{"gene":"FBLN1","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":2002,"finding":"FBLN1 protein is expressed in the extracellular matrix in association with digits during developing limb formation; the FBLN1-D splice variant, but not the FBLN1-C splice variant, shows altered incorporation into ECM and conditioned medium in fibroblasts from patients with synpolydactyly, suggesting splice-variant-specific ECM roles.","method":"Immunolocalization of FBLN1 in developing limb ECM; analysis of FBLN1-C and FBLN1-D splice variant levels in patient fibroblast ECM and conditioned medium","journal":"Journal of medical genetics","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct biochemical fractionation of ECM from patient fibroblasts with two splice variants distinguished; single lab, no reconstitution","pmids":["11836357"],"is_preprint":false},{"year":2005,"finding":"Fbln1 null mice are lethal, establishing FBLN1 as an essential component of blood vessel walls; Fbln1 is expressed in spongiotrophoblast, endothelia of large fetal blood vessels, and a subset of giant cells in the mouse placenta, but Fbln1 alone is not the key cause of placental hyperplasia phenotypes in a congenic interspecies hybrid model.","method":"Homozygous targeted gene deletion (knockout mouse), in situ expression analysis, rescue experiment attempting to normalize Fbln1 expression in hyperplastic placentas","journal":"Placenta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic knockout with defined lethal phenotype and placental expression pattern; rescue experiment showed no rescue (negative result for placental role); single lab","pmids":["16338003"],"is_preprint":false},{"year":2021,"finding":"FBLN1 promotes chondrocyte proliferation in knee cartilage by increasing phosphorylation of Smad2 and upregulating Col2; inhibition of pSmad2 abolishes the FBLN1-driven proliferation effect.","method":"Adenoviral overexpression of FBLN1 in isolated chondrocytes, EdU proliferation assay, Western blotting for pSmad2 and Col2, pharmacological inhibition of pSmad2","journal":"Journal of orthopaedic science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — overexpression with defined signaling readout and inhibitor rescue; single lab, single method per endpoint","pmids":["33610427"],"is_preprint":false},{"year":2024,"finding":"FBLN1 overexpression inhibits the TGF-β/Smad signaling pathway (decreased TGF-β and p-Smad protein levels) and thereby reduces ferroptosis markers (MDA, lipid ROS, Fe2+) in LPS-treated lung epithelial cells; FBLN1 and TGF-β have antagonistic roles in regulating ferroptosis.","method":"Overexpression of FBLN1 and TGF-β1 in MLE-12 cells treated with LPS, Western blotting for TGF-β/Smad pathway proteins and ferroptosis markers, cell viability assay","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — in vitro overexpression with multiple molecular readouts but single lab, no orthogonal validation of TGF-β/Smad interaction","pmids":["39671383"],"is_preprint":false},{"year":2024,"finding":"FBLN1 mRNA stability is regulated by m6A methylation at a site in its 3'UTR; YTHDF2 binds this m6A site and induces FBLN1 mRNA instability, reducing FBLN1 expression and suppressing osteogenic differentiation of WJCMSCs. Mutation of the m6A site enhances FBLN1 mRNA stability and osteogenic differentiation. miR-615-3p also negatively regulates FBLN1 by binding its 3'UTR, and YTHDF2 facilitates miR-615-3p-mediated FBLN1 mRNA decay.","method":"m6A site mutation, YTHDF2 overexpression/knockdown, miR-615-3p gain/loss of function, mRNA stability assays, osteogenic differentiation assays in WJCMSCs, bone regeneration in vivo model","journal":"Cell proliferation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal molecular methods (m6A mutation, RBP binding, miRNA target) in single lab; functional consequence validated in vitro and in vivo","pmids":["38353178"],"is_preprint":false},{"year":2022,"finding":"A missense variant of FBLN1 (p.Arg550His) results in reduced protein stability and decreased FBLN1 expression. In combination with an ARHGAP31 variant, but not alone, FBLN1 mutation decreases cell viability, impairs proliferation, activates apoptosis, reduces Cdc42 activity, and activates the MAPK/ERK pathway in vitro, indicating synergistic disruption of cellular signaling.","method":"Expression vectors with wild-type and mutant FBLN1 transfected in mammalian