Affinage

LYPD3

Ly6/PLAUR domain-containing protein 3 · UniProt O95274

Length
346 aa
Mass
36.0 kDa
Annotated
2026-06-10
34 papers in source corpus 15 papers cited in narrative 16 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 9/9 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

LYPD3 (C4.4A) is a GPI-anchored, two-domain Ly6/uPAR (LU)-module cell surface protein that functions as a pro-invasive, pro-metastatic scaffold coordinating laminin engagement, integrin signalling, and focalized matrix degradation (PMID:15729693, PMID:23727360). Crystallography shows its two LU domains pack face-to-face into a compact globule, atypical for the flexible LU-domain family, with the AGR2-binding site spanning both domains (PMID:32140067). It is heavily N- and O-glycosylated, and glycosylation governs its subcellular localization (PMID:15012588, PMID:39009906). Functionally, C4.4A binds the extracellular ligands laminin-1 and laminin-5 and drives spreading, lamellipodia formation, and migration on these substrates (PMID:15729693); despite sharing domain architecture with uPAR, it does not bind uPA, marking a distinct functional identity (PMID:15012588). At the cell surface it recruits activated alpha6beta4 integrin together with MT1-MMP (MMP14) into lipid rafts to enable localized laminin/matrix degradation and, via alpha6beta4-dependent BAD phosphorylation and Bcl2/BclXl upregulation, drug resistance; this complex persists in exosomes and shed C4.4A retains laminin-degrading activity (PMID:22431918, PMID:23727360). Extracellular AGR2 binds C4.4A directly to drive an autocrine signalling loop requiring laminin-1/5 and integrin beta1 that promotes proliferation, migration, invasion, and chemoresistance (PMID:25646014). C4.4A acts upstream of EMT, with knockdown reversing mesenchymal marker expression and reducing invasion across multiple carcinoma types (PMID:22404718, PMID:29048672). Genetic ablation in mice confirms a physiological role: knockout animals develop normally but show delayed keratinocyte wound-closure migration and reduced invasive bladder carcinoma (PMID:27169360). Its expression is controlled transcriptionally by C/EBPbeta cooperating with JunD/c-Jun (PMID:17278103) and post-transcriptionally by miR-151-5p targeting (PMID:39009906), and it is cleaved by ADAM10 and ADAM17 (PMID:18979631).

Mechanistic history

Synthesis pass · year-by-year structured walk · 14 steps
  1. 2003 Low

    Established the first candidate binding partners for C4.4A, nominating AGR2/AG-3 and alpha-dystroglycan as interactors and framing it as a partner-engaging surface protein.

    Evidence Yeast two-hybrid cloning

    PMID:12592373

    Open questions at the time
    • Single yeast two-hybrid screen without reciprocal Co-IP validation
    • Functional consequence of binding not addressed
    • alpha-dystroglycan interaction not pursued in later timeline entries
  2. 2004 Medium

    Defined C4.4A as a GPI-anchored, heavily glycosylated two-LU-module protein and showed it does NOT bind uPA, distinguishing it functionally from its structural homologue uPAR.

    Evidence Recombinant protein purification, glycosylation mass spectrometry, and ELISA binding with soluble and GPI-anchored C4.4A

    PMID:15012588

    Open questions at the time
    • Did not identify the functional ligand of C4.4A
    • Role of the protease-sensitive Tyr200-Arg204 region not tested functionally
  3. 2005 Medium

    Identified laminin-1 and laminin-5 as bona fide extracellular ligands and linked C4.4A to cell spreading and migration on laminin, giving the protein a concrete adhesion/motility function.

    Evidence ELISA ECM screening, cDNA transfection, migration/spreading assays, and galectin-3 Co-IP

    PMID:15729693

    Open questions at the time
    • Mechanism by which laminin binding drives migration not resolved
    • Role of galectin-3 association not mechanistically detailed
  4. 2007 Medium

    Resolved how C4.4A transcription is controlled, showing C/EBPbeta binding to a TRE/CCAAT element cooperating with JunD/c-Jun drives expression.

