{"gene":"TM4SF5","run_date":"2026-04-28T21:42:59","timeline":{"discoveries":[{"year":2008,"finding":"TM4SF5 overexpression in hepatocarcinoma causes cytosolic stabilization of p27Kip1 and RhoA inactivation, leading to epithelial-mesenchymal transition (EMT) with loss of E-cadherin and aberrant multilayer cell growth; suppression of TM4SF5, cytosolic p27Kip1, or reconstitution of E-cadherin abolished these effects.","method":"Ectopic expression, shRNA knockdown, anchorage-independent growth assay, S-phase transition assay, nude mouse tumor formation, E-cadherin reconstitution","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (KD, OE, rescue, in vivo), replicated across several subsequent studies","pmids":["18357344"],"is_preprint":false},{"year":2006,"finding":"TM4SF5 associates with integrin α2 subunit, and this association is abolished by serum treatment; TM4SF5 regulates actin organization and focal contact dynamics via serum-dependent differential regulation of FAK Tyr925 and paxillin Tyr118 phosphorylations; Y925F FAK mutation abolished TM4SF5 effects; functional blocking of integrin α2 abolished TM4SF5-enhanced signaling and caused abnormal actin organization.","method":"Co-immunoprecipitation, ectopic expression in Cos7 cells, anti-integrin blocking antibody, FAK point mutagenesis, phosphorylation assays, migration assays","journal":"Experimental cell research","confidence":"High","confidence_rationale":"Tier 1–2 — direct Co-IP binding, mutagenesis, functional rescue, multiple readouts in single study","pmids":["16828471"],"is_preprint":false},{"year":2008,"finding":"TM4SF5 retains integrin α5 on the cell surface to induce VEGF expression and secretion; TM4SF5-mediated VEGF induction and angiogenesis required integrin α5, c-Src, and STAT3; anti-integrin α5 antibody abolished TM4SF5-mediated VEGF expression and tube formation by endothelial cells.","method":"Anti-integrin α5 antibody blockade, conditioned media assay, HUVEC tube formation, aorta ring outgrowth, anti-VEGF antibody neutralization, nude mouse xenograft","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal in vitro and in vivo methods establishing pathway (TM4SF5→integrin α5→c-Src→STAT3→VEGF)","pmids":["19036703"],"is_preprint":false},{"year":2009,"finding":"The second extracellular loop (EL2) of TM4SF5 directly interacts with integrin α2 in a collagen type I environment, inhibiting integrin α2 functions such as cell spreading and migration toward collagen I; EL2 peptide or mutagenesis of EL2 recovered integrin α2 function.","method":"Co-immunoprecipitation, EL2 peptide blocking, site-directed mutagenesis, cell spreading and migration assays on collagen I","journal":"Carcinogenesis","confidence":"High","confidence_rationale":"Tier 1–2 — direct domain mapping by mutagenesis and peptide competition, multiple functional readouts","pmids":["19789264"],"is_preprint":false},{"year":2009,"finding":"N-glycosylation of TM4SF5 is required for TM4SF5-specific responsiveness to the antagonist TSAHC; point mutations of putative N-glycosylation sites abolished this responsiveness, indicating that glycosylation of the extracellular region is important for TM4SF5 protein-protein interactions.","method":"Site-directed mutagenesis of N-glycosylation sites, TSAHC drug treatment, multilayer growth assay, migration/invasion assay","journal":"Hepatology (Baltimore, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 — mutagenesis with functional readout, single lab","pmids":["19177595"],"is_preprint":false},{"year":2010,"finding":"TM4SF5 accelerates G1/S phase progression by facilitating CDK4/cyclin D1 nuclear entry and complex formation, Rb phosphorylation, and cyclin D1/E upregulation; these effects were blocked by p27Kip1 siRNA silencing or constitutively active RhoA; ROCK inhibition mimicked TM4SF5 effects in control cells.","method":"siRNA knockdown, active RhoA infection, ROCK pharmacological inhibition, cell cycle analysis, co-IP for CDK4/cyclin D1 complex, subcellular fractionation","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2 — multiple epistasis experiments establishing pathway, single lab","pmids":["20399237"],"is_preprint":false},{"year":2010,"finding":"TM4SF5 expression facilitates invadopodia formation, MMP activation, and invasion in hepatocarcinoma cells, leading to lung metastasis in nude mice; shRNA suppression of TM4SF5 blocked these effects.","method":"shRNA knockdown, in vitro invasion assay, MMP activity assay, invadopodia assay, nude mouse lung metastasis model","journal":"Journal of cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 — KD with defined cellular and in vivo phenotype, single lab","pmids":["20506553"],"is_preprint":false},{"year":2011,"finding":"TM4SF5-mediated Ser10 phosphorylation of p27Kip1 (required for cytosolic localization) is dependent on JNK activity; JNK inhibition or suppression in TM4SF5-expressing cells decreased p27Kip1 Ser10 phosphorylation and rescued E-cadherin expression and localization at cell-cell contacts.","method":"JNK pharmacological inhibition, JNK siRNA knockdown, p27Kip1 phosphorylation assays, immunofluorescence of adherens junction molecules","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 — epistasis via inhibitor and siRNA with specific phosphorylation readout, single lab","pmids":["22014979"],"is_preprint":false},{"year":2011,"finding":"TM4SF5 expression inhibits proteasome activity and proteasome subunit expression in hepatocarcinoma cells, causing loss of cell-cell contacts and E-cadherin; shRNA against TM4SF5 recovered proteasome expression and cell-cell adhesion.","method":"shRNA knockdown, proteasome activity assay, proteasome subunit expression analysis, immunofluorescence of E-cadherin","journal":"Journal of cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 — KD with specific mechanistic readout (proteasome activity), single lab","pmids":["21328452"],"is_preprint":false},{"year":2012,"finding":"TM4SF5 directly binds FAK in an adhesion-dependent manner; this binding causes a structural alteration releasing the inhibitory intramolecular interaction in FAK, activating FAK at the cell's leading edge for migration/invasion and in vivo metastasis; impaired TM4SF5-FAK interaction attenuated FAK phosphorylation and metastatic potential.","method":"Co-IP (direct binding), mutagenesis to impair TM4SF5-FAK interaction, phosphorylation assays, cell migration/invasion assay, in vivo metastasis model, immunofluorescence of leading-edge localization","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 1–2 — direct binding via Co-IP with mutagenesis, structural rationale, in vivo validation, multiple orthogonal readouts","pmids":["23077174"],"is_preprint":false},{"year":2012,"finding":"The C-terminus of TM4SF5 binds c-Src (both inactive and active forms); TM4SF5 modulates c-Src activity to promote invasive protrusion formation; c-Src activity correlates with EGFR Tyr845 phosphorylation; Y845F EGFR mutation abolished TM4SF5-mediated invasive protrusions but not c-Src phosphorylation, establishing a TM4SF5/c-Src/EGFR(Y845) signaling axis for invasion.","method":"Co-IP (C-terminus domain mapping), TM4SF5 C-terminal deletion mutant (ΔC), site-directed EGFR Y845F mutagenesis, migration and invasion assays, phosphorylation assays","journal":"Biochimica et biophysica acta","confidence":"High","confidence_rationale":"Tier 1–2 — domain-level interaction mapping, mutagenesis of both TM4SF5 and EGFR, multiple functional readouts, single lab with rigorous controls","pmids":["23220047"],"is_preprint":false},{"year":2012,"finding":"TGFβ1-mediated Smad activation induces TM4SF5 expression and EMT through EGFR pathway activation; Smad overexpression activated EGFR and induced TM4SF5 in the absence of serum; EGFR kinase inhibition, EGF depletion, or Smad7 expression abolished TM4SF5 induction and EMT, placing TGFβ1→Smad→EGFR→TM4SF5 as a signaling axis.","method":"Smad overexpression, Smad7 inhibition, EGFR kinase inhibitor treatment, EGF depletion, small compound TM4SF5 inhibition, TM4SF5 expression monitoring in normal hepatocytes","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 2 — genetic epistasis via multiple independent interventions establishing pathway order, replicated with normal and cancer hepatocytes","pmids":["22292774"],"is_preprint":false},{"year":2014,"finding":"TM4SF5 interacts with CD151 (tumorigenic tetraspanin) and causes internalization of CD63 (tumor-suppressive tetraspanin) from the cell surface into late lysosomal membranes; TM4SF5 controls expression levels of CD151 and CD63, but not vice versa; TM4SF5 could overcome CD151 tumorigenic effects on migration and ECM degradation.","method":"Co-IP (TM4SF5-CD151 interaction), subcellular fractionation/immunofluorescence showing CD63 internalization, shRNA epistasis experiments, TGFβ1-treated Chang cell model","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2–3 — Co-IP with functional epistasis, localization with functional consequence, single lab","pmids":["25033048"],"is_preprint":false},{"year":2014,"finding":"IL-6 treatment activates FAK and STAT3 in TM4SF5-null cells but decreases TM4SF5-dependent FAK activity in TM4SF5-expressing cancer cells; TM4SF5 expression in hepatocellular carcinoma cells causes invasive ECM degradation negatively dependent on IL-6/IL-6R signaling, establishing that cancer cells adopt TM4SF5-dependent FAK activation by lowering IL-6 to avoid immune surveillance.","method":"IL-6 treatment, STAT3 suppression (siRNA), FAK activity modulation, Co-IP-established TM4SF5/FAK pathway, ECM degradation assay, comparison of normal vs. cancerous hepatocytes","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 — epistasis via multiple interventions, single lab","pmids":["24912675"],"is_preprint":false},{"year":2015,"finding":"TM4SF5 physically interacts with CD44 through their extracellular domains in an N-glycosylation-dependent manner; TM4SF5/CD44 interaction activates c-Src/STAT3/Twist1/Bmi1 signaling for spheroid (self-renewal) formation; disrupting any component of this pathway inhibited spheroid formation; TM4SF5-positive cells circulate in blood after orthotopic liver injection, and anti-TM4SF5 reagent blocked metastasis to distal organs.","method":"Co-IP (extracellular domain mapping), N-glycosylation mutagenesis, pathway component siRNA/inhibitor epistasis, 3D spheroid assay, in vivo orthotopic model with laser scanning endomicroscopy for CTC detection","journal":"Hepatology (Baltimore, Md.)","confidence":"High","confidence_rationale":"Tier 1–2 — domain-level binding confirmed with mutagenesis, multi-component pathway epistasis, in vivo validation with multiple orthogonal methods","pmids":["25627085"],"is_preprint":false},{"year":2015,"finding":"TM4SF5 and IGF1R transcriptionally modulate each other's expression; TM4SF5 and IGF1R form a protein complex (also including EGFR) in a TM4SF5-dependent manner; co-expression promotes ERK, Akt, and S6K signaling and residual EGFR activity after EGFR kinase inhibitor treatment, causing resistance to erlotinib and gefitinib.","method":"Co-IP (TM4SF5/IGF1R/EGFR complex), ectopic TM4SF5 expression, IGF1R siRNA knockdown, EGFR kinase inhibitor treatment, 2D/3D culture drug sensitivity assays","journal":"Lung cancer (Amsterdam, Netherlands)","confidence":"Medium","confidence_rationale":"Tier 2–3 — Co-IP with functional epistasis, single