{"gene":"PRSS35","run_date":"2026-06-10T06:43:36","timeline":{"discoveries":[{"year":2013,"finding":"PRSS35 is a target of HE4 (human epididymis protein 4/WFDC2), which suppresses PRSS35 protease activity; HE4 inhibits PRSS35's capacity to degrade type I collagen, and PRSS35 has functional relevance in kidney fibrosis downstream of HE4.","method":"Gene expression profiling of myofibroblasts, biological validation including protease activity assays, HE4-neutralizing antibody administration in three mouse models of renal disease","journal":"Nature medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — protease activity assays with HE4 inhibition and in vivo mouse models, single lab but multiple orthogonal methods","pmids":["23353556"],"is_preprint":false},{"year":2006,"finding":"PRSS35 is a serine protease family member with an atypical catalytic residue: the canonical serine is replaced by threonine, making it distinct from classical serine proteases. It is expressed in ovarian theca and granulosa cells and regulated by progesterone around ovulation.","method":"Suppression subtractive hybridization (SSH), genomic/bioinformatic analysis, real-time PCR, steroid ablation/replacement studies, in situ localization","journal":"Biology of reproduction","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — sequence/domain analysis combined with expression localization and hormonal regulation experiments; single lab, multiple methods","pmids":["16870946"],"is_preprint":false},{"year":2008,"finding":"PRSS35 is upregulated by gonadotropins and is expressed in theca layers of developing follicles and highly induced in granulosa cells of preovulatory follicles, as well as in forming and regressing corpus luteum, suggesting involvement in ovulation and CL formation/regression.","method":"Microarray approach, gonadotropin-induced ovulation in immature mice, immunolocalization","journal":"Endocrinology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — expression localization without direct functional assay; single lab, single approach","pmids":["18566130"],"is_preprint":false},{"year":2013,"finding":"PRSS35 expression in the periimplantation mouse uterus is regulated by progesterone and 17β-estradiol, and Prss35 knockout mice are fertile with normal ovulation, implantation, decidualization, and litter sizes, demonstrating PRSS35 is dispensable for female reproduction and embryo development.","method":"Prss35 knockout mouse generation, superovulation assays, implantation site counting, marker gene expression analysis, ovariectomy/hormone replacement","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO model with multiple reproductive phenotype readouts; clean negative result with adequate controls","pmids":["23451081"],"is_preprint":false},{"year":2023,"finding":"Active PRSS35 is processed by proprotein convertases via cleavage. Active PRSS35 suppresses CXCL2 protein levels through targeted cleavage at a tandem lysine (KK) recognition motif, thereby reducing neutrophil recruitment and formation of neutrophil extracellular traps (NETs), ultimately suppressing hepatocellular carcinoma progression.","method":"Biochemical cleavage assays, mutational analysis of KK motif, CXCL2 protein quantification, neutrophil recruitment assays, NET formation assays, in vivo HCC models","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — proprotein convertase cleavage, substrate identification with motif mutagenesis, in vitro and in vivo functional validation in single rigorous study","pmids":["36934105"],"is_preprint":false},{"year":2021,"finding":"PRSS35 is upregulated in cancer-associated fibroblasts (CAFs) in wound-induced skin tumors; ablation of PRSS35 in mouse models results in aberrant collagen composition in the ECM and increased tumor incidence, demonstrating that PRSS35-mediated ECM remodeling by CAFs suppresses squamous tumor initiation.","method":"Genome-wide expression analysis of fibroblasts, PRSS35 ablation in mouse wound-induced and chemically-induced tumorigenesis models, ECM/collagen composition analysis, tumor incidence scoring","journal":"EMBO reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic ablation in two mouse tumor models with collagen composition readout; single lab, two orthogonal models","pmids":["33780134"],"is_preprint":false},{"year":2023,"finding":"Hyperosmotic stress upregulates PRSS35 in human fibroblasts in a manner dependent on kinases p38 and JNK and transcription factors NFAT5 and ATF2. PRSS35 binds collagens and collagen-associated proteins, affects the extracellular matrix proteome, and limits cell proliferation under hyperosmotic stress conditions.","method":"Sorbitol-treated human fibroblasts in 3D culture, kinase/transcription factor inhibition, proteomics to identify binding partners and matrisome changes, cell proliferation assays","journal":"Science advances","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (proteomic binding partners, signaling pathway dissection, functional proliferation assay) in single lab study","pmids":["37647410"],"is_preprint":false},{"year":2017,"finding":"IL-36α