cells; immunoblotting for protein stability and expression; cell viability (CCK-8), immunofluorescence, co-transfection with ARHGAP31 mutant; Cdc42 activity assay; Western blotting for MAPK/ERK pathway","journal":"Frontiers in genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — multiple functional assays with mutant protein in vitro; single lab; synergistic effect context-dependent","pmids":["36176297"],"is_preprint":false},{"year":2024,"finding":"fbln1 mutant zebrafish show increased mineralization of cranial elements, altered ceratohyal angle, increased vertebral dimensions and tissue mineral density in adults, with transcriptomic upregulation of collagen biosynthesis genes and downregulation of Fgf8 signaling, placing FBLN1 as a negative regulator of skeletal mineralization upstream of Fgf8 signaling.","method":"CRISPR/Cas9-generated fbln1 zebrafish mutants, micro-CT skeletal analysis, bulk RNA-seq transcriptomics of mutant vs. wild-type","journal":"Biomolecules","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic knockout with quantitative skeletal phenotype and transcriptomic pathway placement; single lab","pmids":["38397376"],"is_preprint":false}],"current_model":"FBLN1 is a secreted extracellular matrix glycoprotein with splice-variant-specific ECM incorporation; it is essential for vascular integrity (Fbln1 null lethality), promotes chondrocyte proliferation and osteogenic differentiation via Smad2 phosphorylation, antagonizes TGF-β/Smad signaling to reduce ferroptosis, and negatively regulates skeletal mineralization upstream of Fgf8; its mRNA stability is post-transcriptionally controlled by YTHDF2-mediated m6A reading and miR-615-3p, and missense variants reduce protein stability and synergize with ARHGAP31 mutations to impair Cdc42 and MAPK/ERK signaling."},"narrative":{"mechanistic_narrative":"FBLN1 is a secreted extracellular matrix glycoprotein essential for vascular wall integrity, with developmental and skeletal regulatory roles that operate through TGF-β/Smad and Fgf8 signaling [PMID:16338003, PMID:33610427, PMID:38397376]. During limb development it is deposited in digit-associated ECM, and its splice variants differ in matrix incorporation, with the FBLN1-D variant showing altered ECM assembly in synpolydactyly patient fibroblasts [PMID:11836357]. Genetic deletion in mice is lethal and identifies FBLN1 as a structural component of blood vessel walls [PMID:16338003]. FBLN1 acts as a context-dependent modulator of TGF-β/Smad signaling: it promotes chondrocyte proliferation and Col2 upregulation by increasing Smad2 phosphorylation [PMID:33610427], yet in injured lung epithelium it suppresses TGF-β/Smad activity to limit ferroptosis [PMID:39671383], indicating its signaling output depends on cellular context. In skeletal tissue FBLN1 is a negative regulator of mineralization, acting upstream of Fgf8 signaling, as zebrafish mutants show increased mineralization and collagen biosynthesis gene upregulation [PMID:38397376]. FBLN1 expression is post-transcriptionally constrained: m6A methylation in its 3'UTR is read by YTHDF2 to destabilize the transcript and, together with miR-615-3p, suppress osteogenic differentiation [PMID:38353178]. A destabilizing missense variant (p.Arg550His) reduces FBLN1 protein levels and, in combination with an ARHGAP31 variant, impairs Cdc42 activity and activates MAPK/ERK signaling [PMID:36176297].","teleology":[{"year":2002,"claim":"Established that FBLN1 is incorporated into developing limb ECM in a splice-variant-specific manner, framing distinct functional roles for the C and D isoforms in matrix assembly.","evidence":"Immunolocalization in developing limb ECM and biochemical fractionation of FBLN1-C/D variants from synpolydactyly patient fibroblasts and conditioned medium","pmids":["11836357"],"confidence":"Medium","gaps":["Molecular basis for differential variant incorporation not defined","No reconstitution of ECM assembly","Link between altered incorporation and synpolydactyly phenotype indirect"]},{"year":2005,"claim":"Demonstrated that FBLN1 is genetically essential for blood vessel wall integrity and defined its placental expression, resolving whether FBLN1 drives placental hyperplasia (it does not).","evidence":"Homozygous knockout mouse with lethal vascular phenotype, in situ expression, and a non-rescuing placental hyperplasia model","pmids":["16338003"],"confidence":"Medium","gaps":["Molecular