Contribution of Cadherins to Directional Cell Migration and Histogenesis in Xenopus Embryos

Perturbation of adhesion mediated by cadherins was achieved by over-expressing truncated forms of E- and EP-cadherins (in which the extracellular domain was deleted) in different blastomeres of stage 6 Xenopus laevis embryos. Injections of mRNA encoding truncated E- and EP-cadherins into A1A2 blastomeres resulted in inhibition of cell adhesion and, at later stages, in morphogenetic defects in the anterior neural tissues to which they mainly contribute. In addition, truncated EP-cadherin mRNA produced a duplication of the dorso-posterior axis in a significant number of cases. The expression of truncated E- and EP-cadherins in blastomeres involved in gastrulation and neural induction (B1B2 and C1), led to the duplication of the dorso-posterior axis as well as to defects in anterior structures. Morphogenetic defects obtained with truncated EP-cadherin were more severe than those induced with truncated E-cadherin. Cells derived from blastomeres injected with truncated EP-cadherin mRNA, dispersed more readily at the blastula and gastrula stages than the cells derived from the blastomeres expressing truncated E-cadherin. Presumptive mesodermal cells expressing truncated cadherins did not engage in coherent directional migration. The alteration of cadherin-mediated cell adhesion led directly to the perturbation of the convergent-extension movements during gastrulation as shown in the animal cap assays and indirectly to perturbation of neural induction. Although the cytoplasmic domains of type I cadherins share a high degree of sequence identity, the over-expression of their cytoplasmic domains induces a distinct pattern of perturbations, strongly suggesting that in vivo, each cadherin may transduce a specific adhesive signal. These graded perturbations may in part result from the relative ability of each cadherin cytoplasmic domain to titer the P-catenin.     

Differential perturbations in the morphogenesis of anterior structures induced by overexpression of truncated XB- and N-cadherins in Xenopus embryos

Cadherins, a family of Ca-dependent adhesion molecules, have been proposed to act as regulators of morphogenetic processes and to be major effectors in the maintenance of tissue integrity. In this study, we have compared the effects of the expression of two truncated cadherins during early neurogenesis in Xenopus laevis. mRNA encoding deleted forms of XB- and N-cadherin lacking most of the extracellular domain were injected into the four animal dorsal blastomeres of 32-cell stage Xenopus embryos. These truncated cadherins altered the cohesion of cells derived from the injected blastomeres and induced morphogenetic defects in the anterior neural tissue to which they chiefly contributed. Truncated XB-cadherin was more efficient than N- cadherin in inducing these perturbations. Moreover, the coexpression of both truncated cadherins had additive perturbation effects on neural development. The two truncated cadherins can interact with the three known catenins, but with distinct affinities. These results suggest that the adhesive signal mediated by cadherins can be perturbed by overexpressing their cytoplasmic domains by competing with different affinity with catenins and/or a common anchor structure. Therefore, the correct regulation of cadherin function through the cytoplasmic domain appears to be a crucial step in the formation of the neural tissue.     

Cadherin-mediated cell sorting not determined by binding or adhesion specificity.

Cadherin adhesion molecules play important roles in the establishment of tissue boundaries. Cells expressing different cadherins sort out from each other in cell aggregation assays. To determine the contribution of cadherin binding and adhesion specificity to the sorting process, we examined the adhesion of cells to different purified cadherin proteins. Chinese hamster ovary cell lines expressing one of four different cadherins were allowed to bind to the purified cadherin extracellular domains of either human E-cadherin or Xenopus C-cadherin, and the specificity of adhesion was compared with cell-sorting assays. None of the different cadherin-expressing cells exhibited any adhesive specificity toward either of the two purified cadherin substrates, even though these cadherins differ considerably in their primary sequence. In addition, all cells exhibited similar strengthening of adhesion on both substrates. However, this lack of adhesive specificity did not determine whether different cadherin-expressing cells would sort from each other, and the tendency to sort was not predictable by the extent of sequence diversity in their extracellular domains. These results show that cadherins are far more promiscuous in their adhesive-binding capacity than had been expected and that the ability to sort out must be determined by mechanisms other than simple adhesive-binding specificity.     

