T-Lineage Specification and Commitment Requires Constraint of Myeloid Gene Expression Programs by Hes1

Early thymic progenitors (ETPs) are not committed to the T cell lineage; however, whether ETPs realize alternative (non T cell) lineage potentials in vivo is not well understood and indeed controversial. Notch signaling induces T cell lineage gene expression and discourages alternative fate outcomes; however, the mechanisms by which this occurs remain unclear. The work described here provides insight into two related questions in the field of early T cell development: first, we address whether ETPs adopt alternative fates in the thymus (Chapter 2) and second, we investigate the mechanisms used to constrain alternative gene expression programs as progenitors commit to the T cell lineage (Chapter 3 and 4). We found that ETPs do in fact access myeloid developmental fates in vivo, since the majority of thymic granulocytes appear to derive from ETPs. Next, we identified the Notch target and transcriptional repressor Hes1 as an important mechanism that constrains myeloid gene expression programs in T cell progenitors. Hes1 deficiency in hematopoietic progenitors severely compromises T cell development; however, this defect can be completely rescued by deletion of the myeloid regulator C/EBPα. Thus, our findings indicate that ETPs are bona fide myelo-lymphoid progenitors and establishes the critical importance of constraining myeloid developmental programs early in T cell development

Runx1 Loss Minimally Impacts Long-Term Hematopoietic Stem Cells

RUNX1 encodes a DNA binding subunit of the core-binding transcription factors and is frequently mutated in acute leukemia, therapy-related leukemia, myelodysplastic syndrome, and chronic myelomonocytic leukemia. Mutations in RUNX1 are thought to confer upon hematopoietic stem cells (HSCs) a pre-leukemic state, but the fundamental properties of Runx1 deficient pre-leukemic HSCs are not well defined. Here we show that Runx1 deficiency decreases both apoptosis and proliferation, but only minimally impacts the frequency of long term repopulating HSCs (LT-HSCs). It has been variously reported that Runx1 loss increases LT-HSC numbers, decreases LT-HSC numbers, or causes age-related HSC exhaustion. We attempt to resolve these discrepancies by showing that Runx1 deficiency alters the expression of several key HSC markers, and that the number of functional LT-HSCs varies depending on the criteria used to score them. Finally, we identify genes and pathways, including the cell cycle and p53 pathways that are dysregulated in Runx1 deficient HSCs

Erythroid/myeloid progenitors and hematopoietic stem cells originate from distinct populations of endothelial cells

SummaryHematopoietic stem cells (HSCs) and an earlier wave of definitive erythroid/myeloid progenitors (EMPs) differentiate from hemogenic endothelial cells in the conceptus. EMPs can be generated in vitro from embryonic or induced pluripotent stem cells, but efforts to produce HSCs have largely failed. The formation of both EMPs and HSCs requires the transcription factor Runx1 and its non-DNA binding partner core binding factor β (CBFβ). Here we show that the requirements for CBFβ in EMP and HSC formation in the conceptus are temporally and spatially distinct. Panendothelial expression of CBFβ in Tek-expressing cells was sufficient for EMP formation, but was not adequate for HSC formation. Expression of CBFβ in Ly6a-expressing cells, on the other hand, was sufficient for HSC, but not EMP, formation. The data indicate that EMPs and HSCs differentiate from distinct populations of hemogenic endothelial cells, with Ly6a expression specifically marking the HSC-generating hemogenic endothelium

T cell development requires constraint of the myeloid regulator C/EBP-α by the Notch target and transcriptional repressor Hes1

International audienceNotch signaling induces gene expression of the T cell lineage and discourages alternative fate outcomes. Hematopoietic deficiency in the Notch target Hes1 results in severe T cell lineage defects; however, the underlying mechanism is unknown. We found here that Hes1 constrained myeloid gene-expression programs in T cell progenitor cells, as deletion of the myeloid regulator C/EBP-α restored the development of T cells from Hes1-deficient progenitor cells. Repression of Cebpa by Hes1 required its DNA-binding and Groucho-recruitment domains. Hes1-deficient multipotent progenitor cells showed a developmental bias toward myeloid cells and dendritic cells after Notch signaling, whereas Hes1-deficient lymphoid progenitor cells required additional cytokine signaling for diversion into the myeloid lineage. Our findings establish the importance of constraining developmental programs of the myeloid lineage early in T cell development