Key Specifications Table
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H, M, R||EMSA, ICC, IP, WB, ChIP||Rb||Purified||Polyclonal Antibody|
|Presentation||Protein A purified rabbit IgG in 200 L of 0.1 M Tris-glycine, pH 7.4, 0.15 M NaCl, 0.05% sodium azide. Frozen at -20°C.|
|Safety Information according to GHS|
|Material Size||200 µg|
|Anti-STAT3 - 2453259||2453259|
|Anti-STAT3 - 15772||15772|
|Anti-STAT3 - 17470||17470|
|Anti-STAT3 - 1951972||1951972|
|Anti-STAT3 - 2019513||2019513|
|Anti-STAT3 - 2040087||2040087|
|Anti-STAT3 - 2189947||2189947|
|Anti-STAT3 - 22775||22775|
|Anti-STAT3 - 2309071||2309071|
|Anti-STAT3 - 24010||24010|
|Anti-STAT3 - 2533166||2533166|
|Anti-STAT3 - 25988||25988|
|Anti-STAT3 - 26460||26460|
|Anti-STAT3 - 29693||29693|
|Anti-STAT3 - 32584||32584|
|Anti-STAT3 - 33508||33508|
|Anti-STAT3 - DAM1661998||DAM1661998|
|Anti-STAT3 - JBC1355757||JBC1355757|
References | 38 Available | See All References
|Reference overview||Application||Species||Pub Med ID|
|Glutamine Deprivation Causes Hydrogen Peroxide-induced Interleukin-8 Expression via Jak1/Stat3 Activation in Gastric Epithelial AGS Cells. |
Lee, YM; Kim, MJ; Kim, Y; Kim, H
Journal of cancer prevention 20 179-84 2015
The Janus kinase (Jak)/Signal transducers of activated transcription (Stat) pathway is an upstream signaling pathway for NF-κB activation in Helicobacter pylori-induced interleukin (IL)-8 production in gastric epithelial AGS cells. H. pylori activates NADPH oxidase and produces hydrogen peroxide, which activates Jak1/Stat3 in AGS cells. Therefore, hydrogen peroxide may be critical for IL-8 production via Jak/Stat activation in gastric epithelial cells. Glutamine is depleted during severe injury and stress and contributes to the formation of glutathione (GSH), which is involved in conversion of hydrogen peroxide into water as a cofactor for GSH peroxidase.We investigated whether glutamine deprivation induces hydrogen peroxide-mediated IL-8 production and whether hydrogen peroxide activates Jak1/Stat3 to induce IL-8 in AGS cells. Cells were cultured in the presence or absence of glutamine or hydrogen peroxide, with or without GSH or a the Jak/Stat specific inhibitor AG490.Glutamine deprivation decreased GSH levels, but increased levels of hydrogen peroxide and IL-8, an effect that was inhibited by treatment with GSH. Hydrogen peroxide induced the activation of Jak1/Stat3 time-dependently. AG490 suppressed hydrogen peroxide- induced activation of Jak1/Stat3 and IL-8 expression in AGS cells, but did not affect levels of reactive oxygen species in AGS cells.In gastric epithelial AGS cells, glutamine deprivation increases hydrogen peroxide levels and IL-8 expression, which may be mediated by Jak1/Stat3 activation. Glutamine supplementation may be beneficial for preventing gastric inflammation by suppressing hydrogen peroxide-mediated Jak1/Stat3 activation and therefore, reducing IL-8 production. Scavenging hydrogen peroxide or targeting Jak1/Stat3 may also prevent oxidant-mediated gastric inflammation.
|Jak1/Stat3 is an upstream signaling of NF-κB activation in Helicobacter pylori-induced IL-8 production in gastric epithelial AGS cells. |
Cha, B; Lim, JW; Kim, H
Yonsei medical journal 56 862-6 2015
Helicobacter pylori (H. pylori) induces the activation of nuclear factor-kB (NF-κB) and cytokine expression in gastric epithelial cells. The Janus kinase/signal transducers and activators of transcription (Jak/Stat) cascade is the inflammatory signaling in various cells. The purpose of the present study is to determine whether H. pylori-induced activation of NF-κB and the expression of interleukin-8 (IL-8) are mediated by the activation of Jak1/Stat3 in gastric epithelial (AGS) cells. Thus, gastric epithelial AGS cells were infected with H. pylori in Korean isolates (HP99) at bacterium/cell ratio of 300:1, and the level of IL-8 in the medium was determined by enzyme-linked immonosorbent assay. Phospho-specific and total forms of Jak1/Stat3 and IκBα were assessed by Western blot analysis, and NF-κB activation was determined by electrophoretic mobility shift assay. The results showed that H. pylori induced the activation of Jak1/Stat3 and IL-8 production, which was inhibited by a Jak/Stat3 specific inhibitor AG490 in AGS cells in a dose-dependent manner. H. pylori-induced activation of NF-κB, determined by phosphorylation of IκBα and NF-κB-DNA binding activity, were inhibited by AG490. In conclusion, Jak1/Stat3 activation may mediate the activation of NF-κB and the expression of IL-8 in H. pylori-infected AGS cells. Inhibition of Jak1/Stat3 may be beneficial for the treatment of H. pylori-induced gastric inflammation, since the activation of NF-κB is inhibited and inflammatory cytokine expression is suppressed.
