Cadherin dimers in cell-cell adhesion

While the critical function of classic cadherin in cell-cell junctions is well established, the molecular mechanism of cadherin-based adhesion remains unclear. The elusive but principal part of this adhesion process is the cadherin-cadherin interaction maintaining the intercellular contacts. This interaction is believed to be weak, suggesting that the adhesive contacts are strengthened by the cytoskeleton-dependent clustering of numerous cadherin molecules. An examination of cadherin homodimers in living cells has shown, however, that cadherin adhesive interaction is surprisingly strong. This observation implies that the strength of the adhesive contacts is regulated by the processes disintegrating cadherin dimers. The molecular structure of these dimers and mechanisms potentially responsible for their dynamics in living cells are discussed in this review.     

Endothelial cell regulation of leukocyte infiltration in inflammatory tissues

Endothelial cells play an important, active role in the onset and regulation of inflammatory and immune reactions. Through the production of chemokines they attract leukocytes and activate their adhesive receptors. This leads to the anchorage of leukocytes to the adhesive molecules expressed on the endothelial surface. Leukocyte adhesion to endothelial cells is frequently followed by their extravasation. The mechanisms which regulate the passage of leukocytes through endothelial clefts remain to be clarified. Many indirect data suggest that leukocytes might transfer signals to endothelial cells both through the release of active agents and adhesion to the endothelial cell surface. Adhesive molecules (such as PECAM) on the endothelial cell surface might also 'direct' leukocytes through the intercellular junction by haptotaxis. The information available on the molecular structure and functional properties of endothelial chemokines, adhesive molecules or junction organization is still fragmentary. Further work is needed to clarify how they interplay in regulating leukocyte infiltration into tissues.     

Desmosomal cadherins utilize distinct kinesins for assembly into desmosomes

The desmosomal cadherins, desmogleins (Dsgs) and desmocollins (Dscs), comprise the adhesive core of intercellular junctions known as desmosomes. Although these adhesion molecules are known to be critical for tissue integrity, mechanisms that coordinate their trafficking into intercellular junctions to regulate their proper ratio and distribution are unknown. We demonstrate that Dsg2 and Dsc2 both exhibit microtubule-dependent transport in epithelial cells but use distinct motors to traffic to the plasma membrane. Functional interference with kinesin-1 blocked Dsg2 transport, resulting in the assembly of Dsg2-deficient junctions with minimal impact on distribution of Dsc2 or desmosomal plaque components. In contrast, inhibiting kinesin-2 prevented Dsc2 movement and decreased its plasma membrane accumulation without affecting Dsg2 trafficking. Either kinesin-1 or -2 deficiency weakened intercellular adhesion, despite the maintenance of adherens junctions and other desmosome components at the plasma membrane. Differential regulation of desmosomal cadherin transport could provide a mechanism to tailor adhesion strength during tissue morphogenesis and remodeling.     

Desmosome structure, composition and function

Desmosomes are intercellular junctions of epithelia and cardiac muscle. They resist mechanical stress because they adopt a strongly adhesive state in which they are said to be hyper-adhesive and which distinguishes them from other intercellular junctions; desmosomes are specialised for strong adhesion and their failure can result in diseases of the skin and heart. They are also dynamic structures whose adhesiveness can switch between high and low affinity adhesive states during processes such as embryonic development and wound healing, the switching being signalled by protein kinase C. Desmosomes may also act as signalling centres, regulating the availability of signalling molecules and thereby participating in fundamental processes such as cell proliferation, differentiation and morphogenesis. Here we consider the structure, composition and function of desmosomes, and their role in embryonic development and disease.     

