scholarly journals The roles of catenins in the cadherin-mediated cell adhesion: functional analysis of E-cadherin-alpha catenin fusion molecules.

1994 ◽  
Vol 127 (1) ◽  
pp. 235-245 ◽  
Author(s):  
A Nagafuchi ◽  
S Ishihara ◽  
S Tsukita
Keyword(s):  
Cell Adhesion ◽  
Cell Movement ◽  
Negative Regulator ◽  
Beta Catenin ◽  
Amino Terminal ◽  
Cytoplasmic Proteins ◽  
The Difference ◽  
Adhesion Activity ◽  
Carboxy Terminal ◽  
E Cadherin

The carboxyl terminus-truncated cadherin (nonfunctional cadherin) has no cell adhesion activity probably because of its failure to associate with cytoplasmic proteins called alpha and beta catenin. To rescue this nonfunctional cadherin as adhesion molecules, we constructed three cDNAs for fusion proteins between nonfunctional E-cadherin and alpha catenin, nE alpha, nE alpha N, and nE alpha C, where the intact, amino-terminal and carboxy-terminal half of alpha catenin, respectively, were directly linked to the nonfunctional E-cadherin, and introduced them into mouse L cells. The subcellular distribution and cell adhesion activity of nE alpha and nE alpha C molecules was similar to those of intact E-cadherin transfectants: they bound to cytoskeletons, were concentrated at cell-cell adhesion sites and showed strong cell adhesion activity. nE alpha N molecules, which also bound to cytoskeletons, showed very poor cell adhesion activity. Taken together, we conclude that in the formation of the cadherin-catenin complex, the mechanical association of alpha catenin, especially its carboxy-terminal half, with E-cadherin is a key step for the cadherin-mediated cell adhesion. Close comparison revealed that the behavior of nE alpha molecules during cytokinesis was quite different from that of intact E-cadherin, and that the intercellular motility, i.e., the cell movement in a confluent sheet, was significantly suppressed in nE alpha transfectants although it was facilitated in E-cadherin transfectants. Considering that nE alpha was not associated with endogenous beta catenin in transfectants, the difference in the nature of cell adhesion between nE alpha and intact E-cadherin transfectants may be explained by the function of beta catenin. The possible functions of beta catenin are discussed with a special reference to its role as a negative regulator for the cadherin-mediated cell adhesion system.

scholarly journals The Catenin p120ctnInteracts with Kaiso, a Novel BTB/POZ Domain Zinc Finger Transcription Factor

1999 ◽  
Vol 19 (5) ◽  
pp. 3614-3623 ◽  
Author(s):  
Juliet M. Daniel ◽  
Albert B. Reynolds
Keyword(s):  
Transcription Factor ◽  
Monoclonal Antibodies ◽  
Cell Adhesion ◽  
Mammalian Cells ◽  
Yeast Two Hybrid ◽  
Amino Terminal ◽  
Juxtamembrane Region ◽  
Carboxy Terminal ◽  
Two Hybrid ◽  
E Cadherin

