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A C Larner

Publications and source records attributed to A C Larner.

At least 37 records · Page 2Linked to original sources
PubMed1996

The SH2 domain-containing tyrosine phosphatase PTP1D is required for interferon alpha/beta-induced gene expression.

Interferons (IFNs) induce early response genes by stimulating Janus family (Jak) tyrosine kinases, leading to tyrosine phosphorylation of Stat (signal transducer and activator of transcription) proteins. Previous studies demonstrated that a protein-tyrosine phosphatase (PTP) is required for activation of the ISGF3 transcription complex by IFNalpha/beta, but the specific PTP responsible remained unidentified. We now show that the SH2 domain containing tyrosine phosphatase PTP1D (also designated as SHPTP2, SHPTP3, PTP2C, or Syp) is constitutively associated with the IFNalpha/beta receptor and becomes tyrosine-phosphorylated in response to ligand. Furthermore, transient expression of a phosphatase-inactive mutant or the COOH-terminal SH2 domain of PTP1D causes a dominant negative effect on IFNalpha/beta-induced early response gene expression. These results provide strong evidence that PTP1D functions as a positive regulator of the IFNalpha/beta-induced Jak/Stat signal transduction pathway.

Base Sequence↗
PubMed1996

STAT activation by epidermal growth factor (EGF) and amphiregulin. Requirement for the EGF receptor kinase but not for tyrosine phosphorylation sites or JAK1.

The epidermal growth factor (EGF) receptor activates several signaling cascades in response to the ligands EGF and amphiregulin (AR). One of these signaling events involves the tyrosine phosphorylation of STATs (signal transducers and activators of transcription), a process believed to require the activation of a tyrosine kinase of the JAK family. In this report we demonstrate that EGF- and AR-induced STAT activation requires the intrinsic kinase activity of the receptor but not the presence of Jak1. We show that both wild type (WT) and truncated EGF receptors lacking all autophosphorylation sites activate STAT 1, 3, and 5 in response to either EGF or AR. Furthermore, relative to cells expressing WT receptor, ligand-induced tyrosine phosphorylation of the STATs was enhanced in cells expressing only the truncated receptor. These results provide the first evidence that (i) EGF receptor-mediated STAT activation occurs in a Jak1-independent manner, (ii) the intrinsic tyrosine kinase activity of the receptor is essential for STAT activation, and (iii) tyrosine phosphorylation sites within the EGF receptor are not required for STAT activation.

Amphiregulin↗
PubMed1996

Prolactin recruits STAT1, STAT3 and STAT5 independent of conserved receptor tyrosines TYR402, TYR479, TYR515 and TYR580.

The present study of prolactin (PRL) receptor-mediated recruitment of signal transducers and activators of transcription (STATs) demonstrates that PRL activates STAT3, in addition to STAT1 and STAT5 as previously reported, and that STAT1, STAT3 and STAT5 are mediators of PRL effects in cells whether of lymphoid, myeloid or mammary epithelial origin. Furthermore, receptor mutants M240 and T280 that do not mediate PRL-induced JAK2 activation and cell proliferation, are also unable to mediate STAT activation, supporting the proposed model of JAK2 as the initial effector protein used by PRL receptors. On the other hand, tyrosine phosphorylation analysis and electrophoretic mobility shift assays showed that receptor mutant G328, which lacks four of the five conserved cytoplasmic tyrosine residues of PRL receptors, retained the ability to activate JAK2 and STAT1, STAT3 and STAT5. These results support the notion that phosphotyrosyl residues other than those of the receptor, i.e., JAK2, are involved in recruiting STAT proteins to the activated PRL receptor complex.

Animals↗
PubMed1996

Activation of protein kinase A inhibits interferon induction of the Jak/Stat pathway in U266 cells.

