{"gene":"CALD1","run_date":"2026-06-09T22:57:17","timeline":{"discoveries":[{"year":2007,"finding":"The low molecular weight caldesmon isoform (Hela l-CaD) localizes to motility-related cell protrusions (filopodia, microspikes, lamellipodia, podosomes, membrane blebs, and membrane ruffles) in endothelial cells/endothelial progenitor cells in tumor vasculature, implicating it in cell migration during angiogenesis and vasculogenesis.","method":"Immunohistochemistry and histological analysis of motility-related protrusions in histologically preserved tumor microenvironment sections","journal":"Cell adhesion & migration","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single method (localization by IHC), no functional rescue or in vitro motility assay","pmids":["19329885"],"is_preprint":false},{"year":2022,"finding":"AHSA1 recruits ERK1/2 to phosphorylate and inactivate CALD1; pharmacological inhibition of ERK1/2 phosphorylation (with SCH772984) reversed AHSA1-promoted proliferation and EMT, and CALD1 knockdown rescued the inhibition of proliferation and EMT caused by AHSA1 knockdown, placing CALD1 downstream of the ERK1/2 axis.","method":"Gain- and loss-of-function studies in vitro and in vivo, ERK1/2 phosphorylation inhibitor treatment, Western blot for phosphorylated CALD1, siRNA knockdown rescue experiments","journal":"Cancers","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, multiple orthogonal methods (KO/KD, pharmacological inhibition, rescue experiment, in vivo validation)","pmids":["36230524"],"is_preprint":false},{"year":2021,"finding":"CALD1 promotes PD-L1 (CD274) expression in bladder cancer via activation of the JAK/STAT signaling pathway; this effect was blocked by the specific JAK inhibitor Ruxolitinib.","method":"CALD1 overexpression/knockdown in bladder cancer cells, GSEA pathway enrichment, pharmacological inhibition with Ruxolitinib, Western blot, flow cytometry","journal":"Annals of translational medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, multiple methods (OE/KD, pharmacological rescue, pathway inhibitor), consistent with GSEA","pmids":["34733993"],"is_preprint":false},{"year":2023,"finding":"miR-1278 directly targets and negatively regulates CALD1 expression (confirmed by dual luciferase reporter assay); CALD1 overexpression activates the MAPK pathway (upregulating Ras, p-P38, p-ERK1/2), promoting gastric cancer cell viability and migration, which is partially rescued by miR-1278 mimic.","method":"Dual luciferase reporter assay, CCK-8, Transwell migration assay, Western blot for MAPK pathway proteins, in vivo xenograft","journal":"Open medicine (Warsaw, Poland)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, direct binding confirmed by luciferase assay, pathway proteins measured, rescue experiment performed","pmids":["38025524"],"is_preprint":false},{"year":2024,"finding":"CALD1 promotes EMT in gastric cancer by activating the PI3K-Akt pathway: CALD1 overexpression increased PI3K, p-AKT, and p-mTOR while decreasing PTEN; PI3K-Akt inhibitor treatment reversed CALD1-mediated effects on proliferation, migration, invasion, and EMT marker expression.","method":"siRNA knockdown and overexpression in AGS and MKN45 cells, CCK-8, wound healing and Transwell assays, Western blot, PI3K-Akt inhibitor treatment, in vivo xenograft","journal":"World journal of gastrointestinal oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, multiple orthogonal methods (KD/OE, pharmacological inhibition, in vivo validation)","pmids":["38577446"],"is_preprint":false},{"year":2023,"finding":"METTL14 methyltransferase increases CALD1 mRNA and m6A methylation levels in oral squamous cell carcinoma; METTL14 silencing depleted CALD1 mRNA and m6A levels, suppressing cell growth and metastasis, which was rescued by CALD1 overexpression.","method":"MeRIP assay for m6A levels, qRT-PCR, Western blot, colony formation and Transwell assays, siRNA knockdown, in vivo tumorigenicity assay","journal":"Journal of environmental pathology, toxicology and oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, MeRIP directly measures m6A on CALD1, rescue experiment confirms pathway placement","pmids":["37017680"],"is_preprint":false},{"year":2025,"finding":"CALD1 knockdown in SK-OV-3 ovarian cancer cells reduces F-actin