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Epithelial cell adhesion molecule (Ep-CAM) modulates cell-cell interactions mediated by classic cadherins.

Litvinov SV, Balzar M, Winter MJ, Bakker HA, Briaire-de Bruijn IH, Prins F, Fleuren GJ, Warnaar SO - J. Cell Biol. (1997)

Bottom Line: Similarly, the detergent-insoluble fractions of alpha- and beta-catenins decreased in cells overexpressing Ep-CAM.While the total beta-catenin content remains unchanged, a reduction in total cellular alpha-catenin is observed as Ep-CAM expression increases.The ability of Ep-CAM to modulate the cadherin-mediated cell-cell interactions, as demonstrated in the present study, suggests a role for this molecule in development of the proliferative, and probably malignant, phenotype of epithelial cells, since an increase of Ep-CAM expression was observed in vivo in association with hyperplastic and malignant proliferation of epithelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Leiden University, Leiden 2300 RC, The Netherlands. slitvinov@pathology.medfac.leidenuniv.nl

ABSTRACT
The contribution of noncadherin-type, Ca2+-independent cell-cell adhesion molecules to the organization of epithelial tissues is, as yet, unclear. A homophilic, epithelial Ca2+-independent adhesion molecule (Ep-CAM) is expressed in most epithelia, benign or malignant proliferative lesions, or during embryogenesis. Here we demonstrate that ectopic Ep-CAM, when expressed in cells interconnected by classic cadherins (E- or N-cadherin), induces segregation of the transfectants from the parental cell type in coaggregation assays and in cultured mixed aggregates, respectively. In the latter assay, Ep-CAM-positive transfectants behave like cells with a decreased strength of cell-cell adhesion as compared to the parental cells. Using transfectants with an inducible Ep-CAM-cDNA construct, we demonstrate that increasing expression of Ep-CAM in cadherin-positive cells leads to the gradual abrogation of adherens junctions. Overexpression of Ep-CAM has no influence on the total amount of cellular cadherin, but affects the interaction of cadherins with the cytoskeleton since a substantial decrease in the detergent-insoluble fraction of cadherin molecules was observed. Similarly, the detergent-insoluble fractions of alpha- and beta-catenins decreased in cells overexpressing Ep-CAM. While the total beta-catenin content remains unchanged, a reduction in total cellular alpha-catenin is observed as Ep-CAM expression increases. As the cadherin-mediated cell-cell adhesions diminish, Ep-CAM-mediated intercellular connections become predominant. An adhesion-defective mutant of Ep-CAM lacking the cytoplasmic domain has no effect on the cadherin-mediated cell-cell adhesions. The ability of Ep-CAM to modulate the cadherin-mediated cell-cell interactions, as demonstrated in the present study, suggests a role for this molecule in development of the proliferative, and probably malignant, phenotype of epithelial cells, since an increase of Ep-CAM expression was observed in vivo in association with hyperplastic and malignant proliferation of epithelial cells.

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Segregation of Ep-CAM–positive LEC cell transfectants from the parental cells in multicellular aggregates. LEC-C  and LEC-Ep cells, labeled with fluorescent dyes PKH-26 and  PKH-2, respectively, were mixed at a 1:1 ratio, sedimented, and  allowed to form an aggregate. This aggregate, in which both cell  types were represented in a random pattern, was mechanically  dispersed, and the smaller aggregates obtained were further cultured in suspension for 24 h, fixed, and analyzed. The micrographs present optical cross sections at the equatorial area of the  aggregates after 24 h, as seen with a confocal microscope. The artificial colors were assigned to the cells depending on the color of  the fluorochrome and the cell type: LEC-C (red); LEC-Ep (white)  cells. The figures show similar cell patterning in different size aggregates in the range of <100 to ∼1,000 cells. A dark area in the  middle of some aggregates is an optically nontransparent zone.
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Figure 3: Segregation of Ep-CAM–positive LEC cell transfectants from the parental cells in multicellular aggregates. LEC-C and LEC-Ep cells, labeled with fluorescent dyes PKH-26 and PKH-2, respectively, were mixed at a 1:1 ratio, sedimented, and allowed to form an aggregate. This aggregate, in which both cell types were represented in a random pattern, was mechanically dispersed, and the smaller aggregates obtained were further cultured in suspension for 24 h, fixed, and analyzed. The micrographs present optical cross sections at the equatorial area of the aggregates after 24 h, as seen with a confocal microscope. The artificial colors were assigned to the cells depending on the color of the fluorochrome and the cell type: LEC-C (red); LEC-Ep (white) cells. The figures show similar cell patterning in different size aggregates in the range of <100 to ∼1,000 cells. A dark area in the middle of some aggregates is an optically nontransparent zone.

Mentions: When mixed as monocellular suspensions and sedimented together, LEC-C and LEC-Ep cells were able to establish connections in the pellet. The resulting large aggregate, formed by randomly distributed cells of both types, was mechanically dispersed into a number of smaller aggregates. After 24 h of culturing these aggregates in suspension, it was found that the LEC-C cells formed the tight core of the aggregates, with LEC-Ep cells forming the external layer. This structure was observed for all aggregates irrespective of their size, with the latter ranging from 100 to more than 1,000 cells (Fig. 3). When either LEC-C or LEC-Ep cells were mixed with the differentially labeled cells of self-type, no cell patterning was observed, indicating that segregation was unrelated to the labeling and other experimental procedures.