The Intracellular Domain of Cadherin-11 is not Required for the Induction of Cell Aggregation,Adhesion or Gap-Junction Formation

The cadherin family of cell adhesion molecules demonstrates calcium-dependent hemophilic binding, leading to cellular recognition and adhesion. The adhesion mediated by the classical type 1 cadherins is strengthened through catenin-mediated coupling of the cytoplasmic domain to the cytoskeleton. This cytoskeletal interaction may not be essential for the adhesion promoted by all cadherins, several of which lack cytosolic catenin-binding sequences. Cadherin-11, a classical cadherin, possesses a cytoplasmic domain that interacts with catenins, but may also occur as a variant form expressing a truncated cytoplasmic domain. To study the role of the cytoplasmic sequence in cadherin-11 mediated adhesion we have constructed and expressed a truncated cadherin-11 protein lacking the cytoplasmic domain and unable to bind β-catenin. Expression of the truncated cadherin-11 in MDA-MB-435S human mammary carcinoma cells reduced their motility and promoted calcium-dependent cell aggregation, frequent cell contacts, and functional gap-junctions. We conclude that the intracellular catenin-binding domain of cadherin-11, and by inference cytoskeletal interaction, is not required for the initiation and formation of cell adhesion.     

The Intracellular Domain of Cadherin-11 is not Required for the Induction of Cell Aggregation, Adhesion or Gap-Junction Formation

The cadherin family of cell adhesion molecules demonstrates calcium-dependent hemophilic binding, leading to cellular recognition and adhesion. The adhesion mediated by the classical type 1 cadherins is strengthened through catenin-mediated coupling of the cytoplasmic domain to the cytoskeleton. This cytoskeletal interaction may not be essential for the adhesion promoted by all cadherins, several of which lack cytosolic catenin-binding sequences. Cadherin-11, a classical cadherin, possesses a cytoplasmic domain that interacts with catenins, but may also occur as a variant form expressing a truncated cytoplasmic domain. To study the role of the cytoplasmic sequence in cadherin-11 mediated adhesion we have constructed and expressed a truncated cadherin-11 protein lacking the cytoplasmic domain and unable to bind β-catenin. Expression of the truncated cadherin-11 in MDA-MB-435S human mammary carcinoma cells reduced their motility and promoted calcium-dependent cell aggregation, frequent cell contacts, and functional gap-junctions. We conclude that the intracellular catenin-binding domain of cadherin-11, and by inference cytoskeletal interaction, is not required for the initiation and formation of cell adhesion.     

Transmembrane control of cadherin-mediated cell adhesion: a 94 kDa protein functionally associated with a specific region of the cytoplasmic domain of E-cadherin.

Cadherins are a family of transmembrane glycoproteins which play a key role in Ca(2+)-dependent cell-cell adhesion. Cytoplasmic domains of these molecules are anchored to the cell cytoskeleton and are required for cadherin function. To elucidate how the function of cadherins is controlled through their cytoplasmic domains, we deleted five different regions in the cytoplasmic domain of E-cadherin. After transfecting L cells with cDNA encoding the mutant polypeptides, we assayed aggregating activity of these transfectants; all these mutant proteins were shown to have an extracellular domain with normal Ca(2+)-sensitivity and molecular weight. Two mutant polypeptides with deletions in the carboxy half of the cytoplasmic domain, however, did not promote cell-cell adhesion and had also lost the ability to bind to the cytoskeleton, whereas the mutant molecules with deletions of other regions retained the ability to promote cell adhesion and to anchor to the cytoskeleton. Thus, the cytoplasmic domain contains a subdomain which was involved in the cell adhesion and cytoskeleton-binding functions. When E-cadherin in F9 cells or in L cells transfected with wild-type or functional mutant cadherin polypeptides was solubilized with nonionic detergents and immunoprecipitated, two additional 94 and 102 kDa components were coprecipitated. The 94 kDa component, however, was not detected in the immunoprecipitates from cells expressing the mutant cadherins which had lost the adhesive function. These results suggest that the interaction of the carboxy half of the cytoplasmic domain with the 94 kDa component regulates the cell binding function of the extracellular domain of E-cadherin.     