The desmosome and pemphigus

Desmosomes are patch-like intercellular adhering junctions ("maculae adherentes"), which, in concert with the related adherens junctions, provide the mechanical strength to intercellular adhesion. Therefore, it is not surprising that desmosomes are abundant in tissues subjected to significant mechanical stress such as stratified epithelia and myocardium. Desmosomal adhesion is based on the Ca(2+)-dependent, homo- and heterophilic transinteraction of cadherin-type adhesion molecules. Desmosomal cadherins are anchored to the intermediate filament cytoskeleton by adaptor proteins of the armadillo and plakin families. Desmosomes are dynamic structures subjected to regulation and are therefore targets of signalling pathways, which control their molecular composition and adhesive properties. Moreover, evidence is emerging that desmosomal components themselves take part in outside-in signalling under physiologic and pathologic conditions. Disturbed desmosomal adhesion contributes to the pathogenesis of a number of diseases such as pemphigus, which is caused by autoantibodies against desmosomal cadherins. Beside pemphigus, desmosome-associated diseases are caused by other mechanisms such as genetic defects or bacterial toxins. Because most of these diseases affect the skin, desmosomes are interesting not only for cell biologists who are inspired by their complex structure and molecular composition, but also for clinical physicians who are confronted with patients suffering from severe blistering skin diseases such as pemphigus. To develop disease-specific therapeutic approaches, more insights into the molecular composition and regulation of desmosomes are required.     

The molecular biology of desmosomes and hemidesmosomes: ′What's in a name?'

Desmosomes are junctions involved in intercellular adhesion of epithelial cells and hemidesmosomes are junctions involved in adhesion of epithelia to basement membranes. Both are characterised at the ultrastructural level by dense cytoplasmic plaques which are linked to the intermediate filament cytoskeleton of the cells. The plaques strongly resemble each other suggesting a relationship between the two kinds of junctions, as implied by their names. Recent characterisation of the molecular components of the junctions shows they are, in fact, quite unrelated implying that structural similarity is fortuitous. The molecular biology raises many fascinating problems relating to their structure and function.     

The extracellular architecture of adherens junctions revealed by crystal structures of type I cadherins

Adherens junctions, which play a central role in intercellular adhesion, comprise clusters of type I classical cadherins that bind via extracellular domains extended from opposing cell surfaces. We show that a molecular layer seen in crystal structures of E- and N-cadherin ectodomains reported here and in a previous C-cadherin structure corresponds to the extracellular architecture of adherens junctions. In all three ectodomain crystals, cadherins dimerize through a trans adhesive interface and are connected by a second, cis, interface. Assemblies formed by E-cadherin ectodomains coated on liposomes also appear to adopt this structure. Fluorescent imaging of junctions formed from wild-type and mutant E-cadherins in cultured cells confirm conclusions derived from structural evidence. Mutations that interfere with the trans interface ablate adhesion, whereas cis interface mutations disrupt stable junction formation. Our observations are consistent with a model for junction assembly involving strong trans and weak cis interactions localized in the ectodomain.     

Adhesive and Signaling Functions of Cadherins and Catenins in Vertebrate Development

Properly regulated intercellular adhesion is critical for normal development of all metazoan organisms. Adherens junctions play an especially prominent role in development because they link the adhesive function of cadherin–catenin protein complexes to the dynamic forces of the actin cytoskeleton, which helps to orchestrate a spatially confined and very dynamic assembly of intercellular connections. Intriguingly, in addition to maintaining intercellular adhesion, cadherin–catenin proteins are linked to several major developmental signaling pathways crucial for normal morphogenesis. In this article we will highlight the key genetic studies that uncovered the role of cadherin–catenin proteins in vertebrate development and discuss the potential role of these proteins as molecular biosensors of external cellular microenvironment that may spatially confine signaling molecules and polarity cues to orchestrate cellular behavior throughout the complex process of normal morphogenesis.Development of any multicellular organism is impossible without a dynamic and properly regulated intercellular adhesion. Adhesive contacts between cells provide a physical anchoring system that is necessary to form highly organized tissues, and these contacts are essential for effective intercellular communication that ensures the homeostasis and survival of the entire organism. A number of unique developmental processes, including such early events as embryonic compaction and first cell fate specification, as well as later tissue morphogenesis and organogenesis, rely on a dynamic balance between cellular adhesion and migration. Cadherin–catenin protein complexes, which constitute the core of a specialized subtype of cellular adhesion structures termed adherens junctions (AJs), play a particularly important role during these processes. Apart from maintaining adhesive contacts at the cell–cell junctions, they are actively involved in epithelial-to-mesenchymal and mesenchymal-to-epithelial transitions, which are crucial to sustain the tissue plasticity during development. Most importantly, the components of cadherin–catenin complexes are tightly linked to several major signaling networks controlling cell division, differentiation, and apoptosis and this feature is crucial for the broad roles of the AJs throughout the vertebrate development (see Cavey and Lecuit 2009).This article will focus on the role of cadherin–catenin proteins in regulating the signaling events critical for vertebrate development. Altering the expression pattern of particular cadherin–catenin complex components in the developing embryo often leads to major developmental defects, which reflect their role in both signaling and mechanical adhesion. In this article, we will highlight crucial findings suggesting that cadherin–catenin complexes provide not only the structural integrity of the tissue, but may also serve as biosensors of the external cellular microenvironment that modulate cellular behavior and make individual cells work together to ensure the fitness of the entire organism.     

