A dominant-negative GFI1B mutation causes autosomal dominant gray platelet syndrome. L. Van Laer1, D. Monteferrario2, N. A. Bolar1, K. Hebeda3, S. Bergevoet2, H. Veenstra2, B. Laros2, M. MacKenzie2, E. Fransen1,5, G. Van Camp1, F. Preijers2, S. Salemink4, W. van Heerde2, G. Huls2, J. H. Jansen2, M. Kempers4, B. A. van der Reijden2, B. L. Loeys1 1) Center of Medical Genetics, Antwerp University Hospital/University of Antwerp, Antwerp, Belgium; 2) Department of Laboratory Medicine, Laboratory of Hematology Nijmegen Centre of Molecular Life sciences, Radboud University Medical Centre, Nijmegen, The Netherlands; 3) Department of Pathology, Radboud University Medical Centre, Nijmegen, The Netherlands; 4) Department of Clinical Genetics, St. Radboud University Medical Centre, Nijmegen, The Netherlands; 5) StatUa Center for Statistics, University of Antwerp, Antwerp, Belgium.

   Gray platelet syndrome (GPS) is a hereditary, usually autosomal recessive bleeding disorder caused by defective production of -granules in platelets. It has been proposed that the -granule deficiency in GPS can be attributed to the failure of megakaryocytes to efficiently route endogenous proteins into -granules, thereby hampering their maturation. Here we describe a large family with an autosomal dominant type of GPS characterized by mild to severe bleeding complications. In addition to large gray platelets lacking -granules, other GPS-associated phenomena like thrombocytopenia, emperipolesis, myelofibrosis and lower expression of platelet factor 4 were observed in affected individuals. To determine the disease causing mutation we first performed linkage analysis and identified a candidate locus on chromosome 9q34 with a LOD score of 3.9. We considered GFI1B (Growth factor independence 1B), located within this region, an excellent candidate gene because of its function as a transcriptional repressor essential in erythroid and megakaryocyte lineage development. Sequence analysis identified a nonsense mutation in exon 6 (c.859C>T, p.Gln287*) that co-segregated with the GPS in this family. The mutated transcript was not targeted for nonsense-mediated mRNA decay, resulting in a protein truncated within its 5th zinc finger, a domain essential for DNA binding. Using luciferase reporter assays we demonstrated that the truncated GFI1B protein was unable to repress gene expression and that it inhibited wild type GFI1B in a dominant-negative manner. Subsequently, we performed an immunophenotypical analysis of the peripheral blood of affected family members. Myeloid and erythroid lineages were unaffected, but we observed aberrant platelets, dysplastic abnormalities and disturbed lineage marker expression in GFI1B-mutated megakaryocytes. These studies define GFI1B to be key to proper megakaryocyte development, platelet production and -granule biogenesis.

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