Previous studies have shown that collagen gel overlay induced selective proteolysis of focal adhesion complex proteins in Madin-Darby canine kidney (MDCK) cells. In this study, we examined whether morphological and biochemical changes were present in cells cultured on collagen gel. We found that focal adhesion complex proteins, including focal adhesion kinase (FAK), talin, paxillin, and p130cas , but not vinculin, were decreased within 1 h when MDCK cells were cultured on collagen gel. Collagen gel-induced selective decrease of focal adhesion proteins was observed in all lines of cells examined, including epithelial, fibroblastic, and cancer cells. Matrigel also induced selective down-regulation of focal adhesion proteins. However, cells cultured on collagen gel-or matrigel-coated dishes did not show any changes of focal adhesion proteins. These data suggest that the physical nature of the gel, i.e. the rigidity, is involved in the expression of focal adhesion proteins. The collagen gel-induced down-regulation of focal adhesion complex proteins was caused by reduction of protein synthesis and activation of proteases such as calpain. Overexpression of a dominant negative mutant of discoidin domain receptor 1 (DDR1) or FAK-related non-kinase (FRNK) did not prevent collagen gel-induced down-regulation of the focal adhesion complex protein, whereas an anti-␣21 integrin-neutralizing antibody completely blocked it. Taken together, our results indicate that the rigidity of collagen gel controls the expression of focal adhesion complex proteins, which is mediated by ␣ 2  1 integrin but not DDR1.
Adhesion of cells to the extracellular matrix (ECM)1 is a crucial event in multi-cellular organisms for the modulation of cellular processes such as cell growth, differentiation, and apoptosis (1-4). The integration of intracellular signaling and the structure of ECM elicited by ECM-integrin engagement may be mediated by focal adhesion complex proteins (5). Focal adhesion kinase (FAK) is a cytoplasmic non-receptor tyrosine kinase located close to focal adhesions and may play a central role in integrating signals from ECM-integrin and growth factors (6 -8). FAK also plays important roles in the assembly of several signaling proteins to focal adhesions via interactions with a number of cellular proteins, including Src, Grb2, phosphatidylinostiol-3 kinase, paxillin, Crk, talin, and p130 cas (6, 9 -13). Recent data have shown that FAK plays important roles in cell cycle progression (14), migration (8, 9, 15), adhesion (12), and the prevention of apoptosis (16, 17).Three-dimensional collagen gel has been used as a cell culture vehicle for the study of morphogenesis by us and many laboratories (18 -20). Collagen fibrils can transduce signals through integrins and the receptor tyrosine kinase discoidin domain receptor 1 (DDR1) (21). Interaction of ECM with its transmembrane receptor integrins causes subsequent cascades of protein-protein interaction and modification at the focal adhesion complex site and the recruitment of several cy...