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F Gouilleux

Publications and source records attributed to F Gouilleux.

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

IL-10 induces DNA binding activity of three STAT proteins (Stat1, Stat3, and Stat5) and their distinct combinatorial assembly in the promoters of selected genes.

Interaction of IL-10 with its receptor leads to the activation of STAT transcription factors. Herein we report the IL-10 dependent simultaneous activation of three STAT transcription factors: Stat1, Stat3, and Stat5. Upon IL-10 treatment multiple Stat proteins become simultaneously activated, and bind to different promoters with equal kinetics but form distinct homo- and heterodimeric transcriptionally active complexes depending on the STAT-consensus elements of a selected gene promoter. Upon IL-10 treatment Stat1, 3, and 5 bind to the GRR of the FcgammaRI gene, activated Stat1 and 3 bind to the SIE sequence of the c-fos promoter and transcriptionally active Stat5 assembles at the PRL-STAT consensus sequence of the beta-casein gene. Thus, functionally relevant STAT dimerization is influenced by the activated cytokine receptor as well as the specific STAT consensus sequence present in a specific gene promoter.

B-Lymphocytes↗
PubMed1996

Identification of tyrosine residues within the intracellular domain of the erythropoietin receptor crucial for STAT5 activation.

FDCP-1 cells are hematopoietic progenitor cells which require interleukin-3 for survival and proliferation. FDCP-1 cells stably transfected with the murine erythropoietin receptor cDNA survive and proliferate in the presence of erythropoietin. Erythropoietin induces the activation of the short forms (80 kDa) of STAT5 in the cells. Erythropoietin-induced activation of STAT5 was strongly reduced in cells expressing mutated variants of the erythropoietin receptors in which tyrosine residues in their intracellular domain have been eliminated. We determined that the erythropoietin receptor tyrosine residues 343 and 401 are independently necessary for STAT5 activation. The amino acid sequences surrounding these two tyrosine residues are very similar. Peptides comprising either phosphorylated Tyr343 or phosphorylated Tyr401, but not their unphosphorylated counterparts, inhibited the STAT5 activation. We propose that these two tyrosine residues of the erythropoietin receptor constitute docking sites for the STAT5 SH2 domain. The growth stimulus mediated by erythropoietin was decreased in cells expressing erythropoietin receptors lacking both Tyr343 and Tyr401. This suggests that STAT5 activation could be involved in the growth control of FDCP-1 cells.

Amino Acid Sequence↗
PubMed1996

Glucocorticoid and progestin receptors are differently involved in the cooperation with a structural element of the mouse mammary tumor virus promoter.

We have previously characterized a regulatory element located between -294 and -200 within the mouse mammary tumor virus (MMTV) long terminal repeat (LTR). This element termed AA element cooperates with the glucocorticoid response elements (GREs) for glucocorticoid activation. Here we show that in a MMTV LTR wild type context, the deletion of this element significantly reduces both glucocorticoid and progestin activation of the promoter. Deletion of the two most distal GREs forces the glucocorticoid receptor (GR) and the progestin receptor (PR) to bind the same response elements and results in a dramatic decrease in the inducibility of the MMTV promoter by the two hormones. The simultaneous deletion of the two distal GREs and of the AA element abolishes completely the glucocorticoid-induced activation of the promoter. In contrast it restores a significant level of progestin-induced activation. This different effect of the double deletion on glucocorticoid- and progestin-induced MMTV promoter activation is not cell specific because it is also observed, and is even stronger, when either GR or PR is expressed in the same cell line (NIH 3T3). This is the first description of a mutated MMTV promoter that, although retaining GREs, is activated by progestins and not by glucocorticoids. This suggests a different functional cooperation between protein(s) interacting with the AA element and GR or PR. Cotransfections with constructs containing wild-type or mutated MMTV LTR with either PR lacking its C-terminal domain or GR/PR chimeras in which the N-terminal domains have been exchanged demonstrate that the N-terminal domains of the receptors specify the different behavior of GR and PR regarding the AA element.

