The structural analysis of adhesions mediated by Ep-CAM

The epithelial cell adhesion molecule Ep-CAM is capable of mediating Ca2+-independent homotypic cell-cell adhesion when introduced into cells lacking their own means of cell-cell interactions. We used (confocal) immunofluorescent and (immuno-) electron microscopy to investigate the structural organization of Ep-CAM-mediated adhesions and their relation to other types of intercellular adhesions. Ep-CAM-transfected cell lines, cells of epithelial origin, and epithelial tissues were analyzed. In transfected L cells Ep-CAM brings the opposing intercellular membranes into a close proximity (approximately 10-14 nm) at sporadic contacts; however, no structures resembling junctional complexes were observed. In L cells cotransfected with Ep-CAM and E-cadherin, both molecules localize at the sites of cell-cell contact, forming independent adhesion sites with no Ep-CAM detectable within the structurally distinguishable cadherin-mediated adherens junctions. In well-differentiated carcinoma cell lines Ep-CAM colocalized with E-cadherin practically along the whole lateral domain; however, no colocalization was observed between Ep-CAM and the components of the tight junction complex (occludin and ZO-1), desmosomes (desmoplakins I/II), or cell-substrate adhesions (beta1 integrins). This was confirmed by analysis of polarized epithelium of normal colon where Ep-CAM was present at the lateral membrane including the adherens junction areas, but was fully excluded from the apical cell membrane (microvilli), tight junctions, and desmosomes. We conclude that (1) Ep-CAM does not form junctional complexes in L cells, (2) in epithelial cells, cell surface Ep-CAM is present at the lateral cell membrane, but is excluded from tight junctions and desmosomes, and (3) in epithelial cells, Ep-CAM is present within adhesions mediated by the classic cadherins (especially E-cadherin) with both types of molecules remaining as independent clusters. The colocalization with cadherins might be important for the modulating effect of Ep-CAM on cadherin-mediated adhesions.