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J Exp Med. 2014 Jun 30;211(7):1349-62. doi: 10.1084/jem.20130477. Epub 2014 Jun 23.

Human CalDAG-GEFI gene (RASGRP2) mutation affects platelet function and causes severe bleeding.

Author information

1
Institut National de la Santé et de la Recherche Médicale (Inserm), UMR_S 1062, 13005 Marseille, France Inra, UMR_INRA 1260, 13005 Marseille, France Aix Marseille Université, 13005 Marseille, France.
2
Post-Genomic Platform of Pitié-Salpêtrière (P3S), Pierre and Marie Curie University, F-75013 Paris, France.
3
Sorbonne Universités, UPMC Univ Paris 06, UMR_S 1166, F-75013 Paris, France Inserm, UMR_S 1166, Team Genomics and Pathophysiology of Cardiovascular Diseases, F-75013 Paris, France ICAN Institute for Cardiometabolism and Nutrition, F-75013 Paris, France.
4
Hématopoïèse Normale et Pathologique, Inserm Médicale U1009, 94805 Villejuif, France.
5
LIRYC, Plateforme Technologique et d'Innovation Biomédicale, Hôpital Xavier Arnozan, Pessac, France Inserm, UMR_1034, 33600 Pessac, France.
6
Inserm, UMR_1034, 33600 Pessac, France.
7
Aix Marseille Université, 13005 Marseille, France Inserm, UMR_1067, 13288 Marseille, France CNRS UMR_7333, 13288 Marseille, France.
8
Department of Blood Cell Research, Sanquin Research and Landsteiner Laboratory, Academic Medical Center, University of Amsterdam, 1105 AZ Amsterdam, Netherlands.
9
Institut National de la Santé et de la Recherche Médicale (Inserm), UMR_S 1062, 13005 Marseille, France Inra, UMR_INRA 1260, 13005 Marseille, France Aix Marseille Université, 13005 Marseille, France marie-christine.alessi@univ-amu.fr.

Abstract

The nature of an inherited platelet disorder was investigated in three siblings affected by severe bleeding. Using whole-exome sequencing, we identified the culprit mutation (cG742T) in the RAS guanyl-releasing protein-2 (RASGRP2) gene coding for calcium- and DAG-regulated guanine exchange factor-1 (CalDAG-GEFI). Platelets from individuals carrying the mutation present a reduced ability to activate Rap1 and to perform proper αIIbβ3 integrin inside-out signaling. Expression of CalDAG-GEFI mutant in HEK293T cells abolished Rap1 activation upon stimulation. Nevertheless, the PKC- and ADP-dependent pathways allow residual platelet activation in the absence of functional CalDAG-GEFI. The mutation impairs the platelet's ability to form thrombi under flow and spread normally as a consequence of reduced Rac1 GTP-binding. Functional deficiencies were confined to platelets and megakaryocytes with no leukocyte alteration. This contrasts with the phenotype seen in type III leukocyte adhesion deficiency caused by the absence of kindlin-3. Heterozygous did not suffer from bleeding and have normal platelet aggregation; however, their platelets mimicked homozygous ones by failing to undergo normal adhesion under flow and spreading. Rescue experiments on cultured patient megakaryocytes corrected the functional deficiency after transfection with wild-type RASGRP2. Remarkably, the presence of a single normal allele is sufficient to prevent bleeding, making CalDAG-GEFI a novel and potentially safe therapeutic target to prevent thrombosis.

PMID:
24958846
PMCID:
PMC4076591
DOI:
10.1084/jem.20130477
[Indexed for MEDLINE]
Free PMC Article

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