3T3 Cells↗
PubMed1996

STAT-related transcription factors are constitutively activated in peripheral blood cells from acute leukemia patients.

A signal transduction pathway activated by many cytokines has recently been elaborated. The JAK kinases and the signal transducers and activators of transcription (STAT) factors have been found to be essential components. In this report, we describe the presence of constitutively activated STAT factors in peripheral blood cells from patients with acute leukemia. We used oligonucleotide probes from the beta-casein and IRF-1 gene promoters and the ISRE probe to detect STAT proteins in nuclear extracts from acute leukemia cells in bandshift assays. Specific DNA protein complex formation was observed with the probes from the beta-casein and IRF-1 gene promoters, but not with the ISRE oligonucleotide probe, when cell extracts from acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) were investigated. We used nonradioactive oligonucleotides as competitors to show the specificity of the complex formation. Specific antibodies directed against the individual STAT proteins were used in supershift experiments. STAT5- and STAT1-related factors were detected in ALL and STAT1-, STAT3-, and STAT5-related proteins were present in nuclear cell extracts from AML. Since the cells were not treated with cytokines before the nuclear proteins were extracted, we conclude that these factors are constitutively activated in vivo. It is likely that the constitutive activation of STAT proteins is a part of the events of leukemogenesis.

Acute Disease↗
PubMed1996

Deletion of the carboxyl-terminal transactivation domain of MGF-Stat5 results in sustained DNA binding and a dominant negative phenotype.

The Stat (signal transducer and activator of transcription) factors transmit cytokine, growth factor, and hormone responses. Seven members of the Stat gene family are known. MGF-Stat5a has been discovered as a mediator of the prolactin response in mammary epithelial cells. Two closely related variants of Stat5, Stat5a and Stat5b, are encoded by distinct genes. We examined the functional properties of the carboxyl termini of these molecules. Wild-type Stat5a (794 amino acids) and the carboxyl-terminal deletion mutant Stat5a delta 772 supported prolactin-induced transcription of a beta-casein promoter-reporter construct in COS7 cells; Stat5a delta 750 did not. Upon prolactin activation, tyrosine phosphorylation and the specificity of DNA binding were indistinguishable among the three Stat5a variants. Tyrosine dephosphorylation and the downregulation of the DNA-binding activity were delayed in the Stat5a delta 750 mutant. The carboxyl-terminal transactivation domain of Stat5a, amino acids 722 to 794, can be conferred to the DNA-binding domain of the yeast transcription factor GAL4. Coexpression of Stat5a or Stat5b and of the carboxyl-terminal deletion mutants resulted in the suppression of transcriptional induction in COS or Ba/F3 cells. We propose that Stat5a delta 750 and Stat5b delta 754 are lacking functional transactivation domains and exert their dominant negative effects by blocking the DNA-binding site in Stat5-responsive gene promoters.

Amino Acid Sequence↗
PubMed1996

Identification of a growth hormone-responsive STAT5-binding element in the rat insulin 1 gene.

GH and PRL stimulate both proliferation and insulin production in pancreatic beta-cells as well as in the rat insulinoma cell line RIN-5AH, We report here that human GH increases insulin mRNA levels in RIN-5AH cells via both somatogenic and lactogenic receptors. GH stimulated the rat insulin 1 promoter activity 2-fold, and this stimulation was abolished by introduction of a block mutation in a gamma-interferon-activated sequence (GAS)-like element (GLE) with the sequence 5'-TTCTGGGAA-3' located in the rat insulin 1 enhancer at position -330 to -322. This element, termed Ins-GLE, was able to confer GH responsiveness to a heterologous promoter. GH induced the binding of two protein complexes to the Ins-GLE. An antibody directed against the transcription factor STAT5 (signal transducer and activator of transcription) supershifted the GH-induced complexes. Furthermore, in COS7 cells transiently transfected with STAT5 and GH receptor cDNAs, it was found that expression of STAT5 was necessary for GH induction of these two DNA-binding complexes. These results suggest that GH stimulates insulin 1 promoter activity by inducing the binding of STAT5 to Ins-GLE.