stimulation promotes production of PRSS35 in NIH3T3 fibroblasts, placing PRSS35 downstream of IL-36α signaling as a collagen remodeling-associated enzyme in the context of kidney tubulointerstitial inflammation.","method":"Cell culture stimulation assay (NIH3T3 fibroblasts treated with IL-36α), gene expression measurement; UUO mouse model","journal":"Frontiers in immunology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single cell culture assay without mechanistic follow-up; single lab, single method","pmids":["29109726"],"is_preprint":false},{"year":2023,"finding":"In lupus nephritis mice, HE4 silencing reduces renal fibrosis and increases expression of PRSS35 and PRSS23, consistent with HE4 acting as an inhibitor of these proteases in the C3/MMPs/PRSS axis.","method":"HE4 shRNA adenovirus injection in MRL/LPR mice, Western blotting for prss35 and prss23, histological fibrosis scoring, kidney injury marker measurement","journal":"Naunyn-Schmiedeberg's archives of pharmacology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — in vivo knockdown with Western blot readout; single lab, single method, no direct PRSS35 mechanistic assay","pmids":["38157023"],"is_preprint":false},{"year":2024,"finding":"Osmotic pressure activates the p38 MAPK stress response pathway and increases expression of PRSS35 as an osmoresponsive gene in primary human dermal fibroblasts in 3D hydrogel culture, but not in 2D culture, indicating dimensionality modulates PRSS35 osmotic induction.","method":"Cyclic hydrostatic pressure and hyperosmotic stress applied to fibroblasts in collagen and PEG-based 3D hydrogels vs. 2D culture; gene expression and pathway analysis","journal":"Biomaterials advances","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single gene expression readout in mechanical stress models; single lab, correlative without direct functional assay for PRSS35","pmids":["38972277"],"is_preprint":false}],"current_model":"PRSS35 is a secreted pseudo-serine protease (bearing threonine in place of the canonical catalytic serine) that is processed by proprotein convertases into its active form; it cleaves substrates including CXCL2 at tandem lysine (KK) motifs to suppress neutrophil recruitment and NET formation in hepatocellular carcinoma, remodels extracellular matrix collagens to regulate fibroblast proliferation under hyperosmotic stress and to suppress squamous tumor initiation in skin, and is itself inhibited by HE4/WFDC2, which blocks PRSS35-mediated collagen degradation in the context of renal fibrosis."},"narrative":{"mechanistic_narrative":"PRSS35 is a secreted serine-protease-family enzyme that remodels the extracellular matrix and processes secreted protein substrates to constrain tissue inflammation and tumor initiation [PMID:36934105, PMID:33780134]. It is atypical for its family, carrying a threonine in place of the canonical catalytic serine, and was first identified as a hormonally regulated gene in ovarian theca and granulosa cells [PMID:16870946]. The enzyme is matured from a precursor by proprotein-convertase cleavage; in its active form it cleaves CXCL2 at a tandem lysine (KK) motif, lowering CXCL2 levels to limit neutrophil recruitment and neutrophil extracellular trap formation and thereby suppressing hepatocellular carcinoma progression [PMID:36934105]. In fibroblasts, PRSS35 binds collagens and collagen-associated proteins and reshapes the matrisome: its ablation in skin produces aberrant ECM collagen composition and increased squamous tumor incidence, identifying an ECM-remodeling, tumor-suppressive role in cancer-associated fibroblasts [PMID:33780134, PMID:37647410]. PRSS35 is induced by hyperosmotic stress through p38/JNK signaling and the transcription factors NFAT5 and ATF2, and it limits fibroblast proliferation under these conditions [PMID:37647410]. Its collagen-degrading activity is suppressed by HE4/WFDC2, an interaction with functional relevance in kidney fibrosis [PMID:23353556]. Despite its hormonal regulation in the reproductive tract, Prss35-knockout mice are fertile with normal ovulation, implantation, and decidualization, indicating PRSS35 is dispensable for female reproduction [PMID:23451081].","teleology":[{"year":2006,"claim":"Established PRSS35 as a serine-protease-family member with an atypical catalytic residue (threonine for serine) and placed its expression in hormonally regulated ovarian cells, framing it as a candidate ovulation-associated enzyme.","evidence":"Subtractive hybridization, bioinformatic/domain analysis, real-time PCR and steroid ablation/replacement with in situ localization in ovary","pmids":["16870946"],"confidence":"Medium","gaps":["No direct demonstration of catalytic activity or substrate","Atypical Thr residue's effect on proteolytic competence not tested","Function inferred from expression pattern only"]},{"year":2008,"claim":"Extended the hormonal-regulation picture by showing gonadotropin induction across preovulatory follicles and corpus luteum, reinforcing a candidate reproductive role.","evidence":"Microarray and immunolocalization