To investigate the role of the Lyn kinase in establishing signaling thresholds in hematopoietic cells, a gain-of-function mutation analogous to the Src Y527F-activating mutation was introduced into the Lyn gene. Intriguingly, although Lyn is widely expressed within the hematopoietic system, these mice displayed no propensity toward hematological malignancy. By contrast, analysis of aging cohorts of both loss- and gain-of-function Lyn mutant mice revealed that Lyn(-/-) mice develop splenomegaly, increased numbers of myeloid progenitors, and monocyte/macrophage (M phi) tumors. Biochemical analysis of cells from these mutants revealed that Lyn is essential in establishing ITIM-dependent inhibitory signaling and for activation of specific protein tyrosine phosphatases within myeloid cells. Loss of such inhibitory signaling may predispose mice lacking this putative protooncogene to tumorigenesis.

Genetic ablation of the Lyn tyrosine kinase has revealed unique inhibitory roles in B lymphocyte signaling. We now report the consequences of sustained activation of Lyn in vivo using a targeted gain-of-function mutation (Lynup/up mice). Lynup/up mice have reduced numbers of conventional B lymphocytes, down-regulated surface immunoglobulin M and costimulatory molecules, and elevated numbers of B1a B cells. Lynup/up B cells are characterized by the constitutive phosphorylation of negative regulators of B cell antigen receptor (BCR) signaling including CD22, SHP-1, and SHIP-1, and display attributes of lymphocytes rendered tolerant by constitutive engagement of the antigen receptor. However, exaggerated positive signaling is also apparent as evidenced by the constitutive phosphorylation of Syk and phospholipase Cγ2 in resting Lynup/up B cells. Similarly, Lynup/up B cells show a heightened calcium flux in response to BCR stimulation. Surprisingly, Lynup/up mice develop circulating autoreactive antibodies and lethal autoimmune glomerulonephritis, suggesting that enhanced positive signaling eventually overrides constitutive negative signaling. These studies highlight the difficulty in maintaining tolerance in the face of chronic stimulation and emphasize the pivotal role of Lyn in B cell signaling.

Lyn-deficient mice develop Ab-mediated autoimmune disease resembling systemic lupus erythematosus where hyperactive B cells are major contributors to pathology. In this study, we show that an inflammatory environment is established in Lyn−/− mice that perturbs several immune cell compartments and drives autoimmune disease. Lyn−/− leukocytes, notably B cells, are able to produce IL-6, which facilitates hyperactivation of B and T cells, enhanced myelopoiesis, splenomegaly, and, ultimately, generation of pathogenic autoreactive Abs. Lyn−/− dendritic cells show increased maturation, but this phenotype is independent of autoimmunity as it is reiterated in B cell-deficient Lyn−/− mice. Genetic deletion of IL-6 on a Lyn-deficient background does not alter B cell development, plasma cell accumulation, or dendritic cell hypermaturation, suggesting that these characteristics are intrinsic to the loss of Lyn. However, hyperactivation of B and T cell compartments, extramedullary hematopoiesis, expansion of the myeloid lineage and autoimmune disease are all ameliorated in Lyn−/−IL-6−/− mice. Importantly, our studies show that although Lyn−/− B cells may be autoreactive, it is the IL-6–dependent inflammatory environment they engender that dictates their disease-causing potential. These findings improve our understanding of the mode of action of anti–IL-6 and B cell-directed therapies in autoimmune and inflammatory disease treatment.

ultimately lead to splenomegaly and myeloid neoplasia. In this study, we demonstrate that loss of Lyn results in a stem/ progenitor cell-intrinsic defect leading to an age-dependent increase in myeloid, erythroid, and primitive hematopoietic progenitor numbers that is independent of autoimmune disease. Despite possessing increased numbers of erythroid progenitors, and a more robust expansion of these cells following phenylhydrazine challenge, Lyn-deficient mice are more severely affected by the chemotherapeutic drug 5-fluorouracil, revealing a greater proportion of cycling progenitors. We also show that mice lacking SHIP-1 have defects in the erythroid and myeloid compartments similar to those in mice lacking Lyn or SHP-1, suggesting an intimate relationship between Lyn, SHP-1, and SHIP-1 in regulating hematopoiesis. IntroductionThe production and lineage commitment of hematopoietic cells is governed by the actions of a multitude of cytokines, hormones, and growth factors that bind to cell surface receptors activating signal transduction cascades that ultimately regulate the expression of genes that control cell fate and effector function. 1 Signal propagation in these cells is actively counterbalanced by several families of inhibitory gene products including protein tyrosine phosphatases, 2 phosphatidyl-inositol phosphatases, 3 the suppressors of cytokine signaling (SOCS) proteins, 4 and receptors bearing immunoreceptor tyrosine-based inhibitory motifs (ITIMs). 5 The central role played by tyrosine phosphorylation is exemplified by mutations in particular genes that lead to deregulation of hematopoiesis. For example, mutational activation of either the Abl 6,7 or Janus tyrosine kinases [8][9][10] leads to leukemogenesis. Loss of appropriate negative regulation of signaling may also have catastrophic consequences. For example, loss-of-function mutations within the inhibitory phosphatase Src homology 2 (SH2)-containing phosphatase-1 (SHP-1) [11][12][13] in motheaten and motheaten viable mice (Me v ), or disruption of the murine SH2-domain containing 5Ј-inositol phosphatase (SHIP-1) 14,15 gene, lead to severe perturbations in hematopoiesis with myeloid cell consolidation of the lungs of deficient mice leading to premature death. [14][15][16] Thus, the appropriate balance of positive and negative elements of signal transduction is essential for maintaining normal hematopoietic cell self-renewal, differentiation, and immune cell function.Although clearly involved in initiating tyrosine-phosphorylation cascades following hematopoietic cell stimulation, 17 Lyn has emerged as a critical enzyme responsible for establishing signaling thresholds in B cells, 18-21 myelomonocytic cells, 22,23 and mast cells. [24][25][26][27][28] Indeed, loss of Lyn kinase leads to defects in activation of inhibitory phosphatases that likely underlies the hypersensitivity of deficient cells to immunoreceptor and cytokine stimulation. 20,22,29,30 In the case of B cells and mast cells, Lyn deficiency is associated with impaired activation of ...