Outside-in signaling of soluble and solid-phase fibrinogen through integrin αIIbβ3 is different and cooperative with each other in a megakaryoblastic leukemia cell line, CMK

The function and the outside-in signaling pathways of αIIbβ3 were examined in relation to cell adhesion using a megakaryoblastic leukemia cell line, CMK. After 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment, the cells adhered to the culture plate and underwent megakaryocytic differentiation with expression of αIIbβ3. Binding of soluble fibrinogen to the cells via αIIbβ3 was dependent on cell adhesion. Cell detaching reduced the affinity of this integrin for soluble fibrinogen, although its surface expression was almost unchanged. In contrast, detached cells became tightly adherent to the fibrinogen-coated plate (solid-phase fibrinogen). The same ligand, fibrinogen, present either in soluble or solid-phase form, triggered differential signaling pathways mediated by αIIbβ3. By the stimulation with soluble fibrinogen, Syk was tyrosine-phosphorylated but FAK was dephosphorylated, whereas solid-phase fibrinogen promptly caused tyrosine phosphorylation of FAK followed by delayed phosphorylation of Syk. In addition, the binding of soluble fibrinogen to the cells adherent to fibrinogen-coated plate resulted in tyrosine phosphorylation of integrin β3 and a complex formation of integrin β3 with Syk. This implies the cooperation of both soluble and solid-phase fibrinogen-mediated signaling pathways.