Animals↗
PubMed1996

Differential activation of Stat3 and Stat5 by distinct regions of the growth hormone receptor.

The GH receptor (GHR) is a member of the cytokine receptor superfamily; its signaling involves the activation of Janus tyrosine kinases (JAK2) and Stat (signal transducers and activators of transcription) transcription factors. Using truncated and tyrosine mutants of the receptor, we show that different receptor domains are essential for the activation of Stat3 and Stat5. GH-dependent phosphorylation of JAK2, Stat3, and Stat5, as well as transactivation studies with reporter genes containing Stat3 and Stat5 DNA-binding elements, was performed in cells expressing the various GHR mutants. The membrane-proximal region of the receptor necessary for JAK2 activation is sufficient for Stat3 activation. In contrast, C-terminal tyrosine residues of GHR are absolutely required for Stat5 activation. The same residues are also involved in the regulation of JAK2 dephosphorylation, possibly through the activation of a phosphatase. Using in vitro experiments with glutathione-S-transferase fusion proteins, we demonstrate that the SH2 domain of Stat5 binds to the carboxy-terminal tyrosine-phosphorylated residues of GHR. Our results show that a cytokine receptor can mediate differently the activation of distinct Stat proteins that could be involved in cytokine-specific effects.

Animals↗
PubMed1995

Cloning and expression of Stat5 and an additional homologue (Stat5b) involved in prolactin signal transduction in mouse mammary tissue.

Prolactin (PRL) induces transcriptional activation of milk protein genes, such as the whey acidic protein (WAP), beta-casein, and beta-lactoglobulin genes, through a signaling cascade encompassing the Janus kinase Jak2 and the mammary gland factor (MGF; also called Stat5), which belongs to the family of proteins of signal transducers and activators of transcription (STAT). We isolated and sequenced from mouse mammary tissue Stat5 mRNA and a previously unreported member, which we named Stat5b (Stat5 is renamed to Stat5a). On the protein level Stat5a and Stat5b show a 96% sequence similarity. The 5' and 3' untranslated regions of the two mRNAs are not conserved. Stat5a comprises 793 amino acids and is encoded by a mRNA of 4.2 kb. The Stat5b mRNA has a size of 5.6 kb and encodes a protein of 786 amino acids. Both Stat5a and Stat5b recognized the GAS site (gamma-interferon-activating sequence; TTCNNNGAA) in vitro and mediated PRL-induced transcription in COS cells transfected with a PRL receptor. Stat5b also induced basal transcription in the absence of PRL. Similar levels of Stat5a and Stat5b mRNAs were found in most tissues of virgin and lactating mice, but a differential accumulation of the Stat5 mRNAs was found in muscle and mammary tissue. The two RNAs are present in mammary tissue of immature virgin mice, and their levels increase up to day 16 of pregnancy, followed by a decline during lactation. The increase of Stat5 expression during pregnancy coincides with the activation of the WAP gene.

Amino Acid Sequence↗
PubMed1995

Signaling through the interleukin 2 receptor beta chain activates a STAT-5-like DNA-binding activity.

