Surface expression of glycoprotein Ibα is dependent on glycoprotein Ibβ: evidence from a novel mutation causing Bernard-Soulier syndrome

Abstract

Bernard-Soulier syndrome is a rare bleeding disorder caused by a quantitative or qualitative defect in the platelet glycoprotein (GP) Ib-IX-V complex. The complex, which serves as a platelet receptor for von Willebrand factor, is composed of 4 subunits: GPIb, GPIbβ, GPIX, and GPV. We here describe the molecular basis of a novel form of Bernard-Soulier syndrome in a patient in whom the components of the GPIb-IX-V complex were undetectable on the platelet surface. Although confocal imaging confirmed that GPIb was not present on the platelet surface, GPIb was readily detectable in the patient's platelets. Moreover, immunoprecipitation of plasma with specific monoclonal antibodies identified circulating, soluble GPIb. DNA-sequence analysis revealed normal sequences for GPIb and GPIX. There was a G to A substitution at position 159 of the gene encoding GPIbβ, resulting in a premature termination of translation at amino acid 21. Studies of transient coexpression of this mutant, W21stop-GPIbβ, together with wild-type GPIb and GPIX, demonstrated a failure of GPIX expression on the surface of HEK 293T cells. Similar results were obtained with Chinese hamster ovary  IX cells, a stable cell line expressing GPIb that retains the capacity to re-express GPIX. Thus, we found that GPIbβ affects the surface expression of the GPIb-IX complex by failing to support the insertion of GPIb and GPIX into the platelet membrane.

Synthesis of GPIbβ with novel transmembrane and cytoplasmic sequences in a Bernard–Soulier patient resulting in GPIb‐defective signaling in CHO cells

“…The abnormal protein was nevertheless detected in total platelet lysates, consistent with confocal microscopy results showing its intracellular accumulation in CHO cells. In previous studies where GPIb β mutations were reproduced in transfected cells, GPIb β was generally either absent (Arg17Cys, Pro74Arg, Trp21Stop, Trp123Stop) [38–41] or detected in very small amounts (Tyr88Cys, Asn64Thr) [25,37]. Two exceptions were the N‐terminal Cys5Tyr and Pro29Leu mutants [36,42].…”

Macrothrombocytopenia in velocardiofacial syndrome

“…The symptomatic patients were homozygotes, because they had a deletion of one chromosome containing the GpIbβ gene and a mutation in the other. Furthermore, BSS associated with the defective beta subunit of GpIb (BSS type B) has been reported in 22q11.2‐deleted patients [4–6] and in homozygotes and heterozygotes for missense or non‐sense mutations in GPIbβ[8–12].…”

Identification and Characterisation of a Platelet GPIb/V/IX‐like Complex on Human Breast Cancers: Implications for the Metastatic Process

“…7,8), and also interacts with actin-binding protein (ABP), 9) linking actin filaments to the cytoskeleton. Functional operation of the complex through the GPIbα-vWf axis requires the associated surface expression of both the GPIbβ and GPIX subunits, 6,10) as well as the cytoplasmic interaction with ABP. 11,12) GPIb/V/IX complex expression was originally thought to be restricted to cells of the megakaryocytic lineage.…”