Ligand-specific, transient interaction between integrins and calreticulin during cell adhesion to extracellular matrix proteins is dependent upon phosphorylation dephosphorylation events

As transmembrane heterodimers, integrins bind to both extracellular ligands and intracellular proteins. We are currently investigating the interaction between integrins and the intracellular protein calreticulin. A prostatic carcinoma cell line (PC-3) was used to demonstrate that calreticulin can be found in the alpha(3) immunoprecipitates of cells plated on collagen type IV, but not when plated on vitronectin. Conversely, alpha(v) immunoprecipit ates contained calreticulin only when cells were plated on vitronectin, i.e . not when plated on collagen IV. The interactions between these integrins and calreticulin were independent of actin cytoskeleton assembly and were t ransient, being maximal approx. 10-30 min after the cells came into contact with the substrates prior to complete cell spreading and formation of firm adhesive contacts. We demonstrate that okadaic acid, an inhibitor of intra cellular serine/threonine protein phosphatases, inhibited the alpha(3)beta( 1)-mediated adhesion of PC-3 cells to collagen EV and the alpha(2)beta(1)-m ediated attachment of Jurkat cells to collagen I. This inhibition by okadai c acid was accompanied by inhibition of the ligand-specific interaction of calreticulin with the respective integrins in the two cell types. Additiona lly, we found that pharmacological inhibition of mitogen-activated protein kinase kinase (MEK) resulted in prolongation of the calreticulin-integrin i nteraction, and enhancement of PC-3 cell attachment to collagen IV. We conc lude that calreticulin interacts transiently with integrins during cell att achment and spreading. This interaction depends on receptor occupation, is ligand-specific, and can be modulated by protein phosphatase and MEK activi ty.