mechanism of vascular wall defect unresolved","Lethality precludes tissue-specific functional dissection","Negative placental result leaves the hyperplasia cause unidentified"]},{"year":2021,"claim":"Connected FBLN1 to active Smad2 signaling, showing it promotes chondrocyte proliferation rather than acting purely as a structural matrix protein.","evidence":"Adenoviral FBLN1 overexpression in chondrocytes with EdU proliferation, pSmad2/Col2 Western blotting, and pSmad2 inhibitor rescue","pmids":["33610427"],"confidence":"Medium","gaps":["Mechanism linking secreted FBLN1 to intracellular Smad2 phosphorylation unknown","No direct receptor or binding partner identified","Single overexpression system"]},{"year":2022,"claim":"Showed a missense variant destabilizes FBLN1 protein and synergizes with ARHGAP31 mutation to disrupt Cdc42 and MAPK/ERK signaling, linking FBLN1 dosage to intracellular signaling defects.","evidence":"Wild-type/mutant FBLN1 transfection with stability immunoblots, viability/apoptosis assays, Cdc42 activity assay, and ARHGAP31 co-transfection","pmids":["36176297"],"confidence":"Medium","gaps":["Effects only manifest in combination with ARHGAP31 variant","Mechanism connecting extracellular FBLN1 to Cdc42 not defined","Single in vitro lab without in vivo confirmation"]},{"year":2024,"claim":"Identified post-transcriptional control of FBLN1 by m6A/YTHDF2 and miR-615-3p, explaining how FBLN1 levels are tuned to govern osteogenic differentiation.","evidence":"m6A site mutation, YTHDF2 gain/loss, miR-615-3p manipulation, mRNA stability and osteogenic assays in WJCMSCs, plus in vivo bone regeneration","pmids":["38353178"],"confidence":"Medium","gaps":["m6A writer/eraser controlling the site not identified","Whether this regulation operates in vascular or skeletal tissues unknown","Single lab"]},{"year":2024,"claim":"Defined a context-dependent role in which FBLN1 antagonizes TGF-β/Smad signaling to limit ferroptosis, contrasting with its Smad2-promoting role in chondrocytes.","evidence":"FBLN1 and TGF-β1 overexpression in LPS-treated MLE-12 lung epithelial cells with TGF-β/Smad and ferroptosis marker readouts","pmids":["39671383"],"confidence":"Medium","gaps":["Direct biochemical interaction with TGF-β/Smad not demonstrated","Reconciliation with Smad2-promoting role in cartilage unresolved","In vitro overexpression only"]},{"year":2024,"claim":"Placed FBLN1 as a negative regulator of skeletal mineralization upstream of Fgf8, establishing an in vivo developmental signaling axis.","evidence":"CRISPR/Cas9 fbln1 zebrafish mutants with micro-CT skeletal phenotyping and bulk RNA-seq pathway analysis","pmids":["38397376"],"confidence":"Medium","gaps":["Direct mechanism linking FBLN1 to Fgf8 downregulation not defined","Mineralization phenotype not yet linked to human skeletal disease","Single model organism"]},{"year":null,"claim":"How FBLN1 produces opposite effects on Smad signaling in different tissues, and what receptors or matrix partners transduce its secreted signal, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No defined receptor or signaling co-factor for FBLN1","Context determinants of Smad2 promotion vs TGF-β/Smad antagonism unknown","No structural basis for variant-driven instability or splice-variant ECM behavior"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,1]}],"localization":[{"term_id":"GO:0031012","term_label":"extracellular matrix","supporting_discovery_ids":[0]},{"term_id":"GO:0005576","term_label":"extracellular region","supporting_discovery_ids":[0,1]}],"pathway":[{"term_id":"R-HSA-1474244","term_label":"Extracellular matrix organization","supporting_discovery_ids":[0]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[1,6]}],"complexes":[],"partners":[],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P23142","full_name":"Fibulin-1","aliases":[],"length_aa":703,"mass_kda":77.2,"function":"Incorporated into fibronectin-containing matrix fibers. May play a role in cell adhesion and migration along protein fibers within the extracellular matrix (ECM). Could be important for certain developmental processes and contribute to the supramolecular organization of ECM architecture, in particular to those of basement membranes. Has been implicated in a role in cellular transformation and tumor invasion, it appears to be a tumor suppressor. May play a role in haemostasis and thrombosis