Thrombopoietin (TPO) Induces Hematopoietic Differentiation of Mouse ES Cells Via HIF-1 Alpha-Dependent Activation of a BMP4 Autoregulatory Loop

Understanding the molecular mechanisms underlying hematopoietic differentiation of embryonic stem (ES) cells may help to ascertain the optimal conditions for the production of hematopoietic cells as a source for transplantation or experimental use. Previously, we found that patients with congenital amegakaryocytic thrombocytopenia (CAMT), who develop pancytopenia early after birth, harbor mutations within the thrombopoietin (TPO) receptor, c-mpl. This knowledge, together with observations in vitro and in animal models in vivo, suggests that TPO/c-mpl signaling promotes early hematopoiesis. However, the downstream mechanisms underlying TPO signaling are not fully elucidated. Here, we describe for the first time a direct connection between the TPO and bone morphogenetic protein 4 (BMP4) signaling pathways in the hematopoietic fate decision of ES cells. BMP4 is a classical morphogen and a well-known inducer of early hematopoietic differentiation of ES cells. Treatment of ES cells with TPO induced the autocrine production of BMP4 by ES cells with concomitant upregulation of the BMP receptor, BMPR1A, phosphorylation of Smad1, 5, and 8 and activation of the specific target genes, Id1, 2, and 3, and Msx1 and 2. This was mediated by TPO-dependent binding of the HIF-1α transcription factor to the BMP4 gene promoter, resulting in further activation of the BMP4-autoregulatory loop. Treatment of ES cells with the BMP antagonist noggin substantially reduced TPO-dependent hematopoietic differentiation of ES cell. Taken together, our findings contribute to the understanding the mechanisms of hematopoietic differentiaiton of ES cells and might help to establish new methods for the efficient production of hematopoietic stem cells in vitro.