phenotype in N-glycosylation disorders: Gillessen-Kaesbach-Nishimura skeletal dysplasia due to pathogenic variants in ALG9.","date":"2015","source":"European journal of human genetics : EJHG","url":"https://pubmed.ncbi.nlm.nih.gov/25966638","citation_count":34,"is_preprint":false},{"pmid":"15945070","id":"PMC_15945070","title":"CDG-IL: an infant with a novel mutation in the ALG9 gene and additional phenotypic features.","date":"2005","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/15945070","citation_count":34,"is_preprint":false},{"pmid":"28932688","id":"PMC_28932688","title":"ALG9-CDG: New clinical case and review of the literature.","date":"2017","source":"Molecular genetics and metabolism reports","url":"https://pubmed.ncbi.nlm.nih.gov/28932688","citation_count":20,"is_preprint":false},{"pmid":"19451548","id":"PMC_19451548","title":"Quality control of glycoproteins bearing truncated glycans in an ALG9-defective (CDG-IL) patient.","date":"2009","source":"Glycobiology","url":"https://pubmed.ncbi.nlm.nih.gov/19451548","citation_count":18,"is_preprint":false},{"pmid":"26453364","id":"PMC_26453364","title":"Further Delineation of the ALG9-CDG Phenotype.","date":"2015","source":"JIMD reports","url":"https://pubmed.ncbi.nlm.nih.gov/26453364","citation_count":16,"is_preprint":false},{"pmid":"39899384","id":"PMC_39899384","title":"Characterization of the Cystic Phenotype Associated with Monoallelic ALG8 and ALG9 Pathogenic Variants.","date":"2025","source":"Journal of the American Society of Nephrology : JASN","url":"https://pubmed.ncbi.nlm.nih.gov/39899384","citation_count":12,"is_preprint":false},{"pmid":"35839600","id":"PMC_35839600","title":"Missense variant c.1460 T > C (p.L487P) enhances protein degradation of ER mannosyltransferase ALG9 in two new ALG9-CDG patients presenting with West syndrome and review of the literature.","date":"2022","source":"Molecular genetics and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/35839600","citation_count":8,"is_preprint":false},{"pmid":"37761895","id":"PMC_37761895","title":"Heterozygosity of ALG9 in Association with Autosomal Dominant Polycystic Liver Disease.","date":"2023","source":"Genes","url":"https://pubmed.ncbi.nlm.nih.gov/37761895","citation_count":3,"is_preprint":false},{"pmid":"16859551","id":"PMC_16859551","title":"Common variations in ALG9 are not associated with bipolar I disorder: a family-based study.","date":"2006","source":"Behavioral and brain functions : BBF","url":"https://pubmed.ncbi.nlm.nih.gov/16859551","citation_count":3,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9144,"output_tokens":2030,"usd":0.028941,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":8945,"output_tokens":2426,"usd":0.052687,"stage2_stop_reason":"end_turn"},"total_usd":0.081628,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1996,\n      \"finding\": \"Deletion of yeast ALG9 leads to accumulation of lipid-linked Man6GlcNAc2 in vivo and hypoglycosylation of secreted proteins, identifying ALG9 as encoding a putative mannosyltransferase required for stepwise assembly of the lipid-linked oligosaccharide in the ER lumen.\",\n      \"method\": \"Yeast gene deletion, in vivo lipid-linked oligosaccharide analysis, secreted protein glycosylation assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct biochemical assay (LLO accumulation) combined with genetic deletion in yeast, foundational mechanistic study replicated by subsequent work\",\n      \"pmids\": [\"8692962\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Human ALG9 encodes an alpha-1,2-mannosyltransferase whose deficiency causes accumulation of lipid-linked GlcNAc2Man6 and GlcNAc2Man8, and transfer of incomplete oligosaccharide precursors to protein; a homozygous E523K missense mutation abolishes function as shown by yeast complementation assay.\",\n      \"method\": \"Lipid-linked oligosaccharide analysis in patient fibroblasts, yeast complementation assay with ALG9-deficient yeast, mutation analysis\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — biochemical LLO accumulation assay, yeast complementation, and mutation identification; replicated across multiple CDG patients in subsequent papers\",\n      \"pmids\": [\"15148656\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"ALG9 mannosyltransferase is required for the addition of two distinct alpha-1,2-linked mannose residues to the lipid-linked oligosaccharide (LLO) using dolichylphosphomannose as donor, making it unique among ER lumen-oriented glycosyltransferases which otherwise each add only a single hexose unit.\",\n      \"method\": \"Biochemical LLO analysis, genetic complementation, in vivo and in vitro glycosyltransferase assays in yeast\",\n      \"journal\": \"Glycobiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct biochemical characterization of substrate specificity, complementary genetic and biochemical methods, published in focused mechanistic study\",\n      \"pmids\": [\"15987956\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"A Y286C mutation in ALG9 causes CDG-IL; patient fibroblasts accumulate DolPP-GlcNAc2Man6 and DolPP-GlcNAc2Man8, and the causal effect of the mutation was confirmed by complementation of alg9-deficient yeast cells.