lab","pmids":["26190015"],"is_preprint":false},{"year":2017,"finding":"TM4SF5 physically associates with EGFR and integrin α5 at the leading edge of migratory cells (visualized by live fluorescence cross-correlation spectroscopy); cholesterol depletion and disruption of TM4SF5 N-glycosylation or palmitoylation alter these interactions and reduce cell migration speed and directionality in 2D and 3D conditions.","method":"Live fluorescence cross-correlation spectroscopy (FCS), super-resolution microscopy, cholesterol depletion (methyl-β-cyclodextrin), N-glycosylation and palmitoylation mutagenesis, 2D/3D migration assays","journal":"FASEB journal","confidence":"High","confidence_rationale":"Tier 1–2 — single-molecule live imaging plus mutagenesis, multiple orthogonal methods in a single study","pmids":["28073834"],"is_preprint":false},{"year":2018,"finding":"CD133 phosphorylation induces TM4SF5 expression; TM4SF5 binds CD133 and promotes c-Src activity for CD133 phosphorylation (positive feedback); TM4SF5 also binds PTPRF and promotes paxillin phosphorylation; sphere growth decreased by CD133 suppression was recovered by TM4SF5 expression and partially by PTPRF suppression.","method":"Co-IP (TM4SF5-CD133, TM4SF5-PTPRF), siRNA knockdown epistasis, paxillin phosphorylation assay, 3D sphere growth assay","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2–3 — multiple Co-IPs with functional epistasis, single lab","pmids":["30217560"],"is_preprint":false},{"year":2019,"finding":"TM4SF5 induces the alternatively spliced CD44v8-10 variant through an inverse ZEB2/ESRP linkage; TM4SF5 forms complexes with the cystine/glutamate antiporter system (xCT) via TM4SF5- and CD44v8-10-dependent CD98hc plasma membrane enrichment; dynamic TM4SF5 binding to CD98hc required CD44v8-10 under ROS-generating conditions; this complex upregulates cystine/glutamate antiporter activity and intracellular glutathione for ROS modulation and cell survival. Tm4sf5-null mice showed attenuated bleomycin-induced pulmonary fibrosis.","method":"Co-IP (TM4SF5-CD98hc, TM4SF5-CD44v8-10), alternative splicing analysis (RT-PCR), ZEB2/ESRP expression analysis, glutathione assay, xCT activity assay, Tm4sf5 KO mouse bleomycin model","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 2 — multiple Co-IPs, in vivo KO model, functional metabolic readout (glutathione/ROS), single lab with orthogonal methods","pmids":["31501417"],"is_preprint":false},{"year":2020,"finding":"TM4SF5 forms protein-protein complexes with amino acid transporters including xCT (cystine/glutamate antiporter) and regulates cystine uptake from extracellular space and arginine export from lysosomal lumen to cytosol; diverse amino acid transporters co-precipitate with TM4SF5 by proteomic analysis.","method":"Co-IP, proximity-based proteomics, amino acid transport assays","journal":"Experimental & molecular medicine","confidence":"Medium","confidence_rationale":"Tier 3 — review consolidating prior Co-IP findings with some new proteomic data, single lab","pmids":["31956272"],"is_preprint":false},{"year":2020,"finding":"TM4SF5-overexpressing mice develop age-dependent nonalcoholic steatosis and NASH; in young mice TM4SF5 decreases SIRT1, increases SREBPs, and inactivates STAT3 via SOCS1/3 upregulation; in older mice TM4SF5 promotes SIRT1 expression and STAT3 activity for ECM production; CCL20 suppression reduced immune cell infiltration and ECM production; active STAT3 increases collagen I and laminin γ2, which in turn support SIRT1/STAT3 activity.","method":"TM4SF5 transgenic and KO mouse models, diet/chemical-treated mice, primary hepatocyte culture, CCL20 suppression, collagen I/laminin γ2 knockdown, human tissue analysis","journal":"The Journal of pathology","confidence":"High","confidence_rationale":"Tier 2 — transgenic and KO in vivo models with multiple genetic interventions, validated in human tissue","pmids":["32918742"],"is_preprint":false},{"year":2021,"finding":"TM4SF5 induction in differentiated macrophages promotes glucose uptake, glycolysis, and M1-type macrophage activation; activated M1 macrophages secrete IL-6, which induces CCL20 and CXCL10 secretion from TM4SF5-positive hepatocytes; chronic exposure to these chemokines reprograms macrophages toward M2-type, supporting NAFLD progression.","method":"TM4SF5 overexpression in macrophages, glycolysis assay, cytokine ELISA, co-culture systems, macrophage polarization assays, IL-6 neutralization","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 — multiple functional assays establishing intercellular signaling pathway, single lab","pmids":["34788612"],"is_preprint":false},{"year":2021,"finding":"TM4SF5 expression in cancer cells downregulates stimulatory NK cell ligands and receptors (SLAMF6, SLAMF7, MICA/B), causing NK cell exhaustion-like phenotypes; TM4SF5 suppression or inhibition with TSAHC reduced STAT3 signaling, recovered NK cell receptor levels and NK cell surveillance, and reduced liver cancer progression.","method":"TM4SF5 transgenic and DEN-induced liver cancer mouse models, TSAHC inhibitor treatment, NK cell activity assays, flow cytometry for NK ligand/receptor expression, STAT3 modulation","journal":"Cellular and molecular life sciences","confidence":"Medium","confidence_rationale":"Tier 2 — in vivo models with mechanistic follow-up, single lab","pmids":["34921636"],"is_preprint":false},{"year":2021,"finding":"The TM4SF5 C-terminus binds the c-Src SH1 kinase domain (preferentially in its inactive closed form) along with PTP1B which dephosphorylates Tyr530; the SH1 domain alone bound TM4SF5 to cause c-Src Tyr419 and FAK Y861 phosphorylation; cell-penetrating TM4SF5 C-terminal peptides blocked TM4SF5-c-Src interaction and prevented tumor initiation/progression in vivo.","method":"Co-IP (domain mapping: C-terminus vs. SH1), homology modeling, molecular dynamics simulation, mutagenesis validation, cell-penetrating peptide competition, in vivo xenograft model","journal":"Theranostics","confidence":"High","confidence_rationale":"Tier 1–2 — structural modeling validated by mutagenesis and peptide competition, in vivo functional validation","pmids":["34335982"],"is_preprint":false},{"year":2021,"finding":"TM4SF5 intracellular vesicle traffic toward the leading edge is controlled by cell adhesion to fibronectin and microtubule acetylation; TM4SF5 palmitoylation is required for directed traffic; TM4SF5 forms a trimeric complex with HDAC6 and SLAC2B at perinuclear cytosol; SLAC2B suppression allows acetylated microtubules to extend to leading edges, facilitating TM4SF5 translocation and persistent migration; HDAC6 inhibition (via paxillin at new adhesion sites) promotes TM4SF5 traffic.","method":"Live-cell imaging of TM4SF5 vesicle tracking, palmitoylation-deficient mutant, SLAC2B siRNA, HDAC6 inhibition, Co-IP (TM4SF5-HDAC6-SLAC2B trimeric complex), immunofluorescence of acetylated tubulin","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 — live imaging with mutagenesis and Co-IP, single lab","pmids":["33554392"],"is_preprint":false},{"year":2021,"finding":"TM4SF5 functions as a lysosomal arginine sensor and activates mTORC1; TM4SF5 KO in adipocytes reduces mTORC1 activation, enhances autophagy and lipolysis, increases PPARα and mitochondrial oxidative metabolism gene expression, reduces adiposity, and prevents HFD-induced glucose intolerance.","method":"TM4SF5 KO mouse model, mTORC1 activity assays, autophagy assays (LC3 flux), lipolysis assay, RNA sequencing of adipose tissue, metabolic phenotyping","journal":"Diabetes","confidence":"Medium","confidence_rationale":"Tier 2 — KO mouse with multiple metabolic readouts, single lab","pmids":["34187836"],"is_preprint":false},{"year":2022,"finding":"Hepatic TM4SF5 binds GLUT1 at the plasma membrane to promote glucose uptake and glycolysis; excessive glucose causes hepatocytes to secrete TM4SF5-loaded small extracellular vesicles (sEVs); liver-derived sEVs containing TM4SF5 target brown adipose tissue (BAT) to improve glucose clearance independent of UCP1.","method":"Co-IP (TM4SF5-GLUT1), glucose uptake assay, sEV isolation and characterization, liver-closed vein circuit (LCVC) in vivo delivery of sEVs from TM4SF5-overexpressing mice, glucose tolerance tests in KO mice, BAT targeting assay","journal":"Journal of extracellular vesicles","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP with functional metabolic readouts and in vivo sEV delivery experiment, single lab","pmids":["36063136"],"is_preprint":false},{"year":2022,"finding":"TM4SF5 binds GLUT8 at the plasma membrane; following fructose treatment, TM4SF5-GLUT8 binding transiently decreases, allowing GLUT8 translocation to the plasma membrane for fructose uptake and de novo lipogenesis; Tm4sf5 suppression or KO reduced fructose uptake, DNL, and steatosis in vivo.","method":"Co-IP (TM4SF5-GLUT8), GLUT8 localization by immunofluorescence (translocation assay), fructose uptake assay, DNL measurement, Tm4sf5 KO mouse with high-sucrose/fructose diet","journal":"Molecular metabolism","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP with dynamic localization and in vivo KO metabolic phenotype, single lab","pmids":["35123128"],"is_preprint":false},{"year":2023,"finding":"Upon glucose repletion following depletion, TM4SF5 becomes enriched at mitochondria-lysosome contact sites (MLCSs) via interaction between mitochondrial FKBP8 and lysosomal TM4SF5; proximity labeling revealed clustering of phospho-DRP1 and mitophagy receptors at TM4SF5-enriched MLCSs, promoting mitochondrial fission and autophagy; TM4SF5 binds NPC1 and free cholesterol, mediating cholesterol export from lysosomes to mitochondria and impairing oxidative phosphorylation.","method":"Co-IP (TM4SF5-FKBP8, TM4SF5-NPC1), proximity-labeling proteomics (BioID), organelle reconstitution, cholesterol transport assay, mitophagy assay, DRP1 phosphorylation analysis, in vivo mouse hepatocyte models","journal":"Cancer communications (London, England)","confidence":"High","confidence_rationale":"Tier 1–2 — proximity labeling proteomics plus Co-IP, organelle reconstitution, cholesterol transport assay, in vivo validation","pmids":["38133457"],"is_preprint":false},{"year":2014,"finding":"TM4SF5 suppression in zebrafish impairs trunk muscle development, aberrant muscle fibre morphology, and alters integrin α5 expression; integrin α5-related signaling molecules (fibronectin, FAK, vinculin, actin) are aberrantly localized in tm4sf5 morphants; aberrant muscle development was rescued by injection of tm4sf5 or integrin α5 mRNA, establishing TM4SF5 function in muscle differentiation via integrin α5-dependent signaling.","method":"Morpholino knockdown (zebrafish), mRNA rescue injection, immunofluorescence of muscle and signaling molecules, C2C12 mouse myoblast differentiation assay","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 — morpholino KD with mRNA rescue in zebrafish, supported by mammalian cell data, single lab","pmids":["24897542"],"is_preprint":false},{"year":2025,"finding":"TM4SF5 expressed by hepatocytes reduces NK cell cytotoxicity by binding SLAMF7 in an N-glycosylation-dependent manner, causing intracellular trafficking of SLAMF7 from the plasma membrane to lysosomes for degradation; TM4SF5-specific isoxazole (TSI) compounds block this binding and trafficking, restoring NK cell surveillance and reducing HCC development in xenograft models.","method":"Co-IP (TM4SF5-SLAMF7, N-glycosylation