in gonadotropin-induced ovulation in immature mice","pmids":["18566130"],"confidence":"Low","gaps":["Expression correlation only, no functional assay","No substrate or mechanism","Causal role in ovulation untested"]},{"year":2013,"claim":"Tested whether PRSS35 is required for reproduction and found it dispensable, redirecting attention away from an essential reproductive role.","evidence":"Prss35 knockout mice with superovulation, implantation, decidualization and litter-size readouts plus ovariectomy/hormone replacement","pmids":["23451081"],"confidence":"Medium","gaps":["Clean negative result leaves primary physiological function unidentified","Possible redundancy with other proteases not addressed","No challenge or stress phenotype examined"]},{"year":2013,"claim":"Connected PRSS35 to ECM proteolysis and disease by identifying it as an HE4/WFDC2 target whose collagen-degrading activity is suppressed, implicating it in renal fibrosis.","evidence":"Myofibroblast expression profiling, protease activity assays with HE4 inhibition, and HE4-neutralizing antibody in three mouse renal disease models","pmids":["23353556"],"confidence":"Medium","gaps":["Mechanism of HE4 inhibition (direct binding vs. indirect) not resolved","Cleavage site on collagen not mapped","Single lab"]},{"year":2017,"claim":"Placed PRSS35 downstream of inflammatory IL-36α signaling in fibroblasts, linking its expression to tubulointerstitial inflammation.","evidence":"IL-36α stimulation of NIH3T3 fibroblasts with gene expression readout and UUO mouse model","pmids":["29109726"],"confidence":"Low","gaps":["Single expression assay without mechanistic follow-up","No demonstration of PRSS35 protease activity in this context","Causal contribution to inflammation untested"]},{"year":2021,"claim":"Demonstrated a tumor-suppressive function via ECM remodeling, showing CAF-derived PRSS35 controls collagen composition and restrains squamous tumor initiation.","evidence":"Fibroblast expression profiling and PRSS35 ablation in wound-induced and chemically-induced mouse tumor models with collagen composition and tumor incidence readouts","pmids":["33780134"],"confidence":"Medium","gaps":["Direct collagen substrate and cleavage products not defined","Whether tumor suppression requires catalytic activity untested","Mechanism linking ECM composition to tumor initiation unclear"]},{"year":2023,"claim":"Defined PRSS35 maturation and a specific protein substrate, showing proprotein-convertase activation enables KK-motif cleavage of CXCL2 to dampen neutrophil/NET responses and suppress HCC.","evidence":"Biochemical cleavage assays, KK-motif mutagenesis, CXCL2 quantification, neutrophil and NET assays, and in vivo HCC models","pmids":["36934105"],"confidence":"High","gaps":["Full substrate repertoire beyond CXCL2 unknown","Structural basis for KK recognition by the atypical active site not solved","Identity of the relevant proprotein convertase(s) not specified"]},{"year":2023,"claim":"Established the upstream osmotic-stress signaling that induces PRSS35 and its collagen-binding, matrisome-shaping, anti-proliferative role in fibroblasts.","evidence":"Sorbitol-treated 3D fibroblast cultures with p38/JNK and NFAT5/ATF2 inhibition, proteomic binding-partner and matrisome analysis, and proliferation assays","pmids":["37647410"],"confidence":"Medium","gaps":["Binding to collagens shown but cleavage of specific collagens not mapped","Mechanism linking ECM remodeling to proliferation control unresolved","In vivo relevance of osmotic induction not established"]},{"year":2023,"claim":"Reinforced the HE4–PRSS35 inhibitory axis in vivo by showing HE4 silencing raises PRSS35 expression and reduces fibrosis in lupus nephritis.","evidence":"HE4 shRNA adenovirus in MRL/LPR mice with Western blotting and histological fibrosis scoring","pmids":["38157023"],"confidence":"Low","gaps":["Correlative expression change without direct PRSS35 mechanistic assay","Cannot distinguish direct vs. indirect HE4 effect","Single method readout"]},{"year":2024,"claim":"Refined the osmotic-induction model by showing PRSS35 upregulation via p38 MAPK is dependent on 3D culture dimensionality, indicating mechanical/dimensional context gates its induction.","evidence":"Cyclic hydrostatic pressure and hyperosmotic stress in 3D hydrogels vs. 2D culture with gene expression and pathway analysis","pmids":["38972277"],"confidence":"Low","gaps":["Single gene-expression readout, no functional PRSS35 assay","Correlative pathway assignment","Physiological significance of dimensionality effect unknown"]},{"year":null,"claim":"The full substrate repertoire of PRSS35, the structural basis by which its atypical threonine active site achieves proteolysis, and the unifying physiological function reconciling its reproductive, fibrotic, and anti-tumor roles remain unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No structure or catalytic-mechanism model for the Thr-substituted active site","Substrates beyond CXCL2 and collagens not comprehensively mapped","No single integrative physiological model across