Dynamic Expression of Id2 Determines Cell Fate Decisions in Multipotent Hematopoietic Cells

Abstract 2369

Inhibitor of DNA binding protein-2 (Id2) is a member of the helix-loop-helix family of transcriptional regulators that is required for the normal development of natural killer (NK) cells, dendritic cells, B cells and erythrocytes. However, little is known about the expression and function of Id2 in hematopoietic stem and progenitor cells (HSPC). Recent evidence suggests that cell fate is resolved in HSPC by regulating the levels transcription factor expression. Therefore, to determine if Id2 functions in cell fate decisions, and in which progenitor populations these decisions are made, we generated an Id2-EYFP reporter mouse model and validated that EYFP accurately reflected Id2 expression. Using this model we mapped Id2 expression levels in purified HSPC and their differentiated progeny. Id2 is highly expressed in differentiated neutrophils, dendritic cells and NK cells, but is suppressed during erythroid development. Id2 expression is dynamically regulated during lymphoid development, with high levels of Id2 expression in lymphoid progenitors that are down regulated during the early stages of T cell (DN2-DN4) and B cell (Pre-B) differentiation. Id2 is then up-regulated and highly expressed in CD4+CD8+ and single positive CD4 and CD8 thymocytes, suggesting that Id2 may have novel functions in differentiated T cells and in neutrophils. Id2 is expressed in HSC and multi-potent progenitors (MPP), is decreased in common myeloid progenitors (CMP), and further decreased in myeloid/erythroid progenitors (MEP), but increased in granulocyte/macrophage progenitors (GMP). We also observed a range of Id2 expression within purified HSPC, suggesting that the levels of Id2 expression in purified HSC, MPP and CMP may correlate with potential of these progenitors for myeloid, erythroid and lymphoid development. To investigate this, CMP Id2-hi and CMP-Id2-low expressing cells were sorted and evaluated for differentiation potential in vitro. CMP-Id2-hi cells showed greatly increased myeloid developmental potential compared to the CMP-Id2-low cells as indicated by flow cytometry and growth in soft agar. Gene expression profiles confirmed these results and showed that the CMP-Id2-hi cell expression profile correlated with the expression profile of GMP, while the CMP-Id2-lo cells showed an expression profile that resembled MEP. Based on these observations we anticipate that MPP-Id2-hi and MPP-Id2-lo cells will show differences in myeloid and lymphoid potential. The expression of Id2 in primitive HSPC suggested that Id2 may be required for the maintenance and fate of HSC. Using the Id2−/− mouse model, we discovered that Id2−/− bone marrow cells have impaired ability to rescue mice after serial transplantation, suggesting that Id2 is required for HSC self-renewal. Competitive repopulation assays showed that Id2−/− HSC have diminished repopulation potential. Thus, these data suggest that Id2 is required for the self-renewal of HSC, and that changes in the levels of Id2 expression are associated with cell fate determination of HSPC.