Characterisation of a novel protein, ANKRD18A, implicated in a severe form of thrombocytopenia

[摘要] Platelets (thrombocytes) mediate clot formation following vessel injury.Inherited thrombocytopenias, characterised by platelet counts below 150 x 109 L-1, havevariable bleeding severity. Whole exome sequencing of two cousins with severe thrombocytopeniaidentified a shared single-codon deletion in ANKRD18A. ANKRD18A’s function is unknown.We hypothesise that this mutation in ANKRD18A is the aetiology of these children’s thrombocytopenia. Despite limited patients’ blood, quantitative real-time polymerase chain reactiondemonstrated reduced ANKRD18A mRNA levels in patient leukocytes compared to controls.Patients’ platelets demonstrated decreased activity when stimulated with adenosine diphosphate, collagen related peptide and the synthetic peptide SFLLRN, which mimics the N-terminal sequence of protease activated receptors.Electron microscopy demonstrated macrothrombocytopenia. To assist characterisation studies, a recombinant fragment of ANKRD18A, the ankyrin repeat domain, was purified to generate a polyclonal antibody, α-25277. Experiments demonstrated possible specificity of α-25277.Cellular functions of ANKRD18A remain unclear, yet the availability of α-25277, once better characterised, will be instrumental in demonstrating the potential link between ANKRD18A and the observed thrombocytopenia.Another in-house antibody, α-21234, may also be effective.Antibody specificity can be proven by immunoblotting lysates of cells, where ANKRD18A is silenced by RNAi.Should both antibodies not be specific, generating a monoclonal antibody is requisite.

[发布机构] University:University of Birmingham;Department:Institute of Cardiovascular Sciences

[关键词] R Medicine;RC Internal medicine