Dissociation of Madin-Darby Canine Kidney Epithelial Cells by the Monoclonal Antibody Anti-Arc-l: Mechanistic Aspects and Identification of the Antigen as a Component Related to Uvomorulin

It has previously been shown that the monoclonal antibody anti-Arc-1 dissociates Madin-Darby canine kidney (MDCK) epithelial cells and changes their morphology in vitro (Imhof, B. A., H. P. Vollmers, S. L. Goodman, and W. Birchmeier, 1983, Cell, 35:667-675). In this article we demonstrate that the anti-Arc-1 antibody recognizes an uvomorulin-like molecule on MDCK cells, i.e., it immunoprecipitates an 84-kD protein fragment from a tryptic digest of cell surfaces in the presence of Ca 2+ (as does anti-uvomorulin antiserum). Furthermore, anti-uvomorulin antiserum prevents the binding of anti-Arc 1 to MDCK cells. The distribution of the Arc-1 antigen is also quite similar to that of uvomorulin: it is enriched at the cell-cell contacts both of MDCK cells and of cells in various canine tissues. In the intestinal epithelium the antigen could be further localized in the region of the junctional complex. To study the mechanism of action of the dissociating antibody, MDCK cells grown on Nuclepore filters in Boyden chambers were exposed to anti-Arc-1 from either the upper or lower compartment. It could be shown that the antibody interfered with cell adhesion only from the basolateral but not from the apical cell surface. Antibody action was inhibited in the presence of colchicine but not cytochalasin B. Furthermore, cell dissociation was prevented when the cellular cAMP level was raised. These findings indicate that the anti-Arc-1 antibody acts on a target below the tight junctions (possibly on the antigen located in the junctional complex), and they confirm that cytoskeleton and metabolic factors are actively involved in the maintenance of junctional integrity. In recent years the process of cell adhesion has been extensively studied. In one line of research, antibodies that specifically inhibit cell-cell contacts have been used to identify the components involved (1-6). The neural cell adhesion molecule (N-CAM) has been so identified, and it seems to form direct bridges between neural cells (1, 7, 8). Different types of adhesion components have been discovered in nonneuronal cells (liver cell adhesion molecule [L-CAM] 1 in chickens [9]; Abbreviations used in this paper. Con A, concanavalin A; L-CAM, liver cell adhesion molecule; MDCK, Madin-Darby canine kidney. THE JOURNAL Of: CELL BIOLOGY • VOLUME 101 OCTOBER 1985 1307-1315 © The Rockefeller University Press . 0021-9525/85/10/1307/09 $1.00 uvomorulin and cadherin in the mouse [2, 10, 11]; and cell adhesion molecule 120/80 in humans [12]). Their biochemical characteristics and their distribution in different tissues indicate that these adhesion components may all be closely related. Thus, L-CAM/uvomorulin-like adhesion molecules are found on cell surfaces and have molecular weights of -120,000, from which 84,000-mol-wt fragments can be cleaved by trypsin in the presence of Ca ~÷ (2, 13). They are enriched at cell contact areas in epithelial tissues (9), and uvomorulin has been localized by immunoelectron microscopy at the intermediate junctions of the intestinal epithelium 1307 on F ebuary 1, 2013 jcb.rress.org D ow nladed fom Published October 1, 1985