stress fibers, loosens cytoskeletal structure, decreases Vinculin expression and focal adhesion number, and enhances cell invasiveness, demonstrating that CALD1 restrains invasion by maintaining cytoskeletal organization and focal adhesion formation.","method":"Stable CALD1 knockdown by shRNA, Transwell invasion assay, immunofluorescence staining of F-actin and Vinculin, Western blot","journal":"Translational cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, direct functional readout (invasion + cytoskeletal imaging), loss-of-function with specific phenotypic readout","pmids":["40104711"],"is_preprint":false},{"year":2024,"finding":"ET-1 treatment increases high-molecular-weight CALD1 expression (a marker of the contractile smooth muscle cell phenotype) in placental vessel explants and HUVSMCs via ETAR/ETBR receptors; elevated CALD1 is associated with stronger contraction in preeclampsia placental chorionic plate veins.","method":"DMT vessel tone measurement, whole-cell patch clamp, Western blot, ELISA, PCR, ET-1 treatment with receptor antagonists in placental explants and HUVSMCs","journal":"Placenta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, pharmacological receptor blockade identifies ETAR/ETBR as writers of CALD1 upregulation, multiple orthogonal methods","pmids":["39476475"],"is_preprint":false},{"year":2025,"finding":"In Helicobacter pylori-positive gastric cancer, TLR signaling in cancer-associated fibroblasts suppresses miR-148a-5p, which in turn de-represses CALD1; elevated CAF CALD1 promotes collagen VI secretion that interacts with tumoral SDC4 receptors to drive GC cell proliferation.","method":"microRNA and transcriptome sequencing, single-cell sequencing, in vitro co-culture, CDX and PDX in vivo models, miR-148a-5p agomir treatment","journal":"Cellular oncology (Dordrecht, Netherlands)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, multiple orthogonal methods (in vitro and in vivo models, sequencing, functional rescue with agomir)","pmids":["41171370"],"is_preprint":false}],"current_model":"CALD1 (caldesmon) is a cytoskeleton-associated actin-binding protein whose high-molecular-weight isoform stabilizes the contractile smooth muscle phenotype (regulated by ET-1 via ETAR/ETBR) and whose non-muscle isoform localizes to actin-rich cell protrusions to support cell migration; CALD1 activity is post-translationally regulated by ERK1/2-mediated phosphorylation (downstream of AHSA1), transcriptionally regulated by METTL14-dependent m6A methylation and by miR-1278/miR-148a-5p targeting, and it signals through the JAK/STAT, MAPK, and PI3K-Akt pathways to promote proliferation, EMT, and immune evasion in multiple cancer types, while in ovarian cancer it acts as a tumor suppressor by maintaining F-actin stress fibers, focal adhesions, and Vinculin expression to restrain invasion."},"narrative":{"mechanistic_narrative":"CALD1 (caldesmon) is an actin-associated cytoskeletal protein whose isoforms govern smooth muscle contractility and cell motility, and which is repeatedly co-opted in cancer to control proliferation, EMT, and invasion [PMID:40104711, PMID:39476475]. Its high-molecular-weight isoform marks the contractile smooth muscle phenotype and is upregulated by endothelin-1 acting through ETAR/ETBR receptors, with elevated CALD1 linked to stronger vascular contraction [PMID:39476475], while the low-molecular-weight isoform localizes to motility-related protrusions such as filopodia, lamellipodia, and podosomes consistent with a role in cell migration [PMID:19329885]. In ovarian cancer CALD1 acts to restrain invasion: its loss reduces F-actin stress fibers, loosens cytoskeletal organization, and decreases Vinculin expression and focal adhesion number [PMID:40104711]. In several other cancers CALD1 instead promotes malignant behavior through downstream signaling, activating the MAPK pathway (Ras, p-P38, p-ERK1/2) [PMID:38025524] and the PI3K-Akt-mTOR axis with concomitant PTEN loss to drive proliferation and EMT [PMID:38577446], and activating JAK/STAT signaling to upregulate PD-L1 (CD274) [PMID:34733993]. CALD1 is post-translationally inactivated by ERK1/2-mediated phosphorylation recruited via AHSA1 [PMID:36230524], and its expression is controlled by METTL14-dependent m6A methylation [PMID:37017680] and by negative regulation through miR-1278 [PMID:38025524] and miR-148a-5p [PMID:41171370]; in H. pylori-positive gastric cancer, de-repressed CALD1 in cancer-associated fibroblasts drives collagen VI secretion that engages tumoral SDC4 to promote tumor growth [PMID:41171370].","teleology":[{"year":2007,"claim":"Established where the low-molecular-weight caldesmon isoform acts in cells, linking it to the migratory machinery rather than only contraction.","evidence":"Immunohistochemistry of motility-related protrusions in tumor vasculature sections","pmids":["19329885"],"confidence":"Low","gaps":["Localization only — no functional motility assay or rescue","Does not establish causal role in migration","Isoform-specific function not dissected mechanistically"]},{"year":2021,"claim":"Connected CALD1 to immune evasion by showing it drives PD-L1 expression through a defined kinase pathway.","evidence":"Overexpression/knockdown in bladder cancer cells with Ruxolitinib JAK inhibition, Western blot, flow cytometry","pmids":["34733993"],"confidence":"Medium","gaps":["Mechanism linking CALD1 to JAK/STAT activation unresolved","Single cancer context","No direct biochemical interaction shown"]},{"year":2022,"claim":"Defined how CALD1 activity is post-translationally controlled, placing it downstream of an AHSA1–ERK1/2 axis that phosphorylates and inactivates it.","evidence":"Gain/loss-of-function in vitro and in vivo, SCH772984 ERK inhibition, Western blot for phospho-CALD1, knockdown rescue","pmids":["36230524"],"confidence":"Medium","gaps":["Phosphosites on CALD1 not mapped","Direct AHSA1–CALD1 vs indirect recruitment not distinguished","Functional consequence of inactivation on actin binding not measured"]},{"year":2023,"claim":"Identified upstream transcriptional/epigenetic regulators of CALD1 (m6A and microRNA) and a downstream MAPK effector pathway.","evidence":"MeRIP/qRT-PCR with METTL14 silencing and rescue (OSCC); dual luciferase miR-1278 binding, CCK-8/Transwell, MAPK Western blots (gastric)","pmids":["37017680","38025524"],"confidence":"Medium","gaps":["m6A reader mediating CALD1 mRNA fate not identified","How CALD1 protein activates MAPK is not mechanistically defined","Single-lab findings per cancer type"]},{"year":2024,"claim":"Extended CALD1's pro-tumor signaling to the PI3K-Akt-mTOR axis driving EMT.","evidence":"siRNA/overexpression in AGS and MKN45 cells, pharmacological PI3K-Akt inhibition, Western blot, in vivo xenograft","pmids":["38577446"],"confidence":"Medium","gaps":["Link between CALD1 and PI3K activation not biochemically defined","Relationship to the MAPK and JAK/STAT effects in other studies unclear"]},{"year":2025,"claim":"Revealed a context-dependent tumor-suppressive role (ovarian) via cytoskeletal/focal adhesion maintenance, and a stromal CAF-driven pro-tumor axis (gastric).","evidence":"shRNA knockdown with invasion assay and F-actin/Vinculin immunofluorescence (ovarian); miR/transcriptome and single-cell sequencing, co-culture, CDX/PDX, miR-148a-5p agomir (gastric)","pmids":["40104711","41171370"],"confidence":"Medium","gaps":["Molecular basis for opposing tumor-suppressor vs oncogenic roles unresolved","Direct CALD1 role in collagen VI secretion vs correlative not established","Cell-type specificity of CALD1 function not reconciled"]},{"year":null,"claim":"It remains unresolved how CALD1's core actin-binding/cytoskeletal function mechanistically dictates its opposing tumor-suppressive versus oncogenic outcomes across tissues, and how the multiple signaling pathways (MAPK, PI3K-Akt, JAK/STAT) are causally linked to its molecular activity.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural or biochemical reconciliation of context-dependent roles","Phosphoregulation site mapping absent","Direct effector mechanism upstream of signaling pathways undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[0,6]}],"localization":[{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0,6]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[2,3,4]},{"term_id":"R-HSA-397014","term_label":"Muscle contraction","supporting_discovery_ids":[7]}],"complexes":[],"partners":["AHSA1","ERK1/2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q05682","full_name":"Caldesmon","aliases":[],"length_aa":793,"mass_kda":93.2,"function":"Actin- and