Abstract
BACKGROUND: Germline RUNX1 haplodeficiency (RHD) is associated with thrombocytopenia, platelet dysfunction and predisposition to myeloid malignancies. Platelet expression profiling of a RHD patient showed decreased F13A1, encoding for the A subunit of factor XIII, a transglutaminase that cross-links fibrin and induces clot stabilization. FXIII-A is synthesized by hematopoietic cells, megakaryocytes and monocytes. AIMS: To understand RUNX1 regulation of F13A1 expression in platelet/megakaryocyte and the mechanisms and consequences of decreased F13A1 in RHD. METHODS: We performed studies in platelets, HEL cells and human CD34+ cell-derived megakaryocytes including on clot contraction in cells following small inhibitor (si)RNA knockdown (KD) of RUNX1 or F13A1 . RESULTS: Platelet F13A1 mRNA and protein were decreased in our index patient and in two siblings from an unrelated family with RHD. Platelet-driven clot contraction was decreased in the patient and affected daughter. Promoter studies in HEL cells showed that RUNX1 regulates F13A1 transcription; RUNX1 overexpression increased and (si)RNA RUNX1 KD reduced F13A1 promoter activity and protein. Following RUNX1 or F13A1 KD clot contraction by HEL cells was decreased as were FXIII-A surface expression, myosin light chain phosphorylation and PAC1 binding upon activation. F13A1 expression and clot contraction were impaired on RUNX1 downregulation in human megakaryocytes. CONCLUSIONS: RUNX1 regulates platelet-megakaryocyte F13A1 expression, which is decreased in RHD, reflecting regulation of a coagulation protein by a hematopoietic transcription factor. Platelet and megakaryocyte clot contraction is decreased in RHD, related to multiple impaired mechanisms including F13A1 expression, myosin phosphorylation and αII (b) β (3) activation. Scientific category - Platelets and thrombopoiesis. ESSENTIALS: RUNX1 regulates expression of FXIII-A chain ( F13A1) in megakaryocytes (MK) and platelets. Platelet and MK F13A1 expression and clot contraction are decreased in RUNX1 deficiency. MK clot contraction, myosin phosphorylation and PAC1-binding are impaired in F13A1 deficiency. Defective clot contraction in RHD arises from defects in multiple platelet-MK mechanisms.