Epithelial cell adhesion molecule (Ep-CAM) modulates cell-cell interactions mediated by classic cadherins.

Litvinov SV, Balzar M, Winter MJ, Bakker HA, Briaire-de Bruijn IH, Prins F, Fleuren GJ, Warnaar SO - J. Cell Biol. (1997)

Segregation of Ep-CAM–positive LEC cell transfectants from the parental cells in multicellular aggregates. LEC-C  and LEC-Ep cells, labeled with fluorescent dyes PKH-26 and  PKH-2, respectively, were mixed at a 1:1 ratio, sedimented, and  allowed to form an aggregate. This aggregate, in which both cell  types were represented in a random pattern, was mechanically  dispersed, and the smaller aggregates obtained were further cultured in suspension for 24 h, fixed, and analyzed. The micrographs present optical cross sections at the equatorial area of the  aggregates after 24 h, as seen with a confocal microscope. The artificial colors were assigned to the cells depending on the color of  the fluorochrome and the cell type: LEC-C (red); LEC-Ep (white)  cells. The figures show similar cell patterning in different size aggregates in the range of <100 to ∼1,000 cells. A dark area in the  middle of some aggregates is an optically nontransparent zone.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2140211&req=5

Figure 3: Segregation of Ep-CAM–positive LEC cell transfectants from the parental cells in multicellular aggregates. LEC-C and LEC-Ep cells, labeled with fluorescent dyes PKH-26 and PKH-2, respectively, were mixed at a 1:1 ratio, sedimented, and allowed to form an aggregate. This aggregate, in which both cell types were represented in a random pattern, was mechanically dispersed, and the smaller aggregates obtained were further cultured in suspension for 24 h, fixed, and analyzed. The micrographs present optical cross sections at the equatorial area of the aggregates after 24 h, as seen with a confocal microscope. The artificial colors were assigned to the cells depending on the color of the fluorochrome and the cell type: LEC-C (red); LEC-Ep (white) cells. The figures show similar cell patterning in different size aggregates in the range of <100 to ∼1,000 cells. A dark area in the middle of some aggregates is an optically nontransparent zone.
Mentions: When mixed as monocellular suspensions and sedimented together, LEC-C and LEC-Ep cells were able to establish connections in the pellet. The resulting large aggregate, formed by randomly distributed cells of both types, was mechanically dispersed into a number of smaller aggregates. After 24 h of culturing these aggregates in suspension, it was found that the LEC-C cells formed the tight core of the aggregates, with LEC-Ep cells forming the external layer. This structure was observed for all aggregates irrespective of their size, with the latter ranging from 100 to more than 1,000 cells (Fig. 3). When either LEC-C or LEC-Ep cells were mixed with the differentially labeled cells of self-type, no cell patterning was observed, indicating that segregation was unrelated to the labeling and other experimental procedures.

Bottom Line: Similarly, the detergent-insoluble fractions of alpha- and beta-catenins decreased in cells overexpressing Ep-CAM.While the total beta-catenin content remains unchanged, a reduction in total cellular alpha-catenin is observed as Ep-CAM expression increases.The ability of Ep-CAM to modulate the cadherin-mediated cell-cell interactions, as demonstrated in the present study, suggests a role for this molecule in development of the proliferative, and probably malignant, phenotype of epithelial cells, since an increase of Ep-CAM expression was observed in vivo in association with hyperplastic and malignant proliferation of epithelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Leiden University, Leiden 2300 RC, The Netherlands. slitvinov@pathology.medfac.leidenuniv.nl

ABSTRACT
The contribution of noncadherin-type, Ca2+-independent cell-cell adhesion molecules to the organization of epithelial tissues is, as yet, unclear. A homophilic, epithelial Ca2+-independent adhesion molecule (Ep-CAM) is expressed in most epithelia, benign or malignant proliferative lesions, or during embryogenesis. Here we demonstrate that ectopic Ep-CAM, when expressed in cells interconnected by classic cadherins (E- or N-cadherin), induces segregation of the transfectants from the parental cell type in coaggregation assays and in cultured mixed aggregates, respectively. In the latter assay, Ep-CAM-positive transfectants behave like cells with a decreased strength of cell-cell adhesion as compared to the parental cells. Using transfectants with an inducible Ep-CAM-cDNA construct, we demonstrate that increasing expression of Ep-CAM in cadherin-positive cells leads to the gradual abrogation of adherens junctions. Overexpression of Ep-CAM has no influence on the total amount of cellular cadherin, but affects the interaction of cadherins with the cytoskeleton since a substantial decrease in the detergent-insoluble fraction of cadherin molecules was observed. Similarly, the detergent-insoluble fractions of alpha- and beta-catenins decreased in cells overexpressing Ep-CAM. While the total beta-catenin content remains unchanged, a reduction in total cellular alpha-catenin is observed as Ep-CAM expression increases. As the cadherin-mediated cell-cell adhesions diminish, Ep-CAM-mediated intercellular connections become predominant. An adhesion-defective mutant of Ep-CAM lacking the cytoplasmic domain has no effect on the cadherin-mediated cell-cell adhesions. The ability of Ep-CAM to modulate the cadherin-mediated cell-cell interactions, as demonstrated in the present study, suggests a role for this molecule in development of the proliferative, and probably malignant, phenotype of epithelial cells, since an increase of Ep-CAM expression was observed in vivo in association with hyperplastic and malignant proliferation of epithelial cells.

Show MeSH
Related in: MedlinePlus