myosin-binding protein implicated in the regulation of actomyosin interactions in smooth muscle and nonmuscle cells (could act as a bridge between myosin and actin filaments). Stimulates actin binding of tropomyosin which increases the stabilization of actin filament structure. In muscle tissues, inhibits the actomyosin ATPase by binding to F-actin. This inhibition is attenuated by calcium-calmodulin and is potentiated by tropomyosin. Interacts with actin, myosin, two molecules of tropomyosin and with calmodulin. Also plays an essential role during cellular mitosis and receptor capping. Involved in Schwann cell migration during peripheral nerve regeneration (By similarity)","subcellular_location":"Cytoplasm, cytoskeleton; Cytoplasm, myofibril; Cytoplasm, cytoskeleton, stress fiber","url":"https://www.uniprot.org/uniprotkb/Q05682/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CALD1","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000122786","cell_line_id":"CID000958","localizations":[{"compartment":"cytoskeleton","grade":3},{"compartment":"cytoplasmic","grade":1}],"interactors":[{"gene":"CAPZB","stoichiometry":10.0},{"gene":"FLOT1","stoichiometry":0.2},{"gene":"AP2A2","stoichiometry":0.2},{"gene":"FLNB","stoichiometry":0.2},{"gene":"RPLP0;RPLP0P6","stoichiometry":0.2},{"gene":"RPLP1","stoichiometry":0.2},{"gene":"ATP1A1","stoichiometry":0.2},{"gene":"OAT","stoichiometry":0.2},{"gene":"VIM","stoichiometry":0.2},{"gene":"ANXA6","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID000958","total_profiled":1310},"omim":[{"mim_id":"608667","title":"NIPPED-B-LIKE; NIPBL","url":"https://www.omim.org/entry/608667"},{"mim_id":"114213","title":"CALDESMON 1; CALD1","url":"https://www.omim.org/entry/114213"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Plasma membrane","reliability":"Enhanced"},{"location":"Actin filaments","reliability":"Enhanced"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/CALD1"},"hgnc":{"alias_symbol":["CDM","H-CAD","L-CAD","h-CD"],"prev_symbol":[]},"alphafold":{"accession":"Q05682","domains":[],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q05682","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q05682-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q05682-F1-predicted_aligned_error_v6.png","plddt_mean":64.44},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CALD1","jax_strain_url":"https://www.jax.org/strain/search?query=CALD1"},"sequence":{"accession":"Q05682","fasta_url":"https://rest.uniprot.org/uniprotkb/Q05682.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q05682/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q05682"}},"corpus_meta":[{"pmid":"15161654","id":"PMC_15161654","title":"Differential expression of splicing variants of the human caldesmon gene (CALD1) in glioma neovascularization versus normal brain microvasculature.","date":"2004","source":"The American journal of pathology","url":"https://pubmed.ncbi.nlm.nih.gov/15161654","citation_count":50,"is_preprint":false},{"pmid":"34070840","id":"PMC_34070840","title":"CALD1 Modulates Gliomas Progression via Facilitating Tumor Angiogenesis.","date":"2021","source":"Cancers","url":"https://pubmed.ncbi.nlm.nih.gov/34070840","citation_count":23,"is_preprint":false},{"pmid":"34733993","id":"PMC_34733993","title":"CALD1 promotes the expression of PD-L1 in bladder cancer via the JAK/STAT signaling pathway.","date":"2021","source":"Annals of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/34733993","citation_count":22,"is_preprint":false},{"pmid":"19329885","id":"PMC_19329885","title":"Hela l-CaD is implicated in the migration of endothelial cells/endothelial progenitor cells in human neoplasms.","date":"2007","source":"Cell adhesion & migration","url":"https://pubmed.ncbi.nlm.nih.gov/19329885","citation_count":18,"is_preprint":false},{"pmid":"34422947","id":"PMC_34422947","title":"Quantitative Expression of TYR, CD34, and CALD1 Discriminates Between Canine Oral Malignant Melanomas and Soft Tissue Sarcomas.","date":"2021","source":"Frontiers in veterinary science","url":"https://pubmed.ncbi.nlm.nih.gov/34422947","citation_count":17,"is_preprint":false},{"pmid":"36230524","id":"PMC_36230524","title":"AHSA1 Promotes Proliferation and EMT by Regulating ERK/CALD1 Axis