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Haematologica. 2010 Jul;95(7):1158-66. doi: 10.3324/haematol.2009.018572. Epub 2010 Jan 15.

L718P mutation in the membrane-proximal cytoplasmic tail of beta 3 promotes abnormal alpha IIb beta 3 clustering and lipid microdomain coalescence, and associates with a thrombasthenia-like phenotype.

Author information

  • 1Department of Cellular and Molecular Medicine, Centro de Investigaciones Biológicas, Ramiro de Maeztu 9, 28040 Madrid, Spain.

Abstract

BACKGROUND:

Support for the role of transmembrane and membrane-proximal domains of alpha IIb beta 3 integrin in the maintenance of receptor low affinity comes from mutational studies showing that activating mutations can induce constitutive bi-directional transmembrane signaling.

DESIGN AND METHODS:

We report the functional characterization of a mutant alpha IIb beta 3 integrin carrying the Leu718Pro mutation in the membrane-proximal region of the beta 3 cytoplasmic domain, identified in heterozygosis in a patient with a severe bleeding phenotype and defective platelet aggregation and adhesion.

RESULTS:

Transiently transfected cells expressed similar levels of normal and mutant alpha IIb beta 3, but surface expression of mutant alpha v beta 3 was reduced due to its retention in intracellular compartments. Cells stably expressing mutant alpha IIb beta 3 showed constitutive binding to soluble multivalent ligands as well as spontaneous fibrinogen-dependent aggregation, but their response to DTT was markedly reduced. Fibrinogen-adherent cells exhibited a peculiar spreading phenotype with long protrusions. Immunofluorescence analysis revealed the formation of alpha IIb beta 3 clusters underneath the entire cell body and the presence of atypical high-density patches of clustered alpha IIb beta 3 containing encircled areas devoid of integrin that showed decreased affinity for the fluorescent lipid analog DiIC(16) and were disrupted in cholesterol-depleted cells.

CONCLUSIONS:

These findings are consistent with an important role of the membrane-proximal region of beta 3 in modulating alpha IIb beta 3 clustering and lateral redistribution of membrane lipids. Since the beta 3 mutant was associated with a thrombasthenic phenotype in a patient carrying one normal beta 3 allele, these results support a dominant role of clustering in regulating integrin alpha IIb beta 3 functions in vivo.

Comment in

PMID:
20081061
[PubMed - indexed for MEDLINE]
PMCID:
PMC2895041
Free PMC Article

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