\",\n      \"method\": \"LLO analysis in patient fibroblasts, yeast complementation assay, mutation analysis\",\n      \"journal\": \"American journal of medical genetics. Part A\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — biochemical LLO accumulation assay plus yeast complementation, consistent with prior CDG-IL characterization\",\n      \"pmids\": [\"15945070\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"In ALG9-deficient (CDG-IL) patient cells, lipid-linked Man6GlcNAc2 and Man8GlcNAc2 are transferred onto proteins with the same efficiency; glycoproteins bearing these truncated structures enter the glucosylation/deglucosylation quality control cycle, and the Man8GlcNAc2 isomer C on patient glycoproteins promotes enhanced degradation of misfolded glycoproteins.\",\n      \"method\": \"Biochemical protein transfer efficiency assay, quality control cycle analysis, glycoprotein degradation assays in patient fibroblasts\",\n      \"journal\": \"Glycobiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — multiple orthogonal biochemical assays in a single study using patient-derived cells with a defined ALG9 mutation\",\n      \"pmids\": [\"19451548\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Inactivation of Alg9 in cell-based assays results in impaired maturation and defective glycosylation of polycystin-1 (PC1), placing ALG9 in the pathway required for proper N-glycosylation-dependent biogenesis of PC1 in the ER.\",\n      \"method\": \"In vitro cell-based assays of PC1 protein maturation and glycosylation following Alg9 inactivation\",\n      \"journal\": \"Journal of the American Society of Nephrology : JASN\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cell-based functional assay with defined readout (PC1 glycosylation/maturation), single lab but specific mechanistic conclusion\",\n      \"pmids\": [\"31395617\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The homozygous missense variant p.L487P in ALG9 causes enhanced protein degradation of the ALG9 enzyme in patient fibroblasts despite elevated mRNA levels, with LLO analysis confirming characteristic accumulation of Man6GlcNAc2-PP-dolichol and Man8GlcNAc2-PP-dolichol.\",\n      \"method\": \"Quantitative RT-PCR, Western blot for protein quantification, LLO analysis in patient fibroblasts\",\n      \"journal\": \"Molecular genetics and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — two orthogonal methods (protein level and LLO biochemistry) but single lab, single variant\",\n      \"pmids\": [\"35839600\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Immunohistochemistry showed absence of ALG9 protein in liver cyst tissue from a patient with a heterozygous ALG9 missense variant, consistent with somatic loss of heterozygosity as the second hit mechanism for cyst formation.\",\n      \"method\": \"Immunohistochemistry on liver cyst tissue, whole exome sequencing, in silico 3D protein modeling\",\n      \"journal\": \"Genes\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single patient, IHC only for protein loss, no functional reconstitution; mechanistic inference is indirect\",\n      \"pmids\": [\"37761895\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ALG9 encodes an ER-localized alpha-1,2-mannosyltransferase that uniquely catalyzes the addition of two distinct alpha-1,2-linked mannose residues to the dolichol-PP-linked oligosaccharide precursor (converting Man6GlcNAc2 to Man8GlcNAc2 and then to Man9GlcNAc2) using dolichylphosphomannose as donor, and its loss results in transfer of truncated oligosaccharides onto nascent glycoproteins—including polycystin-1—disrupting ER quality control and glycoprotein maturation, thereby causing congenital disorder of glycosylation type IL (CDG-IL) and, in heterozygous carriers, autosomal dominant polycystic kidney and liver disease.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ALG9 encodes an ER lumen-oriented alpha-1,2-mannosyltransferase that builds the dolichol-PP-linked oligosaccharide precursor used for protein N-glycosylation [#0, #2]. It is unique among ER-luminal glycosyltransferases in catalyzing the addition of two distinct alpha-1,2-linked mannose residues to the lipid-linked oligosaccharide using dolichylphosphomannose as donor, acting at two separate steps of the assembly pathway [#2]. Loss of ALG9 function arrests precursor maturation, causing accumulation of lipid-linked Man6GlcNAc2 and Man8GlcNAc2 intermediates and transfer of these truncated oligosaccharides onto nascent glycoproteins [#0, #1, #3]; the truncated glycans still enter the glucosylation/deglucosylation ER quality control cycle, and the resulting Man8GlcNAc2 isomer C promotes enhanced degradation of misfolded glycoproteins [#4]. Disrupted N-glycosylation impairs the maturation and biogenesis of the substrate glycoprotein polycystin-1 [#5]. Loss-of-function ALG9 variants cause congenital disorder of glycosylation type IL, established through patient LLO accumulation and yeast complementation of disease-associated missense mutations [#1, #3, #6], and biallelic loss in cyst tissue links ALG9 to polycystic kidney and liver disease via a second-hit mechanism [#5, #7].