    Evidence Reporter deletion/mutation constructs, EMSA, co-transfection, and ChIP

    PMID:17278103

    Open questions at the time
    • Upstream signals controlling C/EBPbeta engagement not defined
    • Single-lab promoter analysis
  5. 2008 Medium

    Identified ADAM10 and ADAM17 as the proteases that cleave C4.4A, providing a mechanism for its shedding during invasion.

    Evidence SILAC proteomics with shRNA knockdown of ADAM10/ADAM17 in MCF7 cells

    PMID:18979631

    Open questions at the time
    • Cleavage site and fate of shed fragment not mapped
    • Functional consequence of shedding inferred rather than directly tested
  6. 2012 Medium

    Placed C4.4A in a lipid-raft complex with alpha6beta4 integrin, MT1-MMP, and TACE that switches cells from laminin adhesion to motility under hypoxia and degrades laminin, including from exosomes.

    Evidence Reciprocal Co-IP, wound-healing assays, exosome isolation, and CoCl2 hypoxia model with knockdown

    PMID:22431918

    Open questions at the time
    • Stoichiometry and assembly order of the raft complex unknown
    • Single-lab evidence
  7. 2012 Medium

    Positioned C4.4A upstream of EMT, showing knockdown reverses mesenchymal marker expression and reduces invasion without affecting proliferation.

    Evidence siRNA knockdown with EMT marker Western blots and invasion assays in HCT116 cells

    PMID:22404718

    Open questions at the time
    • Signalling link between C4.4A and EMT transcriptional program not defined
    • Single cell line
  8. 2013 Medium

    Demonstrated in vivo that C4.4A recruits activated alpha6beta4 integrin into rafts to drive focalized matrix degradation and metastasis, while alpha6beta4-BAD-Bcl2 signalling confers drug resistance.

    Evidence Stable shRNA knockdown in ASML rat carcinoma cells, intrafootpad metastasis model, raft fractionation, and PI3K/Akt pathway analysis

    PMID:23727360

    Open questions at the time
    • Direct vs. indirect nature of C4.4A-integrin coupling not fully resolved
    • Single in vivo model system
  9. 2015 High

    Confirmed a direct AGR2-C4.4A interaction and defined an autocrine signalling loop requiring laminins and integrin beta1 that drives PDAC proliferation, invasion, and chemoresistance.

    Evidence Co-IP from lysates and recombinant protein mixing, siRNA knockdown, Boyden chamber and apoptosis assays, and orthotopic xenografts with blocking antibodies

    PMID:25646014

    Open questions at the time
    • Downstream effectors of AGR2-C4.4A signalling not fully mapped
    • How the loop integrates with the alpha6beta4/MMP14 raft complex unclear
  10. 2016 High

    Provided definitive genetic loss-of-function evidence that C4.4A promotes keratinocyte migration and tumor invasion in vivo.

    Evidence Constitutive C4.4A knockout mice in incisional wound-healing and chemical bladder carcinogenesis models

    PMID:27169360

    Open questions at the time
    • Molecular basis of the wound-healing migration defect not dissected in vivo
    • Normal development indicates functional redundancy not explored
  11. 2017 Medium

    Generalized the pro-migratory/invasive function to hepatocellular carcinoma and validated C4.4A as an internalizing target amenable to antibody-drug conjugate therapy.

    Evidence siRNA knockdown with migration/invasion/proliferation assays in Huh7/HepG2, and ADC (BAY 1129980) efficacy in transfected cells and xenograft/PDX models

    PMID:28292941 PMID:29048672

    Open questions at the time
    • Internalization route and recycling not characterized
    • Single-lab functional studies
  12. 2020 High

    Solved the structure of the two LU domains, revealing an atypical compact face-to-face arrangement and mapping the AGR2-binding/antibody epitope spanning both domains.