Numerical investigation of the role of intercellular interactions on collective epithelial cell migration

During collective cell migration, the intercellular forces will significantly affect the collective migratory behaviors. However, the measurement of mechanical stresses exerted at cell–cell junctions is very challenging. A recent experimental observation indicated that the intercellular adhesion sites within a migrating monolayer are subjected to both normal stress exerted perpendicular to cell–cell junction surface and shear stress exerted tangent to cell–cell junction surface. In this study, an interfacial interaction model was proposed to model the intercellular interactions for the first time. The intercellular interaction model-based study of collective epithelial migration revealed that the direction of cell migration velocity has better alignment with the orientation of local principal stress at higher maximum shear stress locations in an epithelial monolayer sheet. Parametric study of the effects of adhesion strength indicated that normal adhesion strength at the cell–cell junction surface has dominated effect on local alignment between the direction of cell velocity vector and the principal stress orientation, while the shear adhesion strength has little effect, which provides compelling evidence to help explain the force transmission via cell–cell junctions between adjacent cells in collective cell motion and provides new insights into “adhesive belt” effects at cell–cell junction.     

The C-terminal unique region of desmoglein 2 inhibits its internalization via tail–tail interactions

Desmosomal cadherins, desmogleins (Dsgs) and desmocollins, make up the adhesive core of intercellular junctions called desmosomes. A critical determinant of epithelial adhesive strength is the level and organization of desmosomal cadherins on the cell surface. The Dsg subclass of desmosomal cadherins contains a C-terminal unique region (Dsg unique region [DUR]) with unknown function. In this paper, we show that the DUR of Dsg2 stabilized Dsg2 at the cell surface by inhibiting its internalization and promoted strong intercellular adhesion. DUR also facilitated Dsg tail–tail interactions. Forced dimerization of a Dsg2 tail lacking the DUR led to decreased internalization, supporting the conclusion that these two functions of the DUR are mechanistically linked. We also show that a Dsg2 mutant, V977fsX1006, identified in arrhythmogenic right ventricular cardiomyopathy patients, led to a loss of Dsg2 tail self-association and underwent rapid endocytosis in cardiac muscle cells. Our observations illustrate a new mechanism desmosomal cadherins use to control their surface levels, a key factor in determining their adhesion and signaling roles.     

Intercellular adhesive selectivity. II. Properties of embryonic chick liver cell-cell adhesion

Studies directed at understanding the molecular basis of liver cell homotypic adhesion are presented. An assay which measures the rate of adhesion of isotopically labeled (32PO4) embryonic chick liver cells to liver cell aggregates, described in a companion paper, has been used to investigate the problem of intercellular adhesive selectivity. Cation requirements, the effects of various inhibitors of metabolism and protein synthesis, of chelators (EDTA and EGTA), and the effects of temperature on liver cell adhesion are reported. Two mechanisms of inhibition of liver intercellular adhesion are suggested. One involves destruction of cell-surface adhesion receptors (sensitivity to proteases); the other is an energy-dependent step which may involve alterations in plasma membrane conformation and/or membrane fluidity. Finally, a model is suggested for liver cell-cell adhesion that incorporates the early tissue selectivity of intercellular adhesion previously reported, followed by a multistep process which leads to histogenic aggregation.     

