New insights into regulation of αIIbβ3 integrin signaling by filamin A

Abstract

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 αIIbβ3 signaling. Methods Three FLNa deletion mutants were overexpressed in the erythro‐megakaryocytic leukemic cell line HEL: Del1, which lacks the N‐terminal CH1‐CH2 domains mediating the FLNa‐actin interaction; Del2, lacking the Ig‐like repeat 21, which mediates the FLNa‐β3 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‐β3 proximity in cells adherent to immobilized fibrinogen were assessed before and after αIIbβ3 activation by the protein kinase C agonist phorbol 12‐myristate 13‐acetate. Results Our results show that FLNa‐actin and FLNa‐β3 interactions negatively regulate αIIbβ3 activation. Moreover, FLNa‐actin interaction represses Rac activation, contributing to the negative regulation of αIIbβ3 activation. In contrast, the FLNa dimerization domain, which maintains Rho inactive, was found to negatively regulate αIIbβ3 outside‐in signaling. Conclusion We conclude that FLNa negatively controls αIIbβ3 activation by regulating actin polymerization and restraining activation of Rac, as well as outside‐in signaling by repressing Rho.

A gain‐of‐function filamin A mutation in mouse platelets induces thrombus instability

“…Likewise, it has been proposed that the activation of αIIbβ3 integrin requires the dissociation of FLNa from β3 allowing talin binding, but hard evidence is missing as well. Recently, our group showed clearly that the FLNa–β3 interaction exerts a negative regulation on talin–β3 interaction after activation, using a megakaryocytic cell line (HEL) 11 …”

“…Finally, the Rho GTPase, RhoA, which binds constitutively to FLNa via the C‐terminal Ig repeat 24, has recently been reported to be dependent on αIIbβ3. This interaction modulates proplatelet formation in megakaryocytes 14 and outside‐in signaling in platelets 11 . How the interactions of FLNa with its partners integrate cellular signaling and cellular functions remains an open question.…”

“…Recently, our group showed clearly that the FLNa-β3 interaction exerts a negative regulation on talin-β3 interaction after activation, using a megakaryocytic cell line (HEL). 11 The tyrosine kinase Syk involved in the GPVI signaling is another partner of FLNa (Ig repeat 5) highlighted in Flna −/− mice. 6 This interaction, essential in platelet activation under arterial shear, allows Syk recruitment to GPVI.…”

“…Recently, our group showed clearly that the FLNa–β3 interaction exerts a negative regulation on talin–β3 interaction after activation, using a megakaryocytic cell line (HEL). 11 …”

“…This interaction modulates proplatelet formation in megakaryocytes 14 and outside‐in signaling in platelets. 11 How the interactions of FLNa with its partners integrate cellular signaling and cellular functions remains an open question. Investigating FLNA gene variants is of great help.…”