Effect of cytoplasmic domain mutations on the agonist-stimulated ligand binding activity of the platelet integrin αIIbβ3

Abstract

Function of the platelet integrin αIIbβ3 is regulated by agonist- generated signals interacting with its cytoplasmic tails. When αIIbβ3 is expressed in Epstein-Barr virus-transformed B lymphocytes, stimulation of the cells with phorbol 12-myristate 13-acetate results in αIIbβ3-mediated lymphocyte adherence to immobilized fibrinogen, as well as soluble fibrinogen binding to αIIbβ3, indicating that agonists increase the affinity of αIIbβ3 for fibrinogen in these cells. To address the contribution of the αIIb and β3 cytoplasmic tails to this process, we mutated each tall and expressed the mutants in B lymphocytes. Truncation of the αIIb tail did not impair unstimulated or stimulated lymphocyte adherence to fibrinogen, regardless whether the truncation was proximal or distal to the conserved GFFKR sequence. However, deleting GFFKR or replacing it with alanines markedly reduced αIIbβ3 expression due to impaired intracellular assembly of αIIbβ3 heterodimers, probably due to a mutation-induced change in the conformation of αIIb. Introducing β3 mutations known to impair αIIbβ3 function in platelets into the cytoplasmic tail of β3 in lymphocytes also impaired αIIbβ3 function in these cells. These studies demonstrate that the cytoplasmic tail of αIIb is not required for αIIbβ3 function in lymphocytes, although the presence of GFFKR in the αIIb tail is required for αIIb to interact with β3. Additionally, they indicate that signals interacting with the β3 cytoplasmic tail are responsible for the ability of agonists to stimulate αIIbβ3 function.