in Hepatocellular Carcinoma.","date":"2022","source":"Cancers","url":"https://pubmed.ncbi.nlm.nih.gov/36230524","citation_count":16,"is_preprint":false},{"pmid":"38013282","id":"PMC_38013282","title":"MYLK and CALD1 as molecular targets in bladder cancer.","date":"2023","source":"Medicine","url":"https://pubmed.ncbi.nlm.nih.gov/38013282","citation_count":8,"is_preprint":false},{"pmid":"38577446","id":"PMC_38577446","title":"CALD1 facilitates epithelial-mesenchymal transition progression in gastric cancer cells by modulating the PI3K-Akt pathway.","date":"2024","source":"World journal of gastrointestinal oncology","url":"https://pubmed.ncbi.nlm.nih.gov/38577446","citation_count":6,"is_preprint":false},{"pmid":"38025524","id":"PMC_38025524","title":"MiR-1278 targets CALD1 and suppresses the progression of gastric cancer via the MAPK pathway.","date":"2023","source":"Open medicine (Warsaw, Poland)","url":"https://pubmed.ncbi.nlm.nih.gov/38025524","citation_count":5,"is_preprint":false},{"pmid":"37017680","id":"PMC_37017680","title":"METTL14 Promotes Oral Squamous Cell Carcinoma Progression by Regulating the mRNA and m6A Levels of CALD1.","date":"2023","source":"Journal of environmental pathology, toxicology and oncology : official organ of the International Society for Environmental Toxicology and Cancer","url":"https://pubmed.ncbi.nlm.nih.gov/37017680","citation_count":4,"is_preprint":false},{"pmid":"39476475","id":"PMC_39476475","title":"Endothelin-1 potentiated constriction in preeclampsia placental veins: Role of ETAR/ETBR/CaV1.2/CALD1.","date":"2024","source":"Placenta","url":"https://pubmed.ncbi.nlm.nih.gov/39476475","citation_count":4,"is_preprint":false},{"pmid":"28255976","id":"PMC_28255976","title":"Association of rs 3807337 polymorphism of CALD1 gene with diabetic nephropathy occurrence in type 1 diabetes - preliminary results of a family-based study.","date":"2017","source":"Endokrynologia Polska","url":"https://pubmed.ncbi.nlm.nih.gov/28255976","citation_count":4,"is_preprint":false},{"pmid":"40818763","id":"PMC_40818763","title":"CALD1-derived circ-0003746 targeting miR-526b promotes EMT-mediated bladder cancer progression.","date":"2025","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/40818763","citation_count":2,"is_preprint":false},{"pmid":"40104711","id":"PMC_40104711","title":"CALD1 inhibits invasion of human ovarian cancer cells by affecting cytoskeletal structure and the number of focal adhesion.","date":"2025","source":"Translational cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/40104711","citation_count":2,"is_preprint":false},{"pmid":"34708628","id":"PMC_34708628","title":"Identification of Cald1 as a novel regulator of Linggui Zhugan decoction for improving insulin resistance in vivo and in vitro.","date":"2021","source":"Journal of traditional Chinese medicine = Chung i tsa chih ying wen pan","url":"https://pubmed.ncbi.nlm.nih.gov/34708628","citation_count":2,"is_preprint":false},{"pmid":"41171370","id":"PMC_41171370","title":"Cancer-associated fibroblast miR-148a-5p/CALD1/collagen VI pathway promotes proliferation in Helicobacter pylori-positive gastric cancer.","date":"2025","source":"Cellular oncology (Dordrecht, Netherlands)","url":"https://pubmed.ncbi.nlm.nih.gov/41171370","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":11169,"output_tokens":2445,"usd":0.035091,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9557,"output_tokens":2587,"usd":0.05623,"stage2_stop_reason":"end_turn"},"total_usd":0.091321,"stage1_batch_id":"msgbatch_01ACAxJQabDsht39JXeWG22d","stage2_batch_id":"msgbatch_01QEU22A9U5sbPTgsMzTmvSf","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2007,\n      \"finding\": \"The low molecular weight caldesmon isoform (Hela l-CaD) localizes to motility-related cell protrusions (filopodia, microspikes, lamellipodia, podosomes, membrane blebs, and membrane ruffles) in endothelial cells/endothelial progenitor cells in tumor vasculature, implicating it in cell migration during angiogenesis and vasculogenesis.