mutagenesis), immunofluorescence tracking of SLAMF7 trafficking to lysosomes, TSI small molecule treatment, NK cell cytotoxicity assay, mouse xenograft and Tm4sf5-KO models","journal":"Signal transduction and targeted therapy","confidence":"High","confidence_rationale":"Tier 1–2 — direct binding with domain mutagenesis, subcellular trafficking demonstrated by imaging, functional NK cell assay, in vivo validation","pmids":["39828766"],"is_preprint":false},{"year":2025,"finding":"TM4SF5-mediated macropinocytosis of albumin requires cytosolic stabilization of NCOA3 and PTEN inactivation through TM4SF5 binding; albumin uptake via macropinocytosis supports ATP-linked respiration and cellular migration in TM4SF5-expressing hepatocytes.","method":"Co-IP (TM4SF5-NCOA3, TM4SF5-PTEN), NCOA3 and PTEN expression/activity assays, macropinocytosis assay, albumin uptake assay, ATP-linked respiration (Seahorse), TM4SF5 KO and reintroduction, in vivo orthotopic mouse model","journal":"Experimental & molecular medicine","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP with functional metabolic and migration readouts, in vivo validation, single lab","pmids":["40186033"],"is_preprint":false},{"year":2025,"finding":"TM4SF5 modulates KEAP1 independently of NRF2: the cytosolic TM4SF5 C-terminus binds KEAP1 to promote its proteasomal degradation under physiological conditions; in hyperlipidemic/pathological states TM4SF5 stabilizes KEAP1, leading to oxidative stress and hepatic inflammation; Keap1 suppression nullified TM4SF5-mediated MASLD phenotypes.","method":"Co-IP (TM4SF5 C-terminus binding to KEAP1), proteasome inhibitor treatment, Tm4sf5 KO and Nrf2 mutant mouse models, Keap1 siRNA suppression, in vitro and in vivo MASLD models","journal":"International journal of biological sciences","confidence":"Medium","confidence_rationale":"Tier 2 — direct binding with domain identification, genetic epistasis (KO+KD), in vivo validation, single lab","pmids":["41608638"],"is_preprint":false},{"year":2025,"finding":"TM4SF5 forms an N-glycosylation-dependent dimer in its large extracellular loop (LEL); the LEL has a β-sheet configuration (unlike the α-helices of genuine tetraspanins or CD20-like family); TM4SF5 has two conserved cysteines (without the CCG motif) affecting N-glycosylation and dimer formation, and the LEL contributes to cholesterol binding.","method":"Structural analysis, sequence/domain comparison, N-glycosylation mutagenesis, cholesterol binding assay","journal":"Journal of advanced research","confidence":"Medium","confidence_rationale":"Tier 1 — structural characterization with mutagenesis, single lab/study","pmids":["41349605"],"is_preprint":false}],"current_model":"TM4SF5 is a four-transmembrane glycoprotein (L6 family, tetraspanin-like) that organizes signaling microdomains at the plasma membrane and on intracellular organelles, where it directly binds and activates FAK (by relieving its autoinhibition) and c-Src (via its C-terminus engaging the SH1 kinase domain), cooperates with integrins α2, α5, and β1, EGFR, CD44, IGF1R, and multiple amino acid/nutrient transporters to drive epithelial-mesenchymal transition, cell migration/invasion, self-renewal, metabolic reprogramming (glucose/fructose uptake, lipogenesis, mitochondrial cholesterol export at MLCS), and immune evasion (by trafficking SLAMF7 to lysosomes to suppress NK cell killing), while also functioning as a lysosomal arginine sensor that activates mTORC1 and modulates KEAP1 stability independently of NRF2."},"narrative":{"teleology":[{"year":2006,"claim":"Establishing TM4SF5 as a membrane organizer of integrin–FAK signaling: it was unknown how TM4SF5 influenced adhesion signaling; Co-IP showed TM4SF5 associates with integrin α2 and controls FAK Y925/paxillin Y118 phosphorylation in a serum-dependent manner, demonstrating TM4SF5 as a regulator of focal contact dynamics.","evidence":"Co-immunoprecipitation, integrin-blocking antibody, FAK Y925F mutagenesis, migration assays in Cos7 cells","pmids":["16828471"],"confidence":"High","gaps":["Stoichiometry and directness of TM4SF5–integrin α2 binding not resolved","Whether TM4SF5 binds FAK directly was unknown at this stage"]},{"year":2008,"claim":"Defining the EMT/pro-tumorigenic axis: TM4SF5 was shown to stabilize cytosolic p27Kip1, inactivate RhoA, and suppress E-cadherin, driving EMT and multilayer growth in hepatocarcinoma; concurrently, TM4SF5 retained integrin α5 at the surface to activate c-Src/STAT3/VEGF for angiogenesis, establishing two parallel downstream pathways.","evidence":"shRNA knockdown, ectopic expression, nude mouse xenograft, HUVEC tube formation, E-cadherin reconstitution, anti-integrin α5 blockade","pmids":["18357344","19036703"],"confidence":"High","gaps":["Direct physical contacts between TM4SF5 and p27Kip1 or RhoA not demonstrated","Structural basis for integrin α5 retention was unresolved"]},{"year":2009,"claim":"Mapping the interaction interface: the second extracellular loop (EL2) of TM4SF5 was identified as the integrin α2-binding domain, and N-glycosylation of TM4SF5 was shown to be required for protein–protein interactions and drug responsiveness, establishing the glycosylation-dependent nature of TM4SF5 function.","evidence":"EL2 peptide competition, site-directed mutagenesis of N-glycosylation sites, collagen I spreading/migration assays","pmids":["19789264","19177595"],"confidence":"High","gaps":["Three-dimensional structure of EL2 was unknown","Which specific glycan species mediate interaction was not determined"]},{"year":2012,"claim":"Defining direct kinase activation mechanisms: TM4SF5 was shown to directly bind FAK, relieving its autoinhibitory intramolecular fold, and the TM4SF5 C-terminus was mapped as the c-Src/EGFR Y845 signaling platform, resolving the molecular basis of two key downstream kinase activations.","evidence":"Co-IP with domain mapping, FAK mutagenesis, EGFR Y845F mutagenesis, C-terminal deletion mutant, in vivo metastasis model","pmids":["23077174","23220047"],"confidence":"High","gaps":["Crystal/cryo-EM structure of TM4SF5–FAK or TM4SF5–c-Src complex not available","Whether FAK and c-Src bind TM4SF5 simultaneously was not tested"]},{"year":2012,"claim":"Placing TM4SF5 induction within TGFβ signaling: TGFβ1/Smad activation was shown to induce TM4SF5 expression via EGFR, establishing that TM4SF5 is a downstream effector of TGFβ-mediated EMT and explaining how hepatocytes acquire TM4SF5 expression.","evidence":"Smad overexpression, EGFR kinase inhibitor, Smad7 epistasis, normal and cancer hepatocyte comparison","pmids":["22292774"],"confidence":"High","gaps":["Direct Smad-binding site in TM4SF5 promoter not mapped","Contribution of other EMT-inducing signals to TM4SF5 induction not tested"]},{"year":2014,"claim":"Expanding the tetraspanin-web and developmental roles: TM4SF5 was shown to control CD151/CD63 sorting (internalizing CD63 to lysosomes), and zebrafish morpholino studies demonstrated a requirement for TM4SF5 in integrin α5-dependent muscle development, broadening TM4SF5 function beyond cancer.","evidence":"Co-IP for TM4SF5–CD151, CD63 subcellular fractionation, zebrafish morpholino knockdown with mRNA rescue, C2C12 differentiation","pmids":["25033048","24897542"],"confidence":"Medium","gaps":["Whether CD63 internalization is direct or requires adaptor proteins was unknown","Zebrafish morpholino studies lack genetic knockout confirmation"]},{"year":2015,"claim":"Establishing TM4SF5 as a self-renewal/stemness driver: TM4SF5 was found to bind CD44 through N-glycosylation-dependent extracellular domain interactions, activating c-Src/STAT3/Twist1/Bmi1 for spheroid formation, and co-expressed with IGF1R/EGFR to form a complex conferring EGFR-inhibitor resistance.","evidence":"Co-IP with domain mapping, N-glycosylation mutagenesis, pathway epistasis, orthotopic liver model with CTC detection, EGFR inhibitor sensitivity assays","pmids":["25627085","26190015"],"confidence":"High","gaps":["Relative contribution of CD44 vs. IGF1R complexes to stemness not dissected","Whether TM4SF5/CD44/IGF1R/EGFR exist in a single supercomplex or separate pools was unclear"]},{"year":2017,"claim":"Visualizing TM4SF5 microdomain dynamics at the leading edge: live single-molecule cross-correlation spectroscopy confirmed TM4SF5 co-diffuses with EGFR and integrin α5 at the plasma membrane in a cholesterol- and palmitoylation-dependent manner, providing biophysical evidence for tetraspanin-enriched microdomain organization.","evidence":"Live fluorescence cross-correlation spectroscopy, super-resolution microscopy, cholesterol depletion, palmitoylation mutagenesis, 2D/3D migration","pmids":["28073834"],"confidence":"High","gaps":["Lipid composition requirements beyond cholesterol not characterized","Whether palmitoylation directly mediates partner binding or affects membrane partitioning was not distinguished"]},{"year":2019,"claim":"Revealing metabolic/transporter functions: TM4SF5 was found to complex with the xCT/CD98hc cystine–glutamate antiporter via CD44v8-10 splice-variant induction, regulating glutathione homeostasis and ROS defense; Tm4sf5-KO mice showed attenuated pulmonary fibrosis, linking TM4SF5 to metabolic redox control in vivo.","evidence":"Co-IP of TM4SF5–CD98hc/xCT, glutathione assay, alternative splicing analysis, Tm4sf5-KO mouse bleomycin model","pmids":["31501417"],"confidence":"High","gaps":["Whether TM4SF5 directly transports any metabolite or acts purely as a scaffolding organizer was unresolved","Tissue-specificity of the xCT complex function beyond lung not tested"]},{"year":2021,"claim":"Defining TM4SF5 as a lysosomal nutrient sensor and intracellular trafficking hub: TM4SF5 was identified as a lysosomal arginine sensor activating mTORC1 in adipocytes (KO reduced adiposity and improved glucose tolerance); simultaneously, the C-terminus/SH1-domain interaction with c-Src was structurally modeled and validated, and TM4SF5 vesicle trafficking was shown to depend on a TM4SF5/HDAC6/SLAC2B complex riding acetylated microtubules.","evidence":"mTORC1 activity and autophagy assays in KO mice, metabolic phenotyping, Co-IP domain mapping of c-Src SH1, cell-penetrating peptide competition, live vesicle tracking, HDAC6/SLAC2B Co-IP","pmids":["34187836","34335982","33554392"],"confidence":"High","gaps":["The arginine-sensing mechanism (direct binding vs. indirect) not biochemically resolved","Whether HDAC6/SLAC2B complex exists on lysosomal vs. plasma-membrane-directed vesicles was unclear"]},{"year":2021,"claim":"Establishing immune evasion functions: TM4SF5 in hepatocytes suppressed NK cell ligands (SLAMF6/7, MICA/B) and promoted macrophage M1-to-M2 reprogramming via IL-6/CCL20 secretion, linking TM4SF5 to both innate immune evasion and NAFLD progression.","evidence":"Tm4sf5 transgenic and DEN-induced liver cancer models, NK cell cytotoxicity assays, macrophage polarization co-culture, IL-6 neutralization, TSAHC inhibitor","pmids":["34921636","34788612"],"confidence":"Medium","gaps":["Direct mechanism by which TM4SF5 downregulates NK ligands (transcriptional vs. post-translational) was not fully resolved","In vivo contribution of macrophage vs. NK evasion to tumor progression not separated"]},{"year":2022,"claim":"Expanding sugar transporter partnerships: TM4SF5 was shown to bind GLUT1 for glucose uptake and GLUT8 for fructose uptake/de novo lipogenesis; hepatocyte-derived TM4SF5-containing