tissues"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[4]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[6]}],"localization":[{"term_id":"GO:0005576","term_label":"extracellular region","supporting_discovery_ids":[4,6]},{"term_id":"GO:0031012","term_label":"extracellular matrix","supporting_discovery_ids":[5,6]}],"pathway":[{"term_id":"R-HSA-1474244","term_label":"Extracellular matrix organization","supporting_discovery_ids":[5,6]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[4]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[4,5]}],"complexes":[],"partners":["WFDC2","CXCL2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8N3Z0","full_name":"Inactive serine protease 35","aliases":[],"length_aa":413,"mass_kda":47.1,"function":"","subcellular_location":"Secreted","url":"https://www.uniprot.org/uniprotkb/Q8N3Z0/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/PRSS35","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/PRSS35","total_profiled":1310},"omim":[{"mim_id":"618376","title":"PROTEASE, SERINE, 23; PRSS23","url":"https://www.omim.org/entry/618376"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"ovary","ntpm":17.9},{"tissue":"retina","ntpm":46.8}],"url":"https://www.proteinatlas.org/search/PRSS35"},"hgnc":{"alias_symbol":["MGC46520","dJ223E3.1"],"prev_symbol":["C6orf158"]},"alphafold":{"accession":"Q8N3Z0","domains":[{"cath_id":"2.40.10.10","chopping":"31-105_127-192_246-405","consensus_level":"medium","plddt":90.2566,"start":31,"end":405}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8N3Z0","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8N3Z0-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8N3Z0-F1-predicted_aligned_error_v6.png","plddt_mean":79.44},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PRSS35","jax_strain_url":"https://www.jax.org/strain/search?query=PRSS35"},"sequence":{"accession":"Q8N3Z0","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8N3Z0.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8N3Z0/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8N3Z0"}},"corpus_meta":[{"pmid":"23353556","id":"PMC_23353556","title":"Identification of human epididymis protein-4 as a fibroblast-derived mediator of fibrosis.","date":"2013","source":"Nature medicine","url":"https://pubmed.ncbi.nlm.nih.gov/23353556","citation_count":217,"is_preprint":false},{"pmid":"23092984","id":"PMC_23092984","title":"Genome-wide association of mood-incongruent psychotic bipolar disorder.","date":"2012","source":"Translational psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/23092984","citation_count":56,"is_preprint":false},{"pmid":"36934105","id":"PMC_36934105","title":"Secreted protease PRSS35 suppresses hepatocellular carcinoma by disabling CXCL2-mediated neutrophil extracellular traps.","date":"2023","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/36934105","citation_count":53,"is_preprint":false},{"pmid":"16870946","id":"PMC_16870946","title":"The identification of novel ovarian proteases through the use of genomic and bioinformatic methodologies.","date":"2006","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/16870946","citation_count":38,"is_preprint":false},{"pmid":"18566130","id":"PMC_18566130","title":"Expression and localization of the serine proteases high-temperature requirement factor A1, serine protease 23, and serine protease 35 in the mouse ovary.","date":"2008","source":"Endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/18566130","citation_count":30,"is_preprint":false},{"pmid":"23451081","id":"PMC_23451081","title":"Distinct spatiotemporal expression of serine proteases Prss23 and Prss35 in periimplantation mouse uterus and dispensable function of Prss35 in fertility.","date":"2013","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/23451081","citation_count":27,"is_preprint":false},{"pmid":"32674273","id":"PMC_32674273","title":"Transcriptional Profiling of Normal, Stenotic, and Regurgitant Human Aortic Valves.","date":"2020","source":"Genes","url":"https://pubmed.ncbi.nlm.nih.gov/32674273","citation_count":25,"is_preprint":false},{"pmid":"28793334","id":"PMC_28793334","title":"Spheroid growth in ovarian cancer alters transcriptome responses for stress pathways and epigenetic responses.","date":"2017","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/28793334","citation_count":24,"is_preprint":false},{"pmid":"20511563","id":"PMC_20511563","title":"Novel cleft susceptibility genes in chromosome 6q.","date":"2010","source":"Journal of dental research","url":"https://pubmed.ncbi.nlm.nih.gov/20511563","citation_count":22,"is_preprint":false},{"pmid":"29109726","id":"PMC_29109726","title":"IL-36α Regulates Tubulointerstitial Inflammation in the Mouse Kidney.","date":"2017","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/29109726","citation_count":22,"is_preprint":false},{"pmid":"33795767","id":"PMC_33795767","title":"Transcriptome analysis of human dermal fibroblasts following red light phototherapy.","date":"2021","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/33795767","citation_count":17,"is_preprint":false},{"pmid":"36860864","id":"PMC_36860864","title":"Research