\",\n      \"method\": \"Immunohistochemistry and histological analysis of motility-related protrusions in histologically preserved tumor microenvironment sections\",\n      \"journal\": \"Cell adhesion & migration\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single method (localization by IHC), no functional rescue or in vitro motility assay\",\n      \"pmids\": [\"19329885\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"AHSA1 recruits ERK1/2 to phosphorylate and inactivate CALD1; pharmacological inhibition of ERK1/2 phosphorylation (with SCH772984) reversed AHSA1-promoted proliferation and EMT, and CALD1 knockdown rescued the inhibition of proliferation and EMT caused by AHSA1 knockdown, placing CALD1 downstream of the ERK1/2 axis.\",\n      \"method\": \"Gain- and loss-of-function studies in vitro and in vivo, ERK1/2 phosphorylation inhibitor treatment, Western blot for phosphorylated CALD1, siRNA knockdown rescue experiments\",\n      \"journal\": \"Cancers\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, multiple orthogonal methods (KO/KD, pharmacological inhibition, rescue experiment, in vivo validation)\",\n      \"pmids\": [\"36230524\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CALD1 promotes PD-L1 (CD274) expression in bladder cancer via activation of the JAK/STAT signaling pathway; this effect was blocked by the specific JAK inhibitor Ruxolitinib.\",\n      \"method\": \"CALD1 overexpression/knockdown in bladder cancer cells, GSEA pathway enrichment, pharmacological inhibition with Ruxolitinib, Western blot, flow cytometry\",\n      \"journal\": \"Annals of translational medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, multiple methods (OE/KD, pharmacological rescue, pathway inhibitor), consistent with GSEA\",\n      \"pmids\": [\"34733993\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"miR-1278 directly targets and negatively regulates CALD1 expression (confirmed by dual luciferase reporter assay); CALD1 overexpression activates the MAPK pathway (upregulating Ras, p-P38, p-ERK1/2), promoting gastric cancer cell viability and migration, which is partially rescued by miR-1278 mimic.\",\n      \"method\": \"Dual luciferase reporter assay, CCK-8, Transwell migration assay, Western blot for MAPK pathway proteins, in vivo xenograft\",\n      \"journal\": \"Open medicine (Warsaw, Poland)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, direct binding confirmed by luciferase assay, pathway proteins measured, rescue experiment performed\",\n      \"pmids\": [\"38025524\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"CALD1 promotes EMT in gastric cancer by activating the PI3K-Akt pathway: CALD1 overexpression increased PI3K, p-AKT, and p-mTOR while decreasing PTEN; PI3K-Akt inhibitor treatment reversed CALD1-mediated effects on proliferation, migration, invasion, and EMT marker expression.\",\n      \"method\": \"siRNA knockdown and overexpression in AGS and MKN45 cells, CCK-8, wound healing and Transwell assays, Western blot, PI3K-Akt inhibitor treatment, in vivo xenograft\",\n      \"journal\": \"World journal of gastrointestinal oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, multiple orthogonal methods (KD/OE, pharmacological inhibition, in vivo validation)\",\n      \"pmids\": [\"38577446\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"METTL14 methyltransferase increases CALD1 mRNA and m6A methylation levels in oral squamous cell carcinoma; METTL14 silencing depleted CALD1 mRNA and m6A levels, suppressing cell growth and metastasis, which was rescued by CALD1 overexpression.\",\n      \"method\": \"MeRIP assay for m6A levels, qRT-PCR, Western blot, colony formation and Transwell assays, siRNA knockdown, in vivo tumorigenicity assay\",\n      \"journal\": \"Journal of environmental pathology, toxicology and oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, MeRIP directly measures m6A on CALD1, rescue experiment confirms pathway placement\",\n      \"pmids\": [\"37017680\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CALD1 knockdown in SK-OV-3 ovarian cancer cells reduces F-actin stress fibers, loosens cytoskeletal structure, decreases Vinculin expression and focal adhesion number, and enhances cell invasiveness, demonstrating that CALD1 restrains invasion by maintaining cytoskeletal organization and focal adhesion formation.