|A novel role for histone deacetylase 6 in the regulation of the tolerogenic STAT3/IL-10 pathway in APCs. |
Cheng, F; Lienlaf, M; Wang, HW; Perez-Villarroel, P; Lee, C; Woan, K; Rock-Klotz, J; Sahakian, E; Woods, D; Pinilla-Ibarz, J; Kalin, J; Tao, J; Hancock, W; Kozikowski, A; Seto, E; Villagra, A; Sotomayor, EM
Journal of immunology (Baltimore, Md. : 1950) 193 2850-62 2014
APCs are critical in T cell activation and in the induction of T cell tolerance. Epigenetic modifications of specific genes in the APC play a key role in this process, and among them histone deacetylases (HDACs) have emerged as key participants. HDAC6, one of the members of this family of enzymes, has been shown to be involved in regulation of inflammatory and immune responses. In this study, to our knowledge we show for the first time that genetic or pharmacologic disruption of HDAC6 in macrophages and dendritic cells results in diminished production of the immunosuppressive cytokine IL-10 and induction of inflammatory APCs that effectively activate Ag-specific naive T cells and restore the responsiveness of anergic CD4(+) T cells. Mechanistically, we have found that HDAC6 forms a previously unknown molecular complex with STAT3, association that was detected in both the cytoplasmic and nuclear compartments of the APC. By using HDAC6 recombinant mutants we identified the domain comprising amino acids 503-840 as being required for HDAC6 interaction with STAT3. Furthermore, by re-chromatin immunoprecipitation we confirmed that HDAC6 and STAT3 are both recruited to the same DNA sequence within the Il10 gene promoter. Of note, disruption of this complex by knocking down HDAC6 resulted in decreased STAT3 phosphorylation--but no changes in STAT3 acetylation--as well as diminished recruitment of STAT3 to the Il10 gene promoter region. The additional demonstration that a selective HDAC6 inhibitor disrupts this STAT3/IL-10 tolerogenic axis points to HDAC6 as a novel molecular target in APCs to overcome immune tolerance and tips the balance toward T cell immunity.
|Radiation-enhanced lung cancer progression in a transgenic mouse model of lung cancer is predictive of outcomes in human lung and breast cancer. |
Delgado, O; Batten, KG; Richardson, JA; Xie, XJ; Gazdar, AF; Kaisani, AA; Girard, L; Behrens, C; Suraokar, M; Fasciani, G; Wright, WE; Story, MD; Wistuba, II; Minna, JD; Shay, JW
Clinical cancer research : an official journal of the American Association for Cancer Research 20 1610-22 2014
Carcinogenesis is an adaptive process between nascent tumor cells and their microenvironment, including the modification of inflammatory responses from antitumorigenic to protumorigenic. Radiation exposure can stimulate inflammatory responses that inhibit or promote carcinogenesis. The purpose of this study is to determine the impact of radiation exposure on lung cancer progression in vivo and assess the relevance of this knowledge to human carcinogenesis.K-ras(LA1) mice were irradiated with various doses and dose regimens and then monitored until death. Microarray analyses were performed using Illumina BeadChips on whole lung tissue 70 days after irradiation with a fractionated or acute dose of radiation and compared with age-matched unirradiated controls. Unique group classifiers were derived by comparative genomic analysis of three experimental cohorts. Survival analyses were performed using principal component analysis and k-means clustering on three lung adenocarcinoma, three breast adenocarcinoma, and two lung squamous carcinoma annotated microarray datasets.Radiation exposure accelerates lung cancer progression in the K-ras(LA1) lung cancer mouse model with dose fractionation being more permissive for cancer progression. A nonrandom inflammatory signature associated with this progression was elicited from whole lung tissue containing only benign lesions and predicts human lung and breast cancer patient survival across multiple datasets. Immunohistochemical analyses suggest that tumor cells drive predictive signature.These results demonstrate that radiation exposure can cooperate with benign lesions in a transgenic model of cancer by affecting inflammatory pathways, and that clinically relevant similarities exist between human lung and breast carcinogenesis.
|Nucleolar protein trafficking in response to HIV-1 Tat: rewiring the nucleolus. |
Jarboui, MA; Bidoia, C; Woods, E; Roe, B; Wynne, K; Elia, G; Hall, WW; Gautier, VW
PloS one 7 e48702 2012
The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these proteins collectively participate in interconnected networks converging to adapt the nucleolus dynamic activities, which favor host biosynthetic activities and may contribute to create a cellular environment supporting robust HIV-1 production.
|Preclinical characterization of atiprimod, a novel JAK2 AND JAK3 inhibitor. |
Alfonso Quintás-Cardama,Taghi Manshouri,Zeev Estrov,David Harris,Ying Zhang,Amos Gaikwad,Hagop M Kantarjian,Srdan Verstovsek
Investigational new drugs 29 2011
We herein report on the activity of the JAK2/JAK3 small molecule inhibitor atiprimod on mouse FDCP-EpoR cells carrying either wild-type (JAK2 (WT)) or mutant (JAK2 (V617F)) JAK2, human acute megakaryoblastic leukemia cells carrying JAK2 (V617F) (SET-2 cell line), and human acute megakaryocytic leukemia carrying mutated JAK3 (CMK cells). Atiprimod inhibited more efficaciously the proliferation of FDCP-EpoR JAK2 (V617F) (IC(50) 0.42 μM) and SET-2 cells (IC(50) 0.53 μM) than that of CMK (IC(50) 0.79 μM) or FDCP-EpoR JAK2 (WT) cells (IC(50) 0.69 μM). This activity was accompanied by inhibition of the phosphorylation of JAK2 and downstream signaling proteins STAT3, STAT5, and AKT in a dose- and time-dependent manner. Atiprimod-induced cell growth inhibition of JAK2 (V617F)-positive cells was coupled with induction of apoptosis, as evidenced by heightened mitochondrial membrane potential and caspase-3 activity, as well as PARP cleavage, increased turnover of the anti-apoptotic X-linked mammalian inhibitor of apoptosis (XIAP) protein, and inhibition of the pro-apoptotic protein BCL-2 in a time- and dose-dependent manner. Furthermore, atiprimod was more effective at inhibiting the proliferation of peripheral blood hematopoietic progenitors obtained from patients with JAK2 (V617F)-positive polycythemia vera than at inhibiting hematopoietic progenitors from normal individuals (p = 0.001). The effect on primary expanded erythroid progenitors was paralleled by a decrease in JAK2(V617F) mutant allele burden in single microaspirated BFU-E and CFU-GM colonies. Taken together, our data supports the clinical testing of atiprimod in patients with hematologic malignancies driven by constitutive activation of JAK2 or JAK3 kinases.