Regulation of C-cadherin function during activin induced morphogenesis of Xenopus animal caps

Treatment of Xenopus animal pole tissue with activin results in the induction of mesodermal cell types and a dramatic elongation of the tissue. The morphogenetic movements involved in the elongation appear similar to those in normal gastrulation, which is driven by cell rearrangement and cell intercalations. We have used this system to explore the potential regulation of cell-cell adhesion and cadherin function during morphogenesis. Quantitative blastomere aggregation assays revealed that activin induction reduced the calcium-dependent adhesion between blastomeres. Activin-induced blastomeres formed smaller aggregates, and a greater proportion of the population remained as single cells compared to uninduced blastomeres. The aggregation was mediated by C-cadherin because C-cadherin was present in the blastomeres during the aggregation assay, and monoclonal antibodies against C-cadherin inhibited the calcium-dependent aggregation of blastomeres. E-cadherin was not detectable until after the completion of the assay and, therefore, does not explain the adhesive differences between induced and uninduced blastomeres. L cells stably expressing C- cadherin (LC cells) were used to demonstrate that C-cadherin activity was specifically altered after activin induction. Blastomeres induced with activin bound fewer LC cells than uninduced blastomers. L cells not expressing C-cadherin did not adhere to blastomeres. The changes in C-cadherin-mediated adhesion occurred without detectable changes in the steady-state levels of C-cadherin or the amount of C-cadherin present on the surface of the cell. Immunoprecipitation of C-cadherin and its associated catenins revealed that the ratio of C-cadherin and the catenins was not altered by activin induction. These results demonstrate that activin decreases the adhesive function of existing C- cadherin molecules on the surface of blastomeres and suggest that decreased cadherin mediated cell-cell adhesion is associated with increased morphogenetic movement.     

Expression of N-cadherin by human squamous carcinoma cells induces a scattered fibroblastic phenotype with disrupted cell-cell adhesion

E-cadherin is a transmembrane glycoprotein that mediates calcium- dependent, homotypic cell-cell adhesion and plays an important role in maintaining the normal phenotype of epithelial cells. Disruption of E- cadherin activity in epithelial cells correlates with formation of metastatic tumors. Decreased adhesive function may be implemented in a number of ways including: (a) decreased expression of E-cadherin; (b) mutations in the gene encoding E-cadherin; or (c) mutations in the genes that encode the catenins, proteins that link the cadherins to the cytoskeleton and are essential for cadherin mediated cell-cell adhesion. In this study, we explored the possibility that inappropriate expression of a nonepithelial cadherin by an epithelial cell might also result in disruption of cell-cell adhesion. We showed that a squamous cell carcinoma-derived cell line expressed N-cadherin and displayed a scattered fibroblastic phenotype along with decreased expression of E- and P-cadherin. Transfection of this cell line with antisense N- cadherin resulted in reversion to a normal-appearing squamous epithelial cell with increased E- and P-cadherin expression. In addition, transfection of a normal-appearing squamous epithelial cell line with N-cadherin resulted in downregulation of both E- and P- cadherin and a scattered fibroblastic phenotype. In all cases, the levels of expression of N-cadherin and E-cadherin were inversely related to one another. In addition, we showed that some squamous cell carcinomas expressed N-cadherin in situ and those tumors expressing N- cadherin were invasive. These studies led us to propose a novel mechanism for tumorigenesis in squamous epithelial cells; i.e., inadvertent expression of a nonepithelial cadherin.     

Expressed recombinant cadherins mediate cell sorting in model systems

Cadherins are cell-surface glycoproteins responsible for Ca2+-dependent cell-to-cell adhesion. E- or P-cadherin was transfected into L cells, which normally have little cadherin activity, and cellular aggregation of the resulting transfectants was observed to be a function of the cadherin molecule expressed. Transfected cells preferentially adhered to cells expressing the same cadherin subclass. Furthermore, in reconstituted embryonic lung tissue, E-cadherin-expressing L cells were associated with epithelial tubules expressing E-cadherin, while untransfected L cells associated with mesenchymal cells. These results provide the first direct evidence that the differential expression of cadherins can play a role in cell sorting in heterogeneous cell populations.     

EP-cadherin in muscles and epithelia of Xenopus laevis embryos.

EP-cadherin is a novel Xenopus Ca+2-dependent adhesion molecule, which shares comparable homology with mouse E- and P-cadherins (Ginsberg, De Simone and Geiger; 1991, Development 111, 315-325). We report here the patterns of expression of this molecule in Xenopus laevis embryos at different developmental stages ranging from cleavage to postmetamorphic. EP-cadherin is already expressed in the oocyte and egg and can then be detected in close association with the membrane of all blastomeres up to late blastula stages. Starting at late gastrula stages, the level of EP-cadherin expression increases sharply in non-neural ectodermal cells, in the somites and in the notochord; it persists in endodermal cells and decreases rapidly in all migratory cells. During neurulation the level of EP-cadherin expression declines gradually in the nervous system and is undetectable here throughout later development except in the optic nerve and in the neural part of the olfactory organ. This pattern continues during later development so that in the tailbud stage and up to metamorphosis the most prominent staining is detected in the epidermis and skeletal muscle. After metamorphosis, the molecule gradually disappears from the muscle tissue and the major site of expression remains the skin. EP-cadherin is invariably present in close association with the cell membrane. In the muscle it is associated with the sarcolemma at regions of myoblast-myoblast or myotube-myotube contact. In epidermal cells, EP-cadherin is usually coexpressed with E-cadherin. Yet, while E-cadherin staining is always restricted to the basolateral aspects of the cells, EP-cadherin is often distributed throughout the plasmalemma including the apical surface.     

