Evidence for a Role of the Epithelial Glycoprotein 40 (Ep-CAM) in Epithelial Cell-Cell Adhesion

Recently we have demonstrated that a 40kD human epithelium-specific glycoprotein exhibits the features of a homophilic cell-cell adhesion molecule, when expressed in transfected murine cells. We suggested the name Ep-CAM for this molecule (Litvinov et al., J. Cell Biol., 125: 437-446). Here we investigate the possible biological function of Ep-CAM in its natural environment—cells of epithelial origin. Immunolocalization of Ep-CAM in tissues and in cultures of epithelial/carcinoma cells showed that the majority of the Ep-CAM molecules are localized at cell-cell boundaries, predominantly along the whole lateral domain of polarized cells. In vitro, on single cells in suspension, the Ep-CAM molecules are present on the entire cell surface, and when the single cells grow attached, Ep-CAM is present at their pseudo-apical domain. During formation of intercellular contacts by such single cells, the majority of the Ep-CAM molecules are redistributed from the pseudoapical to the lateral domain of the cell membrane. Attachment of cells to the substrate does not cause redistribution of the molecules to the site of substrate attachment irrespective of the adhesive substrate (fibronectin, collagens, laminin, EHS-matrigel were tested). The monoclonal antibody 323/A3, reactive with the extracellular domain of the Ep-CAM molecule, has a strong negative effect on the aggregating behaviour of COV362 ovarian carcinoma cells and RC-6 immortalized mammary epithelial cells. The mAb affected cell aggregation in both cell lines in the presence of Ca⁺⁺, but with RC-6 cells the effect was more pronounced in low-calcium medium. The effects of the 323/A3 mAb on the already established intercellular contacts was not significant. The data presented demonstrate that the Ep-CAM molecules are functionally active in the epithelial and carcinoma cells tested, are capable of mediating Ca¹⁺-independent intercellular adhesions, and are not likely to be involved in cell-substrate adhesion.     

Evidence for a Role of the Epithelial Glycoprotein 40 (Ep-CAM) in Epithelial Cell-Cell Adhesion

Recently we have demonstrated that a 40kD human epithelium-specific glycoprotein exhibits the features of a homophilic cell-cell adhesion molecule, when expressed in transfected murine cells. We suggested the name Ep-CAM for this molecule (Litvinov et al., J. Cell Biol., 125: 437–446). Here we investigate the possible biological function of Ep-CAM in its natural environment—cells of epithelial origin. Immunolocalization of Ep-CAM in tissues and in cultures of epithelial/carcinoma cells showed that the majority of the Ep-CAM molecules are localized at cell-cell boundaries, predominantly along the whole lateral domain of polarized cells. In vitro, on single cells in suspension, the Ep-CAM molecules are present on the entire cell surface, and when the single cells grow attached, Ep-CAM is present at their pseudo-apical domain. During formation of intercellular contacts by such single cells, the majority of the Ep-CAM molecules are redistributed from the pseudoapical to the lateral domain of the cell membrane. Attachment of cells to the substrate does not cause redistribution of the molecules to the site of substrate attachment irrespective of the adhesive substrate (fibronectin, collagens, laminin, EHS-matrigel were tested). The monoclonal antibody 323/A3, reactive with the extracellular domain of the Ep-CAM molecule, has a strong negative effect on the aggregating behaviour of COV362 ovarian carcinoma cells and RC-6 immortalized mammary epithelial cells. The mAb affected cell aggregation in both cell lines in the presence of Ca⁺⁺, but with RC-6 cells the effect was more pronounced in low-calcium medium. The effects of the 323/A3 mAb on the already established intercellular contacts was not significant. The data presented demonstrate that the Ep-CAM molecules are functionally active in the epithelial and carcinoma cells tested, are capable of mediating Ca¹⁺-independent intercellular adhesions, and are not likely to be involved in cell-substrate adhesion.     

