Identification and Characterization of the Integrin α₂β₁ Binding Motif in Chondroadherin Mediating Cell Attachment*

Chondroadherin is a leucine-rich repeat protein known to mediate adhesion of isolated cells via the integrin α₂β₁ and to interact with collagen. In this work, we show that cell adhesion to chondroadherin leads to activation of MAPKs but does not result in cell spreading and division. This is in contrast to the spreading and dividing of cells grown on collagen, although the binding is mediated via the same α₂β₁ receptor. We identified a cell binding motif, CQLRGLRRWLEAK³¹⁸ by mass spectrometry after protease digestion of chondroadherin. Cells adhering to the synthetic peptide CQLRGLRRWLEAK³¹⁸ remained round, as was observed when they bound to the intact protein. The peptide added in solution was able to inhibit cell adhesion to the intact protein in a dose-dependent manner and was also verified to bind to the α₂β₁ integrin. A cyclic peptide, CQLRGLRRWLEAKASRPDATC³²⁶, mimicking the structural constraints of this sequence in the intact protein, showed similar efficiency in inhibiting binding to chondroadherin. The unique peptide motif responsible for cellular binding is primarily located in the octamer sequence LRRWLEAK³¹⁸. Binding of cells to the active peptide or to chondroadherin immobilized on cell culture plates rapidly induces intracellular signaling (i.e. ERK phosphorylation). Thus, chondroadherin interaction with cells may be central for maintaining the adult chondrocyte phenotype and cartilage homeostasis. The peptides, particularly the more stable cyclic peptide, open new opportunities to modulate cell behavior in situations of tissue pathology.