{"gene":"PADI3","run_date":"2026-06-10T05:19:53","timeline":{"discoveries":[{"year":2021,"finding":"PADI3 (together with PADI1) citrullinates pyruvate kinase M2 (PKM2) at arginine residue R106, reprogramming cross-talk between PKM2 ligands: it lowers PKM2 sensitivity to the inhibitors tryptophan, alanine, and phenylalanine and promotes activation by serine, thereby bypassing normal physiological regulation to promote excessive glycolysis.","method":"Mass spectrometry identification of citrullination site, cancer cell overexpression/knockdown experiments, metabolic assays measuring glycolysis","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, multiple orthogonal methods (MS, KD, metabolic assays) but no in vitro reconstitution with purified PADI3 alone","pmids":["33741961"],"is_preprint":false},{"year":2019,"finding":"CCCA-associated missense mutations in PADI3 result in reduced PADI3 expression, abnormal intracellular localization of the protein, and decreased enzymatic (peptidylarginine deiminase) activity, establishing that loss of PADI3 enzymatic function underlies hair-shaft formation defects.","method":"Enzymatic activity assay, immunofluorescence for localization, immunoblotting for expression, protein structural modeling of mutants","journal":"The New England journal of medicine","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — direct enzymatic assay plus localization experiment plus expression analysis in patient-derived samples, replicated across discovery and replication cohorts","pmids":["30763140"],"is_preprint":false},{"year":2006,"finding":"Transcription factors NF-Y and Sp1/Sp3 directly bind the PADI3 promoter region (within 129 bp of the transcription start site) in keratinocytes in vitro and in vivo, and mutation of either binding motif or siRNA knockdown of Sp1 or NF-YA markedly reduces PADI3 promoter activity and expression.","method":"Luciferase reporter assays with deletion/mutation constructs, EMSA, chromatin immunoprecipitation (ChIP), siRNA knockdown","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal methods (reporter assay, EMSA, ChIP, siRNA) in single study with rigorous mutagenesis controls","pmids":["16671893"],"is_preprint":false},{"year":2019,"finding":"ZDHHC13 palmitoylates PADI3 protein, and this palmitoylation is critical for PADI3 protein stability in vivo; loss of ZDHHC13 enzymatic activity reduces PADI3 palmitoylation and protein levels.","method":"Quantitative proteomics to identify palmitoylation candidates, biochemical palmitoylation assay, knock-in mice bearing enzymatically dead ZDHHC13 mutation","journal":"The Journal of investigative dermatology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical palmitoylation assay plus in vivo genetic model, single lab","pmids":["31669413"],"is_preprint":false},{"year":2019,"finding":"PADI3 overexpression in colon cancer cells suppresses Sirt2 and AKT phosphorylation, increases p21 expression, and induces G1 cell cycle arrest; Sirt2 overexpression partially reverses these effects, placing PADI3 upstream of the Sirt2/AKT/p21 axis. PADI3 is localized mainly in the cytoplasm, and the C-terminal domain is required for its antitumor activity.","method":"Western blot, flow cytometry for cell cycle, CCK-8 proliferation assay, colony formation assay, RNA-seq, truncation mutation experiments, immunocytochemistry, xenograft tumor formation","journal":"Cancer biology & medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal cellular assays plus rescue experiment placing PADI3 in pathway, single lab","pmids":["31908891"],"is_preprint":false},{"year":2019,"finding":"PADI3 decreases Hsp90 and CKS1 expression in colon cancer cells, and Hsp90 is required as an intermediary for PADI3-mediated downregulation of CKS1.","method":"Western blot, real-time PCR, CCK-8 proliferation assay, colony formation assay, overexpression and rescue experiments, xenograft mouse model","journal":"Cancer cell international","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, mechanism inferred from overexpression/rescue without direct biochemical interaction assays","pmids":["31708688"],"is_preprint":false},{"year":2024,"finding":"PADI3 promotes ubiquitin-dependent degradation of CKS1, thereby