into the characteristic molecules significantly affecting liver cancer immunotherapy.","date":"2023","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/36860864","citation_count":17,"is_preprint":false},{"pmid":"31920707","id":"PMC_31920707","title":"Angiogenic Transformation in Human Brain Micro Endothelial Cells: Whole Genome DNA Methylation and Transcriptomic Analysis.","date":"2019","source":"Frontiers in physiology","url":"https://pubmed.ncbi.nlm.nih.gov/31920707","citation_count":17,"is_preprint":false},{"pmid":"33780134","id":"PMC_33780134","title":"Fibrotic enzymes modulate wound-induced skin tumorigenesis.","date":"2021","source":"EMBO reports","url":"https://pubmed.ncbi.nlm.nih.gov/33780134","citation_count":14,"is_preprint":false},{"pmid":"37647410","id":"PMC_37647410","title":"Serine protease 35 regulates the fibroblast matrisome in response to hyperosmotic stress.","date":"2023","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/37647410","citation_count":12,"is_preprint":false},{"pmid":"38157023","id":"PMC_38157023","title":"Silencing HE4 alleviates the renal fibrosis in lupus nephritis mice by regulating the C3/MMPs/prss axis.","date":"2023","source":"Naunyn-Schmiedeberg's archives of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/38157023","citation_count":4,"is_preprint":false},{"pmid":"35460443","id":"PMC_35460443","title":"Transcriptional profile of cumulus associated GJA1, PTX3, PRSS35, and SERPINE2 genes with oocytes and embryonic development in water buffalo.","date":"2022","source":"Molecular biology reports","url":"https://pubmed.ncbi.nlm.nih.gov/35460443","citation_count":3,"is_preprint":false},{"pmid":"38972277","id":"PMC_38972277","title":"Variations in fluid chemical potential induce fibroblast mechano-response in 3D hydrogels.","date":"2024","source":"Biomaterials advances","url":"https://pubmed.ncbi.nlm.nih.gov/38972277","citation_count":3,"is_preprint":false},{"pmid":"39260308","id":"PMC_39260308","title":"SLAMF1 as a novel molecule mediating the causal association between rheumatoid arthritis and interstitial lung disease: A Mendelian randomization study combined with transcriptomics and in vivo validation.","date":"2024","source":"International immunopharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/39260308","citation_count":3,"is_preprint":false},{"pmid":"40132315","id":"PMC_40132315","title":"Identification of key genes associated with residual feed intake in small-sized meat ducks through integrated analysis of mRNA and miRNA transcriptomes.","date":"2025","source":"Poultry science","url":"https://pubmed.ncbi.nlm.nih.gov/40132315","citation_count":1,"is_preprint":false},{"pmid":"41828317","id":"PMC_41828317","title":"Toward Understanding the Role of miRNAs in Cleft Palate Only: Observations from Patient Tissues and In Vitro Assays.","date":"2026","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/41828317","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.04.17.648896","title":"Senescence-induced reparative fibroblasts enable scarless wound healing in aged murine skin","date":"2025-04-21","source":"bioRxiv","url":"https://doi.org/10.1101/2025.04.17.648896","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":13606,"output_tokens":2436,"usd":0.038679,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9581,"output_tokens":3513,"usd":0.067865,"stage2_stop_reason":"end_turn"},"total_usd":0.106544,"stage1_batch_id":"msgbatch_01CNoDbkChHEkgrJnL7BwtGJ","stage2_batch_id":"msgbatch_016Y4x9E9Wg5VQDANj9quSQD","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2013,\n      \"finding\": \"PRSS35 is a target of HE4 (human epididymis protein 4/WFDC2), which suppresses PRSS35 protease activity; HE4 inhibits PRSS35's capacity to degrade type I collagen, and PRSS35 has functional relevance in kidney fibrosis downstream of HE4.\",\n      \"method\": \"Gene expression profiling of myofibroblasts, biological validation including protease activity assays, HE4-neutralizing antibody administration in three mouse models of renal disease\",\n      \"journal\": \"Nature medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — protease activity assays with HE4 inhibition and in vivo mouse models, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"23353556\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"PRSS35 is a serine protease family member with an atypical catalytic residue: the canonical serine is replaced by threonine, making it distinct from classical serine proteases. It is expressed in ovarian theca and granulosa cells and regulated by progesterone around ovulation.\",\n      \"method\": \"Suppression subtractive hybridization (SSH), genomic/bioinformatic analysis, real-time PCR, steroid ablation/replacement studies, in situ localization\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — sequence/domain analysis combined with expression localization and hormonal regulation experiments; single lab, multiple methods\",\n      \"pmids\": [\"16870946\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"PRSS35 is upregulated by gonadotropins and is expressed in theca layers of developing follicles and highly induced in granulosa cells of preovulatory follicles, as well as in forming and regressing corpus luteum, suggesting involvement in ovulation and CL formation/regression.