    Evidence X-ray crystallography of C4.4A D1D2 alone and in complex with an anti-C4.4A Fab

    PMID:32140067

    Open questions at the time
    • Structure of full-length glycosylated C4.4A and its complexes with laminin/integrin not solved
    • Conformational dynamics in membrane context unknown
  13. 2020 Medium

    Placed LYPD3 downstream of beta2-adrenoceptor signalling as a pro-migratory effector in breast cancer.

    Evidence Mass spectrometry protein profiling, siRNA knockdown, and migration assays with adrenoceptor agonist/antagonist treatment in MCF-7 cells

    PMID:32098331

    Open questions at the time
    • Signalling intermediates linking ADRbeta2 to LYPD3 induction not defined
    • Single cell line
  14. 2024 Medium

    Established post-transcriptional control of LYPD3 by miR-151-5p (facilitated by METTL3 m6A and hnRNP U) and linked glycosylation to its subcellular localization and a metastasis-suppressive role in HNSCC.

    Evidence 3'-UTR luciferase reporters, m6A analysis, glycosylation subcellular fractionation, and miR/knockdown functional assays

    PMID:39009906

    Open questions at the time
    • Apparent context-dependent metastasis suppression contrasts with pro-invasive role in other tissues and is unresolved
    • Glycoform-specific localization mechanism not detailed

Open questions

Synthesis pass · forward-looking unresolved questions
  • How the AGR2 autocrine loop, the alpha6beta4/MMP14 raft degradation complex, and the context-dependent pro- versus anti-metastatic roles of LYPD3 are mechanistically integrated remains unresolved.
  • No unified model linking signalling input, complex assembly, and tissue-specific outcome
  • Structure of full-length protein with physiological ligands absent
  • Direct catalytic activity of LYPD3 itself not established

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0060090 molecular adaptor activity 3 GO:0008092 cytoskeletal protein binding 1
Localization
GO:0005886 plasma membrane 3 GO:0031410 cytoplasmic vesicle 1
Pathway
R-HSA-1474244 Extracellular matrix organization 2 R-HSA-162582 Signal Transduction 2 R-HSA-1643685 Disease 2
Partners
ADAM10ADAM17AGR2ITGA6ITGB1ITGB4LGALS3MMP14
Complex memberships
C4.4A-alpha6beta4 integrin-MT1-MMP/TACE lipid raft complex