suppressing EMT: reduced CKS1 leads to decreased Snail and N-cadherin levels and restored E-cadherin expression, inhibiting colon cancer cell migration.","method":"Western blot, Transwell and wound healing migration assays, flow cytometry, overexpression and rescue experiments","journal":"Journal of cancer research and therapeutics","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, ubiquitin-dependent degradation inferred from rescue experiments without direct ubiquitination or proteasome assays","pmids":["39206995"],"is_preprint":false},{"year":2024,"finding":"PADI3 knockdown in endometrial cancer cells inhibits ERK signaling pathway activity, reduces glycolysis, and induces apoptosis; PADI3 overexpression reverses the anti-cancer effects of atractylenolide II, placing PADI3 upstream of ERK in this context.","method":"siRNA knockdown, overexpression rescue experiments, proliferation assays, apoptosis assays, glycolysis measurements, Western blot for ERK pathway proteins","journal":"Molecules (Basel, Switzerland)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, pathway placement based on rescue/KD without direct biochemical interaction between PADI3 and ERK","pmids":["38474453"],"is_preprint":false}],"current_model":"PADI3 is a calcium-dependent peptidylarginine deiminase that post-translationally converts arginine to citrulline on substrate proteins including PKM2 (at R106) and hair-shaft proteins (e.g., filaggrin, trichohyalin); its expression in keratinocytes is driven by NF-Y and Sp1/Sp3 transcription factors, its protein stability is regulated by ZDHHC13-mediated palmitoylation, and disease-causing mutations reduce both its enzymatic activity and proper intracellular localization, while in cancer contexts it acts as a tumor suppressor by modulating the Sirt2/AKT/p21 and Hsp90/CKS1 pathways to induce cell cycle arrest and block EMT."},"narrative":{"mechanistic_narrative":"PADI3 is a calcium-dependent peptidylarginine deiminase that catalyzes the post-translational conversion of arginine to citrulline, a modification essential for normal hair-shaft formation: CCCA-associated missense mutations reduce PADI3 enzymatic activity, lower its expression, and mislocalize the protein, establishing loss of deiminase function as the basis of hair-shaft defects [PMID:30763140]. In keratinocytes, PADI3 transcription is driven directly by NF-Y and Sp1/Sp3 binding within the proximal promoter [PMID:16671893], and the resulting protein is stabilized by ZDHHC13-mediated palmitoylation [PMID:31669413]. Beyond hair biology, PADI3 citrullinates pyruvate kinase M2 (PKM2) at R106, rewiring allosteric regulation to promote glycolysis [PMID:33741961]. In cancer cells PADI3 acts as a cytoplasmic tumor suppressor, suppressing the Sirt2/AKT axis to raise p21 and drive G1 arrest, with its C-terminal domain required for antitumor activity [PMID:31908891], and lowering CKS1 to restrain epithelial-mesenchymal transition [PMID:39206995].","teleology":[{"year":2006,"claim":"Established how PADI3 expression is controlled in its native keratinocyte context, identifying the transcription factors that set its baseline levels.","evidence":"Luciferase reporter assays, EMSA, ChIP, and siRNA knockdown of Sp1/NF-YA in keratinocytes","pmids":["16671893"],"confidence":"High","gaps":["Does not address signals that modulate NF-Y/Sp1 activity during hair-shaft differentiation","No link to enzymatic activity or substrate engagement"]},{"year":2019,"claim":"Defined PADI3 loss-of-function as the molecular cause of a hair-shaft disorder by showing disease mutations cripple enzyme activity, expression, and localization.","evidence":"Enzymatic activity assays, immunofluorescence, immunoblotting, and structural modeling of patient-derived mutants across discovery and replication cohorts","pmids":["30763140"],"confidence":"High","gaps":["Specific hair-shaft substrates citrullinated in vivo not directly enumerated here","Mechanism by which mislocalization arises from point mutations unresolved"]},{"year":2019,"claim":"Identified a post-translational control point governing PADI3 protein stability, showing palmitoylation rather than transcription alone sets steady-state protein levels.","evidence":"Quantitative