owing to its ability to bind fibrinogen and incorporate into clots. Could play a significant role in modulating the neurotrophic activities of APP, particularly soluble APP","subcellular_location":"Secreted, extracellular space, extracellular matrix","url":"https://www.uniprot.org/uniprotkb/P23142/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/FBLN1","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":[{"gene":"CANX","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/FBLN1","total_profiled":1310},"omim":[{"mim_id":"621107","title":"HEMICENTIN 2; HMCN2","url":"https://www.omim.org/entry/621107"},{"mim_id":"608548","title":"HEMICENTIN; HMCN1","url":"https://www.omim.org/entry/608548"},{"mim_id":"608231","title":"RAS ASSOCIATION DOMAIN FAMILY, MEMBER 8; RASSF8","url":"https://www.omim.org/entry/608231"},{"mim_id":"608180","title":"SYNPOLYDACTYLY 2; SPD2","url":"https://www.omim.org/entry/608180"},{"mim_id":"604633","title":"EGF-CONTAINING FIBULIN-LIKE EXTRACELLULAR MATRIX PROTEIN 2; EFEMP2","url":"https://www.omim.org/entry/604633"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Endoplasmic reticulum","reliability":"Approved"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"choroid plexus","ntpm":1775.4}],"url":"https://www.proteinatlas.org/search/FBLN1"},"hgnc":{"alias_symbol":["FBLN"],"prev_symbol":[]},"alphafold":{"accession":"P23142","domains":[{"cath_id":"-","chopping":"38-142","consensus_level":"high","plddt":79.6047,"start":38,"end":142},{"cath_id":"2.10.25.10","chopping":"346-400","consensus_level":"medium","plddt":84.664,"start":346,"end":400},{"cath_id":"2.10.25.10","chopping":"406-443","consensus_level":"medium","plddt":92.3108,"start":406,"end":443},{"cath_id":"2.10.25.10","chopping":"446-482","consensus_level":"medium","plddt":92.6114,"start":446,"end":482},{"cath_id":"2.10.25.10","chopping":"491-524","consensus_level":"medium","plddt":89.0462,"start":491,"end":524},{"cath_id":"2.60.40.60","chopping":"535-699","consensus_level":"high","plddt":83.3285,"start":535,"end":699}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P23142","model_url":"https://alphafold.ebi.ac.uk/files/AF-P23142-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P23142-F1-predicted_aligned_error_v6.png","plddt_mean":79.06},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=FBLN1","jax_strain_url":"https://www.jax.org/strain/search?query=FBLN1"},"sequence":{"accession":"P23142","fasta_url":"https://rest.uniprot.org/uniprotkb/P23142.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P23142/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P23142"}},"corpus_meta":[{"pmid":"11836357","id":"PMC_11836357","title":"The fibulin-1 gene (FBLN1) is disrupted in a t(12;22) associated with a complex type of synpolydactyly.","date":"2002","source":"Journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/11836357","citation_count":87,"is_preprint":false},{"pmid":"19913326","id":"PMC_19913326","title":"Aberrant promoter methylation of FBLN-3 gene and clinicopathological significance in non-small cell lung carcinoma.","date":"2009","source":"Lung cancer (Amsterdam, Netherlands)","url":"https://pubmed.ncbi.nlm.nih.gov/19913326","citation_count":26,"is_preprint":false},{"pmid":"20500888","id":"PMC_20500888","title":"A genomics-informed, SNP association study reveals FBLN1 and FABP4 as contributing to resistance to fleece rot in Australian Merino sheep.","date":"2010","source":"BMC veterinary research","url":"https://pubmed.ncbi.nlm.nih.gov/20500888","citation_count":20,"is_preprint":false},{"pmid":"29715435","id":"PMC_29715435","title":"Serum FBLN1 and STK31 as biomarkers of colorectal cancer and their ability to noninvasively differentiate colorectal cancer from benign polyps.","date":"2018","source":"Clinica chimica acta; international journal of clinical chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/29715435","citation_count":19,"is_preprint":false},{"pmid":"33602229","id":"PMC_33602229","title":"Discovery and validation of FBLN1 and ANT3 as potential biomarkers for early detection of cervical cancer.","date":"2021","source":"Cancer cell international","url":"https://pubmed.ncbi.nlm.nih.gov/33602229","citation_count":14,"is_preprint":false},{"pmid":"16338003","id":"PMC_16338003","title":"Expression and functional analysis of fibulin-1 (Fbln1) during normal and abnormal placental development of the