Endocytosis of the Thrombopoietin Receptor Mpl Regulates Both Megakaryocyte and Erythroid Maturation in Mice

The thrombopoietin receptor Mpl plays a critical role in thrombopoiesis, as it is the primary signaling constituent in megakaryocyte (MK) differentiation from hematopoietic stem and progenitor cells (HSPCs) and maintains circulating blood thrombopoietin levels via clathrin-mediated endocytosis. Thus, thrombopoiesis is tightly controlled by Mpl cell-surface expression on HSPCs, MKs, and platelets. Previous work by our group has shown that Dnm2^fl/flPF4-Cre (Dnm2^Plt-/-) mice, lacking the large endocytic GTPase dynamin 2 (DNM2) specifically within MKs and platelets, develop a variety of clinical phenotypes that closely resemble myelofibrosis, such as bone marrow fibrosis, marked HSPC expansion, MK hyperplasia, extramedullary hematopoiesis, and severe splenomegaly (Bender, Giannini, et al. Blood. 2015;125(6):1014-1024). Dnm2^Plt-/- mice also displayed elevated plasma TPO levels and constitutive JAK2 activation in platelets, due to defective Mpl endocytosis. Here, the role of Mpl endocytosis in the maintenance of normal hematopoiesis was assessed by generating Dnm2^Plt-/- mice in the Mpl^-/- background. At 3 weeks of age, Mpl^-/- Dnm2^Plt-/- mice displayed significantly reduced HSPCs, a near complete depletion of bone marrow MKs, similar to Mpl^-/- mice, indicating that Mpl is the primary receptor contributing to the aberrant hyperproliferative phenotype of Dnm2^Plt-/- mice. However, Mpl^-/- Dnm2^Plt-/- mice also showed severe anemia, defects in erythroblast maturation, grossly elevated plasma erythropoietin (EPO) levels, and splenomegaly, resulting in early fatality by 3 to 4 weeks of age, suggesting that Mpl contributes to normal erythropoiesis in young mice. Mpl^-/- and Mpl^+/- Dnm2^Plt-/- mice displayed reduced erythroblast development at 3 weeks of age, which returned to normal with adulthood. Taken together, the data shows that DNM2-dependent Mpl endocytosis is required for steady-state hematopoiesis and provides novel insights into a developmentally-controlled role for Mpl in normal erythropoiesis.