Intracellular domain of desmoglein 3 (pemphigus vulgaris antigen) confers adhesive function on the extracellular domain of E-cadherin without binding catenins

For the extracellular (EC) domain of E-cadherin to function in homophilic adhesion it is thought that its intracytoplasmic (IC) domain must bind alpha- and beta-catenins, which link it to the actin cytoskeleton. However, the IC domain of pemphigus vulgaris antigen (PVA or Dsg3), which is in the desmoglein subfamily of the cadherin gene superfamily, does not bind alpha- or beta-catenins. Because desmogleins have also been predicted to function in the cell adhesion of desmosomes, we speculated that the PVA IC domain might be able to act in a novel way in conferring adhesive function on the EC domain of cadherins. To test this hypothesis we studied aggregation of mouse fibroblast L cell clones that expressed chimeric cDNAs encoding the EC domain of E-cadherin with various IC domains. We show here that the full IC domain of PVA as well as an IC subdomain containing only 40 amino acids of the PVA intracellular anchor (IA) region confer adhesive function on the E-cadherin EC domain without catenin-like associations with cytoplasmic molecules or fractionation with the cell cytoskeleton. This IA region subdomain is evolutionarily conserved in desmogleins, but not classical cadherins. These findings suggest an important cell biologic function for the IA region of desmogleins and demonstrate that strong cytoplasmic interactions are not absolutely necessary for E- cadherin-mediated adhesion.     

Localization of HIV RNA in mitochondria of infected cells: potential role in cytopathogenicity

A novel member of the cadherin family of cell adhesion molecules has been characterized by cloning from rat liver, sequencing of the corresponding cDNA, and functional analysis after heterologous expression in nonadhesive S2 cells. cDNA clones were isolated using a polyclonal antibody inhibiting Ca(2+)-dependent intercellular adhesion of hepatoma cells. As inferred from the deduced amino acid sequence, the novel molecule has homologies with E-, P-, and N-cadherins, but differs from these classical cadherins in four characteristics. Its extracellular domain is composed of five homologous repeated domains instead of four characteristic for the classical cadherins. Four of the five domains are characterized by the sequence motifs DXNDN and DXD or modifications thereof representing putative Ca(2+)-binding sites of classical cadherins. In its NH2-terminal region, this cadherin lacks both the precursor segment and the endogenous protease cleavage site RXKR found in classical cadherins. In the extracellular EC1 domain, the novel cadherin contains an AAL sequence in place of the HAV sequence motif representing the common cell adhesion recognition sequence of E-, P-, and N-cadherin. In contrast to the conserved cytoplasmic domain of classical cadherins with a length of 150-160 amino acid residues, that of the novel cadherin has only 18 amino acids. Examination of transfected S2 cells showed that despite these structural differences, this cadherin mediates intercellular adhesion in a Ca(2+)-dependent manner. The novel cadherin is solely expressed in liver and intestine and was, hence, assigned the name LI-cadherin. In these tissues, LI- cadherin is localized to the basolateral domain of hepatocytes and enterocytes. These results suggest that LI-cadherin represents a new cadherin subtype and may have a role in the morphological organization of liver and intestine.     

The cadherin cytoplasmic domain is unstructured in the absence of beta-catenin. A possible mechanism for regulating cadherin turnover