A role for the epidermal growth factor-like domain of P-selectin in ligand recognition and cell adhesion

The selectin family of adhesion molecules mediates the initial interactions of leukocytes with endothelium. The extracellular region of each selectin contains an amino-terminal C-type lectin domain, followed by an EGF-like domain and multiple short consensus repeat units (SCR). Previous studies have indirectly suggested a role for each of the extracellular domains of the selectins in cell adhesion. In this study, a panel of chimeric selectins created by exchange of domains between L- and P-selectin was used to directly examine the role of the extracellular domains in cell adhesion. Exchange of only the lectin domains between L- and P-selectin conferred the adhesive and ligand recognition functions of the lectin domain of the parent molecule. However, chimeric selectins which contained both the lectin domain of L- selectin and the EGF-like domain of P-selectin exhibited dual ligand- binding specificity. These chimeric proteins supported adhesion both to myeloid cells and to high endothelial venules (HEV) of lymph nodes and mesenteric venules in vivo. Exchange of the SCR domains had no detectable effect on receptor function or specificity. Thus, the EGF- like domain of P-selectin may play a direct role in ligand recognition and leukocyte adhesion mediated by P-selectin, with the lectin plus EGF- like domains collectively forming a functional ligand recognition unit.     

Neurite Fasciculation Mediated by Complexes of Axonin-1 and Ng Cell Adhesion Molecule

Neural cell adhesion molecules composed of immunoglobulin and fibronectin type III-like domains have been implicated in cell adhesion, neurite outgrowth, and fasciculation. Axonin-1 and Ng cell adhesion molecule (NgCAM), two molecules with predominantly axonal expression exhibit homophilic interactions across the extracellular space (axonin- 1/axonin-1 and NgCAM/NgCAM) and a heterophilic interaction (axonin-1–NgCAM) that occurs exclusively in the plane of the same membrane (cis-interaction). Using domain deletion mutants we localized the NgCAM homophilic binding in the Ig domains 1-4 whereas heterophilic binding to axonin-1 was localized in the Ig domains 2-4 and the third FnIII domain. The NgCAM–NgCAM interaction could be established simultaneously with the axonin-1–NgCAM interaction. In contrast, the axonin-1–NgCAM interaction excluded axonin-1/axonin-1 binding. These results and the examination of the coclustering of axonin-1 and NgCAM at cell contacts, suggest that intercellular contact is mediated by a symmetric axonin-1₂/NgCAM₂ tetramer, in which homophilic NgCAM binding across the extracellular space occurs simultaneously with a cis-heterophilic interaction of axonin-1 and NgCAM. The enhanced neurite fasciculation after overexpression of NgCAM by adenoviral vectors indicates that NgCAM is the limiting component for the formation of the axonin-1₂/NgCAM₂ complexes and, thus, neurite fasciculation in DRG neurons.     

Self recognition in the Ig superfamily. Identification of precise subdomains in carcinoembryonic antigen required for intercellular adhesion

The homophilic binding of extracellular domains of membrane-bound immunoglobulin superfamily (IgSF) molecules is often required for intercellular adhesion and signaling. Carcinoembryonic antigen (CEA), a member of the IgSF, is a widely used tumor marker that functions in vitro as a homotypic intercellular adhesion molecule. CEA has also been shown to contribute to tumorigenicity by inhibiting cellular differentiation, an effect that requires the homophilic binding of its extracellular domains. It was of interest, therefore, to identify small subdomain sequences in CEA that could serve as a focus in the design of peptides that disrupt CEA-mediated intercellular adhesion. Three subdomains in the N-terminal domain of CEA, identified by site-directed deletions and point mutations, were shown to be required for intercellular adhesion. Cyclized peptides representing two of these subdomains, (42)NRQII and (80)QNDTG, were found to be effective in blocking CEA-mediated cellular aggregation when added to CEA-expressing transfectants in suspension. Intermolecular binding involving each of these subdomains is therefore essential for intercellular adhesion and cannot be compensated for by known binding contributions of other regions in the CEA molecule. In further support of this assumption, the binding epitope of an anti-CEA monoclonal antibody (monoclonal antibody A20) known to block CEA-mediated adhesion, was shown to bridge two of the three required subdomains: (42)NRQII and (30)GYSWYK.     