|NFĸB is an unexpected major mediator of interleukin-15 signaling in cerebral endothelia. |
Stone, KP; Kastin, AJ; Pan, W
Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology 28 115-24 2011
Interleukin (IL)-15 and its receptors are induced by tumor necrosis factor α (TNF) in the cerebral endothelial cells composing the blood-brain barrier, but it is not yet clear how IL-15 modulates endothelial function. Contrary to the known induction of JAK/STAT3 signaling, here we found that nuclear factor (NF)- κB is mainly responsible for IL-15 actions on primary brain microvessel endothelial cells and cerebral endothelial cell lines. IL-15-induced transactivation of an NFκB luciferase reporter resulted in phosphorylation and degradation of the inhibitory subunit IκB that was followed by phosphorylation and nuclear translocation of the p65 subunit of NFκB. An IκB kinase inhibitor Bay 11-7082 only partially inhibited IL-15-induced NFκB luciferase activity. The effect of IL-15 was mediated by its specific receptor IL-15Rα, since endothelia from IL-15Rα knockout mice showed delayed nuclear translocation of p65, whereas those from knockout mice lacking a co-receptor IL-2Rγ did not show such changes. At the mRNA level, IL-15 and TNF showed similar effects in decreasing the tight junction protein claudin-2 and increasing the p65 subunit of NFκB but exerted different regulation on caveolin-1 and vimentin. Taken together, NFκB is a major signal transducer by which IL-15 affects cellular permeability, endocytosis, and intracellular trafficking at the level of the blood-brain barrier.
|Histone deacetylase inhibitor LAQ824 augments inflammatory responses in macrophages through transcriptional regulation of IL-10. |
Wang, H; Cheng, F; Woan, K; Sahakian, E; Merino, O; Rock-Klotz, J; Vicente-Suarez, I; Pinilla-Ibarz, J; Wright, KL; Seto, E; Bhalla, K; Villagra, A; Sotomayor, EM
Journal of immunology (Baltimore, Md. : 1950) 186 3986-96 2011
APCs are important in the initiation of productive Ag-specific T cell responses and the induction of T cell anergy. The inflammatory status of the APC at the time of encounter with Ag-specific T cells plays a central role in determining such divergent T cell outcomes. A better understanding of the regulation of proinflammatory and anti-inflammatory genes in its natural setting, the chromatin substrate, might provide novel insights to overcome anergic mechanisms mediated by APCs. In this study, we show for the first time, to our knowledge, that treatment of BALB/c murine macrophages with the histone deacetylase inhibitor LAQ824 induces chromatin changes at the level of the IL-10 gene promoter that lead to enhanced recruitment of the transcriptional repressors HDAC11 and PU.1. Such an effect is associated with diminished IL-10 production and induction of inflammatory cells able of priming naive Ag-specific T cells, but more importantly, capable of restoring the responsiveness of anergized Ag-specific CD4(+) T cells.
|Bone marrow stroma-secreted cytokines protect JAK2(V617F)-mutated cells from the effects of a JAK2 inhibitor. |
Manshouri, T; Estrov, Z; Quintás-Cardama, A; Burger, J; Zhang, Y; Livun, A; Knez, L; Harris, D; Creighton, CJ; Kantarjian, HM; Verstovsek, S
Cancer research 71 3831-40 2011
Signals emanating from the bone marrow microenvironment, such as stromal cells, are thought to support the survival and proliferation of the malignant cells in patients with myeloproliferative neoplasms (MPN). To examine this hypothesis, we established a coculture platform [cells cocultured directly (cell-on-cell) or indirectly (separated by micropore membrane)] designed to interrogate the interplay between Janus activated kinase 2-V617F (JAK2(V617F))-positive cells and the stromal cells. Treatment with atiprimod, a potent JAK2 inhibitor, caused marked growth inhibition and apoptosis of human (SET-2) and mouse (FDCP-EpoR) JAK2(V617F)-positive cells as well as primary blood or bone marrow mononuclear cells from patients with polycythemia vera; however, these effects were attenuated when any of these cell types were cocultured (cell-on-cell) with human marrow stromal cell lines (e.g., HS5, NK.tert, TM-R1). Coculture with stromal cells hampered the ability of atiprimod to inhibit phosphorylation of JAK2 and the downstream STAT3 and STAT5 pathways. This protective effect was maintained in noncontact coculture assays (JAK2(V617F)-positive cells separated by 0.4-μm-thick micropore membranes from stromal cells), indicating a paracrine effect. Cytokine profiling of supernatants from noncontact coculture assays detected distinctly high levels of interleukin 6 (IL-6), fibroblast growth factor (FGF), and chemokine C-X-C-motif ligand 10 (CXCL-10)/IFN-γ-inducible 10-kD protein (IP-10). Anti-IL-6, -FGF, or -CXCL-10/IP-10 neutralizing antibodies ablated the protective effect of stromal cells and restored atiprimod-induced apoptosis of JAK2(V617F)-positive cells. Therefore, our results indicate that humoral factors secreted by stromal cells protect MPN clones from JAK2 inhibitor therapy, thus underscoring the importance of targeting the marrow niche in MPN for therapeutic purposes.