proteomics, biochemical palmitoylation assay, and enzymatically dead ZDHHC13 knock-in mice","pmids":["31669413"],"confidence":"Medium","gaps":["Palmitoylated residue(s) on PADI3 not mapped","Whether palmitoylation regulates PADI3 localization or catalytic activity not addressed"]},{"year":2019,"claim":"Placed PADI3 as a cytoplasmic tumor suppressor upstream of a Sirt2/AKT/p21 cell-cycle axis, extending its role beyond epidermal biology.","evidence":"Overexpression/knockdown with Sirt2 rescue, flow cytometry, truncation mutants, and xenografts in colon cancer cells","pmids":["31908891"],"confidence":"Medium","gaps":["Whether the Sirt2/AKT effect depends on PADI3 deiminase activity not established","Direct PADI3 substrate in this axis unidentified"]},{"year":2021,"claim":"Provided the first defined enzymatic substrate-site for PADI3 in a metabolic context, showing citrullination of PKM2-R106 reprograms allosteric regulation toward glycolysis.","evidence":"Mass spectrometry site mapping plus overexpression/knockdown and metabolic assays in cancer cells","pmids":["33741961"],"confidence":"Medium","gaps":["No in vitro reconstitution with purified PADI3 alone (acts with PADI1)","Apparent contradiction between glycolysis-promoting and tumor-suppressive roles not reconciled"]},{"year":2024,"claim":"Connected PADI3 to suppression of EMT through CKS1 downregulation, linking it to migration control in cancer.","evidence":"Migration assays and overexpression/rescue experiments measuring CKS1, Snail, N-cadherin, E-cadherin in colon cancer cells","pmids":["39206995","31708688"],"confidence":"Low","gaps":["Ubiquitin-dependent CKS1 degradation inferred from rescue without direct ubiquitination or proteasome assays","No direct PADI3-CKS1/Hsp90 biochemical interaction demonstrated"]},{"year":2024,"claim":"Extended PADI3's cancer relevance to endometrial cells via ERK signaling and glycolysis.","evidence":"siRNA knockdown and overexpression rescue with proliferation, apoptosis, glycolysis, and ERK-pathway readouts","pmids":["38474453"],"confidence":"Low","gaps":["Pathway placement based on KD/rescue without direct PADI3-ERK biochemical link","Whether deiminase activity drives the ERK effect unknown"]},{"year":null,"claim":"How a single deiminase reconciles glycolysis promotion via PKM2 citrullination with tumor-suppressive cell-cycle arrest, and whether its cancer-context effects require catalytic activity, remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No structure of PADI3 bound to a substrate","Catalytic dependence of the Sirt2/AKT and CKS1 effects untested","Full in vivo hair-shaft substrate repertoire not enumerated"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,1]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,1]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[4]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,1]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[2]}],"complexes":[],"partners":["PKM2","ZDHHC13","SP1","NFYA"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9ULW8","full_name":"Protein-arginine deiminase type-3","aliases":["Peptidylarginine deiminase III","Protein-arginine deiminase type III"],"length_aa":664,"mass_kda":74.7,"function":"Catalyzes the deimination of arginine residues of proteins","subcellular_location":"Cytoplasm","url":"https://www.uniprot.org/uniprotkb/Q9ULW8/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/PADI3","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/PADI3","total_profiled":1310},"omim":[{"mim_id":"618352","title":"CENTRAL CENTRIFUGAL CICATRICIAL ALOPECIA; CCCA","url":"https://www.omim.org/entry/618352"},{"mim_id":"617251","title":"UNCOMBABLE HAIR SYNDROME 2; UHS2","url":"https://www.omim.org/entry/617251"},{"mim_id":"610363","title":"PEPTIDYLARGININE DEIMINASE, TYPE VI; PADI6","url":"https://www.omim.org/entry/610363"},{"mim_id":"606755","title":"PEPTIDYLARGININE DEIMINASE, TYPE III; PADI3","url":"https://www.omim.org/entry/606755"},{"mim_id":"605347","title":"PEPTIDYLARGININE DEIMINASE, TYPE IV; PADI4","url":"https://www.omim.org/entry/605347"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Vesicles","reliability":"Supported"},{"location":"Cytosol","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Group