Domain and functional analysis of a novel platelet-endothelial cell surface protein, SCUBE1

SCUBE1 (signal peptide-CUB-EGF domain-containing protein 1) is a novel, secreted, cell surface glycoprotein expressed during early embryogenesis and found in platelet and endothelial cells. This protein is composed of an N-terminal signal peptide sequence followed by nine tandemly arranged epidermal growth factor (EGF)-like repeats, a spacer region, three cysteine-rich repeat motifs, and one CUB domain at the C terminus. However, little is known about its domain and biological function. Here, we generated a comprehensive panel of domain deletion constructs and a new genetic mouse model with targeted disruption of Scube1 (Scube1(Delta cub/Delta cub)) to investigate the domain function and biological significance. A number of cell-based assays were utilized to define the critical role of the spacer region for membrane association and establish that the EGF-like repeats 7-9 are sufficient for the formation of SCUBE1-mediated homophilic adhesions in a calcium-dependent fashion. Biochemical and molecular analyses showed that the C-terminal cysteine-rich motifs and CUB domain could directly bind and antagonize the bone morphogenetic protein activity. Furthermore, genetic ablation of this C-terminal region resulted in brain malformation in the Scube1(Delta cub/Delta cub) embryos. Together, our results support the dual roles of SCUBE1 on brain morphogenesis and cell-cell adhesions through its distinct domain function.     

Molecular basis for the homophilic activated leukocyte cell adhesion molecule (ALCAM)-ALCAM interaction

Activated leukocyte cell adhesion molecule (ALCAM/CD166), a member of the immunoglobulin superfamily with five extracellular immunoglobulin-like domains, facilitates heterophilic (ALCAM-CD6) and homophilic (ALCAM-ALCAM) cell-cell interactions. While expressed in a wide variety of tissues and cells, ALCAM is restricted to subsets of cells usually involved in dynamic growth and/or migration processes. A structure-function analysis, using two monoclonal anti-ALCAM antibodies and a series of amino-terminally deleted ALCAM constructs, revealed that homophilic cell adhesion depended on ligand binding mediated by the membrane-distal amino-terminal immunoglobulin domain and on avidity controlled by ALCAM clustering at the cell surface involving membrane-proximal immunoglobulin domains. Co-expression of a transmembrane ALCAM deletion mutant, which lacks the ligand binding domain, and endogenous wild-type ALCAM inhibited homophilic cell-cell interactions by interference with ALCAM avidity, while homophilic, soluble ligand binding remained unaltered. The extracellular structures of ALCAM thus provide two structurally and functionally distinguishable modules, one involved in ligand binding and the other in avidity. Functionality of both modules is required for stable homophilic ALCAM-ALCAM cell-cell adhesion.     

The structure of neurexin 1α reveals features promoting a role as synaptic organizer

α-neurexins are essential synaptic adhesion molecules implicated in autism spectrum disorder and schizophrenia. The α-neurexin extracellular domain consists of six LNS domains interspersed by three EGF-like repeats and interacts with many different proteins in the synaptic cleft. To understand how α-neurexins might function as synaptic organizers, we solved the structure of the neurexin 1α extracellular domain (n1α) to 2.65 ?. The L-shaped molecule can be divided into a flexible repeat I (LNS1-EGF-A-LNS2), a rigid horseshoe-shaped repeat II (LNS3-EGF-B-LNS4) with structural similarity to so-called reelin repeats, and an extended repeat III (LNS5-EGF-B-LNS6) with controlled flexibility. A 2.95 ? structure of n1α carrying splice insert SS#3 in LNS4 reveals that SS#3 protrudes as a loop and does not alter the rigid arrangement of repeat II. The global architecture imposed by conserved structural features enables α-neurexins to recruit and organize proteins in distinct and variable ways, influenced by splicing, thereby promoting synaptic function.     