|Different changes in protein and phosphoprotein levels result from serum starvation of high-grade glioma and adenocarcinoma cell lines. |
Levin, VA; Panchabhai, SC; Shen, L; Kornblau, SM; Qiu, Y; Baggerly, KA
Journal of proteome research 9 179-91 2010
Tumor cells undergoing serum starvation in vitro partially mimic metabolically stressed cells trying to adjust to a changed environment in vivo by inducing signal transduction and gene expression so that the tumor continues to grow. Our hypothesis is that the changes in protein and phosphoprotein levels after serum starvation may reflect the adapted phenotype of the tumor, which could be targeted for therapy. We used reverse-phase protein microarrays to interrogate five high-grade glioma cell lines and seven adenocarcinoma cell lines for differences in the level of 81 proteins and 25 phosphoproteins. All cell lines were studied in the well-fed condition of growth with 10% FBS and the starved condition of 0.5% FBS. Protein expression levels were normalized to beta-actin and trichotomized as increased (+1, upper 75th quartile), decreased (-1, lowest 25th quartile), or unchanged (0, others) to focus on the patterns of the biggest (and hopefully most robust) changes in protein and phosphoprotein levels. We examined these trichotomized values to better understand Starved-Fed differences among the cell lines and thereby gain better/clearer insight into the effects of serum starvation on potential cellular responses. In general, the expression of proteins and phosphoproteins 24 h after FBS starvation increased more often in glioma lines than in adenocarcinoma lines, which appeared to have fewer increased protein scores and more decreased scores. Many of the proteins increased in gliomas were downstream targets of the PTEN-PI-3 kinase-AKT, EGFR-MAPK-Stat, and transcription activator-polyamine signaling pathways. In adenocarcinomas, the expression of proteins and phosphoproteins generally increased in apoptosis pathways, while there were minor fluctuations in the other pathways above. Contrawise, gliomas become resistant to apoptosis after 24 h of serum starvation and upregulate transcription activators and polyamines more so than adenocarciomas.
|A Technical Assessment of the Utility of Reverse Phase Protein Arrays for the Study of the Functional Proteome in Non-microdissected Human Breast Cancers. |
Hennessy, BT; Lu, Y; Gonzalez-Angulo, AM; Carey, MS; Myhre, S; Ju, Z; Davies, MA; Liu, W; Coombes, K; Meric-Bernstam, F; Bedrosian, I; McGahren, M; Agarwal, R; Zhang, F; Overgaard, J; Alsner, J; Neve, RM; Kuo, WL; Gray, JW; Borresen-Dale, AL; Mills, GB
Clinical proteomics 6 129-51 2010
The lack of large panels of validated antibodies, tissue handling variability, and intratumoral heterogeneity potentially hamper comprehensive study of the functional proteome in non-microdissected solid tumors. The purpose of this study was to address these concerns and to demonstrate clinical utility for the functional analysis of proteins in non-microdissected breast tumors using reverse phase protein arrays (RPPA).Herein, 82 antibodies that recognize kinase and steroid signaling proteins and effectors were validated for RPPA. Intraslide and interslide coefficients of variability were less than 15%. Multiple sites in non-microdissected breast tumors were analyzed using RPPA after intervals of up to 24 h on the benchtop at room temperature following surgical resection.Twenty-one of 82 total and phosphoproteins demonstrated time-dependent instability at room temperature with most variability occurring at later time points between 6 and 24 h. However, the 82-protein functional proteomic "fingerprint" was robust in most tumors even when maintained at room temperature for 24 h before freezing. In repeat samples from each tumor, intratumoral protein levels were markedly less variable than intertumoral levels. Indeed, an independent analysis of prognostic biomarkers in tissue from multiple tumor sites accurately and reproducibly predicted patient outcomes. Significant correlations were observed between RPPA and immunohistochemistry. However, RPPA demonstrated a superior dynamic range. Classification of 128 breast cancers using RPPA identified six subgroups with markedly different patient outcomes that demonstrated a significant correlation with breast cancer subtypes identified by transcriptional profiling.Thus, the robustness of RPPA and stability of the functional proteomic "fingerprint" facilitate the study of the functional proteome in non-microdissected breast tumors.Full Text Article
|Phase 2 study of CEP-701, an orally available JAK2 inhibitor, in patients with primary or post-polycythemia vera/essential thrombocythemia myelofibrosis. |
Santos FP, Kantarjian HM, Jain N, Manshouri T, Thomas DA, Garcia-Manero G, Kennedy D, Estrov Z, Cortes J, Verstovsek S
Blood 115 1131-6. Epub 2009 Dec 11. 2010
Few treatment options exist for patients with myelofibrosis (MF), and their survival is significantly shortened. Activating mutation of the JAK2 tyrosine kinase (JAK2(V617F)) is found in approximately 50% of MF patients. CEP-701 is a tyrosine kinase inhibitor that inhibits JAK2 in in vitro and in vivo experiments. We conducted a phase 2 clinical study of CEP-701 in 22 JAK2(V617F)-positive MF patients (80 mg orally twice daily), and 6 (27%) responded by International Working Group criteria (clinical improvement in all cases): reduction in spleen size only (n = 3), transfusion independency (n = 2), and reduction in spleen size with improvement in cytopenias (n = 1). Median time to response was 3 months, and duration of response was more than or equal to 14 months. No improvement was seen in bone marrow fibrosis or JAK2(V617F) allele burden. Phosphorylated STAT3 levels decreased from baseline in responders while on therapy. Eight patients (36%) experienced grade 3 or 4 toxicity, and 6 (27%) required dose reduction. Main side effects were myelosuppression (grade 3 or 4 anemia, 14%; and thrombocytopenia, 23%) and gastrointestinal disturbances (diarrhea, any grade, 72%; grade 3 or 4, 9%; nausea, grade 1 or 2 only, 50%; vomiting, grade 1 or 2 only, 27%). In conclusion, CEP-701 resulted in modest efficacy and mild but frequent gastrointestinal toxicity in MF patients. The study was registered at http://clinicaltrials.gov as NCT00494585.