ABSTRACT p120 ctn is an Armadillo repeat domain protein with structural similarity to the cell adhesion cofactors β-catenin and plakoglobin. All three proteins interact directly with the cytoplasmic domain of the transmembrane cell adhesion molecule E-cadherin; β-catenin and plakoglobin bind a carboxy-terminal region in a mutually exclusive manner, while p120 binds the juxtamembrane region. Unlike β-catenin and plakoglobin, p120 does not interact with α-catenin, the tumor suppressor adenomatous polyposis coli (APC), or the transcription factor Lef-1, suggesting that it has unique binding partners and plays a distinct role in the cadherin-catenin complex. Using p120 as bait, we conducted a yeast two-hybrid screen and identified a novel transcription factor which we named Kaiso. Kaiso’s deduced amino acid sequence revealed an amino-terminal BTB/POZ protein-protein interaction domain and three carboxy-terminal zinc fingers of the C2H2 DNA-binding type. Kaiso thus belongs to a rapidly growing family of POZ-ZF transcription factors that include the Drosophila developmental regulators Tramtrak and Bric à brac, and the human oncoproteins BCL-6 and PLZF, which are causally linked to non-Hodgkins’ lymphoma and acute promyelocytic leukemia, respectively. Monoclonal antibodies to Kaiso were generated and used to immunolocalize the protein and confirm the specificity of the p120-Kaiso interaction in mammalian cells. Kaiso specifically coprecipitated with a variety of p120-specific monoclonal antibodies but not with antibodies to α- or β-catenin, E-cadherin, or APC. Like other POZ-ZF proteins, Kaiso localized to the nucleus and was associated with specific nuclear dots. Yeast two-hybrid interaction assays mapped the binding domains to Arm repeats 1 to 7 of p120 and the carboxy-terminal 200 amino acids of Kaiso. In addition, Kaiso homodimerized via its POZ domain but it did not heterodimerize with BCL-6, which heterodimerizes with PLZF. The involvement of POZ-ZF proteins in development and cancer makes Kaiso an interesting candidate for a downstream effector of cadherin and/or p120 signaling.

scholarly journals Role of Nectin in Formation of E-Cadherin–based Adherens Junctions in Keratinocytes: Analysis with the N-Cadherin Dominant Negative Mutant

2003 ◽  
Vol 14 (4) ◽  
pp. 1597-1609 ◽  
Author(s):  
Yoshinari Tanaka ◽  
Hiroyuki Nakanishi ◽  
Shigeki Kakunaga ◽  
Noriko Okabe ◽  
Tomomi Kawakatsu ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Adhesion Molecule ◽  
Cell Adhesion Molecule ◽  
Adherens Junctions ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Adhesion Activity ◽  
Negative Mutant ◽  
E Cadherin ◽  
Cell Cell

E-Cadherin is a Ca2+-dependent cell-cell adhesion molecule at adherens junctions (AJs) of epithelial cells. A fragment of N-cadherin lacking its extracellular region serves as a dominant negative mutant (DN) and inhibits cell-cell adhesion activity of E-cadherin, but its mode of action remains to be elucidated. Nectin is a Ca2+-independent immunoglobulin-like cell-cell adhesion molecule at AJs and is associated with E-cadherin through their respective peripheral membrane proteins, afadin and catenins, which connect nectin and cadherin to the actin cytoskeleton, respectively. We showed here that overexpression of nectin capable of binding afadin, but not a mutant incapable of binding afadin, reduced the inhibitory effect of N-cadherin DN on the cell-cell adhesion activity of E-cadherin in keratinocytes. Overexpressed nectin recruited N-cadherin DN to the nectin-based cell-cell adhesion sites in an afadin-dependent manner. Moreover, overexpression of nectin enhanced the E-cadherin–based cell-cell adhesion activity. These results suggest that N-cadherin DN competitively inhibits the association of the endogenous nectin-afadin system with the endogenous E-cadherin-catenin system and thereby reduces the cell-cell adhesion activity of E-cadherin. Thus, nectin plays a role in the formation of E-cadherin–based AJs in keratinocytes.

Expression of a dominant negative cadherin mutant inhibits proliferation and stimulates terminal differentiation of human epidermal keratinocytes

1996 ◽  
Vol 109 (13) ◽  
pp. 3013-3023 ◽  
Author(s):  
A.J. Zhu ◽  
F.M. Watt
Keyword(s):  
Cell Adhesion ◽  
Terminal Differentiation ◽  
Intercellular Adhesion ◽  
Dominant Negative ◽  
Epidermal Keratinocytes ◽  
Dominant Negative Mutant ◽  
Beta Catenin ◽  
Human Epidermal Keratinocytes ◽  
Negative Mutant ◽  
E Cadherin

