Dysmegakaryopoiesis in FPD/AML is linked to the downregulation of NFE2 and MYL9, two downstream RUNX1 targets

BLUTEAU D.; GLEMBOTSKY AC.; GILLES L.; HELLER PG.; NURDEN P.; DEBILI N.; VAINCHENKER W.; FAVIER R.; RASLOVA H.

Encuentro; Gordon Research Conference: Cell Biology of megakaryocytes and platelets; 2011

FPD/AML is a familial platelet disorder characterized by qualitative and quantitative platelet defects, predisposition to acute myelogenous leukaemia and germline heterozygous RUNX1 alterations. We investigated in detail the in vitro megakaryopoiesis from FPD/AML patients and after knock-down of RUNX1 by shRNA to understand the mechanisms by which RUNX1 regulates platelet production. We first studied megakaryopoiesis derived from CD34+CD38+ progenitors of two pedigrees harboring either R174Q or R139X RUNX1 mutations. A 60-80% decrease in the output of megakaryocyte (MK) cells was detected. RUNX1 knockdown in normal CD34+ cells leads also to a decreased MK progenitor number followed by an impaired maturation. When mature CD41highCD42high MK were sorted, a major defect in proplatelet formation was observed in FPD/AML patients as well as after RUNX1 inhibition. To explain this defect in proplatelet formation we studied expression of two candidate genes i.e. MYL9 and NF-E2. We found a downregulation of these two genes as well as of NF-E2 transcription targets in FPD/AML and RUNX1 knock-down MKs.. Chromatin immunoprecipitation in primary MK cells in vivo and luciferase asays in HEL cells showed that both genes are directly regulated by RUNX1 and thus their deregulation can contribute to the thrombocytopenia in FPD/AML.