Significance TPO is a cytokine that signals through the receptor MPL (or TPO-R), and is essential for megakaryocyte differentiation and maintenance of hematopoietic stem cells (HSCs). TPO signaling is deregulated in essential thrombocythemia (ET). Here, we engineered diabodies (DBs) against the TPO-R ECD as surrogate TPO ligands to manipulate TPO-R signaling, from full to partial agonism, and that show decoupling of the dual functions of TPO/TPO-R (i.e, HSC maintenance versus megakaryopoiesis). We subsequently discovered that partial agonistic DBs, by reducing the strength of the TPO-R signal, not only preserved HSCs in culture, but also blocked oncogenic signaling in ET. This finding has the potential to improve HSC cultures for transplants, as well as serve as a unique therapeutic approach for ET.