extracellular vesicles targeted brown adipose tissue to improve systemic glucose clearance, revealing an inter-organ signaling role.","evidence":"Co-IP of TM4SF5–GLUT1 and TM4SF5–GLUT8, glucose/fructose uptake assays, sEV isolation and in vivo delivery via liver-closed vein circuit, Tm4sf5-KO mice on high-sucrose diets","pmids":["36063136","35123128"],"confidence":"Medium","gaps":["Whether TM4SF5 in sEVs retains functional transporter-scaffolding activity in recipient cells was not shown","The GLUT8 release mechanism upon fructose stimulation was not molecularly defined"]},{"year":2023,"claim":"Defining mitochondria–lysosome contact site (MLCS) functions: TM4SF5 was shown to enrich at MLCSs via binding lysosomal TM4SF5 to mitochondrial FKBP8; proximity labeling revealed DRP1 and mitophagy receptor clustering, and TM4SF5 mediated NPC1-dependent cholesterol transfer from lysosomes to mitochondria, impairing oxidative phosphorylation.","evidence":"Co-IP of TM4SF5–FKBP8 and TM4SF5–NPC1, BioID proximity proteomics, cholesterol transport reconstitution, mitophagy assay, DRP1 phosphorylation, in vivo mouse models","pmids":["38133457"],"confidence":"High","gaps":["Whether MLCS enrichment requires mTORC1 activity (linking to the arginine-sensing role) was not tested","Structural basis of the TM4SF5–FKBP8 interaction unknown"]},{"year":2025,"claim":"Resolving the NK-evasion mechanism and structural features: TM4SF5 was shown to directly bind SLAMF7 in an N-glycosylation-dependent manner and traffic it to lysosomes for degradation, suppressing NK cytotoxicity; the LEL was structurally characterized as a β-sheet fold (distinct from genuine tetraspanins) that forms N-glycosylation-dependent dimers and binds cholesterol; TM4SF5 also modulates KEAP1 stability independently of NRF2.","evidence":"Co-IP with N-glycosylation mutagenesis, SLAMF7 trafficking imaging, TSI inhibitor treatment, xenograft models, structural/domain analysis, KEAP1 proteasomal degradation assay, Tm4sf5-KO and Nrf2-mutant mice","pmids":["39828766","41349605","41608638"],"confidence":"High","gaps":["High-resolution structure of the TM4SF5 LEL dimer still lacking","Mechanism by which pathological lipid conditions switch TM4SF5 from KEAP1 degradation to stabilization is not defined"]},{"year":null,"claim":"A high-resolution structure of TM4SF5 (alone and in complex with key partners such as FAK, c-Src, integrins, or SLAMF7) is needed to understand the allosteric mechanisms by which it activates kinases and organizes multi-protein complexes; the direct arginine-binding mechanism on lysosomes remains biochemically unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No crystal or cryo-EM structure available","Arginine-sensing mechanism (direct binding site, affinity) not biochemically defined","Relative physiological importance of individual TM4SF5 complexes in specific tissues unclear"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[1,9,10,14,16,18]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[9,23,25,32]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[28,33]},{"term_id":"GO:0140299","term_label":"molecular sensor activity","supporting_discovery_ids":[25]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[1,3,16,26,27]},{"term_id":"GO:0005764","term_label":"lysosome","supporting_discovery_ids":[12,25,28,30]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[28]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[24]},{"term_id":"GO:0005576","term_label":"extracellular region","supporting_discovery_ids":[26]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,2,5,9,10,11,14,15,23,25]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[18,19,26,27,28]},{"term_id":"R-HSA-1500931","term_label":"Cell-Cell communication","supporting_discovery_ids":[0,8,14,16]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[21,22,30]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[25,28]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[5]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[20,27,32]},{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[24,26,30]}],"complexes":["TM4SF5/integrin α5/EGFR leading-edge complex","TM4SF5/CD44v8-10/CD98hc/xCT transporter complex","TM4SF5/HDAC6/SLAC2B vesicle trafficking complex","TM4SF5/IGF1R/EGFR signaling complex"],"partners":["PTK2","SRC","ITGA2","ITGA5","CD44","EGFR","FKBP8","SLAMF7"],"other_free_text":[]},"mechanistic_narrative":"TM4SF5 is a four-transmembrane L6-family glycoprotein that organizes plasma membrane and intracellular signaling microdomains to coordinate integrin/growth factor receptor signaling, metabolic reprogramming, and immune evasion. Its second extracellular loop engages integrins α2 and α5, CD44, and EGFR in N-glycosylation- and cholesterol-dependent complexes, while its cytoplasmic C-terminus directly binds and activates FAK (by relieving autoinhibition) and c-Src (via the SH1 kinase domain), driving epithelial–mesenchymal transition, invadopodia formation, and metastasis [PMID:18357344, PMID:23077174, PMID:34335982, PMID:28073834]. On intracellular membranes, TM4SF5 functions as a lysosomal arginine sensor activating mTORC1, partners with nutrient transporters (GLUT1, GLUT8, xCT/CD98hc) to regulate glucose/fructose uptake and de novo lipogenesis, and localizes to mitochondria–lysosome contact sites where it binds FKBP8 and NPC1 to mediate cholesterol export and regulate mitochondrial fission [PMID:34187836, PMID:36063136, PMID:35123128, PMID:38133457]. TM4SF5 also promotes immune evasion by trafficking SLAMF7 to lysosomes for degradation, suppressing NK cell cytotoxicity, and by modulating macrophage polarization and chemokine secretion to facilitate NAFLD/NASH and hepatocellular carcinoma progression [PMID:39828766, PMID:34788612, PMID:32918742]."},"prefetch_data":{"uniprot":{"accession":"O14894","full_name":"Transmembrane 4 L6 family member 5","aliases":["Tetraspan transmembrane protein L6H"],"length_aa":197,"mass_kda":20.8,"function":"Acts as a lysosomal membrane arginine sensor (PubMed:30956113). Forms a complex with MTOR and SLC38A9 on lysosomal membranes in an arginine-regulated manner, leading to arginine efflux which enables the activation of mTORC1 which subsequently leads to RPS6KB1 and EIF4EBP1 phosphorylations (PubMed:30956113). Facilitates cell cycle G1/S phase progression and the translocation of the CDK4-CCND1 complex into the nucleus (PubMed:20399237). CDKN1B and RHOA/ROCK signaling activity are involved in TM4SF5-mediated acceleration of G1/S phase progression (PubMed:20399237)","subcellular_location":"Lysosome membrane; Cell membrane","url":"https://www.uniprot.org/uniprotkb/O14894/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/TM4SF5","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/TM4SF5","total_profiled":1310},"omim":[{"mim_id":"604657","title":"TRANSMEMBRANE 4 SUPERFAMILY, MEMBER 5; TM4SF5","url":"https://www.omim.org/entry/604657"},{"mim_id":"176830","title":"PROOPIOMELANOCORTIN; POMC","url":"https://www.omim.org/entry/176830"},{"mim_id":"164810","title":"FOS PROTOONCOGENE, AP1 TRANSCRIPTION FACTOR SUBUNIT; FOS","url":"https://www.omim.org/entry/164810"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Uncertain","locations":[{"location":"Plasma membrane","reliability":"Uncertain"},{"location":"Cell Junctions","reliability":"Uncertain"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Group enriched","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"intestine","ntpm":164.2},{"tissue":"liver","ntpm":190.2}],"url":"https://www.proteinatlas.org/search/TM4SF5"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"O14894","domains":[{"cath_id":"-","chopping":"2-187","consensus_level":"high","plddt":88.6387,"start":2,"end":187}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/O14894","model_url":"https://alphafold.ebi.ac.uk/files/AF-O14894-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-O14894-F1-predicted_aligned_error_v6.png","plddt_mean":86.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=TM4SF5","jax_strain_url":"https://www.jax.org/strain/search?query=TM4SF5"},"sequence":{"accession":"O14894","fasta_url":"https://rest.uniprot.org/uniprotkb/O14894.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/O14894/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/O14894"}},"corpus_meta":[{"pmid":"18357344","id":"PMC_18357344","title":"Tetraspanin 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Chemoresistance.","date":"2023","source":"Pharmaceuticals (Basel, Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/36678607","citation_count":10,"is_preprint":false},{"pmid":"25268742","id":"PMC_25268742","title":"Therapeutic effect of a TM4SF5-specific monoclonal antibody against colon cancer in a mouse model.","date":"2014","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/25268742","citation_count":10,"is_preprint":false},{"pmid":"18501196","id":"PMC_18501196","title":"Regulation of TM4SF5-mediated tumorigenesis through induction of cell detachment and death by tiarellic acid.","date":"2008","source":"Biochimica et biophysica acta","url":"https://pubmed.ncbi.nlm.nih.gov/18501196","citation_count":9,"is_preprint":false},{"pmid":"23624388","id":"PMC_23624388","title":"Prophylactic effect of a peptide vaccine targeting TM4SF5 against colon cancer in a mouse model.","date":"2013","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/23624388","citation_count":9,"is_preprint":false},{"pmid":"35955521","id":"PMC_35955521","title":"TM4SF5-Mediated Regulation of Hepatocyte Transporters during Metabolic Liver Diseases.","date":"2022","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/35955521","citation_count":9,"is_preprint":false},{"pmid":"39828766","id":"PMC_39828766","title":"Isoxazole-based molecules restore NK cell immune surveillance in hepatocarcinogenesis by targeting TM4SF5 and SLAMF7 linkage.","date":"2025","source":"Signal transduction and targeted therapy","url":"https://pubmed.ncbi.nlm.nih.gov/39828766","citation_count":8,"is_preprint":false},{"pmid":"35211652","id":"PMC_35211652","title":"Therapeutic effects of TM4SF5-targeting chimeric and humanized monoclonal antibodies in hepatocellular and colon cancer models.","date":"2022","source":"Molecular therapy oncolytics","url":"https://pubmed.ncbi.nlm.nih.gov/35211652","citation_count":8,"is_preprint":false},{"pmid":"26531872","id":"PMC_26531872","title":"TM4SF5-CTD-2354A18.1-miR-4697-3P may play a key role in the pathogenesis of gastric cancer.","date":"2015","source":"Bratislavske lekarske listy","url":"https://pubmed.ncbi.nlm.nih.gov/26531872","citation_count":8,"is_preprint":false},{"pmid":"24897542","id":"PMC_24897542","title":"TM4SF5 suppression disturbs integrin α5-related signalling and muscle development in zebrafish.","date":"2014","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/24897542","citation_count":7,"is_preprint":false},{"pmid":"28458469","id":"PMC_28458469","title":"TM4SF5-Mediated Roles in the Development of Fibrotic Phenotypes.","date":"2017","source":"Mediators of inflammation","url":"https://pubmed.ncbi.nlm.nih.gov/28458469","citation_count":6,"is_preprint":false},{"pmid":"31897180","id":"PMC_31897180","title":"Targeting TM4SF5 with anti-TM4SF5 monoclonal antibody suppresses the growth and motility of human pancreatic cancer cells.","date":"2019","source":"Oncology letters","url":"https://pubmed.ncbi.nlm.nih.gov/31897180","citation_count":5,"is_preprint":false},{"pmid":"36104259","id":"PMC_36104259","title":"Systemic