A novel thromboxane A2 receptor N42S variant results in reduced surface expression and platelet dysfunction

Shaista P Nisar, Marie Lordkipanidzé, Matthew L Jones, Ban Dawood, Sherina Murden, Margaret R Cunningham, Andrew D Mumford, Jonathan T Wilde, Steve P Watson, Stuart J Mundell, Gillian C Lowe, UK GAPP study group

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

A small number of thromboxane receptor variants have been described in patients with a bleeding history that result in platelet dysfunction. We have identified a patient with a history of significant bleeding, who expresses a novel heterozygous thromboxane receptor variant that predicts an asparagine to serine substitution (N42S). This asparagine is conserved across all class A GPCRs, suggesting a vital role for receptor structure and function.We investigated the functional consequences of the TP receptor heterozygous N42S substitution by performing platelet function studies on platelet-rich plasma taken from the patient and healthy controls. We investigated the N42S mutation by expressing the wild-type (WT) and mutant receptor in human embryonic kidney (HEK) cells. Aggregation studies showed an ablation of arachidonic acid responses in the patient, whilst there was right-ward shift of the U46619 concentration response curve (CRC). Thromboxane generation was unaffected. Calcium mobilisation studies in cells lines showed a rightward shift of the U46619 CRC in N42S-expressing cells compared to WT. Radioligand binding studies revealed a reduction in BMax in platelets taken from the patient and in N42S-expressing cells, whilst cell studies confirmed poor surface expression. We have identified a novel thromboxane receptor variant, N42S, which results in platelet dysfunction due to reduced surface expression. It is associated with a significant bleeding history in the patient in whom it was identified. This is the first description of a naturally occurring variant that results in the substitution of this highly conserved residue and confirms the importance of this residue for correct GPCR function.

LanguageEnglish
Pages923-932
Number of pages10
JournalThrombosis and haemostasis
Volume111
Issue number5
Early online date23 Jan 2014
DOIs
Publication statusPublished - 5 May 2014

Fingerprint

Prostaglandin H2 Receptors Thromboxane A2
Thromboxane Receptors
Blood Platelets
15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid
Asparagine
Hemorrhage
Platelet-Rich Plasma
Thromboxanes
Arachidonic Acid
Serine
Calcium
Kidney
Cell Line
Mutation

Keywords

  • receptors
  • gene mutations
  • platelet pharmacology
  • platelet disorders
  • platelet pathology

Cite this

Nisar, Shaista P ; Lordkipanidzé, Marie ; Jones, Matthew L ; Dawood, Ban ; Murden, Sherina ; Cunningham, Margaret R ; Mumford, Andrew D ; Wilde, Jonathan T ; Watson, Steve P ; Mundell, Stuart J ; Lowe, Gillian C ; UK GAPP study group. / A novel thromboxane A2 receptor N42S variant results in reduced surface expression and platelet dysfunction. In: Thrombosis and haemostasis. 2014 ; Vol. 111, No. 5. pp. 923-932.
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abstract = "A small number of thromboxane receptor variants have been described in patients with a bleeding history that result in platelet dysfunction. We have identified a patient with a history of significant bleeding, who expresses a novel heterozygous thromboxane receptor variant that predicts an asparagine to serine substitution (N42S). This asparagine is conserved across all class A GPCRs, suggesting a vital role for receptor structure and function.We investigated the functional consequences of the TP receptor heterozygous N42S substitution by performing platelet function studies on platelet-rich plasma taken from the patient and healthy controls. We investigated the N42S mutation by expressing the wild-type (WT) and mutant receptor in human embryonic kidney (HEK) cells. Aggregation studies showed an ablation of arachidonic acid responses in the patient, whilst there was right-ward shift of the U46619 concentration response curve (CRC). Thromboxane generation was unaffected. Calcium mobilisation studies in cells lines showed a rightward shift of the U46619 CRC in N42S-expressing cells compared to WT. Radioligand binding studies revealed a reduction in BMax in platelets taken from the patient and in N42S-expressing cells, whilst cell studies confirmed poor surface expression. We have identified a novel thromboxane receptor variant, N42S, which results in platelet dysfunction due to reduced surface expression. It is associated with a significant bleeding history in the patient in whom it was identified. This is the first description of a naturally occurring variant that results in the substitution of this highly conserved residue and confirms the importance of this residue for correct GPCR function.",
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Nisar, SP, Lordkipanidzé, M, Jones, ML, Dawood, B, Murden, S, Cunningham, MR, Mumford, AD, Wilde, JT, Watson, SP, Mundell, SJ, Lowe, GC & UK GAPP study group 2014, 'A novel thromboxane A2 receptor N42S variant results in reduced surface expression and platelet dysfunction' Thrombosis and haemostasis, vol. 111, no. 5, pp. 923-932. https://doi.org/10.1160/TH13-08-0672