Cell adhesion molecules are not only required for maintenance of tissue integrity, but also regulate many aspects of cell behaviour, including growth and differentiation. While the regulatory functions of integrin extracellular matrix receptors in keratinocytes are well established, such functions have not been investigated for the primary receptors that mediate keratinocyte intercellular adhesion, the cadherins. To examine cadherin function in normal human epidermal keratinocytes we used a retroviral vector to introduce a dominant negative E-cadherin mutant, consisting of the extracellular domain of H-2Kd and the transmembrane and cytoplasmic domains of E-cadherin. As a control a vector containing the same construct, but with the catenin binding site destroyed, was prepared. High levels of expression of the constructs were achieved; the dominant negative mutant, but not the control, formed complexes with alpha-, beta- and gamma-catenin. In cells expressing the dominant negative mutant there was a 5-fold decrease in the level of endogenous cadherins and a 3-fold increase in the level of beta-catenin. Cell-cell adhesion and stratification were inhibited by the dominant negative mutant and desmosome formation was reduced. Expression of the mutant resulted in reduced levels of the alpha 2 beta 1 and alpha 3 beta 1 integrins and increased cell motility, providing further evidence for cross-talk between cadherins and the beta 1 integrins. In view of the widely documented loss of E-cadherin in keratinocyte tumours it was surprising that the dominant negative mutant had an inhibitory effect on keratinocyte proliferation and stimulated terminal differentiation even under conditions in which intercellular adhesion was prevented. These results establish a role for cadherins in regulating keratinocyte growth and differentiation and raise interesting questions as to the relative importance of cell adhesion-dependent and -independent mechanisms.

Tyrosine phosphorylation of p120(ctn) in v-Src transfected L cells depends on its association with E-cadherin and reduces adhesion activity

2001 ◽  
Vol 114 (3) ◽  
pp. 503-512 ◽  
Author(s):  
M. Ozawa ◽  
T. Ohkubo
Keyword(s):  
Cell Adhesion ◽  
Complex Formation ◽  
Tyrosine Phosphorylation ◽  
Chimeric Protein ◽  
Single Amino Acid ◽  
Membrane Localization ◽  
Wild Type ◽  
L Cells ◽  
Adhesion Activity ◽  
E Cadherin

Cadherins are transmembrane glycoproteins involved in Ca2+-dependent cell-cell adhesion. Using L cells expressing one of three functional E-cadherin constructs, the wild-type, a chimeric molecule with alpha-catenin (EalphaC), and a tail-less one, we determined the effect of v-Src expression on E-cadherin-mediated adhesion. The aggregation of L cells expressing the wild-type or EalphaC chimeric protein, which both interact with p120(ctn), was reduced by v-Src expression, whereas that of L cells expressing the tail-less E-cadherin was not affected by the expression. Tyrosine phosphorylation of p120(ctn) was observed in v-Src-transformed L cells expressing the wild-type or EalphaC chimeric protein, but not in ones expressing the tail-less E-cadherin. Thus, tyrosine phosphorylation of p120(ctn) depends on the complex formation with E-cadherin and the resulting membrane localization. Constitutive phosphorylation of p120(ctn) on serine and threonine residues also depends on the complex formation and membrane localization. Coexpression of the p120(ctn) protein with an N-terminal deletion, which eliminates some potential tyrosine phosphorylation sites, or the protein with a single amino acid substitution (tyrosine at 217 to phenylalanine) resulted in an increase in the aggregation of v-Src-transformed EL and EalphaCL cells. These results indicate that tyrosine phosphorylation of p120(ctn) is involved in the v-Src modulation of E-cadherin-mediated cell adhesion.

