αIIbβ3 Integrin: New Allelic Variants in Glanzmann Thrombasthenia, Effects on ITGA2B and ITGB3 mRNA Splicing, Expression, and Structure-Function

Glanzmann thrombasthenia (GT) is an autosomal recessive inherited bleeding disorder characterized by an impaired platelet aggregation due to defects in integrin alpha IIb beta 3 (ITGA2B, ITGB3), a fibrinogen receptor. Mutations from 24 GT patients and two carriers of various origins, Caucasian, North,African and Asian were characterized. Promoter and exon sequences of alpha IIb and beta 3 genes were amplified and directly sequenced. Among 29 identified Mutations, 17 new allelic variants resulting from nonsense, missense and deletion/insertion mutations were described. RNA alterations were evaluated by using Web servers. The alpha IIb p.S926L, p.V903F, and beta 3 p.C38Y, p.M118R, p.G221D substitutions prevented complex expression at the surface of COS-7 cells by altering the alpha IIb or the beta 3 subunit structure. As shown by free energy analyses applied on the resolved structure of alpha IIb beta 3 and structural modeling of the mutant, the p.K253M Substitution of beta 3 helped to define a key role of the K253 in the interaction of the alpha IIb beta-propeller and the beta 3 beta-1 domains. finally, the alpha IIb p.Q595H substitution allowed cell surface expression of the complex but its corresponding c.2800G > T mutation is predicted to alter normal RNA splicing. In conclusion, our study yielded the discovery of 17 new GT allelic variants, revealed the key role of K253 of alpha IIb for the alpha IIb beta 3 complex formation and provides an additional example of an apparently missense mutation causing a splicing defect. Hum Mutat 31:237-246, 2010. (C) 2009 Wiley-Liss, Inc.