mouse.","date":"2005","source":"Placenta","url":"https://pubmed.ncbi.nlm.nih.gov/16338003","citation_count":13,"is_preprint":false},{"pmid":"38353178","id":"PMC_38353178","title":"miR615-3p inhibited FBLN1 and osteogenic differentiation of umbilical cord mesenchymal stem cells by associated with YTHDF2 in a m6A-miRNA interaction manner.","date":"2024","source":"Cell proliferation","url":"https://pubmed.ncbi.nlm.nih.gov/38353178","citation_count":13,"is_preprint":false},{"pmid":"7842734","id":"PMC_7842734","title":"Localization of the human gene for fibulin-1 (FBLN1) to chromosome band 22q13.3.","date":"1995","source":"Cytogenetics and cell genetics","url":"https://pubmed.ncbi.nlm.nih.gov/7842734","citation_count":13,"is_preprint":false},{"pmid":"34424449","id":"PMC_34424449","title":"Circ_FBLN1 promotes the proliferation and osteogenic differentiation of human bone marrow-derived mesenchymal stem cells by regulating let-7i-5p/FZD4 axis and Wnt/β-catenin pathway.","date":"2021","source":"Journal of bioenergetics and biomembranes","url":"https://pubmed.ncbi.nlm.nih.gov/34424449","citation_count":11,"is_preprint":false},{"pmid":"23907575","id":"PMC_23907575","title":"Abnormal hypermethylation and clinicopathological significance of FBLN1 gene in cutaneous melanoma.","date":"2013","source":"Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine","url":"https://pubmed.ncbi.nlm.nih.gov/23907575","citation_count":10,"is_preprint":false},{"pmid":"7806231","id":"PMC_7806231","title":"The fibulin-1 gene (FBLN1) is located on human chromosome 22 and on mouse chromosome 15.","date":"1994","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/7806231","citation_count":8,"is_preprint":false},{"pmid":"38397376","id":"PMC_38397376","title":"The Osteoblast Transcriptome in Developing Zebrafish Reveals Key Roles for Extracellular Matrix Proteins Col10a1a and Fbln1 in Skeletal Development and Homeostasis.","date":"2024","source":"Biomolecules","url":"https://pubmed.ncbi.nlm.nih.gov/38397376","citation_count":5,"is_preprint":false},{"pmid":"33610427","id":"PMC_33610427","title":"FBLN1 promotes chondrocyte proliferation by increasing phosphorylation of Smad2.","date":"2021","source":"Journal of orthopaedic science : official journal of the Japanese Orthopaedic Association","url":"https://pubmed.ncbi.nlm.nih.gov/33610427","citation_count":5,"is_preprint":false},{"pmid":"39671383","id":"PMC_39671383","title":"FBLN1 regulates ferroptosis in acute respiratory distress syndrome by reducing free ferrous iron by inhibiting the TGF-β/Smad pathway.","date":"2024","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/39671383","citation_count":5,"is_preprint":false},{"pmid":"39201719","id":"PMC_39201719","title":"A Bioinformatics-Based Study on Methylation Alterations of the FBLN1 Gene in Hippocampal Tissue of Alzheimer's Disease Model DKO and DTG Mice.","date":"2024","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/39201719","citation_count":2,"is_preprint":false},{"pmid":"36176297","id":"PMC_36176297","title":"Synergistic effects of rare variants of ARHGAP31 and FBLN1 in vitro in terminal transverse limb defects.","date":"2022","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/36176297","citation_count":2,"is_preprint":false},{"pmid":"42250865","id":"PMC_42250865","title":"Integrated multi-omics and experimental validation reveal CSK and FBLN1 as key targets of stanozolol in accelerating atherosclerosis.","date":"2026","source":"Vascular pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/42250865","citation_count":0,"is_preprint":false},{"pmid":"41913707","id":"PMC_41913707","title":"Human Papillomavirus (HPV) Infected Epithelial Cells and FBLN1+ Fibroreticular Cells Govern the Immunogenic Tumor Microenvironment in HPV-Associated Oropharyngeal Squamous Cell Carcinoma.","date":"2026","source":"Clinical and experimental otorhinolaryngology","url":"https://pubmed.ncbi.nlm.nih.gov/41913707","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.05.23.655833","title":"Extracellular Matrix Alterations in Chronic Ischemic Cardiomyopathy Revealed by Quantitative Proteomics","date":"2025-05-28","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.23.655833","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":11685,"output_tokens":2049,"usd":0.032895,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9034,"output_tokens":2836,"usd":0.058035,"stage2_stop_reason":"end_turn"},"total_usd":0.09093,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2002,\n      \"finding\": \"FBLN1 protein is expressed in the extracellular matrix in association with digits during developing limb formation; the FBLN1-D splice variant, but not the FBLN1-C splice variant, shows altered incorporation into ECM and conditioned medium in fibroblasts from patients with synpolydactyly, suggesting splice-variant-specific ECM roles.