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^ΔCTA). Fli-1^ΔCTA/Fli-1^ΔCTA 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 Igα 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^ΔCTA/Fli-1^ΔCTA mice. Proliferation of B cells from Fli-1^ΔCTA/Fli-1^ΔCTA mice was diminished, although intracellular Ca²⁺ flux in B cells from Fli-1^ΔCTA/Fli-1^ΔCTA 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^ΔCTA/Fli-1^ΔCTA mice. Thus, Fli-1 modulates B cell development both centrally and peripherally, resulting in a significant impact on the in vivo immune response.

Increased Fli-1 mRNA is present in PBLs from systemic lupus erythematosus patients, and transgenic overexpression of Fli-1 in normal mice leads to a lupus-like disease. We report in this study that MRL/lpr mice, an animal model of systemic lupus erythematosus, have increased splenic expression of Fli-1 protein compared with BALB/c mice. Using mice with targeted gene disruption, we examined the effect of reduced Fli-1 expression on disease development in MRL/lpr mice. Complete knockout of Fli-1 is lethal in utero. Fli-1 protein expression in heterozygous MRL/lpr (Fli-1^+/−) mice was reduced by 50% compared with wild-type MRL/lpr (Fli-1^+/+) mice. Fli-1^+/− MRL/lpr mice had significantly decreased serum levels of total IgG and anti-dsDNA Abs as disease progressed. Fli-1^+/− MRL/lpr mice had significantly increased splenic CD8⁺ and naive T cells compared with Fli-1^+/+ MRL/lpr mice. Both in vivo and in vitro production of MCP-1 were significantly decreased in Fli-1^+/− MRL/lpr mice. The Fli-1^+/− mice had markedly decreased proteinuria and significantly lower pathologic renal scores. At 48 wk of age, survival was significantly increased in the Fli-1^+/− MRL/lpr mice, as 100% of Fli-1^+/− MRL/lpr mice were alive, in contrast to only 27% of Fli-1^+/+ mice. These findings indicate that Fli-1 expression is important in lupus-like disease development, and that modulation of Fli-1 expression profoundly decreases renal disease and improves survival in MRL/lpr mice.

Objective: The Fli‐1 transcription factor is implicated in the pathogenesis of systemic lupus erythematosus (SLE), both in humans and in animal models. Dysregulation of interleukin‐6 (IL‐6) is also associated with SLE. The purpose of this study was to investigate whether Fli‐1 directly regulates the expression of IL‐6.Methods: Sera were collected from wild‐type and Fli‐1–heterozygous (Fli‐1^+/−) MRL/lpr mice, and the concentration of IL‐6 was measured by enzyme‐linked immunosorbent assay (ELISA). Expression of IL‐6 in the kidney was measured by real‐time polymerase chain reaction analysis. T cells were isolated from wild‐type and Fli‐1^+/− MRL/lpr mice and stimulated with CD3/CD28 beads, and the concentration of IL‐6 in the supernatants was measured by ELISA. MS1 endothelial cells were transfected with Fli‐1 and control small interfering RNA, and the production of IL‐6 was compared after lipopolysaccharide stimulation. A chromatin immunoprecipitation (ChIP) assay was performed to determine whether Fli‐1 binds to the IL‐6 promoter region. Transient transfections with the NIH3T3 cell line were performed to examine whether Fli‐1 regulates the expression of IL‐6.Results: Fli‐1^+/− MRL/lpr mice had significantly decreased IL‐6 levels in sera and reduced expression of IL‐6 in kidneys as compared to their wild‐type littermates. T cells isolated from Fli‐1^+/− MRL/lpr mice produced less IL‐6 than did those from wild‐type mice. Inhibiting the expression of Fli‐1 in endothelial cells resulted in reduced production of IL‐6. The ChIP assay revealed direct binding of Fli‐1 to 3 regions within the IL‐6 promoter. Fli‐1 activated transcription from the IL‐6 promoter in a dose‐dependent manner.Conclusion: The direct regulation of IL‐6 expression by Fli‐1 represents one possible mechanism for the protective effect of decreased Fli‐1 expression in lupus.

Summary: Fli‐1 belongs to the Ets transcription factor family and is expressed in haematopoietic cells, including most of the cells that are active in immunity. The mononuclear phagocytes, i.e. monocytes, macrophages and dendritic cells, originate in haematopoietic stem cells and play an important role in immunity. To assess the role of Fli‐1 in mononuclear phagocyte development in vivo, we generated mice that express a truncated Fli‐1 protein, lacking the C‐terminal transcriptional activation domain (Fli‐1^ΔCTA). Fli‐1^ΔCTA/ΔCTA mice had significantly increased populations of haematopoietic stem cells and common dendritic cell precursors in bone marrow compared with wild‐type littermates. Significantly increased classical dendritic cells, plasmacytoid dendritic cells, and macrophage populations were found in spleens from Fli‐1^{∆CTA/∆CTA} mice compared with wild‐type littermates. Fli‐1^ΔCTA/ΔCTA mice also had increased pre‐classical dendritic cell and monocyte populations in peripheral blood mononuclear cells. Furthermore, bone marrow reconstitution studies demonstrated that expression of Fli‐1 in both haematopoietic cells and stromal cells affected mononuclear phagocyte development in mice. Expression of Fms‐like tyrosine kinase 3 ligand (Flt3L), a haematopoietic growth factor, in multipotent progenitors was statistically significantly increased from Fli‐1^{∆CTA/∆CTA} mice compared with wild‐type littermates. Fli‐1 protein binds directly to the promoter region of the Flt3L gene. Hence, Fli‐1 plays an important role in the mononuclear phagocyte development, and the C‐terminal transcriptional activation domain of Fli‐1 negatively modulates mononuclear phagocyte development.

The friend leukemia insertion site 1 (Fli-1) transcription factor, an Ets family member, is implicated in the pathogenesis of systemic lupus erythematosus in human patients and murine models of lupus. Lupus-prone mice with reduced Fli-1 expression have significantly less nephritis, prolonged survival, and decreased infiltrating inflammatory cells into the kidney. Inflammatory chemokines, including CCL5, are critical for attracting inflammatory cells. In this study, decreased CCL5 mRNA expression was observed in kidneys of lupus-prone NZM2410 mice with reduced Fli-1 expression. CCL5 protein expression was significantly decreased in endothelial cells transfected with Fli-1–specific small interfering RNA compared with controls. Fli-1 binds to endogenous Ets binding sites in the distal region of the CCL5 promoter. Transient transfection assays demonstrate that Fli-1 drives transcription from the CCL5 promoter in a dose-dependent manner. Both Ets1, another Ets family member, and Fli-1 drive transcription from the CCL5 promoter, although Fli-1 transactivation was significantly stronger. Ets1 acts as a dominant-negative transcription factor for Fli-1, indicating that they may have at least one DNA binding site in common. Systematic deletion of DNA binding sites demonstrates the importance of the sites located within a 225-bp region of the promoter. Mutation of the Fli-1 DNA binding domain significantly reduces transactivation of the CCL5 promoter by Fli-1. We identified a novel regulator of transcription for CCL5. These results suggest that Fli-1 is a novel and critical regulator of proinflammatory chemokines and affects the pathogenesis of disease through the regulation of factors that recruit inflammatory cells to sites of inflammation.