Functional characterization of a novel GFI1B mutation causing congenital macrothrombocytopenia. (13th June 2016)

• Record Type: Journal Article
• Authors: Kitamura, K.; Okuno, Y.; Yoshida, K.; Sanada, M.; Shiraishi, Y.; Muramatsu, H.; Kobayashi, R.; Furukawa, K.; Miyano, S.; Kojima, S.; Ogawa, S.; Kunishima, S.
• Is Part Of: Journal of thrombosis and haemostasis. Volume 14:Number 7(2016:Jul.)
• Page Start: 1462
• Page End: 1469
• Publication Date: 2016-06-13
• Subjects: blood platelet disorders -- GFI1B protein -- human -- platelet granule deficiency disorder -- thrombocytopenia -- transcription factors; Thrombosis -- Periodicals; Hemostasis -- Periodicals; Blood coagulation disorders -- Periodicals; 616.1
• DOI: 10.1111/jth.13350
• Languages: English
• ISSNs: 1538-7933

Abstract

Abstract : Essentials Two groups recently reported GFI1B as a novel causative gene for congenital macrothrombocytopenia. We performed functional analysis of a novel GFI1B mutation and previous mutations. An immunofluorescence analysis of the platelet CD34 expression can be useful as a screening test. Mutant‐transduced megakaryocytes produced enlarged proplatelet tips which were reduced in number. Summary: Background: GFI1B is an essential transcription factor for megakaryocyte and erythrocyte development. Two groups have recently identified GFI1B as a novel causative gene for congenital macrothrombocytopenia associated with α‐granule deficiency. Methods: We performed whole exome sequencing and identified a novel GFI1B p.G272fsX274 mutation in a family with macrothrombocytopenia, and a decreased number of platelet α‐granules and abnormally shaped red blood cells. p.G272fsX274 and the previous two mutations all predicted disruption of an essential DNA‐binding domain in GFI1B. We therefore performed functional studies to characterize the biochemical and biological effects of these three patient‐derived mutations. Results: An immunofluorescence analysis revealed decreased thrombospondin‐1 and increased CD34 expression in platelets from our patient. Consistent with the previous studies, the three patient‐derived mutants were unable to repress the expression of the reporter gene and had a dominant‐negative effect over wild‐type GFI1B. In addition, the three mutations abolished recognition of a consensus‐binding site in gel shift assays. Furthermore, transduction of mouse fetal liver‐derived megakaryocytes with the three GFI1B mutants resulted in the production of abnormally large proplatelet tips, which were reduced in number. Conclusions: Our study provides further proof of concept that GFI1B is an essential protein for the normal development of the megakaryocyte lineage.