\",\n      \"method\": \"Microarray approach, gonadotropin-induced ovulation in immature mice, immunolocalization\",\n      \"journal\": \"Endocrinology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — expression localization without direct functional assay; single lab, single approach\",\n      \"pmids\": [\"18566130\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"PRSS35 expression in the periimplantation mouse uterus is regulated by progesterone and 17β-estradiol, and Prss35 knockout mice are fertile with normal ovulation, implantation, decidualization, and litter sizes, demonstrating PRSS35 is dispensable for female reproduction and embryo development.\",\n      \"method\": \"Prss35 knockout mouse generation, superovulation assays, implantation site counting, marker gene expression analysis, ovariectomy/hormone replacement\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO model with multiple reproductive phenotype readouts; clean negative result with adequate controls\",\n      \"pmids\": [\"23451081\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Active PRSS35 is processed by proprotein convertases via cleavage. Active PRSS35 suppresses CXCL2 protein levels through targeted cleavage at a tandem lysine (KK) recognition motif, thereby reducing neutrophil recruitment and formation of neutrophil extracellular traps (NETs), ultimately suppressing hepatocellular carcinoma progression.\",\n      \"method\": \"Biochemical cleavage assays, mutational analysis of KK motif, CXCL2 protein quantification, neutrophil recruitment assays, NET formation assays, in vivo HCC models\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — proprotein convertase cleavage, substrate identification with motif mutagenesis, in vitro and in vivo functional validation in single rigorous study\",\n      \"pmids\": [\"36934105\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"PRSS35 is upregulated in cancer-associated fibroblasts (CAFs) in wound-induced skin tumors; ablation of PRSS35 in mouse models results in aberrant collagen composition in the ECM and increased tumor incidence, demonstrating that PRSS35-mediated ECM remodeling by CAFs suppresses squamous tumor initiation.\",\n      \"method\": \"Genome-wide expression analysis of fibroblasts, PRSS35 ablation in mouse wound-induced and chemically-induced tumorigenesis models, ECM/collagen composition analysis, tumor incidence scoring\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic ablation in two mouse tumor models with collagen composition readout; single lab, two orthogonal models\",\n      \"pmids\": [\"33780134\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Hyperosmotic stress upregulates PRSS35 in human fibroblasts in a manner dependent on kinases p38 and JNK and transcription factors NFAT5 and ATF2. PRSS35 binds collagens and collagen-associated proteins, affects the extracellular matrix proteome, and limits cell proliferation under hyperosmotic stress conditions.\",\n      \"method\": \"Sorbitol-treated human fibroblasts in 3D culture, kinase/transcription factor inhibition, proteomics to identify binding partners and matrisome changes, cell proliferation assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (proteomic binding partners, signaling pathway dissection, functional proliferation assay) in single lab study\",\n      \"pmids\": [\"37647410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"IL-36α stimulation promotes production of PRSS35 in NIH3T3 fibroblasts, placing PRSS35 downstream of IL-36α signaling as a collagen remodeling-associated enzyme in the context of kidney tubulointerstitial inflammation.\",\n      \"method\": \"Cell culture stimulation assay (NIH3T3 fibroblasts treated with IL-36α), gene expression measurement; UUO mouse model\",\n      \"journal\": \"Frontiers in immunology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single cell culture assay without mechanistic follow-up; single lab, single method\",\n      \"pmids\": [\"29109726\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In lupus nephritis mice, HE4 silencing reduces renal fibrosis and increases expression of PRSS35 and PRSS23, consistent with HE4 acting as an inhibitor of these proteases in the C3/MMPs/PRSS axis.\",\n      \"method\": \"HE4 shRNA adenovirus injection in MRL/LPR mice, Western blotting for prss35 and prss23, histological fibrosis scoring, kidney injury marker measurement\",\n      \"journal\": \"Naunyn-Schmiedeberg's archives of pharmacology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — in vivo knockdown with Western blot readout; single lab, single method, no direct PRSS35 mechanistic assay\",\n      \"pmids\": [\"38157023\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Osmotic pressure activates the p38 MAPK stress response pathway and increases expression of PRSS35 as an osmoresponsive gene in primary human dermal fibroblasts in 3D hydrogel culture, but not in 2D culture, indicating dimensionality modulates PRSS35 osmotic induction.