enriched","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"esophagus","ntpm":16.4},{"tissue":"urinary bladder","ntpm":25.6}],"url":"https://www.proteinatlas.org/search/PADI3"},"hgnc":{"alias_symbol":["PDI3"],"prev_symbol":[]},"alphafold":{"accession":"Q9ULW8","domains":[{"cath_id":"2.60.40.1860","chopping":"6-117","consensus_level":"high","plddt":88.8385,"start":6,"end":117},{"cath_id":"2.60.40.1700","chopping":"135-294","consensus_level":"medium","plddt":94.5231,"start":135,"end":294},{"cath_id":"3.75.10.10","chopping":"298-662","consensus_level":"medium","plddt":96.4323,"start":298,"end":662}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9ULW8","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9ULW8-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9ULW8-F1-predicted_aligned_error_v6.png","plddt_mean":93.69},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PADI3","jax_strain_url":"https://www.jax.org/strain/search?query=PADI3"},"sequence":{"accession":"Q9ULW8","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9ULW8.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9ULW8/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9ULW8"}},"corpus_meta":[{"pmid":"30763140","id":"PMC_30763140","title":"Variant PADI3 in Central Centrifugal Cicatricial Alopecia.","date":"2019","source":"The New England journal of medicine","url":"https://pubmed.ncbi.nlm.nih.gov/30763140","citation_count":96,"is_preprint":false},{"pmid":"33741961","id":"PMC_33741961","title":"Citrullination of pyruvate kinase M2 by PADI1 and PADI3 regulates glycolysis and cancer cell proliferation.","date":"2021","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/33741961","citation_count":56,"is_preprint":false},{"pmid":"16671893","id":"PMC_16671893","title":"NF-Y and Sp1/Sp3 are involved in the transcriptional regulation of the peptidylarginine deiminase type III gene (PADI3) in human keratinocytes.","date":"2006","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/16671893","citation_count":32,"is_preprint":false},{"pmid":"31669413","id":"PMC_31669413","title":"Palmitoyl Acyltransferase Activity of ZDHHC13 Regulates Skin Barrier Development Partly by Controlling PADi3 and TGM1 Protein Stability.","date":"2019","source":"The Journal of investigative dermatology","url":"https://pubmed.ncbi.nlm.nih.gov/31669413","citation_count":21,"is_preprint":false},{"pmid":"31908891","id":"PMC_31908891","title":"PADI3 induces cell cycle arrest via the Sirt2/AKT/p21 pathway and acts as a tumor suppressor gene in colon cancer.","date":"2019","source":"Cancer biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/31908891","citation_count":18,"is_preprint":false},{"pmid":"12684055","id":"PMC_12684055","title":"Thioredoxin motif of Caenorhabditis elegans PDI-3 provides Cys and His catalytic residues for transglutaminase activity.","date":"2003","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/12684055","citation_count":14,"is_preprint":false},{"pmid":"31708688","id":"PMC_31708688","title":"PADI3 plays an antitumor role via the Hsp90/CKS1 pathway in colon cancer.","date":"2019","source":"Cancer cell international","url":"https://pubmed.ncbi.nlm.nih.gov/31708688","citation_count":12,"is_preprint":false},{"pmid":"38474453","id":"PMC_38474453","title":"Atractylenolide II Suppresses Glycolysis and Induces Apoptosis by Blocking the PADI3-ERK Signaling Pathway in Endometrial Cancer Cells.","date":"2024","source":"Molecules (Basel, Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/38474453","citation_count":5,"is_preprint":false},{"pmid":"39206995","id":"PMC_39206995","title":"PADI3 inhibits epithelial-mesenchymal transition by targeting CKS1-induced signal transduction in colon cancer.","date":"2024","source":"Journal of cancer research and therapeutics","url":"https://pubmed.ncbi.nlm.nih.gov/39206995","citation_count":3,"is_preprint":false},{"pmid":"37787841","id":"PMC_37787841","title":"Pseudogenization of the Hair-Related Genes PADI3 and S100A3 in Cetaceans and Hippopotamus amphibius.","date":"2023","source":"Journal of molecular evolution","url":"https://pubmed.ncbi.nlm.nih.gov/37787841","citation_count":2,"is_preprint":false},{"pmid":"38155185","id":"PMC_38155185","title":"Citrullinated