Evidence

Reading pass · 16 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2003 C4.4A (LYPD3) was identified as a binding partner for hAG-2 (AGR2) and hAG-3 by yeast two-hybrid cloning, and also binds extracellular alpha-dystroglycan (DAG-1). Yeast two-hybrid cloning British journal of cancer Low 12592373
2004 Human C4.4A is a GPI-anchored protein containing two Ly6/uPAR/alpha-neurotoxin modules, extensively modified by 5-6 N-linked and ~15 O-linked carbohydrates; a protease-sensitive region (Tyr200-Arg204) lies between the N- and O-glycosylation clusters. Recombinant soluble C4.4A and GPI-anchored C4.4A from amnion membranes showed no detectable interaction with uPA, distinguishing C4.4A functionally from uPAR. Recombinant protein expression/purification, mass spectrometry characterization of glycosylation, ELISA-based binding assay, immunohistochemistry The Biochemical journal Medium 15012588
2004 C4.4A does not interact with urokinase-type plasminogen activator (uPA), indicating it lacks functional overlap with its structural homologue uPAR despite shared domain architecture. ELISA-based binding assay with recombinant soluble C4.4A and MCF7 cells expressing GPI-anchored C4.4A The Biochemical journal Medium 15012588
2005 C4.4A binds laminin-1 (LN1) and laminin-5 (LN5) as extracellular ligands; C4.4A-expressing tumor cells show increased spreading, lamellipodia formation, and migration on LN5 and LN1. C4.4A also associates with galectin-3, which influences laminin adhesion. ELISA screening of ECM components with recombinant rat C4.4A, cDNA transfection of BSp73AS cells, cell migration/spreading assays, co-immunoprecipitation for galectin-3 association International journal of cancer Medium 15729693
2007 C4.4A transcription requires C/EBPbeta binding to a TRE/CCAAT composite element (-71 to -88 bp) in the promoter, is enhanced by JunD or c-Jun co-transfection, and the TATA-less GC-rich core promoter alone is insufficient; Sp3 but not Sp1 binding sites are functional. Reporter construct deletions/point mutations, mobility-shift assays (EMSA), co-transfection studies, chromatin immunoprecipitation (ChIP) International journal of cancer Medium 17278103
2008 C4.4A is a substrate for both ADAM10 and ADAM17 metalloproteases, identified by SILAC-based proteomics in MCF7 cells with shRNA knockdown of ADAM10 or ADAM17; cleavage by these proteases likely contributes to tumor invasion. SILAC proteomics, shRNA knockdown of ADAM10/ADAM17 in MCF7 cells Biological chemistry Medium 18979631
2012 C4.4A associates with α6β4 integrin and MT1-MMP (MMP14) as well as TACE in lipid rafts; hypoxia promotes this association and drives a shift from laminin adhesion to cell motility, accompanied by laminin fragmentation. This complex is maintained in exosomes, and shed C4.4A retains laminin-degrading activity. Co-immunoprecipitation, in vitro wound healing assays, exosome isolation, CoCl2-induced hypoxia model, siRNA/knockdown experiments Neoplasia Medium 22431918
2012 C4.4A knockdown in colorectal cancer cells reduces cell invasion (but not proliferation) and correlates with EMT reversal, including increased E-cadherin and decreased vimentin and N-cadherin, placing C4.4A upstream of EMT regulation. siRNA knockdown, invasion assays, Western blot for EMT markers (E-cadherin, vimentin, N-cadherin) in HCT116 cells Cancer science Medium 22404718
2013 C4.4A promotes metastasis by recruiting activated α6β4 integrin into lipid rafts, where C4.4A cooperates with α6β4 and MMP14 (MT1-MMP) to enable focalized matrix degradation; α6β4 also promotes BAD phosphorylation and upregulation of Bcl2/BclXl, mediating drug resistance. C4.4A knockdown in ASML rat pancreatic carcinoma cells strongly reduces metastasis and sensitizes to cisplatin. Stable shRNA knockdown of C4.4A in ASML cells, intrafootpad metastasis model in rats, co-immunoprecipitation, raft fractionation, PI3K/Akt pathway analysis Molecular oncology Medium 23727360