\",\n      \"method\": \"Stable CALD1 knockdown by shRNA, Transwell invasion assay, immunofluorescence staining of F-actin and Vinculin, Western blot\",\n      \"journal\": \"Translational cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, direct functional readout (invasion + cytoskeletal imaging), loss-of-function with specific phenotypic readout\",\n      \"pmids\": [\"40104711\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ET-1 treatment increases high-molecular-weight CALD1 expression (a marker of the contractile smooth muscle cell phenotype) in placental vessel explants and HUVSMCs via ETAR/ETBR receptors; elevated CALD1 is associated with stronger contraction in preeclampsia placental chorionic plate veins.\",\n      \"method\": \"DMT vessel tone measurement, whole-cell patch clamp, Western blot, ELISA, PCR, ET-1 treatment with receptor antagonists in placental explants and HUVSMCs\",\n      \"journal\": \"Placenta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, pharmacological receptor blockade identifies ETAR/ETBR as writers of CALD1 upregulation, multiple orthogonal methods\",\n      \"pmids\": [\"39476475\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In Helicobacter pylori-positive gastric cancer, TLR signaling in cancer-associated fibroblasts suppresses miR-148a-5p, which in turn de-represses CALD1; elevated CAF CALD1 promotes collagen VI secretion that interacts with tumoral SDC4 receptors to drive GC cell proliferation.\",\n      \"method\": \"microRNA and transcriptome sequencing, single-cell sequencing, in vitro co-culture, CDX and PDX in vivo models, miR-148a-5p agomir treatment\",\n      \"journal\": \"Cellular oncology (Dordrecht, Netherlands)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, multiple orthogonal methods (in vitro and in vivo models, sequencing, functional rescue with agomir)\",\n      \"pmids\": [\"41171370\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"CALD1 (caldesmon) is a cytoskeleton-associated actin-binding protein whose high-molecular-weight isoform stabilizes the contractile smooth muscle phenotype (regulated by ET-1 via ETAR/ETBR) and whose non-muscle isoform localizes to actin-rich cell protrusions to support cell migration; CALD1 activity is post-translationally regulated by ERK1/2-mediated phosphorylation (downstream of AHSA1), transcriptionally regulated by METTL14-dependent m6A methylation and by miR-1278/miR-148a-5p targeting, and it signals through the JAK/STAT, MAPK, and PI3K-Akt pathways to promote proliferation, EMT, and immune evasion in multiple cancer types, while in ovarian cancer it acts as a tumor suppressor by maintaining F-actin stress fibers, focal adhesions, and Vinculin expression to restrain invasion.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CALD1 (caldesmon) is an actin-associated cytoskeletal protein whose isoforms govern smooth muscle contractility and cell motility, and which is repeatedly co-opted in cancer to control proliferation, EMT, and invasion [#6, #7]. Its high-molecular-weight isoform marks the contractile smooth muscle phenotype and is upregulated by endothelin-1 acting through ETAR/ETBR receptors, with elevated CALD1 linked to stronger vascular contraction [#7], while the low-molecular-weight isoform localizes to motility-related protrusions such as filopodia, lamellipodia, and podosomes consistent with a role in cell migration [#0]. In ovarian cancer CALD1 acts to restrain invasion: its loss reduces F-actin stress fibers, loosens cytoskeletal organization, and decreases Vinculin expression and focal adhesion number [#6]. In several other cancers CALD1 instead promotes malignant behavior through downstream signaling, activating the MAPK pathway (Ras, p-P38, p-ERK1/2) [#3] and the PI3K-Akt-mTOR axis with concomitant PTEN loss to drive proliferation and EMT [#4], and activating JAK/STAT signaling to upregulate PD-L1 (CD274) [#2]. CALD1 is post-translationally inactivated by ERK1/2-mediated phosphorylation recruited via AHSA1 [#1], and its expression is controlled by METTL14-dependent m6A methylation [#5] and by negative regulation through miR-1278 [#3] and miR-148a-5p [#8]; in H. pylori-positive gastric cancer, de-repressed CALD1 in cancer-associated fibroblasts drives collagen VI secretion that engages tumoral SDC4 to promote tumor growth [#8].\"\n,\n  \"teleology\": [\n    {\n      \"year\": 2007,\n      \"claim\": \"Established where the low-molecular-weight caldesmon isoform acts in cells, linking it to the migratory machinery rather than only contraction.