Cadherins are single pass transmembrane proteins that mediate Ca(2+)-dependent homophilic cell-cell adhesion by linking the cytoskeletons of adjacent cells. In adherens junctions, the cytoplasmic domain of cadherins bind to beta-catenin, which in turn binds to the actin-associated protein alpha-catenin. The physical properties of the E-cadherin cytoplasmic domain and its interactions with beta-catenin have been investigated. Proteolytic sensitivity, tryptophan fluorescence, circular dichroism, and (1)H NMR measurements indicate that murine E-cadherin cytoplasmic domain is unstructured. Upon binding to beta-catenin, the domain becomes resistant to proteolysis, suggesting that it structures upon binding. Cadherin-beta-catenin complex stability is modestly dependent on ionic strength, indicating that, contrary to previous proposals, the interaction is not dominated by electrostatics. Comparison of 18 cadherin sequences indicates that their cytoplasmic domains are unlikely to be structured in isolation. This analysis also reveals the presence of PEST sequences, motifs associated with ubiquitin/proteosome degradation, that overlap the previously identified beta-catenin-binding site. It is proposed that binding of cadherins to beta-catenin prevents recognition of degradation signals that are exposed in the unstructured cadherin cytoplasmic domain, favoring a cell surface population of catenin-bound cadherins capable of participating in cell adhesion.     

N-cadherin is regulated by gonadal steroids in adult sexually dimorphic spinal motoneurons

Gonadal steroids influence the morphology and function of neurons in the adult spinal cord through cellular and molecular mechanisms that are largely unknown. The cadherins are cell adhesion molecules that participate in the formation and organization of the CNS during embryonic development, and recent evidence suggests that the cadherins continue to regulate neural structure and function in adulthood. Using degenerate oligonucleotides coding conserved regions of the catenin-binding domain of classical cadherins in a RT-PCR cloning strategy, we identified several cadherin subtypes, the most frequently cloned being N-, E-, and R-cadherin, suggesting that these are the major classical cadherin subtypes present in the adult male rat lumbosacral spinal cord. We then examined cadherin expression levels of these cadherin subtypes under steroid conditions known to induce plastic changes in spinal motoneurons. Semiquantitative PCR revealed that mRNA levels of N-cadherin, but not E-cadherin or R-cadherin, are elevated in castrated rats treated with testosterone, 17 beta-estradiol, or dihydrotestosterone relative to castrate rats not treated with steroids. Immunolocalization of N-cadherin revealed that steroid treatment increased N-cadherin expression levels in functionally related neural populations whose morphology and function are regulated by steroids. These results suggest a role for N-cadherin in steroid-induced neuroplastic change in the adult lumbar spinal cord.     

Prototypical type I E-cadherin and type II cadherin-7 mediate very distinct adhesiveness through their extracellular domains

Using a dual pipette assay that measures the force required to separate adherent cell doublets, we have quantitatively compared intercellular adhesiveness mediated by Type I (E- or N-cadherin) or Type II (cadherin-7 or -11) cadherins. At similar cadherin expression levels, cells expressing Type I cadherins adhered much more rapidly and strongly than cells expressing Type II cadherins. Using chimeric cadherins, we found that the extracellular domain exerts by far the dominant effect on cell adhesivity, that of E-cadherin conferring high adhesivity, and that of cadherin-7 conferring low adhesivity. Type I cadherins were incorporated to a greater extent into detergent-insoluble cytoskeletal complexes, and their cytoplasmic tails were much more effective in disrupting strong adherent junctions, suggesting that Type II cadherins form less stable complexes with beta-catenin. The present study demonstrates compellingly, for the first time, that cadherins are dramatically different in their ability to promote intercellular adhesiveness, a finding that has profound implications for the regulation of tissue morphogenesis.     

Regulation of embryonic cell adhesion by the cadherin cytoplasmic domain.

Differential adhesion between embryonic cells has been proposed to be mediated by a family of closely related glycoproteins called the cadherins. The cadherins mediate adhesion in part through an interaction between the cadherin cytoplasmic domain and intracellular proteins, called the catenins. To determine whether these interactions could regulate cadherin function in embryos, a form of N-cadherin was generated that lacks an extracellular domain. Expression of this mutant in Xenopus embryos causes a dramatic inhibition of cell adhesion. Analysis of the mutant phenotype shows that at least two regions of the N-cadherin cytoplasmic domain can inhibit adhesion and that the mutant cadherin can inhibit catenin binding to E-cadherin. These results suggest that cadherin-mediated adhesion can be regulated by cytoplasmic interactions and that this regulation may contribute to morphogenesis when emerging tissues coexpress several cadherin types.     

