MYB is an essential regulator of primitive human hematopoiesis in human pluripotent stem cell differentiation cultures.

MYB is well recognized to be a key regulator of definitive hematopoiesis that plays an important role in the maintenance and multilineage differentiation of hematopoietic stem cells (HSCs). In the vertebrate developmental context, MYB is widely regarded dispensable for primitive hematopoiesis but critically required for the development of definitive hematopoiesis. To explore the role of MYB in human hematopoietic development we have inactivated the gene by bi-allelic TALEN-supported gene targeting in several lines of human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs), and subjected these cells to hematopoietic differentiation in well-defined cell culture conditions. Venus gene reporter was inserted into the knock-in allele to monitor MYB expression during the course of the hESC/iPSC differentiation. The gene reporter system showed that MYB is specifically expressed during hematopoietic commitment in the earliest primitive blood cells. Moreover, the level of MYB expression was highest at the commitment stage of differentiation and significantly decreased at the maturations stage. We found that MYB was not required for initial hematopoietic commitment of nascent mesoderm and emergence of primitive, yolk sac-type human hematopoietic progenitors. However, inactivation of MYB severely abrogated proliferation of the primitive erythroid and mixed erythroid-macrophage-megakaryocyte progenitors. In addition, MYB-negative hESC/iPSC lines demonstrated major defects in myeloid cell development and completely failed to generate mature granulocytes. Transposon-mediated rescue of MYB expression in MYB-null cells efficiently restored both the primitive hematopoietic progenitors and immature myeloid cells. Our data indicate that in contrast to its previously attributed exclusive role in definitive hematopoiesis, MYB is indispensable for primitive human hematopoiesis.