Stratifin (14-3-3 σ) Limits Plakophilin-3 Exchange with the Desmosomal Plaque

Desmosomes are prominent cell-cell adhesive junctions in stratified squamous epithelia and disruption of desmosomal adhesion has been shown to have dramatic effects on the function and integrity of these tissues. During normal physiologic processes, such as tissue development and wound healing, intercellular adhesion must be modified locally to allow coordinated cell movements. The mechanisms that control junction integrity and adhesive strength under these conditions are poorly understood. We utilized a proteomics approach to identify plakophilin-3 associated proteins and identified the 14-3-3 family member stratifin. Stratifin interacts specifically with plakophilin-3 and not with other plakophilin isoforms and mutation analysis demonstrated the binding site includes serine 285 in the amino terminal head domain of plakophilin-3. Stratifin interacts with a cytoplasmic pool of plakophilin-3 and is not associated with the desmosome in cultured cells. FRAP analysis revealed that decreased stratifin expression leads to an increase in the exchange rate of cytoplasmic plakophilin-3/GFP with the pool of plakophilin-3/GFP in the desmosome resulting in decreased desmosomal adhesion and increased cell migration. We propose a model by which stratifin plays a role in regulating plakophilin-3 incorporation into the desmosomal plaque by forming a plakophilin-3 stratifin complex in the cytosol and thereby affecting desmosome dynamics in squamous epithelial cells.     

N-cadherin expression level as a critical indicator of invasion in non-epithelial tumors

Cancer cell dissemination away from the primary tumor and their ability to form metastases remain the major causes of death from cancer. Understanding the molecular mechanisms triggering this event could lead to the design of new cancer treatments. The establishment and the maintenance of tissue architecture depend on the coordination of cell behavior within this tissue. Cell-cell interactions must form adhesive structures between neighboring cells while remaining highly dynamic to allow and control tissue renewal or remodeling. Among intercellular junctions, cadherin-based adherens junctions mediate strong physical interactions and transmit information from the cell microenvironment to the cytoplasm. Disruption of these cell-cell contacts perturbs the polarity of epithelial tissues leading to their disorganization and ultimately to aggressive carcinomas. In non-epithelial tissues, the role of cadherins in the development of cancer is still debated. We recently found that downregulation of N-cadherin in malignant glioma—the most frequent primary brain tumor—results in cell polarization defects leading to abnormal motile behavior with increased cell speed and decreased persistence in directionality. Re-expression of N-cadherin in glioma cells restores cell polarity and limits glioma cell migration, providing a potential therapeutic tool for diffuse glioma.     

The LIM protein Ajuba is recruited to cadherin-dependent cell junctions through an association with alpha-catenin

Cell-cell adhesive events affect cell growth and fate decisions and provide spatial clues for cell polarity within tissues. The complete molecular determinants required for adhesive junction formation and their function are not completely understood. LIM domain-containing proteins have been shown to be present at cell-cell contact sites and are known to shuttle into the nucleus where they can affect cell fate and growth; however, their precise localization at cell-cell contacts, how they localize to these sites, and what their functions are at these sites is unknown. Here we show that, in primary keratinocytes, the LIM domain protein Ajuba is recruited to cadherin-dependent cell-cell adhesive complexes in a regulated manner. At cadherin adhesive complexes Ajuba interacts with alpha-catenin, and alpha-catenin is required for efficient recruitment of Ajuba to cell junctions. Ajuba also interacts directly with F-actin. Keratinocytes from Ajuba null mice exhibit abnormal cell-cell junction formation and/or stability and function. These data reveal Ajuba as a new component at cadherin-mediated cell-cell junctions and suggest that Ajuba may contribute to the bridging of the cadherin adhesive complexes to the actin cytoskeleton and as such contribute to the formation or strengthening of cadherin-mediated cell-cell adhesion.     

