Molecular genetic analysis of a variant Bernard-Soulier syndrome due to compound heterozygosity for two novel glycoprotein Ibβ mutations

Bernard-Soulier syndrome (BSS) is a rare bleeding disorder characterized by giant platelets, thrombocytopenia, and prolonged bleeding time. It is caused by abnormalities in the glycoprotein (GP) Ib/IX/V complex, the receptor for von Willebrand factor (vWF). Most cases of BSS described so far involve quantitative rather than qualitative defects in the complex. In this study, we investigated the effects of two naturally occurring mutations in the GPIb beta gene, C122S and 443delG, on the expression of the GPIb/IX complex identified in a variant type of BSS in which the platelets had severely reduced GPIb alpha (similar to 10%) and less markedly reduced GPIb beta and GPIX (similar to 20%) expression. Immunoblot analysis showed the absence of non-reduced GPIb (GPIb alpha/GPIb beta) in the patient's platelets. Transient transfection experiments in 293T cells revealed the expression of GPIb beta Ser122 polypeptide and absence of GPIb beta 443delG polypeptide. Although no disulfide-linked association was observed between GPIb beta Ser122 and GPIb alpha, GPIb beta Ser122 was non-covalently associated with both GPIb alpha and GPIX subunits on the cell surface when cotransfected with wild-type GPIb alpha and GPIX. Chinese hamster ovary cells stably expressing GPIb alpha/Ib beta Ser122/IX had the ability to bind soluble vWF and to aggregate in the presence of ristocetin. These results suggest that despite disruption of the disulfide linkage between GPIb alpha and GPIb beta, GPIb/IX is formed, but its stability may be impaired, resulting in low levels of the complex on the platelet membranes.