Activation of early response genes by interferons (IFNs) requires tyrosine phosphorylation of the Stat transcription factors and is mediated by the Jak family of tyrosine kinases. Recent evidence suggests that ERK2 serine/threonine kinase modulates the IFN-stimulated Jak/Stat pathway. In this report we show that in the myeloma cell line U266 protein kinase A specifically interacts with the cytoplasmic domain of the IFNalpha/beta receptor. Treatment of cells with the adenylate cyclase activator forskolin inhibits IFNbeta-, IFNgamma-, and hydrogen peroxide/vanadate-induced formation of complexes that bind to enhancers known to stimulate the expression of IFN-regulated genes. Immunoprecipitations followed by anti-phosphotyrosine immunoblots indicate that tyrosine phosphorylation of the alpha chain of the IFNalpha/beta receptor, Jak1, Tyk2, as well as Stat1 and Stat2 is reduced as a consequence of incubation of cells with forskolin. In contrast, dideoxyforskolin, which fails to activate adenylate cyclase, has no effect on IFN induction of the Jak/Stat pathway. These results indicate a novel regulatory mechanism by which protein kinase A can modulate the Jak/Stat signaling cascade.

Adenylyl Cyclases↗
PubMed1996

Preassociation of STAT1 with STAT2 and STAT3 in separate signalling complexes prior to cytokine stimulation.

A variety of cytokines and growth factors act through an induction of gene expression mediated by a family of latent transcription factors called STAT (signal transducers and activators of transcription) proteins. Ligand-induced tyrosine phosphorylation of the STATs promotes their homodimer and heterodimer formation and subsequent nuclear translocation. We demonstrate here that STAT protein heterocomplexes exist prior to cytokine treatment. When unstimulated HeLa cells are ruptured in hypotonic buffer without salt or detergent, immunoadsorption of either STAT1 or STAT2 from the resulting cytosol yields coimmunoadsorption of the other STAT protein. Similarly, STAT1-STAT3 heterocomplexes are coimmunoadsorbed from hypotonic cytosol. STAT1 and STAT2 or STAT1 and STAT3 translated in reticulocyte lysate spontaneously form heterocomplexes when the translation lysates are mixed at 0 degrees C. Our data suggest that interferon-alpha /beta-induced tyrosine phosphorylation increases the stability of a preexisting, latent, STAT1-STAT2 signaling complex. Newly translated STAT1 binds in equilibrium fashion to STAT2 and STAT3, but we show that STAT2 and STAT3 exist in separate heterocomplexes with STAT1, consistent with a model in which STAT1 contains a common binding site for other STAT proteins.

3T3 Cells↗
PubMed1996

Inhibition of alpha interferon but not gamma interferon signal transduction by phorbol esters is mediated by a tyrosine phosphatase.

Previous studies have indicated that the expression of viral oncoproteins, cell transformation, or phorbol ester treatment of cells can inhibit alpha/beta interferon (IFN-alpha/beta)-induced gene expression. The mechanisms by which these promoters of cell growth exert their inhibitory effects vary, but in most instances they involve a disruption of the IFN-alpha/beta-induced transcription complex ISGF3 such that the DNA-binding component of this complex (the 48-kDa ISGF3gamma protein) does not bind to the interferon-stimulated response element (ISRE). In this report, we demonstrated that phorbol ester treatment of human peripheral blood monocytes dramatically inhibits activation of IFN-alpha/B-stimulated early response genes but by a mechanism which does not involve abrogation of the ISRE binding of ISGF3gamma. Phorbol ester treatment of monocytes inhibited IFN alpha-stimulated tyrosine phosphorylation of the transcription factors Stat1alpha, Stat2, and Stat3 and of the tyrosine kinase Tyk2 but had no effect on IFN-gamma activation of Stat1alpha. IFNalpha-stimulated tyrosine phosphorylation of Jak1 and the alpha subunit of the IFN-alpha receptor were unaffected by phorbol 12-myristate 13-acetate (PMA). Moreover, PMA caused the dephosphorylation of Tyk2 but not of Jak1, which was activated by IFN. Pretreatment of cells with vanadate prevented the effects of PMA with regard to PMA-induced Tyk2 dephosphorylation. These observations suggest that PMA exerts its inhibitory effects by activation of a tyrosine phosphatase which selectively regulates Tyk2 but not Jak1 activity.

Base Sequence↗
PubMed1995

Requirement for MAP kinase (ERK2) activity in interferon alpha- and interferon beta-stimulated gene expression through STAT proteins.