To explore the possible involvement of STAT factors ("signal transducers and activators of transcription") in the interleukin 2 receptor (IL-2R) signaling cascade, murine HT-2 cells expressing chimeric receptors composed of the extracellular domain of the erythropoietin receptor fused to the cytoplasmic domains of the IL-2R beta or -gamma c chains were prepared. Erythropoietin or IL-2 activation of these cells resulted in rapid nuclear expression of a DNA-binding activity that reacted with select STAT response elements. Based on reactivity with specific anti-STAT antibodies, this DNA-binding activity was identified as a murine homologue of STAT-5. Induction of nuclear expression of this STAT-5-like factor was blocked by the addition of herbimycin A, a tyrosine kinase inhibitor, but not by rapamycin, an immunophilin-binding antagonist of IL-2-induced proliferation. The IL-2R beta chain appeared critical for IL-2-induced activation of STAT-5, since a mutant beta chain lacking all cytoplasmic tyrosine residues was incapable of inducing this DNA binding. In contrast, a gamma c mutant lacking all of its cytoplasmic tyrosine residues proved fully competent for the induction of STAT-5. Physical binding of STAT-5 to functionally important tyrosine residues within IL-2R beta was supported by the finding that phosphorylated, but not nonphosphorylated, peptides corresponding to sequences spanning Y392 and Y510 of the IL-2R beta tail specifically inhibited STAT-5 DNA binding.

Amino Acid Sequence↗
PubMed1995

Interleukin-3, erythropoietin, and prolactin activate a STAT5-like factor in lymphoid cells.

Interleukin-3 (IL-3)-, erythropoietin (EPO)-, and prolactin (PRL)-induced signal transduction via the JAK/STAT pathway was studied in the IL-3-dependent BAF-3 lymphoid cell line. Transfected cells expressing either the long form of the PRL receptor or the EPO receptor were used. We demonstrated that IL-3, EPO, and PRL activated a transcription factor related to the mammary transcription factor STAT5 but not to STAT1, -2, -3, or -4 as opposed to interferon gamma (IFN gamma) which activated STAT1 in the same cells. Similarly, PRL and EPO activated a STAT5-like factor (STAT5-L) in the rat Nb2 and the human UT7 cells expressing endogenous PRL and EPO receptors, respectively. The hematopoietic STAT5-L activated by IL-3, EPO, or PRL was identified as a 97-kDa tyrosine-phosphorylated protein. These results confer to STAT5 a much broader role than previously suggested.

Animals↗
PubMed1995

Thrombopoietin activates a STAT5-like factor in hematopoietic cells.

Thrombopoietin (TPO) is a newly cloned cytokine which is the major regulator of circulating platelet levels, acting on both proliferation and differentiation of megakaryocytes. We have investigated the ability of TPO to activate the JAK/STAT pathway in megakaryocytic cell lines. We used either the granulocyte-macrophage colony-stimulating factor (GM-CSF)- and/or erythropoietin (EPO)-dependent UT7 cell line in which the murine TPO receptor (mumpl) had been transfected (mumpl-UT7 transfectants) or the MO7E and DAMI cells which express endogenous human TPO receptors. We demonstrated that TPO activates the kinase JAK2 and a STAT5-like transcriptional factor but not STAT1, STAT2, STAT3 or STAT4, in a very rapid and transient manner. In order to better ascertain the specificity of the activation of STAT5-related factor by TPO, we investigated the effect of other cytokines/growth factors. Both GM-CSF and EPO activated the STAT5-like factor. In contrast, neither interferon (IFN)-gamma nor the mitogenic stem cell factor (SCF) activated STAT5, although IFN-gamma did activate STAT1 in those cells. The hematopoietic DNA binding activity related to STAT5 was identified as a p97 tyrosine-phosphorylated protein band which exhibited identical gel mobility to the mammary STAT5. Because v-mpl, a truncated form of the TPO receptor c-mpl, was shown to be oncogenic, we tested the activity of v-mpl on STAT5 and found STAT5 constitutively activated in two different v-mpl-expressing cells, the transiently transfected Cos7 cells and the stable v-mpl-UT7 transfectants. Overall, our data indicate that STAT5 is widely expressed in hematopoietic cells and activated by a number of cytokines, including TPO, GM-CSF and EPO, but not by IFN-gamma or SCF.

Animals↗
PubMed1995

Activation of Stat5 by interleukin 2 requires a carboxyl-terminal region of the interleukin 2 receptor beta chain but is not essential for the proliferative signal transmission.

