title: Disruption of the Cys5-Cys7 disulfide bridge in the platelet glycoprotein Ibbeta prevents the normal maturation and surface exposure of GPIb-IX complexes.
abstract: 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 GPIbalpha, GPIbbeta, and GPIX revealed the presence of a single point mutation, a G to A substitution, in codon 30 of GPIbbeta, 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 GPIbalpha and GPIX. Transient transfection of the mutant GPIbalpha subunit failed to render surface expression of GPIbalpha and exerted a dominant-negative effect on the surface exposure of the GPIb-IX complex. Metabolic labelling and immunoprecipitation analysis of transfected cells indicated that [5Tyr]GPIbbeta may associate with GPIX and GPIbalpha, but the maturation of the GPIb-IX complex is impaired. Substitution of either Cys5 or Cys7 by Ala failed to show surface expression of GPIb-IX, suggesting that the Cys5- Cys7 disulfide loop in GPIbbeta is essential for the efficient processing and trafficking of GPIb-IX complexes toward the plasma membrane. Our findings indicate that the identified novel GPIbbeta mutation is responsible for the BSS phenotype of the patients and provide an explanation for the molecular mechanism underlying the reduced platelet content of GPIb-IX complex in the heterozygous individuals.
authors.affiliation: Departamento de Pathophysiology and Human molecular Genetics, Centro de Investigaciones Biológicas (CSIC), Velázquez, Madrid, Spain. cgmanchon@cib.csic.es
authors.forename: Consuelo
authors.initials: C
authors.lastname: González-Manchón
authors.forename: Nora
authors.initials: N
authors.lastname: Butta
authors.forename: Gema
authors.initials: G
authors.lastname: Iruín
authors.forename: Sonia
authors.initials: S
authors.lastname: Alonso
authors.forename: Matilde S
authors.initials: MS
authors.lastname: Ayuso
authors.forename: Roberto
authors.initials: R
authors.lastname: Parrilla
mesh_terms: D000328:Adult; D001606:Bernard-Soulier Syndrome / Q000235:genetics*; D003553:Cystine; D004220:Disulfides; D005192:Family Health; D005260:Female; D006720:Homozygote; D006801:Humans; D008297:Male; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D017510:Protein Folding; D017434:Protein Structure, Tertiary / Q000235:genetics; D021381:Protein Transport; D035781:Siblings
chemical_list: D004220:Disulfides; D019038:Platelet Glycoprotein GPIb-IX Complex; D003553:Cystine