\",\n      \"evidence\": \"Immunohistochemistry of motility-related protrusions in tumor vasculature sections\",\n      \"pmids\": [\"19329885\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Localization only — no functional motility assay or rescue\", \"Does not establish causal role in migration\", \"Isoform-specific function not dissected mechanistically\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Connected CALD1 to immune evasion by showing it drives PD-L1 expression through a defined kinase pathway.\",\n      \"evidence\": \"Overexpression/knockdown in bladder cancer cells with Ruxolitinib JAK inhibition, Western blot, flow cytometry\",\n      \"pmids\": [\"34733993\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking CALD1 to JAK/STAT activation unresolved\", \"Single cancer context\", \"No direct biochemical interaction shown\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined how CALD1 activity is post-translationally controlled, placing it downstream of an AHSA1–ERK1/2 axis that phosphorylates and inactivates it.\",\n      \"evidence\": \"Gain/loss-of-function in vitro and in vivo, SCH772984 ERK inhibition, Western blot for phospho-CALD1, knockdown rescue\",\n      \"pmids\": [\"36230524\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Phosphosites on CALD1 not mapped\", \"Direct AHSA1–CALD1 vs indirect recruitment not distinguished\", \"Functional consequence of inactivation on actin binding not measured\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identified upstream transcriptional/epigenetic regulators of CALD1 (m6A and microRNA) and a downstream MAPK effector pathway.\",\n      \"evidence\": \"MeRIP/qRT-PCR with METTL14 silencing and rescue (OSCC); dual luciferase miR-1278 binding, CCK-8/Transwell, MAPK Western blots (gastric)\",\n      \"pmids\": [\"37017680\", \"38025524\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"m6A reader mediating CALD1 mRNA fate not identified\", \"How CALD1 protein activates MAPK is not mechanistically defined\", \"Single-lab findings per cancer type\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Extended CALD1's pro-tumor signaling to the PI3K-Akt-mTOR axis driving EMT.\",\n      \"evidence\": \"siRNA/overexpression in AGS and MKN45 cells, pharmacological PI3K-Akt inhibition, Western blot, in vivo xenograft\",\n      \"pmids\": [\"38577446\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Link between CALD1 and PI3K activation not biochemically defined\", \"Relationship to the MAPK and JAK/STAT effects in other studies unclear\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Revealed a context-dependent tumor-suppressive role (ovarian) via cytoskeletal/focal adhesion maintenance, and a stromal CAF-driven pro-tumor axis (gastric).\",\n      \"evidence\": \"shRNA knockdown with invasion assay and F-actin/Vinculin immunofluorescence (ovarian); miR/transcriptome and single-cell sequencing, co-culture, CDX/PDX, miR-148a-5p agomir (gastric)\",\n      \"pmids\": [\"40104711\", \"41171370\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis for opposing tumor-suppressor vs oncogenic roles unresolved\", \"Direct CALD1 role in collagen VI secretion vs correlative not established\", \"Cell-type specificity of CALD1 function not reconciled\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how CALD1's core actin-binding/cytoskeletal function mechanistically dictates its opposing tumor-suppressive versus oncogenic outcomes across tissues, and how the multiple signaling pathways (MAPK, PI3K-Akt, JAK/STAT) are causally linked to its molecular activity.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural or biochemical reconciliation of context-dependent roles\", \"Phosphoregulation site mapping absent\", \"Direct effector mechanism upstream of signaling pathways undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [0, 6]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0, 6]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [2, 3, 4]},\n      {\"term_id\": \"R-HSA-397014\", \"supporting_discovery_ids\": [7]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"AHSA1\", \"ERK1/2\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}