\",\n  \"teleology\": [\n    {\n      \"year\": 1996,\n      \"claim\": \"Established ALG9 as a component of stepwise ER lipid-linked oligosaccharide assembly by showing that its loss blocks the pathway at a defined intermediate.\",\n      \"evidence\": \"Yeast gene deletion with in vivo LLO analysis and secreted-protein glycosylation assay\",\n      \"pmids\": [\"8692962\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not directly demonstrate enzymatic activity or donor/acceptor specificity\", \"Human ortholog function not yet addressed\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Connected human ALG9 deficiency to disease by showing that a homozygous missense mutation abolishes mannosyltransferase function and causes transfer of incomplete precursors to protein.\",\n      \"evidence\": \"LLO analysis in patient fibroblasts plus yeast complementation of the E523K variant\",\n      \"pmids\": [\"15148656\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve which specific mannose-addition steps are catalyzed\", \"Substrate glycoproteins affected not identified\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Defined ALG9's distinctive catalytic role as adding two distinct alpha-1,2-mannose residues at separate steps, unlike single-addition ER glycosyltransferases.\",\n      \"evidence\": \"Biochemical LLO analysis with in vivo and in vitro glycosyltransferase assays and genetic complementation in yeast\",\n      \"pmids\": [\"15987956\", \"15945070\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis for dual-step specificity not determined\", \"No high-resolution enzyme structure\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Showed the downstream consequence of truncated glycan transfer: equal-efficiency transfer of Man6 and Man8 intermediates that nonetheless engage ER quality control and bias misfolded glycoprotein degradation.\",\n      \"evidence\": \"Protein transfer efficiency, quality control cycle, and glycoprotein degradation assays in patient fibroblasts\",\n      \"pmids\": [\"19451548\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific endogenous glycoproteins driving phenotype not defined\", \"Quantitative impact on secretory proteome unresolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified polycystin-1 as a glycoprotein whose maturation depends on ALG9, linking the glycosylation defect to a specific clinically relevant substrate.\",\n      \"evidence\": \"Cell-based PC1 maturation and glycosylation assays after Alg9 inactivation\",\n      \"pmids\": [\"31395617\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab cell-based system\", \"In vivo relevance to cystogenesis not directly shown\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Clarified a disease mechanism at the protein level by showing a missense variant destabilizes ALG9 enzyme despite elevated mRNA, producing the characteristic LLO accumulation.\",\n      \"evidence\": \"RT-PCR, Western blot, and LLO analysis in patient fibroblasts for the L487P variant\",\n      \"pmids\": [\"35839600\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single variant and single lab\", \"Degradation pathway for mutant enzyme not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Provided in-tissue evidence for a two-hit mechanism by showing absence of ALG9 protein in cyst tissue from a heterozygous carrier.\",\n      \"evidence\": \"Immunohistochemistry on liver cyst tissue with exome sequencing and in silico modeling\",\n      \"pmids\": [\"37761895\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single patient with IHC only and no functional reconstitution\", \"Somatic loss of heterozygosity inferred indirectly\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ALG9 enzyme architecture enables sequential addition at two distinct LLO positions, and how truncated-glycan transfer mechanistically drives polycystic phenotypes in vivo, remain unresolved.\",\n      \"evidence\": \"No structural or in vivo mechanistic study in the available corpus\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No high-resolution structure of the enzyme\", \"No in vivo animal model linking glycosylation defect to cystogenesis\", \"Full substrate glycoprotein repertoire uncharacterized\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [0, 1, 2]},\n      {\"term_id\": \"GO:0016757\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [0, 2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 4]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [1, 3]}\n    ],\n    \"complexes\": [],\n    \"partners\": [],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}