Effect of cytoplasmic domain mutations on the agonist-stimulated ligand binding activity of the platelet integrin alphaIIbbeta3.
Function of the platelet integrin alphaIIbbeta3 is regulated by agonist-generated signals interacting with its cytoplasmic tails. When alphaIIbbeta3 is expressed in Epstein-Barr virus-transformed B lymphocytes, stimulation of the cells with phorbol 12-myristate 13-acetate results in alphaIIbbeta3-mediated lymphocyte adherence to immobilized fibrinogen, as well as soluble fibrinogen binding to alphaIIbbeta3, indicating that agonists increase the affinity of alphaIIbbeta3 for fibrinogen in these cells. To address the contribution of the alphaIIb and beta3 cytoplasmic tails to this process, we mutated each tail and expressed the mutants in B lymphocytes. Truncation of the alphaIIb 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 alphaIIbbeta3 expression due to impaired intracellular assembly of alphaIIbbeta3 heterodimers, probably due to a mutation-induced change in the conformation of alphaIIb. Introducing beta3 mutations known to impair alphaIIbbeta3 function in platelets into the cytoplasmic tail of beta3 in lymphocytes also impaired alphaIIbbeta3 function in these cells. These studies demonstrate that the cytoplasmic tail of alphaIIb is not required for alphaIIbbeta3 function in lymphocytes, although the presence of GFFKR in the alphaIIb tail is required for alphaIIb to interact with beta3. Additionally, they indicate that signals interacting with the beta3 cytoplasmic tail are responsible for the ability of agonists to stimulate alphaIIbbeta3 function.
Loh
E
E
Department of Medicine, the University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.
Qi
W
W
Vilaire
G
G
Bennett
J S
JS
D000595:Amino Acid Sequence; D002448:Cell Adhesion; D003593:Cytoplasm / Q000378:metabolism*; D005340:Fibrinogen / Q000378:metabolism; D006801:Humans; D008024:Ligands; D008214:Lymphocytes / Q000166:cytology / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding
D008024:Ligands; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D005340:Fibrinogen