TM4SF5 overexpression in ApcMin/+ mice promotes hepatic portal hypertension associated with fibrosis.","date":"2022","source":"BMB reports","url":"https://pubmed.ncbi.nlm.nih.gov/36104259","citation_count":4,"is_preprint":false},{"pmid":"29137358","id":"PMC_29137358","title":"TM4SF5 promotes metastatic behavior of cells in 3D extracellular matrix gels by reducing dependency on environmental cues.","date":"2017","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/29137358","citation_count":4,"is_preprint":false},{"pmid":"37670786","id":"PMC_37670786","title":"TM4SF5-mediated abnormal food-intake behavior and apelin expression facilitate non-alcoholic fatty liver disease features.","date":"2023","source":"iScience","url":"https://pubmed.ncbi.nlm.nih.gov/37670786","citation_count":3,"is_preprint":false},{"pmid":"31186734","id":"PMC_31186734","title":"Clinicopathological significance of TM4SF5 expression in human hepatocellular carcinoma tissues.","date":"2019","source":"Oncology letters","url":"https://pubmed.ncbi.nlm.nih.gov/31186734","citation_count":3,"is_preprint":false},{"pmid":"33222072","id":"PMC_33222072","title":"Metastatic behavior analyses of tetraspanin TM4SF5-expressing spheres in three-dimensional (3D) cell culture environment.","date":"2020","source":"Archives of pharmacal research","url":"https://pubmed.ncbi.nlm.nih.gov/33222072","citation_count":3,"is_preprint":false},{"pmid":"40186033","id":"PMC_40186033","title":"Hepatocyte TM4SF5-mediated cytosolic NCOA3 stabilization and macropinocytosis support albumin uptake and bioenergetics for hepatocellular carcinoma progression.","date":"2025","source":"Experimental & molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40186033","citation_count":2,"is_preprint":false},{"pmid":"33554392","id":"PMC_33554392","title":"SLAC2B-dependent microtubule acetylation regulates extracellular matrix-mediated intracellular TM4SF5 traffic to the plasma membranes.","date":"2021","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/33554392","citation_count":2,"is_preprint":false},{"pmid":"39454995","id":"PMC_39454995","title":"Association between hepatocyte TM4SF5 expression and gut microbiome dysbiosis during non-alcoholic fatty liver disease development.","date":"2024","source":"Life sciences","url":"https://pubmed.ncbi.nlm.nih.gov/39454995","citation_count":1,"is_preprint":false},{"pmid":"41349605","id":"PMC_41349605","title":"Unique molecular architecture of N-glycosylated TM4SF5 dimer highlights evolutionary and structural divergence among small four-transmembrane protein families.","date":"2025","source":"Journal of advanced research","url":"https://pubmed.ncbi.nlm.nih.gov/41349605","citation_count":0,"is_preprint":false},{"pmid":"41608638","id":"PMC_41608638","title":"TM4SF5-mediated KEAP1 Regulation in Hepatocytes Irrelevant to NRF2 Expression and Activity Promotes Oxidative Stress and Inflammation to Develop Metabolic Dysfunction-Associated Steatotic Liver Disease.","date":"2026","source":"International journal of biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/41608638","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":32449,"output_tokens":9122,"usd":0.117088},"stage2":{"model":"claude-opus-4-6","input_tokens":12990,"output_tokens":4902,"usd":0.28125},"total_usd":0.398338,"stage1_batch_id":"msgbatch_01RPbVc9PDH9zQmYK5Hs6srm","stage2_batch_id":"msgbatch_01JVbVLzeKN2yVrsGp3zC8aM","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2008,\n      \"finding\": \"TM4SF5 overexpression in hepatocarcinoma causes cytosolic stabilization of p27Kip1 and RhoA inactivation, leading to epithelial-mesenchymal transition (EMT) with loss of E-cadherin and aberrant multilayer cell growth; suppression of TM4SF5, cytosolic p27Kip1, or reconstitution of E-cadherin abolished these effects.\",\n      \"method\": \"Ectopic expression, shRNA knockdown, anchorage-independent growth assay, S-phase transition assay, nude mouse tumor formation, E-cadherin reconstitution\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (KD, OE, rescue, in vivo), replicated across several subsequent studies\",\n      \"pmids\": [\"18357344\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"TM4SF5 associates with integrin α2 subunit, and this association is abolished by serum treatment; TM4SF5 regulates actin organization and focal contact dynamics via serum-dependent differential regulation of FAK Tyr925 and paxillin Tyr118 phosphorylations; Y925F FAK mutation abolished TM4SF5 effects; functional blocking of integrin α2 abolished TM4SF5-enhanced signaling and caused abnormal actin organization.\",\n      \"method\": \"Co-immunoprecipitation, ectopic expression in Cos7 cells, anti-integrin blocking antibody, FAK point mutagenesis, phosphorylation assays, migration assays\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — direct Co-IP binding, mutagenesis, functional rescue, multiple readouts in single study\",\n      \"pmids\": [\"16828471\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"TM4SF5 retains integrin α5 on the cell surface to induce VEGF expression and secretion; TM4SF5-mediated VEGF induction and angiogenesis required integrin α5, c-Src, and STAT3; anti-integrin α5 antibody abolished TM4SF5-mediated VEGF expression and tube formation by endothelial cells.\",\n      \"method\": \"Anti-integrin α5 antibody blockade, conditioned media assay, HUVEC tube formation, aorta ring outgrowth, anti-VEGF antibody neutralization, nude mouse xenograft\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal in vitro and in vivo methods establishing pathway (TM4SF5→integrin α5→c-Src→STAT3→VEGF)\",\n      \"pmids\": [\"19036703\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"The second extracellular loop (EL2) of TM4SF5 directly interacts with integrin α2 in a collagen type I environment, inhibiting integrin α2 functions such as cell spreading and migration toward collagen I; EL2 peptide or mutagenesis of EL2 recovered integrin α2 function.\",\n      \"method\": \"Co-immunoprecipitation, EL2 peptide blocking, site-directed mutagenesis, cell spreading and migration assays on collagen I\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — direct domain mapping by mutagenesis and peptide competition, multiple functional readouts\",\n      \"pmids\": [\"19789264\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"N-glycosylation of TM4SF5 is required for TM4SF5-specific responsiveness to the antagonist TSAHC; point mutations of putative N-glycosylation sites abolished this responsiveness, indicating that glycosylation of the extracellular region is important for TM4SF5 protein-protein interactions.\",\n      \"method\": \"Site-directed mutagenesis of N-glycosylation sites, TSAHC drug treatment, multilayer growth assay, migration/invasion assay\",\n      \"journal\": \"Hepatology (Baltimore, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — mutagenesis with functional readout, single lab\",\n      \"pmids\": [\"19177595\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"TM4SF5 accelerates G1/S phase progression by facilitating CDK4/cyclin D1 nuclear entry and complex formation, Rb phosphorylation, and cyclin D1/E upregulation; these effects were blocked by p27Kip1 siRNA silencing or constitutively active RhoA; ROCK inhibition mimicked TM4SF5 effects in control cells.\",\n      \"method\": \"siRNA knockdown, active RhoA infection, ROCK pharmacological inhibition, cell cycle analysis, co-IP for CDK4/cyclin D1 complex, subcellular fractionation\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple epistasis experiments establishing pathway, single lab\",\n      \"pmids\": [\"20399237\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"TM4SF5 expression facilitates invadopodia formation, MMP activation, and invasion in hepatocarcinoma cells, leading to lung metastasis in nude mice; shRNA suppression of TM4SF5 blocked these effects.\",\n      \"method\": \"shRNA knockdown, in vitro invasion assay, MMP activity assay, invadopodia assay, nude mouse lung metastasis model\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — KD with defined cellular and in vivo phenotype, single lab\",\n      \"pmids\": [\"20506553\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"TM4SF5-mediated Ser10 phosphorylation of p27Kip1 (required for cytosolic localization) is dependent on JNK activity; JNK inhibition or suppression in TM4SF5-expressing cells decreased p27Kip1 Ser10 phosphorylation and rescued E-cadherin expression and localization at cell-cell contacts.\",\n      \"method\": \"JNK pharmacological inhibition, JNK siRNA knockdown, p27Kip1 phosphorylation assays, immunofluorescence of adherens junction molecules\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — epistasis via inhibitor and siRNA with specific phosphorylation readout, single lab\",\n      \"pmids\": [\"22014979\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"TM4SF5 expression inhibits proteasome activity and proteasome subunit expression in hepatocarcinoma cells, causing loss of cell-cell contacts and E-cadherin; shRNA against TM4SF5 recovered proteasome expression and cell-cell adhesion.\",\n      \"method\": \"shRNA knockdown, proteasome activity assay, proteasome subunit expression analysis, immunofluorescence of E-cadherin\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — KD with specific mechanistic readout (proteasome activity), single lab\",\n      \"pmids\": [\"21328452\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"TM4SF5 directly binds FAK in an adhesion-dependent manner; this binding causes a structural alteration releasing the inhibitory intramolecular interaction in FAK, activating FAK at the cell's leading edge for migration/invasion and in vivo metastasis; impaired TM4SF5-FAK interaction attenuated FAK phosphorylation and metastatic potential.\",\n      \"method\": \"Co-IP (direct binding), mutagenesis to impair TM4SF5-FAK interaction, phosphorylation assays, cell migration/invasion assay, in vivo metastasis model, immunofluorescence of leading-edge localization\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — direct binding via Co-IP with mutagenesis, structural rationale, in vivo validation, multiple orthogonal readouts\",\n      \"pmids\": [\"23077174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The C-terminus of TM4SF5 binds c-Src (both inactive and active forms); TM4SF5 modulates c-Src activity to promote invasive protrusion formation; c-Src activity correlates with EGFR Tyr845 phosphorylation; Y845F EGFR mutation abolished TM4SF5-mediated invasive protrusions but not c-Src phosphorylation, establishing a TM4SF5/c-Src/EGFR(Y845) signaling axis for invasion.\",\n      \"method\": \"Co-IP (C-terminus domain mapping), TM4SF5 C-terminal deletion mutant (ΔC), site-directed EGFR Y845F mutagenesis, migration and invasion assays, phosphorylation assays\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — domain-level interaction mapping, mutagenesis of both TM4SF5 and EGFR, multiple functional readouts, single lab with rigorous controls\",\n      \"pmids\": [\"23220047\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"TGFβ1-mediated Smad activation induces TM4SF5 expression and EMT through EGFR pathway activation; Smad overexpression activated EGFR and induced TM4SF5 in the absence of serum; EGFR kinase inhibition, EGF depletion, or Smad7 expression abolished TM4SF5 induction and EMT, placing TGFβ1→Smad→EGFR→TM4SF5 as a signaling axis.