Enhanced activation of mitogen-activated protein kinase and myosin light chain kinase by the Pro33 polymorphism of integrin β3

Abstract

Integrin β3 is polymorphic at residue 33 (Leu33 or Pro33), and the Pro33 variant exhibits increased outside-in signaling to focal adhesion kinase and greater actin reorganization. Because focal adhesion kinase activation and an intact cytoskeleton are critical links for integrin-mediated signaling to MAPK, we explored the role of integrin αIIbβ3 in this signaling using Chinese hamster ovary and human kidney 293 cell lines expressing either the Leu33 or Pro33 isoform of β3. Compared with Leu33 cells, Pro33 cells demonstrated substantially greater activation of ERK2 (but not MAPK family members JNK and p38) upon adhesion to immobilized fibrinogen (but not fibronectin) and upon integrin cross-linking. ERK2 activation was mediated through MAPK kinase and required phosphoinositide 3-kinase signaling and an intact actin cytoskeleton. Human platelets and Chinese hamster ovary cells expressing the Pro33 isoform showed enhanced activation of the ERK2 substrate myosin light chain kinase (MLCK) upon adhering to fibrinogen. Furthermore, compared with platelets and cells expressing the Leu33 isoform, the Pro33 variant showed greater a-granule release, clot retraction, and adhesion to fibrinogen under shear stress, and these functional differences were abolished by MLCK and MAPK kinase inhibition. Post-integrin occupancy signaling through MAPK and MLCK after αIIbβ3 cross-linking may explain in part the increased adhesive properties of the Pro33 variant of integrin β3.