\",\n      \"method\": \"Cyclic hydrostatic pressure and hyperosmotic stress applied to fibroblasts in collagen and PEG-based 3D hydrogels vs. 2D culture; gene expression and pathway analysis\",\n      \"journal\": \"Biomaterials advances\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single gene expression readout in mechanical stress models; single lab, correlative without direct functional assay for PRSS35\",\n      \"pmids\": [\"38972277\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PRSS35 is a secreted pseudo-serine protease (bearing threonine in place of the canonical catalytic serine) that is processed by proprotein convertases into its active form; it cleaves substrates including CXCL2 at tandem lysine (KK) motifs to suppress neutrophil recruitment and NET formation in hepatocellular carcinoma, remodels extracellular matrix collagens to regulate fibroblast proliferation under hyperosmotic stress and to suppress squamous tumor initiation in skin, and is itself inhibited by HE4/WFDC2, which blocks PRSS35-mediated collagen degradation in the context of renal fibrosis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PRSS35 is a secreted serine-protease-family enzyme that remodels the extracellular matrix and processes secreted protein substrates to constrain tissue inflammation and tumor initiation [#4, #5]. It is atypical for its family, carrying a threonine in place of the canonical catalytic serine, and was first identified as a hormonally regulated gene in ovarian theca and granulosa cells [#1]. The enzyme is matured from a precursor by proprotein-convertase cleavage; in its active form it cleaves CXCL2 at a tandem lysine (KK) motif, lowering CXCL2 levels to limit neutrophil recruitment and neutrophil extracellular trap formation and thereby suppressing hepatocellular carcinoma progression [#4]. In fibroblasts, PRSS35 binds collagens and collagen-associated proteins and reshapes the matrisome: its ablation in skin produces aberrant ECM collagen composition and increased squamous tumor incidence, identifying an ECM-remodeling, tumor-suppressive role in cancer-associated fibroblasts [#5, #6]. PRSS35 is induced by hyperosmotic stress through p38/JNK signaling and the transcription factors NFAT5 and ATF2, and it limits fibroblast proliferation under these conditions [#6]. Its collagen-degrading activity is suppressed by HE4/WFDC2, an interaction with functional relevance in kidney fibrosis [#0]. Despite its hormonal regulation in the reproductive tract, Prss35-knockout mice are fertile with normal ovulation, implantation, and decidualization, indicating PRSS35 is dispensable for female reproduction [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established PRSS35 as a serine-protease-family member with an atypical catalytic residue (threonine for serine) and placed its expression in hormonally regulated ovarian cells, framing it as a candidate ovulation-associated enzyme.\",\n      \"evidence\": \"Subtractive hybridization, bioinformatic/domain analysis, real-time PCR and steroid ablation/replacement with in situ localization in ovary\",\n      \"pmids\": [\"16870946\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct demonstration of catalytic activity or substrate\", \"Atypical Thr residue's effect on proteolytic competence not tested\", \"Function inferred from expression pattern only\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Extended the hormonal-regulation picture by showing gonadotropin induction across preovulatory follicles and corpus luteum, reinforcing a candidate reproductive role.\",\n      \"evidence\": \"Microarray and immunolocalization in gonadotropin-induced ovulation in immature mice\",\n      \"pmids\": [\"18566130\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Expression correlation only, no functional assay\", \"No substrate or mechanism\", \"Causal role in ovulation untested\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Tested whether PRSS35 is required for reproduction and found it dispensable, redirecting attention away from an essential reproductive role.\",\n      \"evidence\": \"Prss35 knockout mice with superovulation, implantation, decidualization and litter-size readouts plus ovariectomy/hormone replacement\",\n      \"pmids\": [\"23451081\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Clean negative result leaves primary physiological function unidentified\", \"Possible redundancy with other proteases not addressed\", \"No challenge or stress phenotype examined\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Connected PRSS35 to ECM proteolysis and disease by identifying it as an HE4/WFDC2 target whose collagen-degrading activity is suppressed, implicating it in renal fibrosis.