Homophilic adhesion between Ig superfamily carcinoembryonic antigen molecules involves double reciprocal bonds

Both carcinoembryonic antigen (CEA) and neural cell adhesion molecule (NCAM) belong to the immunoglobulin supergene family and have been demonstrated to function as homotypic Ca(++)-independent intercellular adhesion molecules. CEA and NCAM cannot associate heterotypically indicating that they have different binding specificities. To define the domains of CEA involved in homotypic interaction, hybrid cDNAs consisting of various domains from CEA and NCAM were constructed and were transfected into a CHO-derived cell line; stable transfectant clones showing cell surface expression of CEA/NCAM chimeric-proteins were assessed for their adhesive properties by homotypic and heterotypic aggregation assays. The results indicate that all five of the Ig(C)-like domains of NCAM are required for intercellular adhesion while the COOH-terminal domain containing the fibronectin-like repeats is dispensable. The results also show that adhesion mediated by CEA involves binding between the Ig(V)-like amino-terminal domain and one of the Ig(C)-like internal repeat domains: thus while transfectants expressing constructs containing either the N domain or the internal domains alone were incapable of homotypic adhesion, they formed heterotypic aggregates when mixed. Furthermore, peptides consisting of both the N domain and the third internal repeat domain of CEA blocked CEA-mediated cell aggregation, thus providing direct evidence for the involvement of the two domains in adhesion. We therefore propose a novel model for interactions between immunoglobulin supergene family members in which especially strong binding is effected by double reciprocal interactions between the V-like domains and C-like domains of antiparallel CEA molecules on apposing cell surfaces.     

Ep-CAM: a human epithelial antigen is a homophilic cell-cell adhesion molecule

The epithelial glycoprotein 40 (EGP40, also known as GA733-2, ESA, KSA, and the 17-1A antigen), encoded by the GA-733-2 gene, is expressed on the baso-lateral cell surface in most human simple epithelia. The protein is also expressed in the vast majority of carcinomas and has attracted attention as a tumor marker. The function of the protein is unknown. We demonstrate here that EGP40 is an epithelium-specific intercellular adhesion molecule. The molecule mediates, in a Ca(2+)- independent manner, a homophilic cell-cell adhesion of murine cells transfected with the complete EGP40 cDNA. Two murine cell lines were tested for the effects of EGP40 expression: fibroblastic L cells and dedifferentiated mammary carcinoma L153S cells. The expression of the EGP40 protein causes morphological changes in cultures of transfected cells--increasing intercellular adhesion of the transfectants--and has a clear effect on cell aggregating behavior in suspension aggregation assays. EGP40 directs sorting in mixed cell populations, in particular, causes segregation of the transfectants from the corresponding parental cells. EGP40 expression suppresses invasive colony growth of L cells in EHS-matrigel providing tight adhesions between cells in growing colonies. EGP40 can thus be considered a new member of the intercellular adhesion molecules. In its biological behavior EGP40 resembles to some extent the molecules of the immunoglobulin superfamily of cell adhesion molecules (CAMs), although no immunoglobulin-like repeats are present in the EGP40 molecule. Certain structural similarities in general organization of the molecule exist between EGP40 and the lin-12/Notch proteins. A possible role of this adhesion molecule in formation of architecture of epithelial tissues is discussed. To reflect the function of the molecule the name Ep-CAM for EGP40 seems appropriate.     

Expression of Ep-CAM shifts the state of cadherin-mediated adhesions from strong to weak

Various adhesion molecules play an important role in defining cell fate and maintaining tissue integrity. Therefore, cross-signaling between adhesion receptors should be a common phenomenon to support the orchestrated changes of cells' connections to the substrate and to the neighboring cells during tissue remodeling. Recently, we have demonstrated that the epithelial cell adhesion molecule Ep-CAM negatively modulates cadherin-mediated adhesions in direct relation to its expression levels. Here, we used E-cadherin/alpha-catenin chimera constructs to define the site of Ep-CAM's negative effect on cadherin-mediated adhesions. Murine L-cells transfected with either E-cadherin/alpha-catenin fusion protein, or E-cadherin fused to the carboxy-terminal half of alpha-catenin, were subsequently supertransfected with an inducible Ep-CAM construct. Introduction of Ep-CAM altered the cell's morphology, weakened the strength of cell-cell interactions, and decreased the cytoskeleton-bound fraction of the cadherin/catenin chimeras in both cell models. Furthermore, expression of Ep-CAM induced restructuring of F-actin, with changes in thickness and orientation of the actin filaments. The results showed that Ep-CAM affects E-cadherin-mediated adhesions without involvement of beta-catenin by disrupting the link between alpha-catenin and F-actin. The latter is likely achieved through remodeling of the actin cytoskeleton by Ep-CAM, possibly through pp120.     