Catenins in Xenopus embryogenesis and their relation to the cadherin-mediated cell-cell adhesion system

Development ◽  
1993 ◽  
Vol 118 (2) ◽  
pp. 629-640 ◽  
Author(s):  
S. Schneider ◽  
K. Herrenknecht ◽  
S. Butz ◽  
R. Kemler ◽  
P. Hausen
Keyword(s):  
Cell Adhesion ◽  
Plasma Membranes ◽  
Early Embryo ◽  
Xenopus Embryo ◽  
Free Form ◽  
Beta Catenin ◽  
Metabolic Labelling ◽  
Adhesion System ◽  
E Cadherin ◽  
Cell Cell

In the course of an analysis of cell-cell adhesion in the Xenopus embryo, antibodies directed against alpha- and beta-catenin were applied to investigate their relation to the cadherins occurring early in this system. The results demonstrate that alpha- and beta-catenin are provided maternally and increase in amount throughout embryogenesis. Immunoprecipitations indicate that both of the catenins are complexed to U-cadherin in the early phase of embryogenesis and to E-cadherin, when it appears during gastrulation. An excess of alpha-catenin occurs in free form in the early embryo, whereas all of the beta-catenin seems to be complexed to cadherin. Synthesis of the two components throughout early embryogenesis and their binding to newly synthesized cadherins were demonstrated by metabolic labelling. The spatial distribution of alpha-catenin was analysed by immunohistology. During cleavage alpha-catenin is deposited evenly along the plasma membranes within the embryo, while the cell peripheries at the surface of the embryo remain devoid of alpha-catenin. At later stages, the pattern of alpha-catenin distribution becomes more complex. Quantitative differences in the intensity of staining along the plasma membranes in the different regions of the embryo can be distinguished. Particularly the appearance of E-cadherin in the gastrula ectoderm is accompanied by conspicuous depositions of alpha-catenin along the respective plasma membranes in this layer. All cells in the later embryo, apart from the neural crest cells, carry alpha-catenin on their plasma membranes indicating the universal character of cadherin-mediated cell-cell adhesion in the Xenopus embryo.

scholarly journals THE FUNCTIONAL DISSECTION OF AN ANTIGEN MOLECULE: SPECIFICITY OF HUMORAL AND CELLULAR IMMUNE RESPONSES TO GLUCAGON

1971 ◽  
Vol 133 (6) ◽  
pp. 1294-1308 ◽  
Author(s):  
George Senyk ◽  
E. Brady Williams ◽  
Danute E. Nitecki ◽  
Joel W. Goodman
Keyword(s):  
Lymphoid Cells ◽  
Delayed Hypersensitivity ◽  
Response Patterns ◽  
Amino Terminal ◽  
Transforming Activity ◽  
The Difference ◽  
Different Response ◽  
Carboxy Terminal ◽  
Cellular Immune ◽  
Immune Assays

Bovine glucagon, a polypeptide of 29 amino acids, was immunogenic in guinea pigs. The immunologic determinants of glucagon were investigated using isolated tryptic peptides of the hormone. Antibodies from virtually all of more than two dozen animals had specificity primarily for the amino-terminal heptadecapeptide (NM) and showed little or no binding with the carboxy-terminal undeca- and dodecapeptides (C). The smallest synthetic peptide of a series initiated at residue 16 which measurably bound antibody comprised residues 5–16 of glucagon. In cellular immune assays, both NM and C elicited delayed cutaneous reactions and inhibited the migration of peritoneal cells from immune animals. However, only intact glucagon and its C fragment stimulated lymphoid cells to synthesize DNA. While glucagon was somewhat more active than C, the addition of NM to C did not enhance its transforming activity. The smallest synthetic carboxy-terminal peptide with discernible transforming activity comprised residues 19–29 of glucagon. In both native and synthetic C peptide preparations, the undecapeptide was generally more active than the dodecapeptide, although cells from different animals gave different response patterns. The difference between the two is the presence of arginine at the amino-terminus of the peptide chain. Thus, the recognition specificity of populations of antigen-reactive cells from different animals displays a variation which is at least superficially analogous to that of populations of antibody molecules. In limited experiments using NM and C peptides as immunogens, neither gave rise to delayed hypersensitivity or to glucagon-binding circulating antibody, following a regimen which invariably provoked these responses when glucagon itself served as the immunogen. These results indicate that glucagon was cleaved by trypsin along functional lines into two parts, one of which housed the major antigenic determinant and the other of which carried the major immunogenic determinant, and they are highly compatible with a two-cell mechanism of immune induction. An apparent dissociation between the capacity to provoke delayed hypersensitivity reactions and to transform antigen-reactive cells in culture was observed.

