Expression and function of cell adhesion molecules in adherent and non-adherent Ehrlich ascites tumor cells.

Abstract

The differences in adhesiveness, laminin and fibronectin synthesis, and cell surface carbohydrates between adherent and non-adherent Ehrlich ascites tumor (EAT) cells have been studied. The adherent EAT (a-EAT) cells adhere to and spread on laminin- or fibronectin-coated plates, whereas the non-adherent EAT (na-EAT) cells do not. The adhesion of a-EAT cells to laminin or fibronectin requires the presence of both Ca$\sp{2+}$ and Mg$\sp{2+}.$ Anti-human fibronectin receptor $(\alpha\sb5\beta\sb1$ integrin) antiserum blocks a-EAT cells from adhering to both laminin- and fibronectin-coated plates. An RGD-containing peptide (GRGDSP) inhibits the adhesion of a-EAT cells to fibronectin-coated but not laminin-coated plates. Taken together these data suggest that a-EAT cells interact with laminin and fibronectin via different receptors which belong to the $\beta$-1 subfamily of integrin receptors. Both na-EAT and a-EAT cells express similar amounts of integrins on their surface. The integrins expressed by these two types of EAT cells appear to have the same molecular weight and affinity toward immobilized laminin and fibronectin, even though na-EAT cells do not adhere to laminin or fibronectin. a-EAT cells adhere to one another in the absence of Ca$\sp{2+}$ and Mg$\sp{2+},$ whereas na-EAT cells do not. N-CAMs play at least a part role, if not all, in mediating the Ca$\sp{2+}$-independent cell-cell adhesion between a-EAT cells. The staining of a-EAT cells with FITC-labeled antibody against anti-N-CAM antibody concentrates at the contact sites between the cells. Although na-EAT cells do not adhere to one another, they express the same amount of N-CAMs on their surface as a-EAT cells. These N-CAMs show the same molecular weight as those expressed on a-EAT cells as revealed by immunoblotting. Both types of EAT cells express $\alpha$-Gal and Neu5Ac $\alpha$ 2,3 Gal residues on their surface. Only a-EAT cells express Neu5Ac $\alpha$ 2,6 Gal units as revealed by the reactivity with the lectins. However, this difference on the cell surface does not appear to be responsible for the differences in cell adhesion between these two types of EAT cells. Both types of EAT cells adhere to MAL-coated surfaces which react with Neu5Ac $\alpha$ 2,3 Gal residues, but only a-EAT cells spread on these surfaces. Each type of EAT cells secrete laminin and fibronectin into the medium. na-EAT cells secrete more laminin and less fibronectin compared with a-EAT cells. Both cell types express surface-bound laminin, and neither type expresses fibronectin on their surface. Immunoblotting of the lysates of both types of cells with monoclonal anti-phosphotyrosine antibody revealed that there are several differences in protein tyrosine phosphorylation between these two types of EAT cells. One phosphorylated protein of $M\sb{r}$ 220 kDa was found exclusively in na-EAT cells. Whether or not these differences in protein phosphorylation is the reason that causes na-EAT cells to lose their adhesiveness remains unknown at this time.