Human colon carcinoma cells use multiple receptors to adhere to laminin: involvement of alpha 6 beta 4 and alpha 2 beta 1 integrins.

Abstract

In this study, we used clone A, a human colon carcinoma cell line, to characterize those integrins that mediate colon carcinoma adhesion to laminin. Monoclonal antibodies specific for the human fi1 subunit inhibited clone A adhesion to laminin. They also precipitated a complex of surface proteins that exhibited an electrophoretic behavior characteristic of a2fll and a3fi1. A monoclonal antibody specific for a2 (PIH5) blocked clone A adhesion to laminin, as well as to collagen 1. An a3-specific antibody (P1 B5) had no effect on clone A adhesion to laminin, even though it can block the adhesion of other cell types to laminin. Thus, the a2,61 integrin can function as both a laminin and collagen I receptor on clone A cells. Although these cells express a31, an established laminin receptor, they do not appear to use it to mediate laminin adhesion. In addition, the monoclonal antibody GoH3, which recognizes the a6 integrin subunit, also inhibited carcinoma adhesion to laminin but not to fibronectin or collagen 1. This antibody precipitated the a6 subunit in association with the fl4 subunit. There was no evidence of a6,B1 association on these cells. In summary, the results obtained in this study indicate that multiple integrin a subunits, in association with two distinct ,@ subunits, are involved in colon carcinoma adhesion to laminin. Based on the behavior of a3,#1 and a2,81, the results also suggest that cells can regulate the ability of a specific integrin to mediate adhesion.

The integrin alpha 6 beta 4 is a laminin receptor.

“…Antibodies to the a6 and 04 Integrin Subunits Inhibit Laminin Adhesion Previously, we reported that antibodies specific for the a6, a2, and /31 integrin subunits blocked clone A adhesion to laminin (Lotz et al ., 1990) . In this study, we examined the ability of the ß4 specific mAb A9 (Kimmel and Carey, 1986 ;van Waes et al ., 1991) to inhibit laminin adhesion .…”

“…1) were performed as described previously (Lotz et al ., 1990) . The antibody inhibition assays (shown in Fig.…”

“…The cells (1-2 x 109) were washed three times with PBS containing 40 mM 0 D-glucose . The pellet was resuspended in 2 ml of the PBS/glucose buffer and the cells were then surface radiolabeled using the lactoperoxidase/ i25 í method at 4°C as described previously (Lotz et al ., 1990) . Radiolabeled cells were solubilized in 50 mM Tris-HCl buffer, pH 7.4, containing 200 MM octyl-ß-D-glucopyranoside (Boehringer Mannheim Biochemicals, Mannheim, Germany), 2 mM phenylmethylsulfonyl fluoride (PMSF), 1 mM each of aprotinin, leupeptin, pepstatin, and 5 mM of the appropriate divalent cation .…”

“…In contrast, these cells adhere poorly, if at all, to tissue culture plastic ( Fig . 1 a) or fibronectin, although they do adhere well to collagen I (Lotz et al ., 1990) . This rapid adhesion of clone A cells to laminin is quite distinct from that which we observed for less invasive carcinoma cell lines (Daneker et al ., 1989), as well as for other cell types including 3T3 fibroblasts, bovine endothelial cells, PC12 cells, and mouse macrophages (not shown) .…”

“…This question has caused considerable controversy in the recent literature. Although we reported that a6ß4 can function as a laminin receptor on colon carcinoma cells based on the ability of an a6 specific antibody to block adhesion (Lotz et al, 1990), this function for a6ß4 has not been widely accepted . Several papers have concluded, for example, that the ligand for a6ß4 must be distinct from that of a6ß1 and have emphasized that a6ß4 has not been shown to bind laminin affinity columns (Deluca et al, 1990;Quaranta and Jones, 1991) .…”