\",\n      \"method\": \"Immunolocalization of FBLN1 in developing limb ECM; analysis of FBLN1-C and FBLN1-D splice variant levels in patient fibroblast ECM and conditioned medium\",\n      \"journal\": \"Journal of medical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct biochemical fractionation of ECM from patient fibroblasts with two splice variants distinguished; single lab, no reconstitution\",\n      \"pmids\": [\"11836357\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Fbln1 null mice are lethal, establishing FBLN1 as an essential component of blood vessel walls; Fbln1 is expressed in spongiotrophoblast, endothelia of large fetal blood vessels, and a subset of giant cells in the mouse placenta, but Fbln1 alone is not the key cause of placental hyperplasia phenotypes in a congenic interspecies hybrid model.\",\n      \"method\": \"Homozygous targeted gene deletion (knockout mouse), in situ expression analysis, rescue experiment attempting to normalize Fbln1 expression in hyperplastic placentas\",\n      \"journal\": \"Placenta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic knockout with defined lethal phenotype and placental expression pattern; rescue experiment showed no rescue (negative result for placental role); single lab\",\n      \"pmids\": [\"16338003\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"FBLN1 promotes chondrocyte proliferation in knee cartilage by increasing phosphorylation of Smad2 and upregulating Col2; inhibition of pSmad2 abolishes the FBLN1-driven proliferation effect.\",\n      \"method\": \"Adenoviral overexpression of FBLN1 in isolated chondrocytes, EdU proliferation assay, Western blotting for pSmad2 and Col2, pharmacological inhibition of pSmad2\",\n      \"journal\": \"Journal of orthopaedic science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — overexpression with defined signaling readout and inhibitor rescue; single lab, single method per endpoint\",\n      \"pmids\": [\"33610427\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"FBLN1 overexpression inhibits the TGF-β/Smad signaling pathway (decreased TGF-β and p-Smad protein levels) and thereby reduces ferroptosis markers (MDA, lipid ROS, Fe2+) in LPS-treated lung epithelial cells; FBLN1 and TGF-β have antagonistic roles in regulating ferroptosis.\",\n      \"method\": \"Overexpression of FBLN1 and TGF-β1 in MLE-12 cells treated with LPS, Western blotting for TGF-β/Smad pathway proteins and ferroptosis markers, cell viability assay\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — in vitro overexpression with multiple molecular readouts but single lab, no orthogonal validation of TGF-β/Smad interaction\",\n      \"pmids\": [\"39671383\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"FBLN1 mRNA stability is regulated by m6A methylation at a site in its 3'UTR; YTHDF2 binds this m6A site and induces FBLN1 mRNA instability, reducing FBLN1 expression and suppressing osteogenic differentiation of WJCMSCs. Mutation of the m6A site enhances FBLN1 mRNA stability and osteogenic differentiation. miR-615-3p also negatively regulates FBLN1 by binding its 3'UTR, and YTHDF2 facilitates miR-615-3p-mediated FBLN1 mRNA decay.\",\n      \"method\": \"m6A site mutation, YTHDF2 overexpression/knockdown, miR-615-3p gain/loss of function, mRNA stability assays, osteogenic differentiation assays in WJCMSCs, bone regeneration in vivo model\",\n      \"journal\": \"Cell proliferation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal molecular methods (m6A mutation, RBP binding, miRNA target) in single lab; functional consequence validated in vitro and in vivo\",\n      \"pmids\": [\"38353178\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"A missense variant of FBLN1 (p.Arg550His) results in reduced protein stability and decreased FBLN1 expression. In combination with an ARHGAP31 variant, but not alone, FBLN1 mutation decreases cell viability, impairs proliferation, activates apoptosis, reduces Cdc42 activity, and activates the MAPK/ERK pathway in vitro, indicating synergistic disruption of cellular signaling.