scholarly journals E-cadherin to P-cadherin switching in lobular breast cancer with tubular elements

2020 ◽  
Vol 33 (12) ◽  
pp. 2483-2498 ◽  
Author(s):  
Matthias Christgen ◽  
Stephan Bartels ◽  
Jana L. van Luttikhuizen ◽  
Janin Bublitz ◽  
Luisa U. Rieger ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Adhesion ◽  
Tumor Cells ◽  
Lobular Carcinoma ◽  
Beta Catenin ◽  
Lobular Breast Cancer ◽  
Mutational Status ◽  
Invasive Lobular Breast Cancer ◽  
Reference Cohort ◽  
E Cadherin

AbstractLoss of E-cadherin expression due to mutation of the CDH1 gene is a characteristic feature of invasive lobular breast cancer (ILBC). Beta-catenin, which binds to the cytoplasmic domain of E-cadherin, is simultaneously downregulated, reflecting disassembly of adherens junctions (AJs) and loss of cell adhesion. E-cadherin to P-cadherin expression switching can rescue AJs and cell adhesion. However, P-cadherin has not been implicated in ILBC, so far. We aimed to characterize 13 ILBCs with exceptional histomorphology, which we termed ILBCs with tubular elements. The CDH1 mutational status was determined by next generation sequencing and whole-genome copy number (CN) profiling. Expression of cadherins was assessed by immunohistochemistry. ILBCs with tubular elements were ER-positive (13/13) and HER2-negative (13/13) and harbored deleterious CDH1 mutations (11/13) accompanied by loss of heterozygosity due to deletion of chromosome 16q22.1 (9/11). E-cadherin expression was lost or reduced in noncohesive tumor cells and in admixed tubular elements (13/13). Beta-catenin expression was lost in noncohesive tumor cells, but was retained in tubular elements (11/13), indicating focal rescue of AJ formation. N-cadherin and R-cadherin were always negative (0/13). Strikingly, P-cadherin was commonly positive (12/13) and immunoreactivity was accentuated in tubular elements. Adjacent lobular carcinoma in situ (LCIS) was always P-cadherin-negative (0/7). In a reference cohort of LCIS specimens, P-cadherin was constantly not expressed (0/25). In a reference cohort of invasive mammary carcinomas, P-cadherin-positive cases (36/268, 13%) were associated with triple-negative nonlobular breast cancer (P < 0.001). Compared with ILBCs from the reference cohort, P-cadherin expression was more common in ILBCs with tubular elements (12/13 versus 7/84, P < 0.001). In summary, E-cadherin to P-cadherin switching occurs in a subset of ILBCs. P-cadherin is the molecular determinant of a mixed-appearing histomorphology in ILBCs with tubular elements.

Characterization of the interactions of alpha-catenin with alpha-actinin and beta-catenin/plakoglobin

1997 ◽  
Vol 110 (8) ◽  
pp. 1013-1022 ◽  
Author(s):  
J.E. Nieset ◽  
A.R. Redfield ◽  
F. Jin ◽  
K.A. Knudsen ◽  
K.R. Johnson ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Protein Interactions ◽  
Beta Catenin ◽  
Protein Protein Interactions ◽  
Yeast Two Hybrid ◽  
Calcium Dependent ◽  
Cytoplasmic Proteins ◽  
Dependent Cell ◽  
Two Hybrid ◽  
Cell Cell

