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Title: Identification and characterisation of the underlying defects in patients with inherited platelet bleeding disorders
Author: Aldossary, Maryam
ISNI:       0000 0004 7961 2367
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2019
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The underlying genetic defects remain unknown in about 50% of inherited platelet bleeding disorders (IPDs). This study investigated the use of whole exome sequencing (WES) to identify candidate gene defects in 34 index cases enrolled in the UK Genotyping and Phenotyping of Platelets study with a history of bleeding, whose platelets demonstrated defects in agonist-induced dense granule secretion or Gisignalling. WES analysis identified a median of 98 candidate disease-causing variants per index case highlighting the complexity of IPDs. Sixteen variants were in genes that had previously been associated with IPDs, two of which were selected for further characterisation. Two novel FLI1 alterations, predicting p.Arg340Cys/His substitutions in the DNA binding domain of FLI1 were shown to reduce transcriptional activity and nuclear accumulation of FLI1, suggesting that these variants interfere with the regulation of essential megakaryocyte genes by FLI1 and may explain the bleeding tendency in affected patients. Expression of a novel truncated p.Arg430* variant of ETV6 revealed it to be stably expressed, possessing normal repressor activity in HEK 293T cells and a slight reduction in repressor activity in megakaryocytic cells. Further studies are required to confirm the pathogenicity of this variant. To identify novel genes involved in platelet granule biogenesis and secretion, gene expression was examined in megakaryocytic cells before and after knockdown of FLI1, defects in which are associated with platelet granule abnormalities. Comparison of the gene expression data with that from platelets from patients with FLI1 defects and with the results of WES analysis in patients with secretion defects highlighted several genes of interest, including C18orf32, IGFBP2, PLCG2, SCFD2, SLC24A3, ST8SIA6 and ZBTB45 which, between them, harboured ten candidate causative variants among the patients with defects in platelet secretion. Further work is warranted to explore the contribution of these genes to platelet secretory pathways.
Supervisor: Daly, Martina ; Makris, Michael Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available