2015 Extracellular AGR2 binds C4.4A directly (co-immunoprecipitation from cell lysates and with recombinant proteins); AGR2 signaling through C4.4A requires laminins 1 or 5 and integrin β1 to stimulate PDAC cell proliferation, migration, invasion, and chemoresistance. C4.4A knockdown reduces migration and gemcitabine resistance. Co-immunoprecipitation from cell lysates, recombinant protein mixing assay, siRNA knockdown, Boyden chamber assays, FACS apoptosis assay, orthotopic xenograft tumor model with blocking monoclonal antibodies Molecular cancer therapeutics High 25646014
2016 C4.4A-deficient mice (gene ablation) develop normally with intact squamous epithelia but show delayed keratinocyte migration in wound closure (incisional skin wound model in male mice) and reduced incidence of invasive lesions in chemically induced bladder carcinomas, establishing C4.4A as a functional promoter of keratinocyte migration and tumor cell invasion in vivo. Constitutive C4.4A knockout mouse generation, in vivo wound-healing assay, chemical carcinogenesis bladder tumor model Scientific reports High 27169360
2017 C4.4A knockdown in hepatocellular carcinoma (Huh7 and HepG2) cells reduces migration and invasion but does not affect proliferation or apoptosis, confirming a specific pro-migratory/invasive function of C4.4A. siRNA knockdown, migration and invasion assays, proliferation and apoptosis assays Oncology reports Medium 29048672
2017 C4.4A (LYPD3) is an internalizing cell surface protein; an ADC targeting C4.4A (BAY 1129980) demonstrates selective antiproliferative activity in C4.4A-expressing cells and requires C4.4A surface expression for efficacy, confirming receptor-mediated internalization. ADC efficacy assay in C4.4A-transfected vs. non-transfected cells, xenograft and PDX tumor models Molecular cancer therapeutics Medium 28292941
2020 Crystal structures of the two LU domains of human C4.4A were solved (alone and in complex with a Fab fragment of a monoclonal anti-C4.4A antibody). The structure reveals that C4.4A forms a compact globule with both LU domains packed face-to-face, contrasting with the flexible arrangement typical of most LU-domain proteins. The Fab combining site spans both LU domains and overlaps with the AGR2-binding site. X-ray crystallography (crystal structures of C4.4A D1D2 domains and C4.4A–Fab complex) International journal of biological sciences High 32140067
2020 β2-adrenoceptor (ADRβ2) signalling upregulates LYPD3 protein expression in breast cancer cells; LYPD3 knockdown significantly reduces both basal and norepinephrine-induced migration activity of MCF-7 cells, placing LYPD3 downstream of β2-adrenoceptor signalling as a pro-migratory effector. Protein profiling (mass spectrometry), siRNA knockdown, migration assays, adrenoceptor agonist/antagonist treatment Biology Medium 32098331
2024 Glycosylation of LYPD3 modulates its subcellular localization and reinforces its role in suppressing HNSCC metastasis; miR-151-5p directly targets the 3'-UTR of LYPD3 mRNA to reduce LYPD3 expression, and this targeting is facilitated by METTL3-mediated m6A modification and hnRNP U-mediated miR-151-5p maturation. 3'-UTR luciferase reporter assays, m6A modification analysis, glycosylation experiments with subcellular fractionation/localization, miR-151-5p overexpression and LYPD3 knockdown functional assays Molecular biomedicine Medium 39009906