and MMP-degraded vimentin is associated with chronic pulmonary diseases and genetic variants in PADI3/PADI4 and CFH in postmenopausal women.","date":"2023","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/38155185","citation_count":1,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":7769,"output_tokens":2026,"usd":0.026849,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":8979,"output_tokens":2486,"usd":0.053523,"stage2_stop_reason":"end_turn"},"total_usd":0.080372,"stage1_batch_id":"msgbatch_013a8chtv3AWyTqv4z4sEF4C","stage2_batch_id":"msgbatch_01AHsuW63vinsvuTgzmQ3n62","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2021,\n      \"finding\": \"PADI3 (together with PADI1) citrullinates pyruvate kinase M2 (PKM2) at arginine residue R106, reprogramming cross-talk between PKM2 ligands: it lowers PKM2 sensitivity to the inhibitors tryptophan, alanine, and phenylalanine and promotes activation by serine, thereby bypassing normal physiological regulation to promote excessive glycolysis.\",\n      \"method\": \"Mass spectrometry identification of citrullination site, cancer cell overexpression/knockdown experiments, metabolic assays measuring glycolysis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, multiple orthogonal methods (MS, KD, metabolic assays) but no in vitro reconstitution with purified PADI3 alone\",\n      \"pmids\": [\"33741961\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"CCCA-associated missense mutations in PADI3 result in reduced PADI3 expression, abnormal intracellular localization of the protein, and decreased enzymatic (peptidylarginine deiminase) activity, establishing that loss of PADI3 enzymatic function underlies hair-shaft formation defects.\",\n      \"method\": \"Enzymatic activity assay, immunofluorescence for localization, immunoblotting for expression, protein structural modeling of mutants\",\n      \"journal\": \"The New England journal of medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — direct enzymatic assay plus localization experiment plus expression analysis in patient-derived samples, replicated across discovery and replication cohorts\",\n      \"pmids\": [\"30763140\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Transcription factors NF-Y and Sp1/Sp3 directly bind the PADI3 promoter region (within 129 bp of the transcription start site) in keratinocytes in vitro and in vivo, and mutation of either binding motif or siRNA knockdown of Sp1 or NF-YA markedly reduces PADI3 promoter activity and expression.\",\n      \"method\": \"Luciferase reporter assays with deletion/mutation constructs, EMSA, chromatin immunoprecipitation (ChIP), siRNA knockdown\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal methods (reporter assay, EMSA, ChIP, siRNA) in single study with rigorous mutagenesis controls\",\n      \"pmids\": [\"16671893\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ZDHHC13 palmitoylates PADI3 protein, and this palmitoylation is critical for PADI3 protein stability in vivo; loss of ZDHHC13 enzymatic activity reduces PADI3 palmitoylation and protein levels.\",\n      \"method\": \"Quantitative proteomics to identify palmitoylation candidates, biochemical palmitoylation assay, knock-in mice bearing enzymatically dead ZDHHC13 mutation\",\n      \"journal\": \"The Journal of investigative dermatology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical palmitoylation assay plus in vivo genetic model, single lab\",\n      \"pmids\": [\"31669413\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PADI3 overexpression in colon cancer cells suppresses Sirt2 and AKT phosphorylation, increases p21 expression, and induces G1 cell cycle arrest; Sirt2 overexpression partially reverses these effects, placing PADI3 upstream of the Sirt2/AKT/p21 axis. PADI3 is localized mainly in the cytoplasm, and the C-terminal domain is required for its antitumor activity.