Activation of early response genes by interferons (IFNs) requires tyrosine phosphorylation of STAT (signal transducers and activators of transcription) proteins. It was found that the serine-threonine kinase mitogen-activated protein kinase (MAPK) [specifically, the 42-kilodalton MAPK or extracellular signal-regulated kinase 2 (ERK2)] interacted with the alpha subunit of IFN-alpha/beta receptor in vitro and in vivo. Treatment of cells with IFN-beta induced tyrosine phosphorylation and activation of MAPK and caused MAPK and Stat1 alpha to coimmunoprecipitate. Furthermore, expression of dominant negative MAPK inhibited IFN-beta-induced transcription. Therefore, MAPK appears to regulate IFN-alpha and IFN-beta activation of early response genes by modifying the Jak-STAT signaling cascade.

Binding Sites↗
PubMed1995

Mapping of the cytoplasmic domain of the human growth hormone receptor required for the activation of Jak2 and Stat proteins.

Incubation of cells with growth hormone (GH) stimulates both tyrosine phosphorylation of the Jak2 tyrosine kinase and, in some cells, the transcription factor Stat1 alpha (1-4). When the promyeloid cell line FDC-P1 is transfected with the human growth hormone receptor, these cells can grow in the presence of GH and in the absence of interleukin-3. Growth hormone treatment of cells expressing the human growth hormone receptor did not activate Stat1 alpha. However, a complex is present in extracts prepared from growth hormone-treated cells that binds to the gamma response region, an enhancer present in the promoter of the high affinity Fc gamma R1 receptor to which cytokine-activated Stat complexes bind. When truncations of the cytoplasmic domain of the receptor are expressed in FDC-P1 cells only the membrane-proximal 80 amino acids (containing box 1 and box 2) are required for activation of both a GH-stimulated binding activity (GHSF) and tyrosine phosphorylation of Jak2. Activation of GHSF can be inhibited in a cell-free system by the addition of a glutathione S-transferase fusion protein containing these 80 amino acids. Replacement of the one tyrosine in this region of the receptor with a phenylalanine does not alter the activation of either GHSF or Jak2, suggesting that tyrosine phosphorylation of the receptor is not required for GH activation of GHSF. Moreover, a cell line expressing a receptor with only the 54 membrane-proximal amino acids of the intracellular domain (including box 1) shows constitutively tyrosine-phosphorylated Jak2 as well as GHSF binding. With this truncated receptor, there is little if any additional GH-induced tyrosine phosphorylation of Jak2 or induced binding to the gamma response region. These results define the importance of the membrane-proximal 80 amino acids of the GH receptor (with the conserved box 1 and box 2 domains) with regard to GH activation of both Jak2 and Stat(s). They also suggest that within these domains there may be positive and negative elements that regulate Jak2 function.

Base Sequence↗
PubMed1995

Interleukin 12 induces tyrosine phosphorylation and activation of STAT4 in human lymphocytes.

Interleukin 12 (IL-12) is an important immunoregulatory cytokine whose receptor is a member of the hematopoietin receptor superfamily. We have recently demonstrated that stimulation of human T and natural killer cells with IL-12 induces tyrosine phosphorylation of the Janus family tyrosine kinase JAK2 and Tyk2, implicating these kinases in the immediate biochemical response to IL-12. Recently, transcription factors known as STATs (signal transducers and activators of transcription) have been shown to be tyrosine phosphorylated and activated in response to a number of cytokines that bind hematopoietin receptors and activate JAK kinases. In this report we demonstrate that IL-12 induces tyrosine phosphorylation of a recently identified STAT family member, STAT4, and show that STAT4 expression is regulated by T-cell activation. Furthermore, we show that IL-12 stimulates formation of a DNA-binding complex that recognizes a DNA sequence previously shown to bind STAT proteins and that this complex contains STAT4. These data, and the recent demonstration of JAK phosphorylation by IL-12, identify a rapid signal-transduction pathway likely to mediate IL-12-induced gene expression.

Base Sequence↗
PubMed1995

The Jak kinases differentially associate with the alpha and beta (accessory factor) chains of the interferon gamma receptor to form a functional receptor unit capable of activating STAT transcription factors.