Source papers

Stage 0 corpus · 34 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2003 hAG-2 and hAG-3, human homologues of genes involved in differentiation, are associated with oestrogen receptor-positive breast tumours and interact with metastasis gene C4.4a and dystroglycan. British journal of cancer 158 12592373
2015 New Blocking Antibodies against Novel AGR2-C4.4A Pathway Reduce Growth and Metastasis of Pancreatic Tumors and Increase Survival in Mice. Molecular cancer therapeutics 59 25646014
2004 Structural analysis and tissue localization of human C4.4A: a protein homologue of the urokinase receptor. The Biochemical journal 55 15012588
2012 Membrane-bound and exosomal metastasis-associated C4.4A promotes migration by associating with the α(6)β(4) integrin and MT1-MMP. Neoplasia (New York, N.Y.) 46 22431918
2020 CircCBFB-mediated miR-28-5p facilitates abdominal aortic aneurysm via LYPD3 and GRIA4. Life sciences 42 32151690
2018 Long non-coding RNA OGFRP1 regulates LYPD3 expression by sponging miR-124-3p and promotes non-small cell lung cancer progression. Biochemical and biophysical research communications 42 30274775
2001 Cloning of the human homologue of the metastasis-associated rat C4.4A. Gene 39 11179665
2005 Ly6 family member C4.4A binds laminins 1 and 5, associates with galectin-3 and supports cell migration. International journal of cancer 36 15729693
2017 Preclinical Antitumor Efficacy of BAY 1129980-a Novel Auristatin-Based Anti-C4.4A (LYPD3) Antibody-Drug Conjugate for the Treatment of Non-Small Cell Lung Cancer. Molecular cancer therapeutics 34 28292941
2008 Altered expression of the urokinase receptor homologue, C4.4A, in invasive areas of human esophageal squamous cell carcinoma. International journal of cancer 34 17849475
2007 C4.4A as a candidate marker in the diagnosis of colorectal cancer. British journal of cancer 33 17912244
2020 β2-Adrenergic Signalling Promotes Cell Migration by Upregulating Expression of the Metastasis-Associated Molecule LYPD3. Biology 31 32098331
2008 Metastasis-associated C4.4A, a GPI-anchored protein cleaved by ADAM10 and ADAM17. Biological chemistry 31 18979631
2001 Upregulation of C4.4A expression during progression of melanoma. The Journal of investigative dermatology 29 11180013
2012 C4.4A is associated with tumor budding and epithelial-mesenchymal transition of colorectal cancer. Cancer science 25 22404718
2011 Expression of C4.4A, a structural uPAR homolog, reflects squamous epithelial differentiation in the adult mouse and during embryogenesis. The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 18 21339181
2015 C4.4A as a biomarker of head and neck squamous cell carcinoma and correlated with epithelial mesenchymal transition. American journal of cancer research 17 26885441
2011 Expression of C4.4A in precursor lesions of pulmonary adenocarcinoma and squamous cell carcinoma. International journal of cancer 17 21792890
2024 m6A-dependent mature miR-151-5p accelerates the malignant process of HNSCC by targeting LYPD3. Molecular biomedicine 12 39009906
2013 The metastasis-associated molecule C4.4A promotes tissue invasion and anchorage independence by associating with the alpha6beta4 integrin. Molecular oncology 12 23727360
2007 CEBPbeta, JunD and c-Jun contribute to the transcriptional activation of the metastasis-associated C4.4A gene. International journal of cancer 12 17278103
2022 LYPD3, a New Biomarker and Therapeutic Target for Acute Myelogenous Leukemia. Frontiers in genetics 11 35360840
2016 C4.4A gene ablation is compatible with normal epidermal development and causes modest overt phenotypes. Scientific reports 11 27169360
2014 Expression of the Ly6/uPAR-domain proteins C4.4A and Haldisin in non-invasive and invasive skin lesions. The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 10 25414274
2023 LY6/PLAUR domain containing 3 (LYPD3) maintains melanoma cell stemness and mediates an immunosuppressive microenvironment. Biology direct 9 37924160
2014 C4.4A as a biomarker in pulmonary adenocarcinoma and squamous cell carcinoma. World journal of clinical oncology 7 25302166
2020 Crystal Structures of Human C4.4A Reveal the Unique Association of Ly6/uPAR/α-neurotoxin Domain. International journal of biological sciences 6 32140067
2012 Distinct expression of C4.4A in colorectal cancer detected by different antibodies. International journal of oncology 5 23175173
2006 [Expression and diagnostic application of C4.4A protein in squamous cell carcinoma and adenocarcinoma]. Zhonghua bing li xue za zhi = Chinese journal of pathology 4 16776998
2017 De novo synthesis of C4.4A in hepatocellular carcinoma promotes migration and invasion of tumor cells. Oncology reports 3 29048672
2017 Expression of C4.4A in an In Vitro Human Tissue-Engineered Skin Model. BioMed research international 2 29075641
2017 Expression and crystallographic studies of the D1D2 domains of C4.4A, a homologous protein to the urokinase receptor. Acta crystallographica. Section F, Structural biology communications 1 28777093
2026 Exploring the molecular function of LYPD3 from pan-cancer to lung cancer: based on bioinformatics and cellular experiments. Mammalian genome : official journal of the International Mammalian Genome Society 0 41553571
2016 [Corrigendum] Distinct expression of C4.4A in colorectal cancer detected by different antibodies. International journal of oncology 0 27634111

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