A novel thromboxane A2 receptor N42S variant results in reduced surface expression and platelet dysfunction. / Nisar, Shaista P; Lordkipanidzé, Marie; Jones, Matthew L; Dawood, Ban; Murden, Sherina; Cunningham, Margaret R; Mumford, Andrew D; Wilde, Jonathan T; Watson, Steve P; Mundell, Stuart J; Lowe, Gillian C; UK GAPP study group.

In: Thrombosis and haemostasis, Vol. 111, No. 5, 05.05.2014, p. 923-932.

Research output: Contribution to journalArticle

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T1 - A novel thromboxane A2 receptor N42S variant results in reduced surface expression and platelet dysfunction

AU - Nisar, Shaista P

AU - Lordkipanidzé, Marie

AU - Jones, Matthew L

AU - Dawood, Ban

AU - Murden, Sherina

AU - Cunningham, Margaret R

AU - Mumford, Andrew D

AU - Wilde, Jonathan T

AU - Watson, Steve P

AU - Mundell, Stuart J

AU - Lowe, Gillian C

AU - UK GAPP study group

PY - 2014/5/5

Y1 - 2014/5/5

N2 - A small number of thromboxane receptor variants have been described in patients with a bleeding history that result in platelet dysfunction. We have identified a patient with a history of significant bleeding, who expresses a novel heterozygous thromboxane receptor variant that predicts an asparagine to serine substitution (N42S). This asparagine is conserved across all class A GPCRs, suggesting a vital role for receptor structure and function.We investigated the functional consequences of the TP receptor heterozygous N42S substitution by performing platelet function studies on platelet-rich plasma taken from the patient and healthy controls. We investigated the N42S mutation by expressing the wild-type (WT) and mutant receptor in human embryonic kidney (HEK) cells. Aggregation studies showed an ablation of arachidonic acid responses in the patient, whilst there was right-ward shift of the U46619 concentration response curve (CRC). Thromboxane generation was unaffected. Calcium mobilisation studies in cells lines showed a rightward shift of the U46619 CRC in N42S-expressing cells compared to WT. Radioligand binding studies revealed a reduction in BMax in platelets taken from the patient and in N42S-expressing cells, whilst cell studies confirmed poor surface expression. We have identified a novel thromboxane receptor variant, N42S, which results in platelet dysfunction due to reduced surface expression. It is associated with a significant bleeding history in the patient in whom it was identified. This is the first description of a naturally occurring variant that results in the substitution of this highly conserved residue and confirms the importance of this residue for correct GPCR function.

AB - A small number of thromboxane receptor variants have been described in patients with a bleeding history that result in platelet dysfunction. We have identified a patient with a history of significant bleeding, who expresses a novel heterozygous thromboxane receptor variant that predicts an asparagine to serine substitution (N42S). This asparagine is conserved across all class A GPCRs, suggesting a vital role for receptor structure and function.We investigated the functional consequences of the TP receptor heterozygous N42S substitution by performing platelet function studies on platelet-rich plasma taken from the patient and healthy controls. We investigated the N42S mutation by expressing the wild-type (WT) and mutant receptor in human embryonic kidney (HEK) cells. Aggregation studies showed an ablation of arachidonic acid responses in the patient, whilst there was right-ward shift of the U46619 concentration response curve (CRC). Thromboxane generation was unaffected. Calcium mobilisation studies in cells lines showed a rightward shift of the U46619 CRC in N42S-expressing cells compared to WT. Radioligand binding studies revealed a reduction in BMax in platelets taken from the patient and in N42S-expressing cells, whilst cell studies confirmed poor surface expression. We have identified a novel thromboxane receptor variant, N42S, which results in platelet dysfunction due to reduced surface expression. It is associated with a significant bleeding history in the patient in whom it was identified. This is the first description of a naturally occurring variant that results in the substitution of this highly conserved residue and confirms the importance of this residue for correct GPCR function.

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KW - gene mutations

KW - platelet pharmacology

KW - platelet disorders

KW - platelet pathology

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JO - Thrombosis and haemostasis

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