Cytoplasmic Tail Regulates the Intercellular Adhesion Function of the Epithelial Cell Adhesion Molecule

Ep-CAM, an epithelium-specific cell-cell adhesion molecule (CAM) not structurally related to the major families of CAMs, contains a cytoplasmic domain of 26 amino acids. The chemical disruption of the actin microfilaments, but not of the microtubuli or intermediate filaments, affected the localization of Ep-CAM at the cell-cell boundaries, suggesting that the molecule interacts with the actin-based cytoskeleton. Mutated forms of Ep-CAM were generated with the cytoplasmic domain truncated at various lengths. All of the mutants were transported to the cell surface in the transfectants; however, the mutant lacking the complete cytoplasmic domain was not able to localize to the cell-cell boundaries, in contrast to mutants with partial deletions. Both the disruption of the actin microfilaments and a complete truncation of the cytoplasmic tail strongly affected the ability of Ep-CAM to mediate aggregation of L cells. The capability of direct aggregation was reduced for the partially truncated mutants but remained cytochalasin D sensitive. The tail truncation did not affect the ability of the transfectants to adhere to solid-phase-adsorbed Ep-CAM, suggesting that the ability to form stable adhesions and not the ligand specificity of the molecule was affected by the truncation. The formation of intercellular adhesions mediated by Ep-CAM induced a redistribution to the cell-cell boundaries of α-actinin, but not of vinculin, talin, filamin, spectrin, or catenins. Coprecipitation demonstrated direct association of Ep-CAM with α-actinin. Binding of α-actinin to purified mutated and wild-type Ep-CAMs and to peptides representing different domains of the cytoplasmic tail of Ep-CAM demonstrates two binding sites for α-actinin at positions 289 to 296 and 304 to 314 of the amino acid sequence. The results demonstrate that the cytoplasmic domain of Ep-CAM regulates the adhesion function of the molecule through interaction with the actin cytoskeleton via α-actinin.     

Cancer-Derived Mutations in the Fibronectin III Repeats of PTPRT/PTPρ Inhibit Cell-Cell Aggregation

Abstract

The receptor protein tyrosine phosphatase T PTPρ is the most frequently mutated tyrosine phosphatase in human cancer. PTPρ mediates homophilic cell-cell aggregation. In its extracellular region, PTPρ has cell adhesion molecule–like motifs, including a MAM domain, an immunoglobulin domain, and four fibronectin type III (FNIII) repeats. Tumor-derived mutations have been identified in all of these extracellular domains. Previously, the authors determined that tumor-derived mutations in the MAM and immunoglobulin domains of PTPρ reduce homophilic cell-cell aggregation. In this paper, the authors describe experiments in which the contribution of the FNIII repeats to PTPρ-mediated cell-cell adhesion was evaluated. The results demonstrate that deletion of the FNIII repeats of PTPρ result in defective cell-cell aggregation. Furthermore, all of the tumor-derived mutations in the FNIII repeats of PTPρ also disrupt cell-cell aggregation. These results further support the hypothesis that mutational inactivation of PTPρ may lead to cancer progression by disrupting cell-cell adhesion.     

The JAM family of junctional adhesion molecules

Junctional adhesion molecules are a family of glycoproteins characterised by two immunoglobulin folds (VH- and C2-type) in the extracellular domain. Junctional adhesion molecule proteins localise to intercellular junctions of polarised endothelial and epithelial cells but can also be expressed on circulating leukocytes and platelets. In addition, they bind several ligands, in both a homophilic and heterophilic manner, and associate with several cytoplasmic partners. All these features represent the likely determinants for the role of junctional adhesion molecule proteins in processes as diverse as junction assembly, leukocyte transmigration and platelet activation.     

Axonin-1/TAG-1 mediates cell-cell adhesion by a cis-assisted trans-interaction.