|A beta(3)-Adrenergic-Leptin-Melanocortin Circuit Regulates Behavioral and Metabolic Changes Induced by Chronic Stress. |
Chuang JC, Krishnan V, Yu HG, Mason B, Cui H, Wilkinson MB, Zigman JM, Elmquist JK, Nestler EJ, Lutter M
Biological psychiatry 2010
BACKGROUND: Obesity has been associated with an increased risk of developing several psychiatric illnesses, including major depression and posttraumatic stress disorder. Likewise, these stress-related disturbances are associated with a higher rate of obesity; yet, the neurobiological mechanisms linking obesity and stress remain incompletely understood. METHODS: Following exposure to chronic social defeat stress (CSDS), mice were given free access to either regular chow or a Western-style diet high in triglycerides and cholesterol. Comprehensive metabolic and behavioral testing was then conducted. RESULTS: Mice subjected to CSDS and then fed a high-fat diet for 30 days display severe behavioral deficits accompanied by redistribution of body fat. Stressed mice have decreased adipose tissue as well as decreased serum leptin levels compared with control mice. Pharmacological inhibition of beta(3)-adrenergic signaling during CSDS normalizes these metabolic abnormalities but worsens behavioral symptoms. Furthermore, mice subjected to CSDS display central leptin resistance including reduced expression of pro-opiomelanocortin in hypothalamus. Administration of a central melanocortin agonist worsens stress-induced behavioral deficits, while mice lacking the melanocortin-4 receptor display attenuated symptoms. CONCLUSIONS: These results indicate that chronic signaling through beta(3)-adrenergic receptors during social stress is an adaptive response that improves behavioral function. However, these responses come at the expense of central leptin resistance and melanocortin signaling alterations that contribute to significant and long-lasting metabolic abnormalities. Copyright Â© 2010 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.
|Defining the ATM-mediated barrier to tumorigenesis in somatic mammary cells following ErbB2 activation. |
Reddy JP, Peddibhotla S, Bu W, Zhao J, Haricharan S, Du YC, Podsypanina K, Rosen JM, Donehower LA, Li Y
Proc Natl Acad Sci U S A 107 3728-33. Epub 2010 Feb 3. 2010
p53, apoptosis, and senescence are frequently activated in preneoplastic lesions and are barriers to progression to malignancy. These barriers have been suggested to result from an ATM-mediated DNA damage response (DDR), which may follow oncogene-induced hyperproliferation and ensuing DNA replication stress. To elucidate the currently untested role of DDR in breast cancer initiation, we examined the effect of oncogene expression in several murine models of breast cancer. We did not observe a detectable DDR in early hyperplastic lesions arising in transgenic mice expressing several different oncogenes. However, DDR signaling was strongly induced in preneoplastic lesions arising from individual mammary cells transduced in vivo by retroviruses expressing either PyMT or ErbB2. Thus, activation of an oncogene after normal tissue development causes a DDR. Furthermore, in this somatic ErbB2 tumor model, ATM, and thus DDR, is required for p53 stabilization, apoptosis, and senescence. In palpable tumors in this model, p53 stabilization and apoptosis are lost, but unexpectedly senescence remains in many tumor cells. Thus, this murine model fully recapitulates early DDR signaling; the eventual suppression of its endpoints in tumorigenesis provides compelling evidence that ErbB2-induced aberrant mammary cell proliferation leads to an ATM-mediated DDR that activates apoptosis and senescence, and at least the former must be overcome to progress to malignancy. This in vivo study also uncovers an unexpected effect of ErbB2 activation previously known for its prosurvival roles, and suggests that protection of the ATM-mediated DDR-p53 signaling pathway may be important in breast cancer prevention.
