Functional analysis of Asb-1 using genetic modification in mice.

The Asbs are a family of ankyrin repeat proteins that, along with four other protein families, contain a C-terminal SOCS box motif, which was first identified in the suppressor of cytokine signaling (SOCS) proteins. While it is clear that the SOCS proteins are involved in the negative regulation of cytokine signaling, the biological roles of the other SOCS box-containing families are unknown. We have investigated Asb-1 function by generating mice that lack this protein, as well as mice that overexpress full-length or truncated Asb-1 in a wide range of tissues.

Placental defects and embryonic lethality in mice lacking suppressor of cytokine signaling 3.

Mice lacking suppressor of cytokine signaling 3 (SOCS3) exhibited embryonic lethality with death occurring between days 11 and 13 of gestation. At this stage, SOCS3(-/-) embryos were slightly smaller than wild type but appeared otherwise normal, and histological analysis failed to detect any anatomical abnormalities responsible for the lethal phenotype. Rather, in all SOCS3(-/-) embryos examined, defects were evident in placental development that would account for their developmental arrest and death.

Identification of ligand-binding site III on the immunoglobulin-like domain of the granulocyte colony-stimulating factor receptor.

The granulocyte colony-stimulating factor receptor (G-CSF-R) forms a tetrameric complex with G-CSF containing two ligand and two receptor molecules. The N-terminal Ig-like domain of the G-CSF-R is required for receptor dimerization, but it is not known whether it binds G-CSF or interacts elsewhere in the complex. Alanine scanning mutagenesis was used to show that residues in the Ig-like domain of the G-CSF-R (Phe(75), Gln(87), and Gln(91)) interact with G-CSF.

Negative regulators of cytokine signaling.

The interaction of a cytokine with its specific cell surface receptor triggers the activation of intracellular signaling pathways that ultimately program the cellular response. Although the specific components and actions of the pathways driving these responses, such as the Janus kinase (JAK)/signal transducers and activators of transcription (STAT) pathway, are relatively well defined, it is becoming clear that important mechanisms exist to restrain these signaling cascades. This review discusses the key biochemical actions and biological roles of the phosphatase SHP-1, the protein inhibitors of activated STATs (PIAS) and the suppressor of cytokine signaling (SOCS) protein family in the negative regulation of cytokine signal transduction.

Cloning and characterization of the genes encoding the ankyrin repeat and SOCS box-containing proteins Asb-1, Asb-2, Asb-3 and Asb-4.

Members of the suppressor of cytokine signalling (SOCS) family of proteins have been shown to inhibit cytokine signalling via direct interactions with JAK kinases or activated cytokine receptors. In addition to their novel amino-terminal regions and SH2 domains that mediate these interactions, the SOCS proteins also contain carboxy-terminal regions of homology called the SOCS box. The SOCS box serves to couple SOCS proteins and their binding partners with the elongin B and C complex, possibly targeting them for degradation.

The suppressors of cytokine signaling (SOCS) proteins: important feedback inhibitors of cytokine action.

While positive effectors of cytokine signaling pathways are relatively well defined, negative regulation can be just as important but is poorly understood. The recently discovered suppressor of cytokine signaling (SOCS) family of proteins has been implicated in the negative regulation of several cytokine pathways, particularly the receptor-associated tyrosine kinase/signal transducer and activator of transcription (AK/STAT) pathways of transcriptional activation. Biochemical studies revealed that inhibition can occur via a variety of mechanisms.

The development of fatal myocarditis and polymyositis in mice heterozygous for IFN-gamma and lacking the SOCS-1 gene.

Mice lacking the gene encoding the suppressor of cytokine signaling-1 (SOCS-1 -/-) and heterozygous for the IFN-gamma gene (IFN-gamma +/-) avoided the IFN-gamma-dependent preweaning death of SOCS-1 -/- IFN-gamma +/+ mice but did not exhibit the good health of young adult SOCS-1 -/- IFN-gamma -/- mice. SOCS-1 -/- IFN-gamma +/- mice died within 160 days of birth with massive T lymphocyte, macrophage, and eosinophil infiltration of all skeletal muscles and a similar severe myocarditis. The cornea also developed inflammatory infiltration and often a corneal ulcer.

Gigantism in mice lacking suppressor of cytokine signalling-2.

Suppressor of cytokine signalling-2 (SOCS-2) is a member of the suppressor of cytokine signalling family, a group of related proteins implicated in the negative regulation of cytokine action through inhibition of the Janus kinase (JAK) signal transducers and activators of transcription (STAT) signal-transduction pathway. Here we use mice unable to express SOCS-2 to examine its function in vivo. SOCS-2(-/-) mice grew significantly larger than their wild-type littermates.