Interferon gamma (IFN gamma) induces the expression of early response genes by tyrosine phosphorylation of Jak kinases and transcription factors referred to as STAT proteins. The topology of the IFN gamma receptor is partially understood and the relationship between the alpha chain that binds the ligand and the beta chain that is required for signal transduction is undefined. In a cell line which expresses only the human alpha chain, we show that these cells did not activate Jak kinases or STAT proteins with human IFN gamma, even though Jak1 co-immunoprecipitated with the alpha chain. In cells unexposed to IFN gamma, Jak1 preferentially associated with the alpha chain, while Jak2 associated with the beta chain. There was evidence for Jak1 kinase activity in untreated cells. For Jak2, kinase activity was IFN gamma-dependent. Although the alpha chain was tyrosine-phosphorylated in response to ligand, we found no evidence for tyrosine phosphorylation of the beta chain. These data are consistent with a model of the IFN gamma receptor in which Jak1 associates with the alpha chain, whereas Jak2 associates with the beta chain. IFN gamma clusters at least two receptor units which results in the tyrosine phosphorylation of Jak1 and Jak2, the activation of Jak2 kinase activity, and the recruitment of STAT1 alpha resulting in its activation by tyrosine phosphorylation.

Base Sequence↗
PubMed1995

Characterization of a Stat-like DNA binding activity in Drosophila melanogaster.

The cytokine signaling pathways that activate the Janus family of tyrosine kinases (Jaks) and the "signal transducers and activators of transcription" (Stats) have been well characterized in mammalian systems. Work shown here provides evidence that an analogous signaling pathway exists in Drosophila melanogaster. Because many of the ligand-receptor pairs in Drosophila have not been fully characterized, it was necessary to bypass the receptor stimulation event that normally triggers intracellular Jak/Stat activation. This was done by treating Drosophila Schneider 2 cells with vanadate/peroxide, which has been shown to closely mimic some signaling events triggered by interferon gamma, including the activation of Jak1, Jak2, and the Stat1 alpha protein. Evidence presented here demonstrates that vanadate/peroxide can induce a gamma response region binding complex in Drosophila Schneider 2 cells. This complex contains two phosphoproteins of 100 and 150 kDa, respectively, and shares many features with the vanadate/peroxide-stimulated binding complex in the mammalian system. Southern blot analysis of genomic DNA using the src homology domain 2 (SH2) of Stat1 alpha confirms the presence of a related gene in the Drosophila genome.

Animals↗
PubMed1995

Enhanced DNA-binding activity of a Stat3-related protein in cells transformed by the Src oncoprotein.

Cytokines and growth factors induce tyrosine phosphorylation of signal transducers and activators of transcription (STATs) that directly activate gene expression. Cells stably transformed by the Src oncogene tyrosine kinase were examined for STAT protein activation. Assays of electrophoretic mobility, DNA-binding specificity, and antigenicity indicated that Stat3 or a closely related STAT family member was constitutively activated by the Src oncoprotein. Induction of this DNA-binding activity was accompanied by tyrosine phosphorylation of Stat3 and correlated with Src transformation. These findings demonstrate that Src can activate STAT signaling pathways and raise the possibility that Stat3 contributes to oncogenesis by Src.

Animals↗
PubMed1995

Protein tyrosine phosphorylation as a mechanism which regulates cytokine activation of early response genes.

Two well-defined rapid responses which occur as a consequence of growth factors binding to their cell surface receptors involve tyrosine phosphorylation of cellular proteins and the induction of the transcription of cellular genes. Recent advances have been made in purification and cloning of Src homology 2 and 3 (SH2/SH3) domain-containing transcription factors which are required for the activation of early response genes by interferons. These transcription factors are covalently modified by tyrosine phosphorylation such that they interact with enhancers needed for interferon-stimulated gene expression. The Jak family of tyrosine kinases are also an integral component in these signalling cascades. The information gained concerning interferon signalling has now been extended to include a broad network of cytokine-regulated signalling systems which use tyrosine phosphorylation of a family of structurally related proteins to activate transcription of early response genes.

Base Sequence↗
PubMed1995

Differential regulation of the alpha/beta interferon-stimulated Jak/Stat pathway by the SH2 domain-containing tyrosine phosphatase SHPTP1.