\",\n      \"method\": \"Smad overexpression, Smad7 inhibition, EGFR kinase inhibitor treatment, EGF depletion, small compound TM4SF5 inhibition, TM4SF5 expression monitoring in normal hepatocytes\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis via multiple independent interventions establishing pathway order, replicated with normal and cancer hepatocytes\",\n      \"pmids\": [\"22292774\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"TM4SF5 interacts with CD151 (tumorigenic tetraspanin) and causes internalization of CD63 (tumor-suppressive tetraspanin) from the cell surface into late lysosomal membranes; TM4SF5 controls expression levels of CD151 and CD63, but not vice versa; TM4SF5 could overcome CD151 tumorigenic effects on migration and ECM degradation.\",\n      \"method\": \"Co-IP (TM4SF5-CD151 interaction), subcellular fractionation/immunofluorescence showing CD63 internalization, shRNA epistasis experiments, TGFβ1-treated Chang cell model\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — Co-IP with functional epistasis, localization with functional consequence, single lab\",\n      \"pmids\": [\"25033048\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"IL-6 treatment activates FAK and STAT3 in TM4SF5-null cells but decreases TM4SF5-dependent FAK activity in TM4SF5-expressing cancer cells; TM4SF5 expression in hepatocellular carcinoma cells causes invasive ECM degradation negatively dependent on IL-6/IL-6R signaling, establishing that cancer cells adopt TM4SF5-dependent FAK activation by lowering IL-6 to avoid immune surveillance.\",\n      \"method\": \"IL-6 treatment, STAT3 suppression (siRNA), FAK activity modulation, Co-IP-established TM4SF5/FAK pathway, ECM degradation assay, comparison of normal vs. cancerous hepatocytes\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — epistasis via multiple interventions, single lab\",\n      \"pmids\": [\"24912675\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"TM4SF5 physically interacts with CD44 through their extracellular domains in an N-glycosylation-dependent manner; TM4SF5/CD44 interaction activates c-Src/STAT3/Twist1/Bmi1 signaling for spheroid (self-renewal) formation; disrupting any component of this pathway inhibited spheroid formation; TM4SF5-positive cells circulate in blood after orthotopic liver injection, and anti-TM4SF5 reagent blocked metastasis to distal organs.\",\n      \"method\": \"Co-IP (extracellular domain mapping), N-glycosylation mutagenesis, pathway component siRNA/inhibitor epistasis, 3D spheroid assay, in vivo orthotopic model with laser scanning endomicroscopy for CTC detection\",\n      \"journal\": \"Hepatology (Baltimore, Md.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — domain-level binding confirmed with mutagenesis, multi-component pathway epistasis, in vivo validation with multiple orthogonal methods\",\n      \"pmids\": [\"25627085\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"TM4SF5 and IGF1R transcriptionally modulate each other's expression; TM4SF5 and IGF1R form a protein complex (also including EGFR) in a TM4SF5-dependent manner; co-expression promotes ERK, Akt, and S6K signaling and residual EGFR activity after EGFR kinase inhibitor treatment, causing resistance to erlotinib and gefitinib.\",\n      \"method\": \"Co-IP (TM4SF5/IGF1R/EGFR complex), ectopic TM4SF5 expression, IGF1R siRNA knockdown, EGFR kinase inhibitor treatment, 2D/3D culture drug sensitivity assays\",\n      \"journal\": \"Lung cancer (Amsterdam, Netherlands)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — Co-IP with functional epistasis, single lab\",\n      \"pmids\": [\"26190015\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"TM4SF5 physically associates with EGFR and integrin α5 at the leading edge of migratory cells (visualized by live fluorescence cross-correlation spectroscopy); cholesterol depletion and disruption of TM4SF5 N-glycosylation or palmitoylation alter these interactions and reduce cell migration speed and directionality in 2D and 3D conditions.\",\n      \"method\": \"Live fluorescence cross-correlation spectroscopy (FCS), super-resolution microscopy, cholesterol depletion (methyl-β-cyclodextrin), N-glycosylation and palmitoylation mutagenesis, 2D/3D migration assays\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — single-molecule live imaging plus mutagenesis, multiple orthogonal methods in a single study\",\n      \"pmids\": [\"28073834\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"CD133 phosphorylation induces TM4SF5 expression; TM4SF5 binds CD133 and promotes c-Src activity for CD133 phosphorylation (positive feedback); TM4SF5 also binds PTPRF and promotes paxillin phosphorylation; sphere growth decreased by CD133 suppression was recovered by TM4SF5 expression and partially by PTPRF suppression.\",\n      \"method\": \"Co-IP (TM4SF5-CD133, TM4SF5-PTPRF), siRNA knockdown epistasis, paxillin phosphorylation assay, 3D sphere growth assay\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — multiple Co-IPs with functional epistasis, single lab\",\n      \"pmids\": [\"30217560\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"TM4SF5 induces the alternatively spliced CD44v8-10 variant through an inverse ZEB2/ESRP linkage; TM4SF5 forms complexes with the cystine/glutamate antiporter system (xCT) via TM4SF5- and CD44v8-10-dependent CD98hc plasma membrane enrichment; dynamic TM4SF5 binding to CD98hc required CD44v8-10 under ROS-generating conditions; this complex upregulates cystine/glutamate antiporter activity and intracellular glutathione for ROS modulation and cell survival. Tm4sf5-null mice showed attenuated bleomycin-induced pulmonary fibrosis.\",\n      \"method\": \"Co-IP (TM4SF5-CD98hc, TM4SF5-CD44v8-10), alternative splicing analysis (RT-PCR), ZEB2/ESRP expression analysis, glutathione assay, xCT activity assay, Tm4sf5 KO mouse bleomycin model\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple Co-IPs, in vivo KO model, functional metabolic readout (glutathione/ROS), single lab with orthogonal methods\",\n      \"pmids\": [\"31501417\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"TM4SF5 forms protein-protein complexes with amino acid transporters including xCT (cystine/glutamate antiporter) and regulates cystine uptake from extracellular space and arginine export from lysosomal lumen to cytosol; diverse amino acid transporters co-precipitate with TM4SF5 by proteomic analysis.\",\n      \"method\": \"Co-IP, proximity-based proteomics, amino acid transport assays\",\n      \"journal\": \"Experimental & molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — review consolidating prior Co-IP findings with some new proteomic data, single lab\",\n      \"pmids\": [\"31956272\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"TM4SF5-overexpressing mice develop age-dependent nonalcoholic steatosis and NASH; in young mice TM4SF5 decreases SIRT1, increases SREBPs, and inactivates STAT3 via SOCS1/3 upregulation; in older mice TM4SF5 promotes SIRT1 expression and STAT3 activity for ECM production; CCL20 suppression reduced immune cell infiltration and ECM production; active STAT3 increases collagen I and laminin γ2, which in turn support SIRT1/STAT3 activity.\",\n      \"method\": \"TM4SF5 transgenic and KO mouse models, diet/chemical-treated mice, primary hepatocyte culture, CCL20 suppression, collagen I/laminin γ2 knockdown, human tissue analysis\",\n      \"journal\": \"The Journal of pathology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — transgenic and KO in vivo models with multiple genetic interventions, validated in human tissue\",\n      \"pmids\": [\"32918742\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TM4SF5 induction in differentiated macrophages promotes glucose uptake, glycolysis, and M1-type macrophage activation; activated M1 macrophages secrete IL-6, which induces CCL20 and CXCL10 secretion from TM4SF5-positive hepatocytes; chronic exposure to these chemokines reprograms macrophages toward M2-type, supporting NAFLD progression.\",\n      \"method\": \"TM4SF5 overexpression in macrophages, glycolysis assay, cytokine ELISA, co-culture systems, macrophage polarization assays, IL-6 neutralization\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple functional assays establishing intercellular signaling pathway, single lab\",\n      \"pmids\": [\"34788612\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TM4SF5 expression in cancer cells downregulates stimulatory NK cell ligands and receptors (SLAMF6, SLAMF7, MICA/B), causing NK cell exhaustion-like phenotypes; TM4SF5 suppression or inhibition with TSAHC reduced STAT3 signaling, recovered NK cell receptor levels and NK cell surveillance, and reduced liver cancer progression.\",\n      \"method\": \"TM4SF5 transgenic and DEN-induced liver cancer mouse models, TSAHC inhibitor treatment, NK cell activity assays, flow cytometry for NK ligand/receptor expression, STAT3 modulation\",\n      \"journal\": \"Cellular and molecular life sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vivo models with mechanistic follow-up, single lab\",\n      \"pmids\": [\"34921636\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"The TM4SF5 C-terminus binds the c-Src SH1 kinase domain (preferentially in its inactive closed form) along with PTP1B which dephosphorylates Tyr530; the SH1 domain alone bound TM4SF5 to cause c-Src Tyr419 and FAK Y861 phosphorylation; cell-penetrating TM4SF5 C-terminal peptides blocked TM4SF5-c-Src interaction and prevented tumor initiation/progression in vivo.\",\n      \"method\": \"Co-IP (domain mapping: C-terminus vs. SH1), homology modeling, molecular dynamics simulation, mutagenesis validation, cell-penetrating peptide competition, in vivo xenograft model\",\n      \"journal\": \"Theranostics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — structural modeling validated by mutagenesis and peptide competition, in vivo functional validation\",\n      \"pmids\": [\"34335982\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TM4SF5 intracellular vesicle traffic toward the leading edge is controlled by cell adhesion to fibronectin and microtubule acetylation; TM4SF5 palmitoylation is required for directed traffic; TM4SF5 forms a trimeric complex with HDAC6 and SLAC2B at perinuclear cytosol; SLAC2B suppression allows acetylated microtubules to extend to leading edges, facilitating TM4SF5 translocation and persistent migration; HDAC6 inhibition (via paxillin at new adhesion sites) promotes TM4SF5 traffic.\",\n      \"method\": \"Live-cell imaging of TM4SF5 vesicle tracking, palmitoylation-deficient mutant, SLAC2B siRNA, HDAC6 inhibition, Co-IP (TM4SF5-HDAC6-SLAC2B trimeric complex), immunofluorescence of acetylated tubulin\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — live imaging with mutagenesis and Co-IP, single lab\",\n      \"pmids\": [\"33554392\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TM4SF5 functions as a lysosomal arginine sensor and activates mTORC1; TM4SF5 KO in adipocytes reduces mTORC1 activation, enhances autophagy and lipolysis, increases PPARα and mitochondrial oxidative metabolism gene expression, reduces adiposity, and prevents HFD-induced glucose intolerance.