Suppressor of cytokine signaling-3 preferentially binds to the SHP-2-binding site on the shared cytokine receptor subunit gp130.

Suppressor of cytokine signaling-3 (SOCS-3) is one member of a family of intracellular inhibitors of signaling pathways initiated by cytokines that use, among others, the common receptor subunit gp130. The SH2 domain of SOCS-3 has been shown to be essential for this inhibitory activity, and we have used a quantitative binding analysis of SOCS-3 to synthetic phosphopeptides to map the potential sites of interaction of SOCS-3 with different components of the gp130 signaling pathway. The only high-affinity ligand found corresponded to the region of gp130 centered around phosphotyrosine-757 (pY757), previously shown to be a docking site for the tyrosine phosphatase SHP-2.

The residual megakaryocyte and platelet production in c-mpl-deficient mice is not dependent on the actions of interleukin-6, interleukin-11, or leukemia inhibitory factor.

Mice lacking thrombopoietin (TPO) or its receptor c-Mpl are severely thrombocytopenic, consistent with a dominant physiological role for this cytokine in megakaryocytopoiesis. However, these mice remain healthy and show no signs of spontaneous hemorrhage, implying that TPO-independent mechanisms for platelet production exist and are sufficient for hemostasis. To investigate the roles of cytokines that act through the gp130 signaling chain in the residual platelet production of mpl (-/-) mice, mpl (-/-)IL-6(-/-), mpl(-/-)LIF(-/-), and mpl(-/-)IL-11Ralpha(-/-) double-mutant mice were generated.

SCL expression in the mouse embryo detected with a targeted lacZ reporter gene demonstrates its localization to hematopoietic, vascular, and neural tissues.

The helix-loop-helix transcription factor SCL (TAL1) is indispensable for blood cell formation in the mouse embryo. We have explored the localization and developmental potential of cells fated to express SCL during murine development using SCL-lacZ mutant mice in which the Escherichia coli lacZ reporter gene was 'knocked in' to the SCL locus. In addition to the hematopoietic defect associated with SCL deficiency, the yolk sac blood vessels in SCL(lacZ/lacZ) embryos formed an abnormal primary vascular plexus, which failed to undergo subsequent remodeling and formation of large branching vessels.

Suppressors of cytokine signaling (SOCS): negative regulators of signal transduction.

SOCS-1 was originally identified as an inhibitor of interleukin-6 signal transduction and is a member of a family of proteins (SOCS-1 to SOCS-7 and CIS) that contain an SH2 domain and a conserved carboxyl-terminal SOCS box motif. Mutation studies have established that critical contributions from both the amino-terminal and SH2 domains are essential for SOCS-1 and SOCS-3 to inhibit cytokine signaling. Inhibition of cytokine-dependent activation of STAT3 occurred in cells expressing either SOCS-1 or SOCS-3, but unlike SOCS-1, SOCS-3 did not directly interact with or inhibit the activity of JAK kinases.

The oncostatin M signalling pathway: reversing the neoplastic phenotype?

Oncostatin M (OSM) is a member of the interleukin 6 (IL-6) family of cytokines and was originally identified by its ability to inhibit proliferation of melanoma cells but augment the growth of normal fibroblasts. OSM has pleiotropic effects on many different cell types, but here we focus on its ability to inhibit the proliferation of cell lines derived from several tumour types, including breast carcinoma, ovarian cancer, melanoma, glioma and lung carcinoma. The inhibition of proliferation of several cancer cell lines by OSM is associated with alterations in cellular morphology and with phenotypic changes that are consistent with the induction of differentiation of these cells.

A method for computational combinatorial peptide design of inhibitors of Ras protein.

A computational combinatorial approach is proposed for the design of a peptide inhibitor of Ras protein. The procedure involves three steps. First, a 'Multiple Copy Simultaneous Search' identifies the location of specific functional groups on the Ras surface.

Suckling defect in mice lacking the soluble haemopoietin receptor NR6.

Cytokines control a variety of cellular responses including proliferation, differentiation, survival and functional activation, via binding to specific receptors expressed on the surface of target cells [1]. The cytokine receptors of the haemopoietin family are defined by the presence of a conserved 200 amino acid extracellular domain known as the haemopoietin domain [2]. We report here the isolation of NR6, a haemopoietin receptor that, like the p40 subunit of interleukin-12 (IL-12) [3] and the EBI3 gene induced by Epstein-Barr virus infection in lymphocytes [4], contains a typical haemopoietin domain but lacks transmembrane and cytoplasmic domains.