Cell adhesion: More than just glue (Review)

The ability of cells to interact with each other and their surroundings in a co-ordinated manner depends on multiple adhesive interactions between neighbouring cells and their extracellular environment. These adhesive interactions are mediated by a family of cell surface proteins, termed cell adhesion molecules. Fortunately these adhesion molecules fall into distinct families with adhesive interactions varying in strength from strong binding involved in the maintenance of tissue architecture to more transient, less avid, dynamic interactions observed in leukocyte biology. Adhesion molecules are extremely versatile cell surface receptors which not only stick cells together but provide biochemical and physical signals that regulate a range of diverse functions, such as cell proliferation, gene expression, differentiation, apoptosis and migration. In addition, like many other cell surface molecules, they have been usurped as portals of entry for pathogens, including prions. How the mechanical and chemical messages generated from adhesion molecules are integrated with other signalling pathways (such as receptor tyrosine kinases and phosphatases) and the role that aberrant cell adhesion plays in developmental defects and disease pathology are currently very active areas of research. This review focuses on the biochemical features that define whether a cell surface molecule can act as an adhesion molecule, and discusses five specific examples of how cell adhesion molecules function as more than just 'sticky’ receptors. The discussion is confined to the signalling events mediated by members of the integrin, cadherin and immunoglobulin gene superfamilies. It is suggested that, by controlling the membrane organization of signalling receptors, by imposing spatial organization, and by regulating the local concentration of cytosolic adapter proteins, intercellular and cell-matrix adhesion is more than just glue holding cells together. Rather dynamic ‘conversations’ and the formation of multi-protein complexes between adhesion molecules, growth factor receptors and matrix macromolecules can now provide a molecular explanation for the long-observed but poorly understood requirement for a number of seemingly distinct cell surface molecules to be engaged for efficient cell function to occur.     

Cell adhesion: more than just glue (review)

The ability of cells to interact with each other and their surroundings in a co-ordinated manner depends on multiple adhesive interactions between neighbouring cells and their extracellular environment. These adhesive interactions are mediated by a family of cell surface proteins, termed cell adhesion molecules. Fortunately these adhesion molecules fall into distinct families with adhesive interactions varying in strength from strong binding involved in the maintenance of tissue architecture to more transient, less avid, dynamic interactions observed in leukocyte biology. Adhesion molecules are extremely versatile cell surface receptors which not only stick cells together but provide biochemical and physical signals that regulate a range of diverse functions, such as cell proliferation, gene expression, differentiation, apoptosis and migration. In addition, like many other cell surface molecules, they have been usurped as portals of entry for pathogens, including prions. How the mechanical and chemical messages generated from adhesion molecules are integrated with other signalling pathways (such as receptor tyrosine kinases and phosphatases) and the role that aberrant cell adhesion plays in developmental defects and disease pathology are currently very active areas of research. This review focuses on the biochemical features that define whether a cell surface molecule can act as an adhesion molecule, and discusses five specific examples of how cell adhesion molecules function as more than just 'sticky' receptors. The discussion is confined to the signalling events mediated by members of the integrin, cadherin and immunoglobulin gene superfamilies. It is suggested that, by controlling the membrane organization of signalling receptors, by imposing spatial organization, and by regulating the local concentration of cytosolic adapter proteins, intercellular and cell-matrix adhesion is more than just glue holding cells together. Rather dynamic 'conversations' and the formation of multi-protein complexes between adhesion molecules, growth factor receptors and matrix macromolecules can now provide a molecular explanation for the long-observed but poorly understood requirement for a number of seemingly distinct cell surface molecules to be engaged for efficient cell function to occur.     