\",\n      \"method\": \"Expression vectors with wild-type and mutant FBLN1 transfected in mammalian cells; immunoblotting for protein stability and expression; cell viability (CCK-8), immunofluorescence, co-transfection with ARHGAP31 mutant; Cdc42 activity assay; Western blotting for MAPK/ERK pathway\",\n      \"journal\": \"Frontiers in genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — multiple functional assays with mutant protein in vitro; single lab; synergistic effect context-dependent\",\n      \"pmids\": [\"36176297\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"fbln1 mutant zebrafish show increased mineralization of cranial elements, altered ceratohyal angle, increased vertebral dimensions and tissue mineral density in adults, with transcriptomic upregulation of collagen biosynthesis genes and downregulation of Fgf8 signaling, placing FBLN1 as a negative regulator of skeletal mineralization upstream of Fgf8 signaling.\",\n      \"method\": \"CRISPR/Cas9-generated fbln1 zebrafish mutants, micro-CT skeletal analysis, bulk RNA-seq transcriptomics of mutant vs. wild-type\",\n      \"journal\": \"Biomolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic knockout with quantitative skeletal phenotype and transcriptomic pathway placement; single lab\",\n      \"pmids\": [\"38397376\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"FBLN1 is a secreted extracellular matrix glycoprotein with splice-variant-specific ECM incorporation; it is essential for vascular integrity (Fbln1 null lethality), promotes chondrocyte proliferation and osteogenic differentiation via Smad2 phosphorylation, antagonizes TGF-β/Smad signaling to reduce ferroptosis, and negatively regulates skeletal mineralization upstream of Fgf8; its mRNA stability is post-transcriptionally controlled by YTHDF2-mediated m6A reading and miR-615-3p, and missense variants reduce protein stability and synergize with ARHGAP31 mutations to impair Cdc42 and MAPK/ERK signaling.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"FBLN1 is a secreted extracellular matrix glycoprotein essential for vascular wall integrity, with developmental and skeletal regulatory roles that operate through TGF-β/Smad and Fgf8 signaling [#1, #2, #6]. During limb development it is deposited in digit-associated ECM, and its splice variants differ in matrix incorporation, with the FBLN1-D variant showing altered ECM assembly in synpolydactyly patient fibroblasts [#0]. Genetic deletion in mice is lethal and identifies FBLN1 as a structural component of blood vessel walls [#1]. FBLN1 acts as a context-dependent modulator of TGF-β/Smad signaling: it promotes chondrocyte proliferation and Col2 upregulation by increasing Smad2 phosphorylation [#2], yet in injured lung epithelium it suppresses TGF-β/Smad activity to limit ferroptosis [#3], indicating its signaling output depends on cellular context. In skeletal tissue FBLN1 is a negative regulator of mineralization, acting upstream of Fgf8 signaling, as zebrafish mutants show increased mineralization and collagen biosynthesis gene upregulation [#6]. FBLN1 expression is post-transcriptionally constrained: m6A methylation in its 3'UTR is read by YTHDF2 to destabilize the transcript and, together with miR-615-3p, suppress osteogenic differentiation [#4]. A destabilizing missense variant (p.Arg550His) reduces FBLN1 protein levels and, in combination with an ARHGAP31 variant, impairs Cdc42 activity and activates MAPK/ERK signaling [#5].\",\n  \"teleology\": [\n    {\n      \"year\": 2002,\n      \"claim\": \"Established that FBLN1 is incorporated into developing limb ECM in a splice-variant-specific manner, framing distinct functional roles for the C and D isoforms in matrix assembly.\",\n      \"evidence\": \"Immunolocalization in developing limb ECM and biochemical fractionation of FBLN1-C/D variants from synpolydactyly patient fibroblasts and conditioned medium\",\n      \"pmids\": [\"11836357\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis for differential variant incorporation not defined\", \"No reconstitution of ECM assembly\", \"Link between altered incorporation and synpolydactyly phenotype indirect\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Demonstrated that FBLN1 is genetically essential for blood vessel wall integrity and defined its placental expression, resolving whether FBLN1 drives placental hyperplasia (it does not).