|The 5-HT 2A serotonin receptor enhances cell viability, affects cell cycle progression and activates MEK-ERK1/2 and JAK2-STAT3 signalling pathways in human choriocarcinoma cell lines. |
Oufkir T, Arseneault M, Sanderson JT, Vaillancourt C
Placenta 31 439-47. Epub 2010 Mar 25. 2010
Previous results from our group have demonstrated the expression of the 5-HT(2A) receptor and a mitogenic effect of serotonin in human trophoblast. The objectives of the present study were to investigate the role of the 5-HT(2A) receptor in trophoblast cells and to determine the signalling pathways activated by this receptor. We investigated the effect of (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI), a selective 5-HT(2A) agonist, on cell cycle progression and cell viability in BeWo and JEG-3 cells. We also investigated, by co-immunoprecipitation and western blot analysis, the involvement of the MEK-ERK1/2 and JAK2-STAT3 signalling pathways following activation of the placental 5-HT(2A) receptor. Our results showed a concentration-dependent increase of cell viability by DOI, which was reversed by ketanserin, a selective 5-HT(2A) receptor antagonist. Furthermore, activation of the 5-HT(2A) receptor by DOI increased cell entry into the G2/M and S phase (DNA synthesis) in BeWo and JEG-3 cells, respectively. In addition, stimulation of BeWo and JEG-3 cells by DOI activated both the MEK-ERK1/2 and the JAK2-STAT3 signalling pathways. This study demonstrated that the 5-HT(2A) receptor increases cell viability and affects cell cycle progression in human trophoblast cell lines as well as activates the MEK-ERK1/2 and JAK2-STAT3 intracellular signalling pathways, which are related to survival, differentiation, migration and invasion. These findings indicate that serotonin through the activation of the 5-HT(2A) receptor is a key regulator of placentation and may play a role in the pathophysiology of certain pregnancy disorders associated with alterations in placental development, such as preeclampsia, gestational diabetes and preterm birth.
|Nuclear expression of a group II intron is consistent with spliceosomal intron ancestry. |
Chalamcharla VR, Curcio MJ, Belfort M
Genes Dev 24 827-36. Epub 2010 Mar 29. 2010
Group II introns are self-splicing RNAs found in eubacteria, archaea, and eukaryotic organelles. They are mechanistically similar to the metazoan nuclear spliceosomal introns; therefore, group II introns have been invoked as the progenitors of the eukaryotic pre-mRNA introns. However, the ability of group II introns to function outside of the bacteria-derived organelles is debatable, since they are not found in the nuclear genomes of eukaryotes. Here, we show that the Lactococcus lactis Ll.LtrB group II intron splices accurately and efficiently from different pre-mRNAs in a eukaryote, Saccharomyces cerevisiae. However, a pre-mRNA harboring a group II intron is spliced predominantly in the cytoplasm and is subject to nonsense-mediated mRNA decay (NMD), and the mature mRNA from which the group II intron is spliced is poorly translated. In contrast, a pre-mRNA bearing the Tetrahymena group I intron or the yeast spliceosomal ACT1 intron at the same location is not subject to NMD, and the mature mRNA is translated efficiently. Thus, a group II intron can splice from a nuclear transcript, but RNA instability and translation defects would have favored intron loss or evolution into protein-dependent spliceosomal introns, consistent with the bacterial group II intron ancestry hypothesis.Full Text Article
|Disruption of the growth hormone--signal transducer and activator of transcription 5--insulinlike growth factor 1 axis severely aggravates liver fibrosis in a mouse model of cholestasis. |
Leander Blaas,Jan-Wilhelm Kornfeld,Daniel Schramek,Monica Musteanu,Gernot Zollner,Judith Gumhold,Franziska van Zijl,Doris Schneller,Harald Esterbauer,Gerda Egger,Markus Mair,Lukas Kenner,Wolfgang Mikulits,Robert Eferl,Richard Moriggl,Josef Penninger,Michael Trauner,Emilio Casanova
Hepatology (Baltimore, Md.) 51 2010
Growth hormone (GH) resistance and low serum levels of insulinlike growth factor 1 (IGF-1) are common features in human liver fibrosis and cirrhosis. Signal transducer and activator of transcription 5 (STAT5) controls several vital functions in the liver, including GH-mediated transcription of IGF-1. To investigate the role of STAT5 in liver fibrogenesis, we specifically deleted the Stat5a/b locus both in hepatocytes and cholangiocytes in the multidrug resistance gene 2 knockout (Mdr2(-/-)) mouse model of cholestasis. Double knockout mice develop an early and severe liver fibrosis phenotype, accompanied by perturbed expression of key regulators of bile acid homeostasis. Deletion of Stat5 resulted in GH resistance, and IGF-1 levels in serum were undetectable. We could observe reduced expression of important hepatoprotective genes, such as epidermal growth factor receptor (Egfr), hepatocyte nuclear factor 6 (Hnf6), prolactin receptor (Prlr), and leukemia inhibitory factor receptor (Lifr) as well as increased numbers of apoptotic hepatocytes. Conclusion: Our data suggest that loss of STAT5 sensitizes hepatocytes to bile acid-induced damage and apoptosis caused by disruption of GH-induced transcription of Igf-1 and down-regulation of hepatoprotective genes. These findings could contribute to the understanding of liver fibrosis and future treatment strategies for liver fibrosis.Full Text Article
|The histone deacetylase HDAC11 regulates the expression of interleukin 10 and immune tolerance. |
Alejandro Villagra, Fengdong Cheng, Hong-Wei Wang, Ildelfonso Suarez, Michelle Glozak, Michelle Maurin, Danny Nguyen, Kenneth L Wright, Peter W Atadja, Kapil Bhalla, Javier Pinilla-Ibarz, Edward Seto, Eduardo M Sotomayor, Alejandro Villagra, Fengdong Cheng, Hong-Wei Wang, Ildelfonso Suarez, Michelle Glozak, Michelle Maurin, Danny Nguyen, Kenneth L Wright, Peter W Atadja, Kapil Bhalla, Javier Pinilla-Ibarz, Edward Seto, Eduardo M Sotomayor, Alejandro Villagra, Fengdong Cheng, Hong-Wei Wang, Ildelfonso Suarez, Michelle Glozak, Michelle Maurin, Danny Nguyen, Kenneth L Wright, Peter W Atadja, Kapil Bhalla, Javier Pinilla-Ibarz, Edward Seto, Eduardo M Sotomayor, Alejandro Villagra, Fengdong Cheng, Hong-Wei Wang, Ildelfonso Suarez, Michelle Glozak, Michelle Maurin, Danny Nguyen, Kenneth L Wright, Peter W Atadja, Kapil Bhalla, Javier Pinilla-Ibarz, Edward Seto, Eduardo M Sotomayor
Nature immunology 10 92-100 2009
Antigen-presenting cells (APCs) induce T cell activation as well as T cell tolerance. The molecular basis of the regulation of this critical 'decision' is not well understood. Here we show that HDAC11, a member of the HDAC histone deacetylase family with no prior defined physiological function, negatively regulated expression of the gene encoding interleukin 10 (IL-10) in APCs. Overexpression of HDAC11 inhibited IL-10 expression and induced inflammatory APCs that were able to prime naive T cells and restore the responsiveness of tolerant CD4+ T cells. Conversely, disruption of HDAC11 in APCs led to upregulation of expression of the gene encoding IL-10 and impairment of antigen-specific T cell responses. Thus, HDAC11 represents a molecular target that influences immune activation versus immune tolerance, a critical 'decision' with substantial implications in autoimmunity, transplantation and cancer immunotherapy.
