SCL and Its Partner E2A Are Required for Long-Term Stem Cell Activity.
Authors: Lacombe Julie , Herblot Sabine , Sauvageau Guy , Hoang Trang 
Published: April 2020

SCL/TAL1 is a member of the basic helix-loop-helix family of transcription factors that also includes E2A and HEB. SCL is essential at the onset of hematopoiesis and activates transcription through heterodimerization with E2A. Both genes are expressed in long-term and short-term repopulating hematopoietic stem cells (HSCs) as assessed by expression profiling. To determine whether these genes regulate lineage commitment in HSCs and possibly other HSCs functions, we proceeded to serial transplantation of E2A-deficient or SCL-impaired HSCs. All mice were on a C57Bl/6 background and transplanted donor cells were discriminated from host cells by differential expression of CD45 isoforms. Since SCL function is dosage-sensitive, we used bone marrow cells from SCL LacZ/wt mice in which one SCL allele has been invalidated. When transplanted mice were analyzed at 4 months for reconstitution in all hematopoietic organs, SCLLacZ/wt HSCs (CD45.2) showed normal repopulating capacity in competition with wild type cells (CD45.1) as expected. Strikingly, analysis at 8 months revealed that SCLLacZ/wt HSCs were markedly impaired compared to normal HSCs. The contribution of SCLLacZ/wt cells to the Kit+Sca+Lin− (KSL) and all myeloid progenitor (CMP, MEP and GMP) populations was less than 5%. This long term defect was also observed in secondary transplantation assays. Despite this near absence of HSCs and CMPs, common lymphoid progenitors (CLPs) were almost equally composed of SCLLacZ/wt and wild type cells, and the generation of B22O positive cells was expanded when SCL gene dosage was reduced. Together, our results indicate that long-term stem cell activity requires full SCL gene dosage. Furthermore, SCL operates at the branchpoint of the myeloid and lymphoid lineages to favor the myeloid lineages. Since E2A is an obligate SCL partner, we next addressed E2A function in HSCs. As E2A−/− embryos die before birth, we therefore transplanted E14.5 fetal liver cells. E2A deficiency did not affect the number of HSCs, as assessed by limiting dilution analysis. Nonetheless, serial transplantation assays revealed that HSCs from wild type littermates could be serially transplanted beyond the fourth passage while there was an early exhaustion of E2A-deficient HSCs at the third transplantation. Together, our analyses are consistent with the view that E2A is the functional partner of SCL in HSCs and that SCL/E2A does not control the size of the HSC pool but is required for long-term HSC activity. Similarly to SCL impairment, all progenitors were decreased by E2A deficiency. There were, however, two major differences. First, CLPs were severely impaired indicating that E2A activity is required for specification of the lymphoid lineages. Second, CD11b positive cells were expanded by E2A-deficiency and this was gene dosage-dependent, consistent with the view that E2A inhibits the monocyte-granulocyte lineage. Taken together, our results indicate that SCL collaborates with E2A in HSCs to maintain their long-term activities. In contrast, at the myeloid-lymphoid branchpoint, the functions of SCL and E2A are opposite: SCL favors the myeloid lineage at the expense of the lymphoid lineage, while E2A specifies the lymphoid lineage and inhibits granulocyte-macrophage differentiation.