Haematopoietic stem cells are widely used in clinical applications but their low abundance severely limits their use. Therefore, strategies that induce either expansion of HSC or the generation of repopulating HSC from alternative sources are highly desirable. To be able to direct reprogramming of non-haematopietic stem cells into HSC, transcription factors that induce the hematopoietic stem cell transcriptional network in the target cells have to identified. We and others have previously shown that the transcription factor HOXB4 instructs differentiating mouse embryonic stem cells to give rise to haematopoietic stem and progenitor cells able to reconstitute lethally irradiated mice in all haematopoietic lineages. We also found expression levels of HOXB4 to be deterministic for haematopoietic lineage fate, with high HOXB4 levels favoring HSC and myeloid fate and low levels being permissive for lymphoid and erythroid differentiation.

Here, we tested HOXB4 as a reprogramming factor, instructing committed blood cells to de-differentiate into repopulating HSC. Ectopic expression of HOXB4 in common lymphoid progenitors (CLP), common myeloid progenitors (CMP) and megakaryocyte-erythroid progenitors (MEP) lead to reprogramming into HSC: Strikingly, HOXB4-infected CLP were still detectable 6 months after transplantation in lethally irradiated mice, developed into lineage-CD150+CD117+Sca1+ stem cells and gave rise to lymphoid as well as myeloid cells. We obtained similar results with HOXB4+ CMP and MEP cells. Importantly, GFP-only infected controls were lost within the first months and were never detected in the stem cell population. To exclude any contamination of our sorted populations with HSC, we performed a lineage-tracing experiment, confirming successful reprogramming of committed cells into multipotent HSC. In summary, we identified HOXB4 as a reprogramming factor for the generation of induced haematopoietic stem cells.