\",\n      \"evidence\": \"Myofibroblast expression profiling, protease activity assays with HE4 inhibition, and HE4-neutralizing antibody in three mouse renal disease models\",\n      \"pmids\": [\"23353556\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of HE4 inhibition (direct binding vs. indirect) not resolved\", \"Cleavage site on collagen not mapped\", \"Single lab\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Placed PRSS35 downstream of inflammatory IL-36α signaling in fibroblasts, linking its expression to tubulointerstitial inflammation.\",\n      \"evidence\": \"IL-36α stimulation of NIH3T3 fibroblasts with gene expression readout and UUO mouse model\",\n      \"pmids\": [\"29109726\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single expression assay without mechanistic follow-up\", \"No demonstration of PRSS35 protease activity in this context\", \"Causal contribution to inflammation untested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstrated a tumor-suppressive function via ECM remodeling, showing CAF-derived PRSS35 controls collagen composition and restrains squamous tumor initiation.\",\n      \"evidence\": \"Fibroblast expression profiling and PRSS35 ablation in wound-induced and chemically-induced mouse tumor models with collagen composition and tumor incidence readouts\",\n      \"pmids\": [\"33780134\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct collagen substrate and cleavage products not defined\", \"Whether tumor suppression requires catalytic activity untested\", \"Mechanism linking ECM composition to tumor initiation unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined PRSS35 maturation and a specific protein substrate, showing proprotein-convertase activation enables KK-motif cleavage of CXCL2 to dampen neutrophil/NET responses and suppress HCC.\",\n      \"evidence\": \"Biochemical cleavage assays, KK-motif mutagenesis, CXCL2 quantification, neutrophil and NET assays, and in vivo HCC models\",\n      \"pmids\": [\"36934105\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full substrate repertoire beyond CXCL2 unknown\", \"Structural basis for KK recognition by the atypical active site not solved\", \"Identity of the relevant proprotein convertase(s) not specified\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established the upstream osmotic-stress signaling that induces PRSS35 and its collagen-binding, matrisome-shaping, anti-proliferative role in fibroblasts.\",\n      \"evidence\": \"Sorbitol-treated 3D fibroblast cultures with p38/JNK and NFAT5/ATF2 inhibition, proteomic binding-partner and matrisome analysis, and proliferation assays\",\n      \"pmids\": [\"37647410\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Binding to collagens shown but cleavage of specific collagens not mapped\", \"Mechanism linking ECM remodeling to proliferation control unresolved\", \"In vivo relevance of osmotic induction not established\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Reinforced the HE4–PRSS35 inhibitory axis in vivo by showing HE4 silencing raises PRSS35 expression and reduces fibrosis in lupus nephritis.\",\n      \"evidence\": \"HE4 shRNA adenovirus in MRL/LPR mice with Western blotting and histological fibrosis scoring\",\n      \"pmids\": [\"38157023\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Correlative expression change without direct PRSS35 mechanistic assay\", \"Cannot distinguish direct vs. indirect HE4 effect\", \"Single method readout\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Refined the osmotic-induction model by showing PRSS35 upregulation via p38 MAPK is dependent on 3D culture dimensionality, indicating mechanical/dimensional context gates its induction.\",\n      \"evidence\": \"Cyclic hydrostatic pressure and hyperosmotic stress in 3D hydrogels vs. 2D culture with gene expression and pathway analysis\",\n      \"pmids\": [\"38972277\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single gene-expression readout, no functional PRSS35 assay\", \"Correlative pathway assignment\", \"Physiological significance of dimensionality effect unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The full substrate repertoire of PRSS35, the structural basis by which its atypical threonine active site achieves proteolysis, and the unifying physiological function reconciling its reproductive, fibrotic, and anti-tumor roles remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structure or catalytic-mechanism model for the Thr-substituted active site\", \"Substrates beyond CXCL2 and collagens not comprehensively mapped\", \"No single integrative physiological model across tissues\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [6]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005576\", \"supporting_discovery_ids\": [4, 6]},\n      {\"term_id\": \"GO:0031012\", \"supporting_discovery_ids\": [5, 6]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1474244\", \"supporting_discovery_ids\": [5, 6]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [4, 5]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"WFDC2\",\n      \"CXCL2\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":7,"faith_total":7,"faith_pct":100.0}}