Evidence for a differential functional regulation of the two β3-integrins αVβ3 and αIIbβ3

The functional regulation of integrins is a major determinant of cell adhesion, migration and tissue maintenance. The binding of cytoskeletal proteins to various sites of integrin cytoplasmic domains is a key mechanism of this functional regulation. Expression of recombinant integrin αIIbβ3 and αMβ2 lacking the GFFKR-region in CHO cells results in constitutively activated integrins. In contrast, CHO cells stably expressing either a GFFKR-deleted αV(del)β3 or a FF to AA-substituted αV(AA)β3 do not reveal a constitutively activated integrin. Adhesion to immobilized fibrinogen is strongly impaired in αV(del)β3 or αV(AA)β3-expressing cells, whereas it is not impaired in αIIbβ3 and αMβ2, both lacking the GFFKR-region. In a parallel plate flow chamber assay, αVβ3-expressing cells adhere firmly to fibrinogen and spread even at shear rates of 15 to 20 dyn/cm2, whereas αV(del)β3 or αV(AA)β3 cells are detached at 15 dyn/cm2. Actin stress fiber formation and focal adhesion plaques containing αVβ3 are observed in αVβ3 cells but not in αV(del)β3 or αV(AA)β3-expressing cells. As an additional manifestation of impaired outside-in signaling, phosphorylation of pp125FAK was reduced in these cells. In summary, we report that the GFFKR-region of the αV-cytoplasmic domain and in particular two phenylalanines are essential for integrin αVβ3 function, especially for outside-in signaling. Our results suggest that the two β3-integrins αIIbβ3 and αVβ3 are differentially regulated via their GFFKR-region.