\",\n      \"method\": \"Western blot, flow cytometry for cell cycle, CCK-8 proliferation assay, colony formation assay, RNA-seq, truncation mutation experiments, immunocytochemistry, xenograft tumor formation\",\n      \"journal\": \"Cancer biology & medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal cellular assays plus rescue experiment placing PADI3 in pathway, single lab\",\n      \"pmids\": [\"31908891\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PADI3 decreases Hsp90 and CKS1 expression in colon cancer cells, and Hsp90 is required as an intermediary for PADI3-mediated downregulation of CKS1.\",\n      \"method\": \"Western blot, real-time PCR, CCK-8 proliferation assay, colony formation assay, overexpression and rescue experiments, xenograft mouse model\",\n      \"journal\": \"Cancer cell international\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, mechanism inferred from overexpression/rescue without direct biochemical interaction assays\",\n      \"pmids\": [\"31708688\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PADI3 promotes ubiquitin-dependent degradation of CKS1, thereby suppressing EMT: reduced CKS1 leads to decreased Snail and N-cadherin levels and restored E-cadherin expression, inhibiting colon cancer cell migration.\",\n      \"method\": \"Western blot, Transwell and wound healing migration assays, flow cytometry, overexpression and rescue experiments\",\n      \"journal\": \"Journal of cancer research and therapeutics\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, ubiquitin-dependent degradation inferred from rescue experiments without direct ubiquitination or proteasome assays\",\n      \"pmids\": [\"39206995\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PADI3 knockdown in endometrial cancer cells inhibits ERK signaling pathway activity, reduces glycolysis, and induces apoptosis; PADI3 overexpression reverses the anti-cancer effects of atractylenolide II, placing PADI3 upstream of ERK in this context.\",\n      \"method\": \"siRNA knockdown, overexpression rescue experiments, proliferation assays, apoptosis assays, glycolysis measurements, Western blot for ERK pathway proteins\",\n      \"journal\": \"Molecules (Basel, Switzerland)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, pathway placement based on rescue/KD without direct biochemical interaction between PADI3 and ERK\",\n      \"pmids\": [\"38474453\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PADI3 is a calcium-dependent peptidylarginine deiminase that post-translationally converts arginine to citrulline on substrate proteins including PKM2 (at R106) and hair-shaft proteins (e.g., filaggrin, trichohyalin); its expression in keratinocytes is driven by NF-Y and Sp1/Sp3 transcription factors, its protein stability is regulated by ZDHHC13-mediated palmitoylation, and disease-causing mutations reduce both its enzymatic activity and proper intracellular localization, while in cancer contexts it acts as a tumor suppressor by modulating the Sirt2/AKT/p21 and Hsp90/CKS1 pathways to induce cell cycle arrest and block EMT.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PADI3 is a calcium-dependent peptidylarginine deiminase that catalyzes the post-translational conversion of arginine to citrulline, a modification essential for normal hair-shaft formation: CCCA-associated missense mutations reduce PADI3 enzymatic activity, lower its expression, and mislocalize the protein, establishing loss of deiminase function as the basis of hair-shaft defects [#1]. In keratinocytes, PADI3 transcription is driven directly by NF-Y and Sp1/Sp3 binding within the proximal promoter [#2], and the resulting protein is stabilized by ZDHHC13-mediated palmitoylation [#3]. Beyond hair biology, PADI3 citrullinates pyruvate kinase M2 (PKM2) at R106, rewiring allosteric regulation to promote glycolysis [#0]. In cancer cells PADI3 acts as a cytoplasmic tumor suppressor, suppressing the Sirt2/AKT axis to raise p21 and drive G1 arrest, with its C-terminal domain required for antitumor activity [#4], and lowering CKS1 to restrain epithelial-mesenchymal transition [#6].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established how PADI3 expression is controlled in its native keratinocyte context, identifying the transcription factors that set its baseline levels.