|Changes related to phosphatidylinositol 3-kinaseAkt signaling in leiomyomas: possible involvement of glycogen synthase kinase 3alpha and cyclin D2 in the pathophysiology. |
L Karra, A Shushan, A Ben-Meir, N Rojansky, BY Klein, D Shveiky, R Levitzki, H Ben-Bassat
Fertility and sterility 2009
OBJECTIVE: To identify changes in the expression and phosphorylation of phosphatidylinositol 3-kinase (PI3K)/Akt protein kinases controlling survival and/or apoptosis of in vitro cell cultures of uterine leiomyomas. DESIGN: Establishment of paired cell cultures of leiomyoma and myometrial specimens. SETTING: Hadassah gynecology research laboratory. PATIENT(S): Eleven white premenopausal women, 35 to 50 years of age, undergoing hysterectomy because of symptomatic uterine leiomyomas. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Immunochemical analysis of expression and phosphorylation of relevant PI3K/Akt and BCL2 proteins. RESULT(S): Analysis of total phosphatase and tensin homologue deleted on chromosome 10 (PTEN) and of nonphosphorylated and phosphorylated (p) PDK1, Akt, glycogen synthase kinase 3 (GSK3), FKHR, tuberin (TSC2) and hamartin (TSC1) complex, and cyclin D2 proteins indicated that  the level of pGSK3alpha and cyclin D2 proteins was elevated significantly in the leiomyoma compared with the normal myometrium,  there was a significant interaction between PTEN- PDK1 and between pAkt-pGSK3beta in the leiomyoma compared with the myometrial cells, and  there was a significant interaction between pAkt-pGSK3alpha in the paired leiomyoma and myometrial cultures. CONCLUSION(S): Our study suggests that the downstream signaling components of the PI3K/Akt pathway, GSK3 (a regulator of apoptosis), and cyclin D2 (a promoter of G1/S progression), as well as the significant interaction between PTEN-PDK and between pAkt-pGSK3beta, are involved in the survival and proliferation of leiomyomas.,
|Src-dependent STAT-3-mediated expression of monocyte chemoattractant protein-1 is required for 15(S)-hydroxyeicosatetraenoic acid-induced vascular smooth muscle cell migration. |
HS Potula, D Wang, DV Quyen, NK Singh, V Kundumani-Sridharan, M Karpurapu, EA Park, WC Glasgow, GN Rao
The Journal of biological chemistry 284 31142-55 2009
To understand the role of human 15-lipoxygenase 1 (15-LOX1) in vascular wall remodeling, we have studied the effect of the major 15-LOX1 metabolite of arachidonic acid, 15(S)-hydroxyeicosatetraenoic acid (15(S)-HETE), on vascular smooth muscle cell (VSMC) migration both in vitro and in vivo. Among 5(S)-HETE, 12(S)-HETE, and 15(S)-HETE, 15(S)-HETE potentially stimulated more vascular smooth muscle cell (VSMC) migration. In addition, 15(S)-HETE-induced VSMC migration was dependent on Src-mediated activation of signal transducer and activator of transcription-3 (STAT-3). 15(S)-HETE also induced monocyte chemoattractant protein-1 (MCP-1) expression via Src-STAT-3 signaling, and neutralizing anti-MCP-1 antibodies completely negated 15(S)-HETE-induced VSMC migration. Cloning and characterization of a 2.6-kb MCP-1 promoter revealed the presence of four putative STAT-binding sites, and the site that is proximal to the transcription start site was found to be essential for 15(S)-HETE-induced Src-STAT-3-mediated MCP-1 expression. Rat carotid arteries that were subjected to balloon injury and transduced with Ad-15-LOX1 upon exposure to [(3)H]arachidonic acid ex vivo produced 15-HETE as a major eicosanoid and enhanced balloon injury-induced expression of MCP-1 in smooth muscle cells in Src and STAT-3-dependent manner in vivo. Adenovirus-mediated delivery of 15-LOX1 into rat carotid artery also led to recruitment and homing of macrophages to medial region in response to injury. In addition, transduction of Ad-15-LOX1 into arteries enhanced balloon injury-induced smooth muscle cell migration from media to intima and neointima formation. These results show for the first time that 15-LOX1-15(S)-HETE axis plays a major role in vascular wall remodeling after balloon angioplasty.Full Text Article
|Oxysterol and diabetes activate STAT3 and control endothelial expression of profilin-1 via OSBP1. |
Romeo, GR; Kazlauskas, A
The Journal of biological chemistry 283 9595-605 2008
Endothelial dysfunction plays a central role in diabetic vascular disease, but its molecular bases are not completely defined. We showed previously that the actin-binding protein proflin-1 was increased in the diabetic endothelium and that attenuated expression of profilin-1 protected against atherosclerosis. Also 7-ketocholesterol up-regulated profilin-1 in endothelial cells via transcriptional mechanisms. The present study addressed the pathways responsible for profilin-1 gene expression in 7-ketocholesterol-stimulated endothelial cells and in the diabetic aorta. In luciferase reporter assays, the response to 7-ketocholesterol within the 5'-flanking region of profilin-1 was dependent on a single STAT response element. In aortic endothelial cells, 7-ketocholesterol enhanced STAT3 activation, which required JAK2 and tyrosine 394 phosphorylation of oxysterol-binding protein-1. These changes were recapitulated in the aorta of diabetic rats. Also 7-ketocholesterol in cultured endothelial cells and diabetes in the aorta elicited the recruitment of STAT3 and relevant coregulatory factors to the oxysterol-responsive region of the profilin-1 promoter. These events were required for profilin-1 up-regulation. These studies identify a previously unrecognized oxysterol-binding protein-mediated mode of activation of STAT3 that controls the expression of the proatherogenic protein profilin-1 in response to 7-ketocholesterol and the diabetic milieu.