Junctional adhesion molecule C (JAM-C) dimerization aids cancer cell migration and metastasis

Most cancer deaths result from metastasis, which is the dissemination of cells from a primary tumor to distant organs. Metastasis involves changes to molecules that are essential for tumor cell adhesion to the extracellular matrix and to endothelial cells. Junctional Adhesion Molecule C (JAM-C) localizes at intercellular junctions as homodimers or more affine heterodimers with JAM-B. We previously showed that the homodimerization site (E66) in JAM-C is also involved in JAM-B binding. Here we show that neoexpression of JAM-C in a JAM-C-negative carcinoma cell line induced loss of adhesive property and pro-metastatic capacities. We also identify two critical structural sites (E66 and K68) for JAM-C/JAM-B interaction by directed mutagenesis of JAM-C and studied their implication on tumor cell behavior. JAM-C mutants did not bind to JAM-B or localize correctly to junctions. Moreover, mutated JAM-C proteins increased adhesion and reduced proliferation and migration of lung carcinoma cell lines. Carcinoma cells expressing mutant JAM-C grew slower than with JAM-C WT and were not able to establish metastatic lung nodules in mice. Overall these data demonstrate that the dimerization sites E66-K68 of JAM-C affected cell adhesion, polarization and migration and are essential for tumor cell metastasis.     

Cell adhesion in rat fibroblasts: effect of tumor promoters

In a study performed on transformed (SGS/3A) and normal syngeneic rat cells (FG/2) to identify the molecular mechanisms which regulate cell adhesion and contact inhibition in cell transformation, we investigated the effects of tumor promoters on cell to cell adhesion of rat fibroblasts. As tumor promoters we used 12-tetradecanoyl-phorbole-13-acetate (TPA) and the melittin, a polypeptide from bee venom, both substances capable of stimulating the neoplastic transformation. The intercellular adhesion assay consists in determining the percent of single cells labeled with 3H-L-leucine adhering to a confluent monolayer of unlabeled cells at different incubation times. The increase of cell-cell adhesion caused by TPA and melittin confirms what we have constantly observed in other experiments, i.e. that neoplastic cells SGS/3A always have a higher intercellular adhesion capacity than corresponding normal syngenic cells FG/2. Since one of the effects of the tumor promoters is also induction of a reversible alteration of the cytoskeleton, it is likely that their action on intercellular adhesion is regulated by a mechanism analogous to that proposed for explaining the increased intercellular adhesion observed after treatment with antimicrotubular compounds.     

Requirements for the Ca2+-independent component in the initial intercellular adhesion of C2 myoblasts

Using a sensitive and quantitative adhesion assay, we have studied the initial stages of the intercellular adhesion of the C2 mouse myoblast line. After dissociation in low levels of trypsin in EDTA, C2 cells can rapidly reaggregate by Ca2+-independent mechanisms to form large multicellular aggregates. If cells are allowed to recover from dissociation by incubation in defined media, this adhesive system is augmented by a Ca2+-dependent mechanism with maximum recovery seen after 4 h incubation. The Ca2+-independent adhesion system is inhibited by preincubation of cell monolayers with cycloheximide before dissociation. Aggregation is also reduced after exposure to monensin, implicating a role for surface-translocated glycoproteins in this mechanism of adhesion. In coaggregation experiments using C2 myoblasts and 3T3 fibroblasts in which the Ca2+-dependent adhesion system was inactivated, no adhesive specificity between the two cell types was seen. Although synthetic peptides containing the RGD sequence are known to inhibit cell-substratum adhesion in various cell types, incubation of C2 myoblasts with the integrin-binding tetrapeptide, RGDS, greatly stimulated the Ca2+-independent aggregation of these cells while control analogs had no effect. These results show that a Ca2+- independent mechanism alone is sufficient to allow for the rapid formation of multicellular aggregates in a mouse myoblast line, and that many of the requirements and perturbants of the Ca2+-independent system of intercellular myoblast adhesion are similar to those of the Ca2+-dependent adhesion mechanisms.