Disruption of the Cys-5-Cys7 disulfide bridge in the platelet glycoprotein Ibβ prevents the normal maturation and surface exposure of GPIb-IX complexes

This work aimed at elucidating the molecular genetic defect in two related patients with Bernard-Soulier syndrome (BSS) phenotype. Flow cytometric analysis revealed undetectable levels of platelet glycoproteins (GP), Ibalpha and IX, although plasma glycocalicin was detectable in both cases. The complete sequencing of GPlbalpha, GPlbbeta, and GPIX revealed the presence of a single point mutation, a G to A substitution, in codon 30 of GPlbbeta, that changes Cys5 to Tyr. The parents and sibling of the patients, heterozygotes for this mutation, were asymptomatic and they all showed a reduced platelet content of GPlbalpha and GPIX Transient transfection of the mutant GPlbbeta subunit failed to render surface expression of GPlbalpha and exerted a dominant-negative effect on the surface exposure of the GPlb-IX complex. Metabolic labelling and immunoprecipitation analysis of transfected cells indicated that [5Tyr]GPlbbeta may associate with GPIX and GPlba, but the maturation of the GPlb-IX complex is impaired. Substitution of either Cys5 or Cys7 by Ala failed to show surface expression of GPlb-IX, suggesting that the Cys5-Cys7 disulfide loop in GPlbbeta is essential for the efficient processing and trafficking of. GPlb-IX complexes toward the plasma membrane. Our findings indicate that the identified novel GPlbbeta mutation is responsible for the BSS phenotype of the patients and provide an explanation for the molecular mechanism underlying the reduced platelet content of GPlb-IX complex in the heterozygous individuals.