\",\n      \"evidence\": \"Homozygous knockout mouse with lethal vascular phenotype, in situ expression, and a non-rescuing placental hyperplasia model\",\n      \"pmids\": [\"16338003\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular mechanism of vascular wall defect unresolved\", \"Lethality precludes tissue-specific functional dissection\", \"Negative placental result leaves the hyperplasia cause unidentified\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Connected FBLN1 to active Smad2 signaling, showing it promotes chondrocyte proliferation rather than acting purely as a structural matrix protein.\",\n      \"evidence\": \"Adenoviral FBLN1 overexpression in chondrocytes with EdU proliferation, pSmad2/Col2 Western blotting, and pSmad2 inhibitor rescue\",\n      \"pmids\": [\"33610427\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking secreted FBLN1 to intracellular Smad2 phosphorylation unknown\", \"No direct receptor or binding partner identified\", \"Single overexpression system\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Showed a missense variant destabilizes FBLN1 protein and synergizes with ARHGAP31 mutation to disrupt Cdc42 and MAPK/ERK signaling, linking FBLN1 dosage to intracellular signaling defects.\",\n      \"evidence\": \"Wild-type/mutant FBLN1 transfection with stability immunoblots, viability/apoptosis assays, Cdc42 activity assay, and ARHGAP31 co-transfection\",\n      \"pmids\": [\"36176297\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Effects only manifest in combination with ARHGAP31 variant\", \"Mechanism connecting extracellular FBLN1 to Cdc42 not defined\", \"Single in vitro lab without in vivo confirmation\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified post-transcriptional control of FBLN1 by m6A/YTHDF2 and miR-615-3p, explaining how FBLN1 levels are tuned to govern osteogenic differentiation.\",\n      \"evidence\": \"m6A site mutation, YTHDF2 gain/loss, miR-615-3p manipulation, mRNA stability and osteogenic assays in WJCMSCs, plus in vivo bone regeneration\",\n      \"pmids\": [\"38353178\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"m6A writer/eraser controlling the site not identified\", \"Whether this regulation operates in vascular or skeletal tissues unknown\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined a context-dependent role in which FBLN1 antagonizes TGF-β/Smad signaling to limit ferroptosis, contrasting with its Smad2-promoting role in chondrocytes.\",\n      \"evidence\": \"FBLN1 and TGF-β1 overexpression in LPS-treated MLE-12 lung epithelial cells with TGF-β/Smad and ferroptosis marker readouts\",\n      \"pmids\": [\"39671383\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct biochemical interaction with TGF-β/Smad not demonstrated\", \"Reconciliation with Smad2-promoting role in cartilage unresolved\", \"In vitro overexpression only\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Placed FBLN1 as a negative regulator of skeletal mineralization upstream of Fgf8, establishing an in vivo developmental signaling axis.\",\n      \"evidence\": \"CRISPR/Cas9 fbln1 zebrafish mutants with micro-CT skeletal phenotyping and bulk RNA-seq pathway analysis\",\n      \"pmids\": [\"38397376\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct mechanism linking FBLN1 to Fgf8 downregulation not defined\", \"Mineralization phenotype not yet linked to human skeletal disease\", \"Single model organism\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How FBLN1 produces opposite effects on Smad signaling in different tissues, and what receptors or matrix partners transduce its secreted signal, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No defined receptor or signaling co-factor for FBLN1\", \"Context determinants of Smad2 promotion vs TGF-β/Smad antagonism unknown\", \"No structural basis for variant-driven instability or splice-variant ECM behavior\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0031012\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005576\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1474244\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [1, 6]}\n    ],\n    \"complexes\": [],\n    \"partners\": [],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}