Fli-1 belongs to the Ets transcription factor family and is expressed primarily in hematopoietic cells, including most cells active in immunity. To assess the role of Fli-1 in lymphocyte development in vivo, we generated mice that express a truncated Fli-1 protein, lacking the C-terminal transcriptional activation domain (Fli-1(DeltaCTA)). Fli-1(DeltaCTA)/Fli-1(DeltaCTA) mice had significantly fewer splenic follicular B cells, and an increased number of transitional and marginal zone B cells, compared with wild-type controls. Bone marrow reconstitution studies demonstrated that this phenotype is the result of lymphocyte intrinsic effects. Expression of Igalpha and other genes implicated in B cell development, including Pax-5, E2A, and Egr-1, are reduced, while Id1 and Id2 are increased in Fli-1(DeltaCTA)/Fli-1(DeltaCTA) mice. Proliferation of B cells from Fli-1(DeltaCTA)/Fli-1(DeltaCTA) mice was diminished, although intracellular Ca(2+) flux in B cells from Fli-1(DeltaCTA)/Fli-1(DeltaCTA) mice was similar to that of wild-type controls after anti-IgM stimulation. Immune responses and in vitro class switch recombination were also altered in Fli-1(DeltaCTA)/Fli-1(DeltaCTA) mice. Thus, Fli-1 modulates B cell development both centrally and peripherally, resulting in a significant impact on the in vivo immune response.