The high-affinity interleukin 2 (IL-2) receptor (IL-2R) consists of three subunits: the IL-2R alpha, IL-2R beta c, and IL-2R gamma c chains. Two members of the Janus kinase family, Jak1 and Jak3, are associated with IL-2R beta c and IL-2R gamma c, respectively, and they are activated upon IL-2 stimulation. The cytokine-mediated Jak kinase activation usually results in the activation of a family of latent transcription factors termed Stat (signal transducer and activator of transcription) proteins. Recently, the IL-2-induced Stat protein was purified from human lymphocytes and found to be the homologue of sheep Stat5/mammary gland factor. We demonstrate that the human Stat5 is activated by IL-2 and that Jak3 is required for the efficient activation. The cytoplasmic region of the IL-2R beta c chain required for activation of Stat5 is mapped within the carboxyl-terminal 147 amino acids. On the other hand, this region is not essential for IL-2-induced cell proliferation.

3T3 Cells↗
PubMed1995

Prolactin, growth hormone, erythropoietin and granulocyte-macrophage colony stimulating factor induce MGF-Stat5 DNA binding activity.

The molecular components which mediate cytokine signaling from the cell membrane to the nucleus were studied. Upon the interaction of cytokines with their receptors, members of the janus kinase (Jak) family of cytoplasmic protein tyrosine kinases and of the signal transducers and activators of transcription (Stat) family of transcription factors are activated through tyrosine phosphorylation. It has been suggested that the Stat proteins are substrates of the Jak protein tyrosine kinases. MGF-Stat5 is a member of the Stat family which has been found to confer the prolactin response. MGF-Stat5 can be phosphorylated and activated in its DNA binding activity by Jak2. The activation of MGF-Stat5 is not restricted to prolactin. Erythropoietin (EPO) and growth hormone (GH) stimulate the DNA binding activity of MGF-Stat5 in COS cells transfected with vectors encoding EPO receptor and MGF-Stat5 or vectors encoding GH receptor and MGF-Stat5. The activation of DNA binding by prolactin, EPO and GH requires the phosphorylation of tyrosine residue 694 of MGF-Stat5. The transcriptional induction of a beta-casein promoter luciferase construct in transiently transfected COS cells is specific for the prolactin activation of MGF-Stat5; it is not observed in EPO- and GH-treated cells. In the UT7 human hematopoietic cell line, EPO and granulocyte-macrophage colony stimulating factor activate the DNA binding activity of a factor closely related to MGF-Stat5 with respect to its immunological reactivity, DNA binding specificity and molecular weight. These results suggest that MGF-Stat5 regulates physiological processes in mammary epithelial cells, as well as in hematopoietic cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Mediation of growth hormone-dependent transcriptional activation by mammary gland factor/Stat 5.

Previous observations have shown that binding of growth hormone to its receptor leads to activation of transcription factors via a mechanism involving phosphorylation on tyrosine residues. In order to establish whether the prolactin-activated transcription factor Stat 5 (mammary gland factor) is also activated by growth hormone, nuclear extracts were prepared from COS-7 cells transiently expressing transfected Stat 5 and growth hormone receptor cDNA. Gel electrophoresis mobility shift analyses revealed the growth hormone-dependent presence of specific DNA-binding proteins in these extracts. The complexes formed could be supershifted by polyclonal anti-Stat 5 antiserum. In other experiments nuclear extracts from growth hormone-treated Chinese hamster ovary cells stably expressing transfected growth hormone receptor cDNA and liver from growth hormone-treated hypophysectomized rats were used for gel electrophoresis mobility shift analyses. These also revealed the presence of specific DNA-binding proteins sharing antigenic determinants with Stat 5. Stat 5 cDNA was shown to be capable of complementing the growth hormone-dependent activation of transcription of a reporter gene in the otherwise unresponsive COS-7 cell line. This complementation was dependent on the presence of Stat 5 tyrosine 694, suggesting a role for phosphorylation of this residue in growth hormone-dependent activation of DNA-binding and transcription.