The neural cell adhesion molecule axonin-1/TAG-1 mediates cell-cell interactions via homophilic and heterophilic contacts. It consists of six Ig and four fibronectin type III domains anchored to the membrane by glycosylphosphatidylinositol. The recently solved crystal structure indicates a module composed of the four N-terminal Ig domains as the contact site between trans-interacting axonin-1 molecules from apposed membranes. Here, we have tested domain-specific monoclonal antibodies for their capacity to interfere with homophilic binding in a cell aggregation assay. The results confirmed the existence of a binding region within the N-terminal Ig domains and identified a second region contributing to homophilic binding on the third and fourth fibronectin domains near the C terminus. The perturbation of each region alone resulted in a complete loss of cell aggregation, suggesting that axonin-1-mediated cell-cell contact results from a cooperative action of two homophilic binding regions. The data support that axonin-1-mediated cell-cell contact is formed by cis-assisted trans-binding. The N-terminal binding regions of axonin-1 establish a linear zipper-like string of trans-interacting axonin-1 molecules alternately provided by the two apposed membranes. The C-terminal binding regions strengthen the cell-cell contact by enhancing the expansion of the linear string into a two-dimensional array via cis-interactions. Cis-assisted trans-binding may be a basic binding mechanism common to many cell adhesion molecules.     

Multiple cadherin extracellular repeats mediate homophilic binding and adhesion.

The extracellular homophilic-binding domain of the cadherins consists of 5 cadherin repeats (EC1-EC5). Studies on cadherin specificity have implicated the NH(2)-terminal EC1 domain in the homophilic binding interaction, but the roles of the other extracellular cadherin (EC) domains have not been evaluated. We have undertaken a systematic analysis of the binding properties of the entire cadherin extracellular domain and the contributions of the other EC domains to homophilic binding. Lateral (cis) dimerization of the extracellular domain is thought to be required for adhesive function. Sedimentation analysis of the soluble extracellular segment of C-cadherin revealed that it exists in a monomer-dimer equilibrium with an affinity constant of approximately 64 microm. No higher order oligomers were detected, indicating that homophilic binding between cis-dimers is of significantly lower affinity. The homophilic binding properties of a series of deletion constructs, lacking successive or individual EC domains fused at the COOH terminus to an Fc domain, were analyzed using a bead aggregation assay and a cell attachment-based adhesion assay. A protein with only the first two NH(2)-terminal EC domains (CEC1-2Fc) exhibited very low activity compared with the entire extracellular domain (CEC1-5Fc), demonstrating that EC1 alone is not sufficient for effective homophilic binding. CEC1-3Fc exhibited high activity, but not as much as CEC1-4Fc or CEC1-5Fc. EC3 is not required for homophilic binding, however, since CEC1-2-4Fc and CEC1-2-4-5Fc exhibited high activity in both assays. These and experiments using additional EC combinations show that many, if not all, the EC domains contribute to the formation of the cadherin homophilic bond, and specific one-to-one interaction between particular EC domains may not be required. These conclusions are consistent with a previous study on direct molecular force measurements between cadherin ectodomains demonstrating multiple adhesive interactions (Sivasankar, S., W. Brieher, N. Lavrik, B. Gumbiner, and D. Leckband. 1999. PROC: Natl. Acad. Sci. USA. 96:11820-11824; Sivasankar, S., B. Gumbiner, and D. Leckband. 2001. Biophys J. 80:1758-68). We propose new models for how the cadherin extracellular repeats may contribute to adhesive specificity and function.     

Segmented Helical Structure of the Neck Region of the Glycan-Binding Receptor DC-SIGNR

Carbohydrate-recognition domains (CRDs) in the glycan-binding receptors DC-SIGN (dendritic-cell-specific intercellular adhesion molecule 1-grabbing nonintegrin; CD209) and DC-SIGNR (DC-SIGN-related receptor, also known as L-SIGN and variously designated CD209L and CD299) are projected from the membrane surface by extended neck domains containing multiple repeats of a largely conserved 23-amino-acid sequence motif. Crystals of a fragment of the neck domain of DC-SIGNR containing multiple repeats in which each molecule extends through multiple unit cells, such that the observed crystallographic asymmetric unit represents one repeat averaged over six repeats of the protein, have been obtained. The repeats are largely α-helical. Based on the structure and arrangement of the repeats in the crystal, the neck region can be described as a series of four-helix bundles connected by short, non-helical linkers. Combining the structure of the isolated neck domain with a previously determined overlapping structure of the distal end of the neck region with the CRDs attached provides a model of the almost-complete extracellular portion of the receptor. The results are consistent with previous characterization of the extended structure for the isolated neck region and the extracellular domain. The organization of the neck suggests how CRDs may be disposed differently in DC-SIGN compared with DC-SIGNR and in variant forms of DC-SIGNR assembled from polypeptides with different numbers of repeats in the neck domain.     