Association of p120, a tyrosine kinase substrate, with E- cadherin/catenin complexes

p120 was originally identified as a substrate of pp60src and several receptor tyrosine kinases, but its function is not known. Recent studies revealed that this protein shows homology to a group of proteins, beta-catenin/Armadillo and plakoglobin (gamma-catenin), which are associated with the cell adhesion molecules cadherins. In this study, we examined whether p120 is associated with E-cadherin using the human carcinoma cell line HT29, as well as other cell lines, which express both of these proteins. When proteins that copurified with E- cadherin were analyzed, not only alpha-catenin, beta-catenin, and plakoglobin but also p120 were detected. Conversely, immunoprecipitates of p120 contained E-cadherin and all the catenins, although a large subpopulation of p120 was not associated with E-cadherin. Analysis of these immunoprecipitates suggests that 20% or less of the extractable E- cadherin is associated with p120. When p120 immunoprecipitation was performed with cell lysates depleted of E-cadherin, beta-catenin was no longer coprecipitated, and the amount of plakoglobin copurified was greatly reduced. This finding suggests that there are various forms of p120 complexes, including p120/E-cadherin/beta-catenin and p120/E- cadherin/plakoglobin complexes; this association profile contrasts with the mutually exclusive association of beta-catenin and plakoglobin with cadherins. When the COOH-terminal catenin binding site was truncated from E-cadherin, not only beta-catenin but also p120 did not coprecipitate with this mutated E-cadherin. Immunocytological studies showed that p120 colocalized with E-cadherin at cell-cell contact sites, even after non-ionic detergent extraction. Treatment of cells with hepatocyte growth factor/scatter factor altered the level of tyrosine phosphorylation of p120 as well as of beta-catenin and plakoglobin. These results suggest that p120 associates with E-cadherin at its COOH-terminal region, but the mechanism for this association differs from that for the association of beta-catenin and plakoglobin with E-cadherin, and thus, that p120, whose function could be modulated by growth factors, may play a unique role in regulation of the cadherin- catenin adhesion system.     

A Short Core Region of E-cadherin is Essential for Catenin Binding and is Highly Phosphorylated

Classical cadherins associate with three cytoplasmic proteins, termed α,-β-and γ-catenin. This association mediates the attachment of cadherins to the microfilament network, which is believed to be of major importance for cadherin function. Deletion of the carboxyterminal 72-amino acid residues of E-cadherin had been previously shown to prevent catenin binding. Here we have analyzed additional mutants of E-cadherin with deletions within this region and identified a core region of 30 amino acids (E-cadherin pos. 832–862) essential for the interaction with catenins. Phosphorylation analysis of wild-type and mutant E-cadherin indicates that the catenin-binding domain is highly phosphorylated. In particular, the 30 amino acid region contains 8 serine residues which are well conserved among cadherins. To elucidate whether phosphorylation might be important for cadherin-catenin complex formation, site-directed mutagenesis experiments were performed. Partial substitutions of up to 5 of the 8 serine residues in the cluster had no influence on E-cadherin-catenin complex formation and E-cadherin mediated cell adhesion, although phosphorylation of E-cadherin was reduced. In contrast, substitution of the whole serine cluster completely abolished phosphorylation and affected complex formation with catenins. These results suggest that E-cadherin-catenin interaction may be regulated by phosphorylation of the catenin-binding domain, which might represent one molecular mechanism to regulate cadherin mediated cell adhesion.     

A novel cadherin cell adhesion molecule: its expression patterns associated with implantation and organogenesis of mouse embryos

The Ca2+-dependent cell adhesion molecules, termed cadherins, were previously divided into two subclasses, E- and N-types, with different adhesive specificity. In this study, we identified a novel class of cadherin, termed P-cadherin, using a visceral endoderm cell line PSA5- E. This cadherin was a 118,000-D glycoprotein and distinct from E- and N-cadherins in immunological specificity and molecular mass. In accord with these findings, cells with P-cadherin did not cross-adhere with cells with E-cadherin. P-Cadherin first appeared in developing mouse embryos in the extraembryonic ectoderm and the visceral endoderm at the egg cylinder stage and later was expressed in various tissues. The placenta and the uterine decidua most abundantly expressed this cadherin. The expression of P-cadherin was transient in many tissues, and its permanent expression was limited to certain tissues such as the epidermis, the mesothelium, and the corneal endothelium. When the tissue distribution of P-cadherin was compared with that of E-cadherin, we found that: each cadherin displayed a unique spatio-temporal pattern of expression; P-cadherin was co-expressed with E-cadherin in local regions of various tissues; and onset or termination of expression of P- cadherin was closely associated with connection or segregation of cell layers, as found with other cadherins. These results suggested that differential expression of multiple classes of cadherins play a role in implantation and morphogenesis of embryos by providing cells with heterogenous adhesive specificity.