Interferons (IFNs) induce early-response genes by stimulating Janus family (Jak) tyrosine kinases, leading to tyrosine phosphorylation of Stat transcription factors. Previous studies implicated protein-tyrosine phosphatase (PTP) activity in the control of IFN-regulated Jak/Stat signaling, but the specific PTPs responsible remained unidentified. We have found that SH2 domain-containing PTP1 (SHPTP1; also called PTP1C, HCP, or SHP) reversibly associates with the IFN-alpha receptor complex upon IFN addition. Compared with macrophages from normal littermate controls, macrophages from motheaten mice, which lack SHPTP1, show dramatically increased Jak1 and Stat1 alpha tyrosine phosphorylation, whereas Tyk2 and Stat2 activation is largely unaffected. These findings correlate with selectively increased complex formation on a gamma response element, but not an IFN-stimulated response element, in motheaten macrophages. Our results establish that SHPTP1 selectively regulates distinct components of Jak/Stat signal transduction pathways in vivo.

Animals↗
PubMed1994

Prolactin activates the interferon-regulated p91 transcription factor and the Jak2 kinase by tyrosine phosphorylation.

The prolactin (PRL) receptor is a member of the family of cytokine receptors that lack intrinsic tyrosine kinase activity but contain two conserved cysteines in their N-terminal regions and a WSXWS motif adjacent to their transmembrane domains. In a manner similar to the interferons (IFNs), exposure of cells to PRL results in tyrosine phosphorylation of several cellular proteins and the rapid transcriptional induction of the IFN regulatory factor 1 gene. In this communication, we demonstrate that treatment of rat Nb2 lymphoma cells with PRL activates a latent protein factor so that it binds to an enhancer in the IFN regulatory factor 1 gene. This enhancer has been shown to be required for IFN-gamma-activated expression of this gene. PRL-induced assembly of the DNA binding complex, PRL-stimulated factor, required tyrosine phosphorylation. PRL-stimulated factor contained at least one protein that was antigenically similar to the p91 transcription factor, a component of several transcription complexes required for cytokine-activated gene expression. PRL not only induced the tyrosine phosphorylation of p91 but also induced tyrosine phosphorylation of Jak2, a tyrosine kinase required for IFN-gamma-activated gene expression. These results provide evidence for a signaling mechanism, some of whose components are shared by both PRL and IFN-gamma receptors, that results in the expression of early response genes.

Animals↗
PubMed1994

Interferon-gamma induces tyrosine phosphorylation of interferon-gamma receptor and regulated association of protein tyrosine kinases, Jak1 and Jak2, with its receptor.

Interferon-gamma (IFN-gamma) induces the expression of a set of early response genes by tyrosine phosphorylation of latent transcription factors such as p91. Although the tyrosine kinases, Jak1 and Jak2, have recently been shown to be critical for signal transduction by IFN-gamma, evidence is lacking for both tyrosine phosphorylation of the IFN-gamma receptor (IFN-gamma R) and the interaction between Jak1, Jak2, and the IFN-gamma R. In this report, we show that binding of IFN-gamma to HeLa cells initiated a series of events that resulted in the extremely rapid (15 s) tyrosine phosphorylation of not only Jak1, Jak2, and p91 but also the IFN-gamma R. Coimmunoprecipitation experiments revealed that Jak1 was associated with the IFN-gamma R prior to ligand binding, whereas Jak2 became part of the IFN-gamma R-Jak1 complex immediately after ligand binding. H2O2/vanadate treatment of cells for 15 min resulted in only the tyrosine phosphorylation of Jak1 and IFN-gamma R. Only after 60 min of this treatment did we observe tyrosine phosphorylation of Jak2 and p91 and assembly of the transcription factor complex FcRF gamma that binds to the promoter of the fcgr1 gene. These data suggest that JAK1 associates with the IFN-gamma R prior to ligand binding. IFN-gamma treatment of cells results in recruitment of JAK2 into the IFN-gamma R-Jak1 complex followed by assembly of the transcription factor FcRF gamma complex.

Electrophoresis, Polyacrylamide Gel↗