IGSF4: a new intercellular adhesion molecule that is called by three names,TSLC1, SgIGSF and SynCAM,by virtue of its diverse function

Members of the immunoglobulin superfamily often play key roles in intercellular adhesion. IGSF4 is a novel immunoglobulin (Ig)-like intercellular adhesion molecule. Three Ig-like domains are included in the extracellular domain of IGSF4 and mediate homophilic or heterophilic interactions independently of Ca2+. The cytoplasmic domain of IGSF4 contains the binding motifs that connect to actin fibers. Since IGSF4 has been characterized by several independent research groups, this molecule is called by three names, TSLC1, SgIGSF and SynCAM. IGSF4 was first characterized as a tumor suppressor of non-small cell lung cancer and termed TSLC1, although how IGSF4 suppresses tumor growth remains unknown. Silencing of the IGSF4 gene was primarily achieved by allelic loss and promoter methylation in this type of cancers. Soon after this discovery, IGSF4 was found to have roles in adhesion of spermatogenic cells to Sertoli cells and mast cells to fibroblasts and termed SgIGSF. Other researchers revealed that IGSF4 drives synaptic formation of neural cells and termed it SynCAM.     

The formation and stability of DC-SIGN microdomains require its extracellular moiety

Dendritic cell-specific intercellular adhesion molecule (ICAM)-3-grabbing non-integrin (DC-SIGN) is a Ca(2+) -dependent transmembrane lectin that binds a large variety of pathogens and facilitates their uptake for subsequent antigen presentation. This receptor is present in cell surface microdomains, but factors involved in microdomain formation and their exceptional stability are not clear. To determine which domain/motif of DC-SIGN facilitates its presence in microdomains, we studied mutations at key locations including truncation of the cytoplasmic tail, and ectodomain mutations that resulted in the removal of the N-linked glycosylation site, the tandem repeats and the carbohydrate recognition domain (CRD), as well as modification of the calcium sites in the CRD required for carbohydrate binding. Confocal imaging and fluorescence recovery after photobleaching measurements showed that the cytoplasmic domain and the N-linked glycosylation site do not affect the ability of DC-SIGN to form stable microdomains. However, truncation of the CRD results in complete loss of visible microdomains and subsequent lateral diffusion of the mutants. Apart from cell adhesions, membrane domains are thought to be localized primarily via the cytoskeleton. By contrast, we propose that interactions between the CRD of DC-SIGN and the extracellular matrix and/or cis interactions with transmembrane scaffolding protein(s) play an essential role in organizing these microdomains.     

The cadherin-catenin complex as a focal point of cell adhesion and signalling: new insights from three-dimensional structures

Cadherins are a large family of single-pass transmembrane proteins principally involved in Ca2+-dependent homotypic cell adhesion. The cadherin molecules comprise three domains, the intracellular domain, the transmembrane domain and the extracellular domain, and form large complexes with a vast array of binding partners (including cadherin molecules of the same type in homophilic interactions and cellular protein catenins), orchestrating biologically essential extracellular and intracellular signalling processes. While current, contrasting models for classic cadherin homophilic interaction involve varying numbers of specific repeats found in the extracellular domain, the structure of the domain itself clearly remains the main determinant of cell stability and binding specificity. Through intracellular interactions, cadherin enhances its adhesive properties binding the cytoskeleton via cytoplasmic associated factors alpha- catenin, beta-catenin and p120ctn. Recent structural studies on classic cadherins and these catenin molecules have provided new insight into the essential mechanisms underlying cadherin-mediated cell interaction and catenin-mediated cellular signalling. Remarkable structural diversity has been observed in beta-catenin recognition of other cellular factors including APC, Tcf and ICAT, proteins that contribute to or compete with cadherin/catenin functioning.