Enforced Nr4a2 expression Drives HSCs into Quiescence.

Hematopoietic stem cells (HSCs) are used as a paradigm for understanding somatic stem cell biology, yet the factors that regulate their self-renewal and differentiation are poorly understood. Our lab has recently discovered that within the hematopoietic lineage, Nr4a2 expression is restricted to the stem and progenitor cell compartment. Furthermore, we have observed a dramatic decrease in its expression when the bone marrow cells are exposed to 5-fluorouracil which forces quiescent HSCs into cycle. We therefore hypothesize that Nr4a2 may be involved in maintaining stem cell quiescence.

To better understand the role of Nr4a2 in hematopoiesis, we have performed gain-of-function studies in which Nr4a2 is overexpressed in HSCs. Bone marrow transplantation (BMT) experiments revealed that Nr4a2 overexpressing cells contributed significantly less to the peripheral blood compared to the control (1% vs 35%, see fig). This suggests that enforced Nr4a2 expression negatively affects HSC regeneration potential. We also found that Nr4a2 overexpression has a negative effect on the overall proliferation capacity with an in vitro methocult assay. Together, these results support our hypothesis that the high level expression of Nr4a2 regulates HSC quiescence by inhibiting HSC self-renewal and/or differentiation.

Cell cycle analysis demonstrates that enforced expression of Nr4a2 causes an increase in the number of cells in quiescence. These data are supported further by analysis at a molecular level revealing that overexpression of Nr4a2 increases expression of cell cycle inhibitors such as p18 and p19, both of which are important for Go to G1 entry. By uncovering the role of Nr4a2 in hematopoiesis we aim to achieve not only a better understanding of the regulation of stem cell self-renewal and differentiation but also of the pathways involved in HSC cell cycle regulation.

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