| Integrin-dependent cell spreading and retraction are required for cell adhesion, migration, and proliferation, and thus are important in thrombosis, wound repair, immunity, and cancer development. It remains unknown how integrin outside-in signaling induces and controls these two opposite processes. This study reveals that calpain cleavage of integrin beta3 at Tyr759 switches the functional outcome of integrin signaling from cell spreading to retraction. Expression of a calpain cleavage-resistant beta3 causes defective clot retraction and RhoA-mediated retractile signaling but enhances cell spreading. Conversely, a calpain-cleaved form of beta3 fails to mediate cell spreading, but inhibition of the RhoA signaling pathway corrects this defect. Interestingly, the calpain-cleaved beta3 also fails to bind c-Src, which is required for integrin-induced cell spreading, and this requirement of beta3-associated c-Src results from its inhibition of RhoA-dependent contractile signals. Thus, calpain cleavage of beta3 at Tyr759 relieves c-Src-mediated RhoA inhibition, activating the RhoA pathway that confines cell spreading and causes cell retraction. In this study, we also show that that integrin outside-in signaling results in a late but sustained activation of mitogen activated protein kinases (MAPKs). This late integrin-mediated MAPK activation plays a important role in integrin-mediated cellular and clot retraction, and MAPK-dependent simulatory effect on clot retraction is mediated by increased myosin light chain (MLC) phosphorylation. Importantly, integrin-dependent MAPK activation, MAPK-dependent MLC phosphorylation, and MAPK-dependent clot retraction are downstream of integrin-dependent Rac1 signaling, but not integrin-mediated RhoA-dependent retractile signaling. Thus, our results reveal two different activation mechanisms of integrin mediated retraction, a calpain-dependent RhoA-mediated retractile signaling pathway, and a novel Rac1-MAPK-dependent cell retractile signaling pathway. |
