title: αIIbβ3 variants defined by next-generation sequencing: predicting variants likely to cause Glanzmann thrombasthenia.
abstract: Next-generation sequencing is transforming our understanding of human genetic variation but assessing the functional impact of novel variants presents challenges. We analyzed missense variants in the integrin αIIbβ3 receptor subunit genes ITGA2B and ITGB3 identified by whole-exome or -genome sequencing in the ThromboGenomics project, comprising ∼32,000 alleles from 16,108 individuals. We analyzed the results in comparison with 111 missense variants in these genes previously reported as being associated with Glanzmann thrombasthenia (GT), 20 associated with alloimmune thrombocytopenia, and 5 associated with aniso/macrothrombocytopenia. We identified 114 novel missense variants in ITGA2B (affecting ∼11% of the amino acids) and 68 novel missense variants in ITGB3 (affecting ∼9% of the amino acids). Of the variants, 96% had minor allele frequencies (MAF) < 0.1%, indicating their rarity. Based on sequence conservation, MAF, and location on a complete model of αIIbβ3, we selected three novel variants that affect amino acids previously associated with GT for expression in HEK293 cells. αIIb P176H and β3 C547G severely reduced αIIbβ3 expression, whereas αIIb P943A partially reduced αIIbβ3 expression and had no effect on fibrinogen binding. We used receiver operating characteristic curves of combined annotation-dependent depletion, Polyphen 2-HDIV, and sorting intolerant from tolerant to estimate the percentage of novel variants likely to be deleterious. At optimal cut-off values, which had 69-98% sensitivity in detecting GT mutations, between 27% and 71% of the novel αIIb or β3 missense variants were predicted to be deleterious. Our data have implications for understanding the evolutionary pressure on αIIbβ3 and highlight the challenges in predicting the clinical significance of novel missense variants.
authors.affiliation: Allen and Frances Adler Laboratory of Blood and Vascular Biology and.
authors.forename: Lorena
authors.initials: L
authors.lastname: Buitrago
authors.affiliation: Department of Haematology, University of Cambridge, Cambridge CB2 0PT, United Kingdom; Medical Research Council Biostatistics Unit, Cambridge Biomedical Campus, Cambridge Institute of Public Health, Cambridge, United Kingdom; National Health Service Blood & Transplant, Cambridge, United Kingdom;
authors.forename: Augusto
authors.initials: A
authors.lastname: Rendon
authors.affiliation: Research Bioinformatics, The Rockefeller University, New York, NY 10065;
authors.forename: Yupu
authors.initials: Y
authors.lastname: Liang
authors.affiliation: Department of Haematology, University of Cambridge, Cambridge CB2 0PT, United Kingdom; National Health Service Blood & Transplant, Cambridge, United Kingdom;
authors.forename: Ilenia
authors.initials: I
authors.lastname: Simeoni
authors.affiliation: Department of Structural and Chemical Biology, Icahn School of Medicine at Mount Sinai, New York, NY 10029; and.
authors.forename: Ana
authors.initials: A
authors.lastname: Negri
authors.affiliation: Department of Structural and Chemical Biology, Icahn School of Medicine at Mount Sinai, New York, NY 10029; and.
authors.forename: Marta
authors.initials: M
authors.lastname: Filizola
authors.affiliation: Department of Haematology, University of Cambridge, Cambridge CB2 0PT, United Kingdom; National Health Service Blood & Transplant, Cambridge, United Kingdom; Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, United Kingdom.
authors.forename: Willem H
authors.initials: WH
authors.lastname: Ouwehand
authors.affiliation: Allen and Frances Adler Laboratory of Blood and Vascular Biology and collerb@rockefeller.edu.
authors.forename: Barry S
authors.initials: BS
authors.lastname: Coller
mesh_terms: D000483:Alleles; D030561:Databases, Nucleic Acid; D059472:Exome / Q000235:genetics; D005340:Fibrinogen / Q000737:chemistry / Q000378:metabolism; D005787:Gene Frequency; D057809:HEK293 Cells; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D015151:Immunoblotting; D008958:Models, Molecular; D020125:Mutation, Missense; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013915:Thrombasthenia / Q000235:genetics* / Q000378:metabolism
chemical_list: D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D005340:Fibrinogen
keywords: Glanzmann; integrin; molecular modeling; next-generation sequencing; single-nucleotide variants
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