Animals↗
PubMed1995

Colony-stimulating factors and interferon-gamma activate a protein related to MGF-Stat 5 to cause formation of the differentiation-induced factor in myeloid cells.

The Jak-Stat pathway of intracellular signals is used by growth factor- and cytokine receptors to induce gene transcription. We have recently reported that differentiation of myeloid cells, induced by phorbol ester, interferon-gamma (IFN-gamma) or colony-stimulating factor-1 (CSF-1) is accompanied by the activation of the differentiation-induced factor (DIF). Activated DIF specifically associates with a subclass of gamma-interferon activation site (GAS)-like DNA elements. We now report that GM-CSF, which like CSF-1 promotes the generation of mature macrophages, activates DIF. No activation was observed after treatment with the granulocyte growth and differentiation factor G-CSF. Antibodies raised against a Stat family protein, designated mammary gland factor-Stat 5 (MGF-Stat 5), reacted with DIF induced by either CSF-1, GM-CSF or IFN-gamma. Antisera to other known Stats were without effect on the DIF complex in electrophoretic mobility shift assays (EMSA). A 112 kDa protein could be isolated from either GM-CSF- or IFN-gamma-treated cells by GAS oligonucleotide precipitation. This protein reacted with antibodies to both MGF-Stat 5 and phosphotyrosine. MGF-Stat 5 and closely related proteins thus define a subfamily of Stat transcription factors that are present in a variety of cell types and are required for the onset of immediate gene expression in response to differentiating stimuli.

Blotting, Western↗
PubMed1995

Prolactin-mediated gene activation in mammary epithelial cells.

Mammary epithelial cells grow and develop with the onset of sexual maturity. In addition, lobular alveolar structures are formed during pregnancy, and quiescent differentiated cells secrete high levels of milk proteins after parturition. These events are governed by multiple hormones and growth factors and involve the sequential and synergistic action of functionally distinct signal transduction pathways. Milk protein genes have been analyzed and composite response elements have been identified in the promoter sequences. Transcription factors, which relay the hormonal signals, bind to these sequences. The factor that confers prolactin simulation to milk protein gene transcription has recently been identified. MGF/Stat5 is a latent transcription factor that becomes activated by a tyrosine-specific protein kinase, Jak2, associated with the prolactin receptor. Tyrosine phosphorylation converts the latent factor into one with DNA-binding and transcriptional activation potential. The regulation of MGF/Stat5 in vitro and in vivo indicates that it is a central component of the lactogenic hormone signaling pathway. Involvement of MGF/Stat5 in the signaling by other cytokines indicates that the same factor might be involved in regulation of growth-promoting genes, primarily in hematopoietic cells.

Animals↗
PubMed1995

Prolactin and interleukin-2 receptors in T lymphocytes signal through a MGF-STAT5-like transcription factor.

The cell surface receptors for PRL and interleukin-2 (IL-2) are structurally distinct, but share regulatory tasks in T lymphocytes. They can stimulate proliferation and activate transcription of over-lapping sets of genes of T cells. PRL and IL-2 receptor activation are both linked to the Jak/Stat (signal transducer and activator of transcription) pathway. We investigated the ability of PRL and IL-2 to activate Stat proteins in different T cell lines. The DNA binding specificities, the reactivities toward Stat-specific antisera, and the mol wt of IL-2- and PRL-induced DNA-binding proteins in Nb2 and C196 T cell lines were investigated. A comparison with the Stat proteins induced by interferon-gamma, PRL, and IL-6 in T47D mammary tumor cells was made. We found that these parameters were indistinguishable for one of the PRL- and IL-2-induced factors. A transcription factor closely related to mammary gland factor-Stat5 is rapidly activated upon interaction of IL-2 and PRL with their respective receptors. Activation of a second protein related to Stat1 was also observed. Our results emphasize the role of PRL as a regulator of the immune response and indicate that the Stat factors mammary gland factor-Stat5 and Stat1 play a role in the regulation of gene expression during T cell development.

Animals↗