Maturation of erythroid cells and erythroleukemia development are affectedby the kinase activity of Lyn

This study examined the impact of the tyrosine kinase Lyn on erythropoietin -induced intracellular signaling in erythroid cells. In J2E erythroleukemic cells, Lyn coimmunoprecipitated with numerous proteins, including SHP-1, S HP-2, ras-GTPase-activating protein, signal transducers and activators of t ranscription (STAT) 5a, STAT5b, and mitogen-activated protein kinase; howev er, introduction of a dominant-negative Lyn (Y397F Lyn) inhibited the inter action of Lyn with all of these molecules except SHP-1. Cells containing th e dominant-negative Lyn displayed altered intracellular phosphorylation pat terns, including mitogen-activated protein kinase, but not erythropoietin r eceptor, Janus-activated kinase (JAK) 2, or STAT5. As a consequence, erythr opoietin-initiated differentiation and basal proliferation were severely im paired. Y397F Lyn reduced the protein levels of erythroid transcription fac tors erythroid Kruppel-like factor and GATA-1 up to 90%, which accounts for the inability of J2E cells expressing Y397F Lyn to synthesize hemoglobin. Although Lyn was shown to bind several sites on the cytoplasmic domain of t he erythropoietin receptor, it was not activated when a receptor mutated at the JAK2 binding site was ectopically expressed in J2E cells, indicating t hat JAK2 is the primary kinase in erythropoietin signaling and that Lyn is a secondary kinase. In normal erythroid progenitors, erythropoietin enhance d phosphorylation of Lyn; moreover, exogenous Lyn increased colony forming unit-erythroid, but not burst forming unit-erythroid, colonies from normal progenitors, demonstrating a stage-specific effect of the kinase. Significa ntly, altering Lyn activity in J2E cells had a profound effect on the devel opment of erythroleukemias in vivo: the mortality rate was markedly reduced and latent period extended when either wild-type Lyn or Y397F Lyn was intr oduced into these cells. Taken together, these data show that Lyn plays an important role in intracellular signaling in nontransformed and leukemic er ythroid cells.