Programmed cell death plays an important role in erythropoiesis under physiological and pathological conditions. In this study, we show that the Notch/RBPjkappa signaling pathway induces erythroid apoptosis in different hematopoietic tissues, including yolk sac and bone marrow as well as in murine erythroleukemia cells. In RBPjkappa(-/-) yolk sacs, erythroid cells have a decreased rate of cell death that results in increased number of Ter119(+) cells. A similar effect is observed when Notch activity is abrogated by incubation with the gamma-secretase inhibitors, DAPT or L685,458. We demonstrate that incubation with Jagged1-expressing cells has a proapoptotic effect in erythroid cells from adult bone marrow that is prevented by blocking Notch activity. Finally, we show that the sole expression of the activated Notch1 protein is sufficient to induce apoptosis in hexametilene-bisacetamide-differentiating murine erythroleukemia cells. Together these results demonstrate that Notch regulates erythroid homeostasis by inducing apoptosis.