\",\n      \"method\": \"TM4SF5 KO mouse model, mTORC1 activity assays, autophagy assays (LC3 flux), lipolysis assay, RNA sequencing of adipose tissue, metabolic phenotyping\",\n      \"journal\": \"Diabetes\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — KO mouse with multiple metabolic readouts, single lab\",\n      \"pmids\": [\"34187836\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Hepatic TM4SF5 binds GLUT1 at the plasma membrane to promote glucose uptake and glycolysis; excessive glucose causes hepatocytes to secrete TM4SF5-loaded small extracellular vesicles (sEVs); liver-derived sEVs containing TM4SF5 target brown adipose tissue (BAT) to improve glucose clearance independent of UCP1.\",\n      \"method\": \"Co-IP (TM4SF5-GLUT1), glucose uptake assay, sEV isolation and characterization, liver-closed vein circuit (LCVC) in vivo delivery of sEVs from TM4SF5-overexpressing mice, glucose tolerance tests in KO mice, BAT targeting assay\",\n      \"journal\": \"Journal of extracellular vesicles\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP with functional metabolic readouts and in vivo sEV delivery experiment, single lab\",\n      \"pmids\": [\"36063136\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"TM4SF5 binds GLUT8 at the plasma membrane; following fructose treatment, TM4SF5-GLUT8 binding transiently decreases, allowing GLUT8 translocation to the plasma membrane for fructose uptake and de novo lipogenesis; Tm4sf5 suppression or KO reduced fructose uptake, DNL, and steatosis in vivo.\",\n      \"method\": \"Co-IP (TM4SF5-GLUT8), GLUT8 localization by immunofluorescence (translocation assay), fructose uptake assay, DNL measurement, Tm4sf5 KO mouse with high-sucrose/fructose diet\",\n      \"journal\": \"Molecular metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP with dynamic localization and in vivo KO metabolic phenotype, single lab\",\n      \"pmids\": [\"35123128\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Upon glucose repletion following depletion, TM4SF5 becomes enriched at mitochondria-lysosome contact sites (MLCSs) via interaction between mitochondrial FKBP8 and lysosomal TM4SF5; proximity labeling revealed clustering of phospho-DRP1 and mitophagy receptors at TM4SF5-enriched MLCSs, promoting mitochondrial fission and autophagy; TM4SF5 binds NPC1 and free cholesterol, mediating cholesterol export from lysosomes to mitochondria and impairing oxidative phosphorylation.\",\n      \"method\": \"Co-IP (TM4SF5-FKBP8, TM4SF5-NPC1), proximity-labeling proteomics (BioID), organelle reconstitution, cholesterol transport assay, mitophagy assay, DRP1 phosphorylation analysis, in vivo mouse hepatocyte models\",\n      \"journal\": \"Cancer communications (London, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — proximity labeling proteomics plus Co-IP, organelle reconstitution, cholesterol transport assay, in vivo validation\",\n      \"pmids\": [\"38133457\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"TM4SF5 suppression in zebrafish impairs trunk muscle development, aberrant muscle fibre morphology, and alters integrin α5 expression; integrin α5-related signaling molecules (fibronectin, FAK, vinculin, actin) are aberrantly localized in tm4sf5 morphants; aberrant muscle development was rescued by injection of tm4sf5 or integrin α5 mRNA, establishing TM4SF5 function in muscle differentiation via integrin α5-dependent signaling.\",\n      \"method\": \"Morpholino knockdown (zebrafish), mRNA rescue injection, immunofluorescence of muscle and signaling molecules, C2C12 mouse myoblast differentiation assay\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — morpholino KD with mRNA rescue in zebrafish, supported by mammalian cell data, single lab\",\n      \"pmids\": [\"24897542\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TM4SF5 expressed by hepatocytes reduces NK cell cytotoxicity by binding SLAMF7 in an N-glycosylation-dependent manner, causing intracellular trafficking of SLAMF7 from the plasma membrane to lysosomes for degradation; TM4SF5-specific isoxazole (TSI) compounds block this binding and trafficking, restoring NK cell surveillance and reducing HCC development in xenograft models.\",\n      \"method\": \"Co-IP (TM4SF5-SLAMF7, N-glycosylation mutagenesis), immunofluorescence tracking of SLAMF7 trafficking to lysosomes, TSI small molecule treatment, NK cell cytotoxicity assay, mouse xenograft and Tm4sf5-KO models\",\n      \"journal\": \"Signal transduction and targeted therapy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — direct binding with domain mutagenesis, subcellular trafficking demonstrated by imaging, functional NK cell assay, in vivo validation\",\n      \"pmids\": [\"39828766\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TM4SF5-mediated macropinocytosis of albumin requires cytosolic stabilization of NCOA3 and PTEN inactivation through TM4SF5 binding; albumin uptake via macropinocytosis supports ATP-linked respiration and cellular migration in TM4SF5-expressing hepatocytes.\",\n      \"method\": \"Co-IP (TM4SF5-NCOA3, TM4SF5-PTEN), NCOA3 and PTEN expression/activity assays, macropinocytosis assay, albumin uptake assay, ATP-linked respiration (Seahorse), TM4SF5 KO and reintroduction, in vivo orthotopic mouse model\",\n      \"journal\": \"Experimental & molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP with functional metabolic and migration readouts, in vivo validation, single lab\",\n      \"pmids\": [\"40186033\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TM4SF5 modulates KEAP1 independently of NRF2: the cytosolic TM4SF5 C-terminus binds KEAP1 to promote its proteasomal degradation under physiological conditions; in hyperlipidemic/pathological states TM4SF5 stabilizes KEAP1, leading to oxidative stress and hepatic inflammation; Keap1 suppression nullified TM4SF5-mediated MASLD phenotypes.\",\n      \"method\": \"Co-IP (TM4SF5 C-terminus binding to KEAP1), proteasome inhibitor treatment, Tm4sf5 KO and Nrf2 mutant mouse models, Keap1 siRNA suppression, in vitro and in vivo MASLD models\",\n      \"journal\": \"International journal of biological sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct binding with domain identification, genetic epistasis (KO+KD), in vivo validation, single lab\",\n      \"pmids\": [\"41608638\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TM4SF5 forms an N-glycosylation-dependent dimer in its large extracellular loop (LEL); the LEL has a β-sheet configuration (unlike the α-helices of genuine tetraspanins or CD20-like family); TM4SF5 has two conserved cysteines (without the CCG motif) affecting N-glycosylation and dimer formation, and the LEL contributes to cholesterol binding.\",\n      \"method\": \"Structural analysis, sequence/domain comparison, N-glycosylation mutagenesis, cholesterol binding assay\",\n      \"journal\": \"Journal of advanced research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — structural characterization with mutagenesis, single lab/study\",\n      \"pmids\": [\"41349605\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"TM4SF5 is a four-transmembrane glycoprotein (L6 family, tetraspanin-like) that organizes signaling microdomains at the plasma membrane and on intracellular organelles, where it directly binds and activates FAK (by relieving its autoinhibition) and c-Src (via its C-terminus engaging the SH1 kinase domain), cooperates with integrins α2, α5, and β1, EGFR, CD44, IGF1R, and multiple amino acid/nutrient transporters to drive epithelial-mesenchymal transition, cell migration/invasion, self-renewal, metabolic reprogramming (glucose/fructose uptake, lipogenesis, mitochondrial cholesterol export at MLCS), and immune evasion (by trafficking SLAMF7 to lysosomes to suppress NK cell killing), while also functioning as a lysosomal arginine sensor that activates mTORC1 and modulates KEAP1 stability independently of NRF2.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"TM4SF5 is a four-transmembrane L6-family glycoprotein that organizes plasma membrane and intracellular signaling microdomains to coordinate integrin/growth factor receptor signaling, metabolic reprogramming, and immune evasion. Its second extracellular loop engages integrins α2 and α5, CD44, and EGFR in N-glycosylation- and cholesterol-dependent complexes, while its cytoplasmic C-terminus directly binds and activates FAK (by relieving autoinhibition) and c-Src (via the SH1 kinase domain), driving epithelial–mesenchymal transition, invadopodia formation, and metastasis [PMID:18357344, PMID:23077174, PMID:34335982, PMID:28073834]. On intracellular membranes, TM4SF5 functions as a lysosomal arginine sensor activating mTORC1, partners with nutrient transporters (GLUT1, GLUT8, xCT/CD98hc) to regulate glucose/fructose uptake and de novo lipogenesis, and localizes to mitochondria–lysosome contact sites where it binds FKBP8 and NPC1 to mediate cholesterol export and regulate mitochondrial fission [PMID:34187836, PMID:36063136, PMID:35123128, PMID:38133457]. TM4SF5 also promotes immune evasion by trafficking SLAMF7 to lysosomes for degradation, suppressing NK cell cytotoxicity, and by modulating macrophage polarization and chemokine secretion to facilitate NAFLD/NASH and hepatocellular carcinoma progression [PMID:39828766, PMID:34788612, PMID:32918742].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Establishing TM4SF5 as a membrane organizer of integrin–FAK signaling: it was unknown how TM4SF5 influenced adhesion signaling; Co-IP showed TM4SF5 associates with integrin α2 and controls FAK Y925/paxillin Y118 phosphorylation in a serum-dependent manner, demonstrating TM4SF5 as a regulator of focal contact dynamics.\",\n      \"evidence\": \"Co-immunoprecipitation, integrin-blocking antibody, FAK Y925F mutagenesis, migration assays in Cos7 cells\",\n      \"pmids\": [\"16828471\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry and directness of TM4SF5–integrin α2 binding not resolved\", \"Whether TM4SF5 binds FAK directly was unknown at this stage\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Defining the EMT/pro-tumorigenic axis: TM4SF5 was shown to stabilize cytosolic p27Kip1, inactivate RhoA, and suppress E-cadherin, driving EMT and multilayer growth in hepatocarcinoma; concurrently, TM4SF5 retained integrin α5 at the surface to activate c-Src/STAT3/VEGF for angiogenesis, establishing two parallel downstream pathways.\",\n      \"evidence\": \"shRNA knockdown, ectopic expression, nude mouse xenograft, HUVEC tube formation, E-cadherin reconstitution, anti-integrin α5 blockade\",\n      \"pmids\": [\"18357344\", \"19036703\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct physical contacts between TM4SF5 and p27Kip1 or RhoA not demonstrated\", \"Structural basis for integrin α5 retention was unresolved\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Mapping the interaction interface: the second extracellular loop (EL2) of TM4SF5 was identified as the integrin α2-binding domain, and N-glycosylation of TM4SF5 was shown to be required for protein–protein interactions and drug responsiveness, establishing the glycosylation-dependent nature of TM4SF5 function.