\",\n      \"evidence\": \"Luciferase reporter assays, EMSA, ChIP, and siRNA knockdown of Sp1/NF-YA in keratinocytes\",\n      \"pmids\": [\"16671893\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not address signals that modulate NF-Y/Sp1 activity during hair-shaft differentiation\", \"No link to enzymatic activity or substrate engagement\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Defined PADI3 loss-of-function as the molecular cause of a hair-shaft disorder by showing disease mutations cripple enzyme activity, expression, and localization.\",\n      \"evidence\": \"Enzymatic activity assays, immunofluorescence, immunoblotting, and structural modeling of patient-derived mutants across discovery and replication cohorts\",\n      \"pmids\": [\"30763140\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific hair-shaft substrates citrullinated in vivo not directly enumerated here\", \"Mechanism by which mislocalization arises from point mutations unresolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified a post-translational control point governing PADI3 protein stability, showing palmitoylation rather than transcription alone sets steady-state protein levels.\",\n      \"evidence\": \"Quantitative proteomics, biochemical palmitoylation assay, and enzymatically dead ZDHHC13 knock-in mice\",\n      \"pmids\": [\"31669413\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Palmitoylated residue(s) on PADI3 not mapped\", \"Whether palmitoylation regulates PADI3 localization or catalytic activity not addressed\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Placed PADI3 as a cytoplasmic tumor suppressor upstream of a Sirt2/AKT/p21 cell-cycle axis, extending its role beyond epidermal biology.\",\n      \"evidence\": \"Overexpression/knockdown with Sirt2 rescue, flow cytometry, truncation mutants, and xenografts in colon cancer cells\",\n      \"pmids\": [\"31908891\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether the Sirt2/AKT effect depends on PADI3 deiminase activity not established\", \"Direct PADI3 substrate in this axis unidentified\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Provided the first defined enzymatic substrate-site for PADI3 in a metabolic context, showing citrullination of PKM2-R106 reprograms allosteric regulation toward glycolysis.\",\n      \"evidence\": \"Mass spectrometry site mapping plus overexpression/knockdown and metabolic assays in cancer cells\",\n      \"pmids\": [\"33741961\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No in vitro reconstitution with purified PADI3 alone (acts with PADI1)\", \"Apparent contradiction between glycolysis-promoting and tumor-suppressive roles not reconciled\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Connected PADI3 to suppression of EMT through CKS1 downregulation, linking it to migration control in cancer.\",\n      \"evidence\": \"Migration assays and overexpression/rescue experiments measuring CKS1, Snail, N-cadherin, E-cadherin in colon cancer cells\",\n      \"pmids\": [\"39206995\", \"31708688\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Ubiquitin-dependent CKS1 degradation inferred from rescue without direct ubiquitination or proteasome assays\", \"No direct PADI3-CKS1/Hsp90 biochemical interaction demonstrated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Extended PADI3's cancer relevance to endometrial cells via ERK signaling and glycolysis.\",\n      \"evidence\": \"siRNA knockdown and overexpression rescue with proliferation, apoptosis, glycolysis, and ERK-pathway readouts\",\n      \"pmids\": [\"38474453\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Pathway placement based on KD/rescue without direct PADI3-ERK biochemical link\", \"Whether deiminase activity drives the ERK effect unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How a single deiminase reconciles glycolysis promotion via PKM2 citrullination with tumor-suppressive cell-cycle arrest, and whether its cancer-context effects require catalytic activity, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structure of PADI3 bound to a substrate\", \"Catalytic dependence of the Sirt2/AKT and CKS1 effects untested\", \"Full in vivo hair-shaft substrate repertoire not enumerated\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"PKM2\", \"ZDHHC13\", \"SP1\", \"NFYA\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":4,"faith_total":4,"faith_pct":100.0}}