Alteration of collagen-dependent adhesion, motility, and morphogenesis by the expression of antisense alpha 2 integrin mRNA in mammary cells

Abstract

Although integrins are known to mediate adhesive binding of cells to the extracellular matrix, their role in mediating cellular growth, morphology, and differentiation is less clear. To determine more directly the role of the alpha 2 beta 1 integrin, a collagen and laminin receptor, in mediating the collagen-dependent differentiation of mammary cells, we reduced expression of the integrin by the well differentiated human breast carcinoma cell line, T47D, by stably expressing alpha 2 integrin antisense mRNA. Flow cytometry demonstrated that the antisense-expressing clones had levels of alpha 2 beta 1 integrin on their surfaces that were decreased by 30–70%. Adhesion of antisense-expressing clones to both collagens I and IV was decreased relative to controls in a manner that correlated with the level of cell surface alpha 2 beta 1 integrin expression. Adhesion to fibronectin and laminin were not affected. Motility across collagen-coated filters in haptotaxis assays was increased for only those clones that exhibited intermediate levels of adhesion to collagen, suggesting that an intermediate density of cell-surface alpha 2 beta 1 integrin optimally supports cell motility. When cultured in three-dimensional collagen gels, T47D cells organized in a manner suggestive of a glandular epithelium. In contrast, antisense-expressing clones with decreased alpha 2 beta 1 integrin were not able to organize in three-dimensional collagen gels. The growth rate of T47D cells was reduced when the cells were cultured in three-dimensional collagen gels. Unlike adhesion, motility, and morphogenesis, growth rates were unaffected by reduction of alpha 2 beta 1 integrin expression. Our results suggest that adhesive interactions mediated by a critical level of surface alpha 2 beta 1 integrin expression are key determinants of the collagen-dependent morphogenetic capacity of mammary epithelial cells.