The high-affinity receptor for granulocyte-macrophage colony-stimulating factor (GM-CSF) consists of a unique a chain and a Pc subunit that is shared with the receptors for interleukin-3 (IL3) and . Two regions of the Pc chain have been defined; these include a membrane-proximal region of the cytoplasmic domain that is required for mitogenesis and a membrane-distal region that is required for activation of Ras, Raf-1, mitogen-activated protein kinase, and S6 kinase. Recent studies have implicated the cytoplasmic protein tyrosine kinase JAK2 in signalling through a number of the cytokine receptors, including the IL-3 and erythropoietin receptors. In the studies described here, we demonstrate that GM-CSF stimulation of cells induces the tyrosine phosphorylation of JAK2 and activates its in vitro kinase activity. Hematopoiesis is regulated through the interaction of one or more of a variety of cytokines with their cognate receptors. Granulocyte-macrophage colony-stimulating factor (GM-CSF) regulates the proliferation and differentiation of cells at various stages of differentiation along the myeloid lineages (1, 17). The functional, high-affinity receptor for GM-CSF consists of two subunits, each of which is a member of the cytokine receptor superfamily (19,20). The binding of GM-CSF occurs through an a subunit of 60 to 80 kDa, which alone binds GM-CSF with low affinity. Association of the a chain with a 140-kDa 1c chain results in the formation of a high-affinity binding site for GM-CSF. The 1c chain also associates with 'a subunits that specifically bind interleukin-3 (IL-3) and IL-5 and similarly contributes to the formation of high-affinity binding sites for these ligands.Stimulation of growth factor-dependent cell lines with GM-CSF has been shown to induce a variety of immediate cellular responses, including the rapid tyrosine phosphorylation of the 1c subunit and a number of cellular substrates (7, 16, 30); induction of transcription of several immediate-early genes (5); and activation of components of the Ras signalling pathway, including SHC phosphorylation, increases in GTP-bound Ras, activation of Raf-1 kinase, and activation of mitogenactivated protein (MAP) kinase (4,8,23,26,27 inducing mitogenesis, while a distal region is required for activation of Ras, Raf-1, MAP kinase, and S6 kinase.Although considerable evidence demonstrates that induction of protein tyrosine phosphorylation is critical to cytokine function, only recently have the JAK family of kinases been implicated in coupling ligand binding to tyrosine phosphorylation. The JAK family consists of JAK1 (15), JAK2 (29), and TYK2 (9) and is characterized by proteins of approximately 130 kDa that contain a carboxyl kinase domain and a second kinase-like domain and lack SH2 or SH3 domains. JAK2 has been shown to be activated in the responses to erythropoietin (Epo) (36), , growth hormone (2), G-CSF (35a), and prolactin (3). In contrast, IL-6, ciliary neurotrophic factor (CNTF), and leukemia-inhibitory factor activate JAK1, JAK2, and to some extent T...

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The high‐affinity receptors for granulocyte‐macrophage colony stimulating factor (GM‐CSF), interleukin 3 (IL‐3) and IL‐5 consist of two subunits, alpha and beta. The alpha subunits are specific to each cytokine and the same beta subunit (beta c) is shared by these three receptors. Although none of these receptor subunits has intrinsic kinase activity, these cytokines induce protein tyrosine phosphorylation, activation of Ras, Raf‐1 and MAP kinase, and transcriptional activation of nuclear proto‐oncogenes such as c‐myc, c‐fos and c‐jun. In this paper, we describe a detailed analysis of the signaling potential of the beta c subunit by using a series of cytoplasmic deletion mutants. The human beta c consists of 881 amino acid residues. A C‐terminal deletion mutant of beta c at amino acid 763 (beta 763) induced phosphorylation of Shc and activation of Ras, Raf‐1, MAP kinase and p70 S6 kinase, whereas a deletion at amino acid 626 (beta 626) induced none of these effects. The beta 763 mutant, as well as the full‐length beta c, induced transcription of c‐myc, c‐fos and c‐jun. Deletions at amino acid 517 (beta 517) and 626 (beta 626) induced c‐myc and pim‐1, but no induction of c‐fos and c‐jun was observed. GM‐CSF increased phosphatidylinositol 3 kinase (PI3‐K) activity in anti‐phosphotyrosine immunoprecipitates from cells expressing beta 763 as well as beta c, whereas it was only marginally increased from cells expressing beta 517 or beta 626. Thus, there are at least two distinct regions within the cytoplasmic domain of beta c that are responsible for different signals, i.e. a membrane proximal region of approximately 60 amino acid residues upstream of Glu517 is essential for induction of c‐myc and pim‐1, and a distal region of approximately 140 amino acid residues (between Leu626 and Ser763) is required for activation of Ras, Raf‐1, MAP kinase and p70 S6 kinase, as well as induction of c‐fos and c‐jun.