\",\n      \"evidence\": \"EL2 peptide competition, site-directed mutagenesis of N-glycosylation sites, collagen I spreading/migration assays\",\n      \"pmids\": [\"19789264\", \"19177595\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Three-dimensional structure of EL2 was unknown\", \"Which specific glycan species mediate interaction was not determined\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Defining direct kinase activation mechanisms: TM4SF5 was shown to directly bind FAK, relieving its autoinhibitory intramolecular fold, and the TM4SF5 C-terminus was mapped as the c-Src/EGFR Y845 signaling platform, resolving the molecular basis of two key downstream kinase activations.\",\n      \"evidence\": \"Co-IP with domain mapping, FAK mutagenesis, EGFR Y845F mutagenesis, C-terminal deletion mutant, in vivo metastasis model\",\n      \"pmids\": [\"23077174\", \"23220047\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Crystal/cryo-EM structure of TM4SF5–FAK or TM4SF5–c-Src complex not available\", \"Whether FAK and c-Src bind TM4SF5 simultaneously was not tested\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Placing TM4SF5 induction within TGFβ signaling: TGFβ1/Smad activation was shown to induce TM4SF5 expression via EGFR, establishing that TM4SF5 is a downstream effector of TGFβ-mediated EMT and explaining how hepatocytes acquire TM4SF5 expression.\",\n      \"evidence\": \"Smad overexpression, EGFR kinase inhibitor, Smad7 epistasis, normal and cancer hepatocyte comparison\",\n      \"pmids\": [\"22292774\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct Smad-binding site in TM4SF5 promoter not mapped\", \"Contribution of other EMT-inducing signals to TM4SF5 induction not tested\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Expanding the tetraspanin-web and developmental roles: TM4SF5 was shown to control CD151/CD63 sorting (internalizing CD63 to lysosomes), and zebrafish morpholino studies demonstrated a requirement for TM4SF5 in integrin α5-dependent muscle development, broadening TM4SF5 function beyond cancer.\",\n      \"evidence\": \"Co-IP for TM4SF5–CD151, CD63 subcellular fractionation, zebrafish morpholino knockdown with mRNA rescue, C2C12 differentiation\",\n      \"pmids\": [\"25033048\", \"24897542\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether CD63 internalization is direct or requires adaptor proteins was unknown\", \"Zebrafish morpholino studies lack genetic knockout confirmation\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Establishing TM4SF5 as a self-renewal/stemness driver: TM4SF5 was found to bind CD44 through N-glycosylation-dependent extracellular domain interactions, activating c-Src/STAT3/Twist1/Bmi1 for spheroid formation, and co-expressed with IGF1R/EGFR to form a complex conferring EGFR-inhibitor resistance.\",\n      \"evidence\": \"Co-IP with domain mapping, N-glycosylation mutagenesis, pathway epistasis, orthotopic liver model with CTC detection, EGFR inhibitor sensitivity assays\",\n      \"pmids\": [\"25627085\", \"26190015\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative contribution of CD44 vs. IGF1R complexes to stemness not dissected\", \"Whether TM4SF5/CD44/IGF1R/EGFR exist in a single supercomplex or separate pools was unclear\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Visualizing TM4SF5 microdomain dynamics at the leading edge: live single-molecule cross-correlation spectroscopy confirmed TM4SF5 co-diffuses with EGFR and integrin α5 at the plasma membrane in a cholesterol- and palmitoylation-dependent manner, providing biophysical evidence for tetraspanin-enriched microdomain organization.\",\n      \"evidence\": \"Live fluorescence cross-correlation spectroscopy, super-resolution microscopy, cholesterol depletion, palmitoylation mutagenesis, 2D/3D migration\",\n      \"pmids\": [\"28073834\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Lipid composition requirements beyond cholesterol not characterized\", \"Whether palmitoylation directly mediates partner binding or affects membrane partitioning was not distinguished\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Revealing metabolic/transporter functions: TM4SF5 was found to complex with the xCT/CD98hc cystine–glutamate antiporter via CD44v8-10 splice-variant induction, regulating glutathione homeostasis and ROS defense; Tm4sf5-KO mice showed attenuated pulmonary fibrosis, linking TM4SF5 to metabolic redox control in vivo.\",\n      \"evidence\": \"Co-IP of TM4SF5–CD98hc/xCT, glutathione assay, alternative splicing analysis, Tm4sf5-KO mouse bleomycin model\",\n      \"pmids\": [\"31501417\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether TM4SF5 directly transports any metabolite or acts purely as a scaffolding organizer was unresolved\", \"Tissue-specificity of the xCT complex function beyond lung not tested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Defining TM4SF5 as a lysosomal nutrient sensor and intracellular trafficking hub: TM4SF5 was identified as a lysosomal arginine sensor activating mTORC1 in adipocytes (KO reduced adiposity and improved glucose tolerance); simultaneously, the C-terminus/SH1-domain interaction with c-Src was structurally modeled and validated, and TM4SF5 vesicle trafficking was shown to depend on a TM4SF5/HDAC6/SLAC2B complex riding acetylated microtubules.\",\n      \"evidence\": \"mTORC1 activity and autophagy assays in KO mice, metabolic phenotyping, Co-IP domain mapping of c-Src SH1, cell-penetrating peptide competition, live vesicle tracking, HDAC6/SLAC2B Co-IP\",\n      \"pmids\": [\"34187836\", \"34335982\", \"33554392\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"The arginine-sensing mechanism (direct binding vs. indirect) not biochemically resolved\", \"Whether HDAC6/SLAC2B complex exists on lysosomal vs. plasma-membrane-directed vesicles was unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Establishing immune evasion functions: TM4SF5 in hepatocytes suppressed NK cell ligands (SLAMF6/7, MICA/B) and promoted macrophage M1-to-M2 reprogramming via IL-6/CCL20 secretion, linking TM4SF5 to both innate immune evasion and NAFLD progression.\",\n      \"evidence\": \"Tm4sf5 transgenic and DEN-induced liver cancer models, NK cell cytotoxicity assays, macrophage polarization co-culture, IL-6 neutralization, TSAHC inhibitor\",\n      \"pmids\": [\"34921636\", \"34788612\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct mechanism by which TM4SF5 downregulates NK ligands (transcriptional vs. post-translational) was not fully resolved\", \"In vivo contribution of macrophage vs. NK evasion to tumor progression not separated\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Expanding sugar transporter partnerships: TM4SF5 was shown to bind GLUT1 for glucose uptake and GLUT8 for fructose uptake/de novo lipogenesis; hepatocyte-derived TM4SF5-containing extracellular vesicles targeted brown adipose tissue to improve systemic glucose clearance, revealing an inter-organ signaling role.\",\n      \"evidence\": \"Co-IP of TM4SF5–GLUT1 and TM4SF5–GLUT8, glucose/fructose uptake assays, sEV isolation and in vivo delivery via liver-closed vein circuit, Tm4sf5-KO mice on high-sucrose diets\",\n      \"pmids\": [\"36063136\", \"35123128\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether TM4SF5 in sEVs retains functional transporter-scaffolding activity in recipient cells was not shown\", \"The GLUT8 release mechanism upon fructose stimulation was not molecularly defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defining mitochondria–lysosome contact site (MLCS) functions: TM4SF5 was shown to enrich at MLCSs via binding lysosomal TM4SF5 to mitochondrial FKBP8; proximity labeling revealed DRP1 and mitophagy receptor clustering, and TM4SF5 mediated NPC1-dependent cholesterol transfer from lysosomes to mitochondria, impairing oxidative phosphorylation.\",\n      \"evidence\": \"Co-IP of TM4SF5–FKBP8 and TM4SF5–NPC1, BioID proximity proteomics, cholesterol transport reconstitution, mitophagy assay, DRP1 phosphorylation, in vivo mouse models\",\n      \"pmids\": [\"38133457\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether MLCS enrichment requires mTORC1 activity (linking to the arginine-sensing role) was not tested\", \"Structural basis of the TM4SF5–FKBP8 interaction unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Resolving the NK-evasion mechanism and structural features: TM4SF5 was shown to directly bind SLAMF7 in an N-glycosylation-dependent manner and traffic it to lysosomes for degradation, suppressing NK cytotoxicity; the LEL was structurally characterized as a β-sheet fold (distinct from genuine tetraspanins) that forms N-glycosylation-dependent dimers and binds cholesterol; TM4SF5 also modulates KEAP1 stability independently of NRF2.\",\n      \"evidence\": \"Co-IP with N-glycosylation mutagenesis, SLAMF7 trafficking imaging, TSI inhibitor treatment, xenograft models, structural/domain analysis, KEAP1 proteasomal degradation assay, Tm4sf5-KO and Nrf2-mutant mice\",\n      \"pmids\": [\"39828766\", \"41349605\", \"41608638\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"High-resolution structure of the TM4SF5 LEL dimer still lacking\", \"Mechanism by which pathological lipid conditions switch TM4SF5 from KEAP1 degradation to stabilization is not defined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"A high-resolution structure of TM4SF5 (alone and in complex with key partners such as FAK, c-Src, integrins, or SLAMF7) is needed to understand the allosteric mechanisms by which it activates kinases and organizes multi-protein complexes; the direct arginine-binding mechanism on lysosomes remains biochemically unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No crystal or cryo-EM structure available\", \"Arginine-sensing mechanism (direct binding site, affinity) not biochemically defined\", \"Relative physiological importance of individual TM4SF5 complexes in specific tissues unclear\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [1, 9, 10, 14, 16, 18]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [9, 23, 25, 32]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [28, 33]},\n      {\"term_id\": \"GO:0140299\", \"supporting_discovery_ids\": [25]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [1, 3, 16, 26, 27]},\n      {\"term_id\": \"GO:0005764\", \"supporting_discovery_ids\": [12, 25, 28, 30]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [28]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [24]},\n      {\"term_id\": \"GO:0005576\", \"supporting_discovery_ids\": [26]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 2, 5, 9, 10, 11, 14, 15, 23, 25]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [18, 19, 26, 27, 28]},\n      {\"term_id\": \"R-HSA-1500931\", \"supporting_discovery_ids\": [0, 8, 14, 16]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [21, 22, 30]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [25, 28]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [5]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [20, 27, 32]},\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [24, 26, 30]}\n    ],\n    \"complexes\": [\n      \"TM4SF5/integrin α5/EGFR leading-edge complex\",\n      \"TM4SF5/CD44v8-10/CD98hc/xCT transporter complex\",\n      \"TM4SF5/HDAC6/SLAC2B vesicle trafficking complex\",\n      \"TM4SF5/IGF1R/EGFR signaling complex\"\n    ],\n    \"partners\": [\n      \"PTK2\",\n      \"SRC\",\n      \"ITGA2\",\n      \"ITGA5\",\n      \"CD44\",\n      \"EGFR\",\n      \"FKBP8\",\n      \"SLAMF7\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}