A gradual restriction in lineage potential of multipotent stem/progenitor cells is a hallmark of adult hematopoiesis, but the underlying molecular events governing these processes remain incompletely understood. Here, we identified robust expression of the leukemia-associated transcription factor hepatic leukemia factor (Hlf) in normal multipotent hematopoietic progenitors, which was rapidly downregulated upon differentiation. Interference with its normal downregulation revealed Hlf as a strong negative regulator of lymphoid development, while remaining compatible with myeloid fates. Reciprocally, we observed rapid lymphoid commitment upon reduced Hlf activity. The arising phenotypes resulted from Hlf binding to active enhancers of myeloid-competent cells, transcriptional induction of myeloid, and ablation of lymphoid gene programs, with Hlf induction of nuclear factor I C (Nfic) as a functionally relevant target gene. Thereby, our studies establish Hlf as a key regulator of the earliest lineage-commitment events at the transition from multipotency to lineage-restricted progeny, with implications for both normal and malignant hematopoiesis.This work was generously supported by project grants to DB from the Swedish Cancer Society, the Swedish Medical Research Council, the Swedish Pediatric Leukemia Foundation, Knut and Alice Wallenberg foundation and an ERC consolidator grant (615068). We would like to acknowledge Tom Serwold, Ewa Sitnicka and Mikael Sigvardsson for valuable scientific discussions, and Eva Erlandsson and Gerd Sten for expert technical assistance. The Genome Technology Access Center, Department of Genetics, Washington University School of Medicine,
assisted with genomic analysis and is partially supported by NCI Cancer Center Support Grant P30 CA91842 to the Siteman Cancer Center, ICTS/CTSA Grant UL1TR000448 from the National Center for Research Resources (NCRR, a component of the NIH), and the NIH Roadmap for Medical Research
