New Insights into Regulation of Αiibβ3 Integrin Signaling by Filamin A

Background: Filamin (FLN) regulates many cell functions through its scaffolding activity cross-linking cytoskeleton and integrins. FLN was shown to inhibit integrin activity, but the exact mechanism remains unclear. Objectives: The aim of this study was to evaluate the role of filamin A (FLNa) subdomains on the regulation of integrin alpha IIb beta 3 signaling. Methods: Three FLNa deletion mutants were overexpressed in the erythromegakaryocytic leukemic cell line HEL: Dell, which lacks the N-terminal CH1-CH2 domains mediating the FLNa-actin interaction; Del2, lacking the Ig-like repeat 21, which mediates the FLNa-beta 3 interaction; and Del3, lacking the C-terminal Ig repeat 24, responsible for FLNa dimerization and interaction with the small Rho guanosine triphosphatase involved in actin cytoskeleton reorganisation. Fibrinogen binding to HEL cells in suspension and talin-beta 3 proximity in cells adherent to immobilized fibrinogen were assessed before and after alpha IIb beta 3 activation by the protein kinase C agonist phorbol 12-myristate 13-acetate. Results: Our results show that FLNa-actin and FLNa-beta 3 interactions negatively regulate alpha IIb beta 3 activation. Moreover, FLNa-actin interaction represses Rac activation, contributing to the negative regulation of alpha IIb beta 3 activation. In contrast, the FLNa dimerization domain, which maintains Rho inactive, was found to negatively regulate alpha IIb beta 3 outside-in signaling. Conclusion: We conclude that FLNa negatively controls allb133 activation by regulating actin polymerization and restraining activation of Rac, as well as outside-in signaling by repressing Rho.