A p.Arg127Gln variant in GPIbα LRR5 allosterically enhances affinity for VWF: a novel form of platelet-type VWD

Abstract

Gain-of-function (GoF) variants in GP1BA cause platelet-type von Willebrand disease (PT-VWD), a rare inherited autosomal dominant bleeding disorder characterized by enhanced platelet GPIbα-von Willebrand factor (VWF) interaction and thrombocytopenia. To date, only 6 variants causing PT-VWD have been described, 5 in the C-terminal disulfide loop of the VWF-binding domain of GPIbα and 1 in the macroglycopeptide. GoF GP1BA variants generate a high affinity conformation of the C-terminal disulfide loop with a consequent allosteric conformational change on another region of GPIbα, the leucine-rich-repeat (LRR) domain. We identified a novel GP1BA variant (p.Arg127Gln) affecting the LRR5 domain of GPIbα in a boy with easy bruising and laboratory test results suggestive of PT-VWD. We thus aimed to investigate the impact of the p.Arg127Gln variant on GPIbα affinity for VWF and on GPIbα structure. CHO cells expressing p.Arg127Gln GPIbα showed increased binding of VWF induced by ristocetin and enhanced tethering on immobilized VWF as compared with cells expressing wild-type (WT) GPIbα. Surface plasmon resonance confirmed that p.Arg127Gln enhances the binding affinity of GPIbα for VWF. Hydrogen-deuterium exchange mass spectrometryshowed that p.Arg127Gln of LRR, while having little effect on the dynamics of the LRR locally, enhances the conformational dynamics of the GPIbα C-terminal disulfide loop structure. Our data demonstrate for the first time that GoF variants outside the GPIbα C-terminal disulfide loop may be pathogenic and that aminoacidic changes in the LRR may cause allosterically conformational changes in the C-terminal disulfide loop of GPIbα inducing a conformation with high affinity for VWF.

Biological, clinical features and modelling of heterozygous variants of glycoprotein Ib platelet subunit alpha (GP1BA) and glycoprotein Ib platelet subunit beta (GP1BB) genes responsible for constitutional thrombocytopenia

“…Their haemorrhagic symptomatology is discrete or non‐existent and mainly affects mucosal sites. This is compatible with the previous reports 3–23 . Moreover, as expected, VWF levels of the patients were normal, without discrepancy between activity and antigen assays.…”

“…In our series, similarly to these cases of 22q11 deletion, the patient presenting a complete deletion of GP1BB was diagnosed with MTP. The mechanism by which only some mutations of GP1BA or GP1BB with hemizygosity for these genes present with MTP still remains unclear although interactions between GPIb‐IX‐V complex subunits are probably essential 16,32–34 . One hypothesis would be that the concentration of the GPIB‐IX‐V complex on the platelet surface is involved in the terminal phases of megakaryopoiesis.…”

“…The latter are called monoallelic BSS with an autosomal dominant transmission. 4 To date, 13 GP1BA variants [4][5][6][7][8][9][10][11][12][13][14][15][16] and 14 GP1BB variants, [15][16][17][18][19][20][21][22] considered to be responsible for monoallelic BSS, have been described in the literature (Table 1). [4][5][6][7][8][9][10][11][13][14][15][16][17][18][19][20][21]23 No variant responsible for monoallelic BSS has been identified to date in the GP9 gene.…”

“…Clinically, patients present with moderate mucocutaneous bleeding, such as epistaxis, ecchymosis, menorrhagia, and sometimes prolonged post-surgical bleeding. [4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23] Platelet counts show moderate thrombocytopenia with an increase in mean platelet volume (MPV). Morphological analysis discloses giant platelets in variable proportions.…”

“…Clinically, patients present with moderate mucocutaneous bleeding, such as epistaxis, ecchymosis, menorrhagia, and sometimes prolonged post‐surgical bleeding 4–23 . Platelet counts show moderate thrombocytopenia with an increase in mean platelet volume (MPV).…”