Negative regulation of the JAK/STAT pathway.

Cytokines induce a variety of biological responses by binding to specific cell surface receptors and activating cytoplasmic signal transduction pathways, such as the JAK/STAT pathway. Although these responses are generally transient, few molecules have been characterised that switch the signal off. Several different steps of the signal transduction pathway appear to be targeted by negative regulators, including the receptor/ligand complex, JAK kinases, and STAT transcription factors.

The conserved SOCS box motif in suppressors of cytokine signaling binds to elongins B and C and may couple bound proteins to proteasomal degradation.

The suppressors of cytokine signaling (SOCS) family of proteins act as intracellular inhibitors of several cytokine signal transduction pathways. Their expression is induced by cytokine activation of the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway and they act as a negative feedback loop by subsequently inhibiting the JAK/STAT pathway either by direct interaction with activated JAKs or with the receptors. These interactions are mediated at least in part by the SH2 domain of SOCS proteins but these proteins also contain a highly conserved C-terminal homology domain termed the SOCS box.

Expression of BCR - ABL in M1 myeloid leukemia cells induces differentiation without arresting proliferation.

The mechanism leading to the expanding population of maturing myeloid cells which characterises chronic myeloid leukemia (CML) remains obscure. Because of its ability to mimic the proliferative and cell survival functions of hematopoietic growth factors, we hypothesized that the oncogene activated in CML, BCR-ABL, might also influence differentiation. To test this hypothesis, we examined the effects of expressing BCR-ABL on the myeloid differentiation of murine M1 leukemic cells, which cease dividing and differentiate into macrophages in the presence of the cytokines leukemia inhibitory factor (LIF) or interleukin (IL)-6.

Hemopoietic growth factor receptor abnormalities in leukemia.

Growth factor and cytokine control of hemopoiesis, the process of blood cell development, is mediated by specific interactions with cell-surface receptors. Hemopoietic growth factor receptors belong to two major families, the transmembrane protein tyrosine kinases and the hemopoietin receptors. Ligand binding stimulates receptor aggregation and activation resulting in transduction of signals that induce diverse cellular responses including proliferation, maturation, prevention of apoptosis and/or functional activation.

The box-1 region of the leukemia inhibitory factor receptor alpha-chain cytoplasmic domain is sufficient for hemopoietic cell proliferation and differentiation.

Leukemia inhibitory factor (LIF) is a pleiotropic cytokine that acts on a variety of cell types and regulates cell proliferation and differentiation. The functional receptor for LIF is composed of LIFR alpha-chain (LIFRalpha) and gp130 both of which are shared in the functional receptors for oncostatin M, ciliary neurotrophic factor, and cardiotrophin-1. By using stable transfection of wild-type or cytoplasmic deletion mutants of LIFRalpha together with full-length gp130 into Ba/F3 cells, we found that cells expressing gp130 and an extensively deleted mutant LIFRalpha containing only the box-1 region were capable of proliferating in response to LIF, although LIF-dependent long term growth of these cells was seriously impaired.

Functional analysis of mature hematopoietic cells from mice lacking the betac chain of the granulocyte-macrophage colony-stimulating factor receptor.

Mice with a null mutation of the betac chain of the granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5 receptors (betac-null mice) develop an alveolar proteinosis-like lung disease. The pathogenesis of this disease is uncertain and, although a defect in alveolar macrophage function has been postulated, no previous analysis of mature hematopoietic cells in mice with alveolar proteinosis has been reported. Therefore, we undertook a functional analysis of the mature hematopoietic cell compartment in betac-null mice.

Liver degeneration and lymphoid deficiencies in mice lacking suppressor of cytokine signaling-1.

SOCS-1, a member of the suppressor of cytokine signaling (SOCS) family, was identified in a genetic screen for inhibitors of interleukin 6 signal transduction. SOCS-1 transcription is induced by cytokines, and the protein binds and inhibits Janus kinases and reduces cytokine-stimulated tyrosine phosphorylation of signal transducers and activators of transcription 3 and the gp130 component of the interleukin 6 receptor. Thus, SOCS-1 forms part of a feedback loop that modulates signal transduction from cytokine receptors.

SOCS: suppressors of cytokine signalling.

Regulation of many aspects of cell behaviour occurs through the interaction of cytokines with specific cell surface receptors, resulting in the activation of cytoplasmic signal transduction pathways. Although cellular responses to cytokines are tightly controlled, few molecules have been identified which are able to switch these signals off. The suppressors of cytokine signalling (SOCS) proteins are a new family of negative regulators of cytokine signal transduction.