Cadherins are calcium-dependent, cell surface glycoproteins involved in cell-cell adhesion. To function in cell-cell adhesion, the transmembrane cadherin molecule must be associated with the cytoskeleton via cytoplasmic proteins known as catenins. Three catenins, alpha-catenin, beta-catenin and gamma-catenin (also known as plakoglobin), have been identified. beta-catenin or plakoglobin is associated directly with the cadherin; alpha-catenin binds to beta-catenin/plakoglobin and serves to link the cadherin/catenin complex to the actin cytoskeleton. The domains on the cadherin and betacatenin/plakoglobin that are responsible for protein-protein interactions have been mapped. However, little is known about the molecular interactions between alpha-catenin and beta-catenin/plakoglobin or about the interactions between alpha-catenin and the cytoskeleton. In this study we have used the yeast two-hybrid system to map the domains on alpha-catenin that allow it to associate with beta-catenin/plakoglobin and with alpha-actinin. We also identify a region on alpha-actinin that is responsible for its interaction with alpha-catenin. The yeast two-hybrid data were confirmed with biochemical studies.

scholarly journals Embryonic axis induction by the armadillo repeat domain of beta-catenin: evidence for intracellular signaling.

1995 ◽  
Vol 128 (5) ◽  
pp. 959-968 ◽  
Author(s):  
N Funayama ◽  
F Fagotto ◽  
P McCrea ◽  
B M Gumbiner
Keyword(s):  
Cell Adhesion ◽  
Intracellular Signaling ◽  
Ventral Side ◽  
Xenopus Embryo ◽  
Protein Interaction Data ◽  
Beta Catenin ◽  
Repeat Region ◽  
Amino Terminal ◽  
Repeat Domain ◽  
Armadillo Repeat

beta-catenin was identified as a cytoplasmic cadherin-associated protein required for cadherin adhesive function (Nagafuchi, A., and M. Takeichi. 1989. Cell Regul. 1:37-44; Ozawa, M., H. Baribault, and R. Kemler. 1989. EMBO [Eur. Mol. Biol. Organ.] J. 8:1711-1717). Subsequently, it was found to be the vertebrate homologue of the Drosophila segment polarity gene product Armadillo (McCrea, P. D., C. W. Turck, and B. Gumbiner. 1991. Science [Wash. DC]. 254:1359-1361; Peifer, M., and E. Wieschaus. 1990. Cell. 63:1167-1178). Also, antibody perturbation experiments implicated beta-catenin in axial patterning of the early Xenopus embryo (McCrea, P. D., W. M. Brieher, and B. M. Gumbiner. 1993. J. Cell Biol. 123:477-484). Here we report that overexpression of beta-catenin in the ventral side of the early Xenopus embryo, by injection of synthetic beta-catenin mRNA, induces the formation of a complete secondary body axis. Furthermore, an analysis of beta-catenin deletion constructs demonstrates that the internal armadillo repeat region is both necessary and sufficient to induce axis duplication. This region interacts with C-cadherin and with the APC tumor suppressor protein, but not with alpha-catenin, that requires the amino-terminal region of beta-catenin to bind to the complex. Since alpha-catenin is required for cadherin-mediated adhesion, the armadillo repeat region alone probably cannot promote cell adhesion, making it unlikely that beta-catenin induces axis duplication by increasing cell adhesion. We propose, rather, that beta-catenin acts in this circumstance as an intracellular signaling molecule. Subcellular fractionation demonstrated that all of the beta-catenin constructs that contain the armadillo repeat domain were present in both the soluble cytosolic and the membrane fraction. Immunofluorescence staining confirmed the plasma membrane and cytoplasmic localization of the constructs containing the armadillo repeat region, but revealed that they also accumulate in the nucleus, especially the construct containing only the armadillo repeat domain. These findings and the beta-catenin protein interaction data offer several intriguing possibilities for the site of action or the protein targets of beta-catenin signaling activity.

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