|Transcriptional activation of the suppressor of cytokine signaling-3 (SOCS-3) gene via STAT3 is increased in F9 REX1 (ZFP-42) knockout teratocarcinoma stem cells relative to wild-type cells. |
Xu, J; Sylvester, R; Tighe, AP; Chen, S; Gudas, LJ
Journal of molecular biology 377 28-46 2008
Rex1 (Zfp42), first identified as a gene that is transcriptionally repressed by retinoic acid (RA), encodes a zinc finger transcription factor expressed at high levels in F9 teratocarcinoma stem cells, embryonic stem cells, and other stem cells. Loss of both alleles of Rex1 by homologous recombination alters the RA-induced differentiation of F9 cells, a model of pluripotent embryonic stem cells. We identified Suppressor of Cytokine Signaling-3 (SOCS-3) as a gene that exhibits greatly increased transcriptional activation in RA, cAMP, and theophylline (RACT)-treated F9 Rex1(-/-) cells (approximately 25-fold) as compared to wild-type (WT) cells ( approximately 2.5-fold). By promoter deletion, mutation, and transient transfection analyses, we have shown that this transcriptional increase is mediated by the STAT3 DNA-binding elements located between -99 to -60 in the SOCS-3 promoter. Overexpression of STAT3 dominant-negative mutants greatly diminishes this SOCS-3 transcriptional increase in F9 Rex1(-/-) cells. This increase in SOCS-3 transcription is associated with a four- to fivefold higher level of tyrosine-phosphorylated STAT3 in the RACT-treated F9 Rex1(-/-) cells as compared to WT. Dominant-negative Src tyrosine kinase, Jak2, and protein kinase A partially reduce the transcriptional activation of the SOCS 3 gene in RACT-treated F9 Rex1 null cells. In contrast, parathyroid hormone peptide enhances the effect of RA in F9 Rex1(-/-) cells, but not in F9 WT. Thus, Rex1, which is highly expressed in stem cells, inhibits signaling via the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway, thereby modulating the differentiation of F9 cells.
|Developmental changes in the human GH receptor and its signal transduction pathways. |
Kenth, G; Mergelas, JA; Goodyer, CG
The Journal of endocrinology 198 71-82 2008
We previously reported the presence of functional human GH receptors (hGHRs) in the human fetal hepatocyte (FH) as early as the first trimester. Interestingly, fetal serum levels of hGH are in the acromegalic range, yet certain hGH-dependent factors are expressed at very low levels (IGF-I, IGF-binding protein-3), suggesting that fetal liver has limited responsiveness to hGH. To determine whether this is due to the fetal tissue levels of hGHR or factors in the hGH/hGHR axis that might influence hGHR function, we compared hGHR isoforms and downstream signaling proteins in FH versus human adult liver (HAL). Immunoprecipitation/immunoblotting (IB) analyses found similar precursor and mature hGHR forms while RT-PCR assays of truncated (T) hGHR(1-279), dominant negative for the full-length (FL) receptor, showed similar T/FL mRNA ratios in FH and HAL. IB demonstrated that Janus kinase (JAK) 2, signal transducers and activators of transcription (STAT(1, 3, 5A/B)), and suppressors of cytokine signaling (SOCS(1, 2, 3, cytokine-inducible SH2-containing protein (CIS))) proteins were detectable in all FH and HAL tested (12 weeks of fetal age to 60 years); the levels were similar (STAT5B) or lower (JAK2/STAT1/STAT3/STAT5A: 38-53%, SOCS/CIS: 58-76%) in FH compared with HAL. Our studies to date demonstrate that, during hepatocyte development, hGHR levels are lower in the fetal cells but the hGHR isoforms, including the relative amount of truncated versus FL, remain unchanged. The JAK2/STAT/SOCS signaling molecules are present in the FH as early as the first trimester. However, they are generally at less than 50% level in postnatal liver. These data suggest that low expression of both hGHR and major hGHR signaling components may explain the limited responsiveness of the fetal cells to the high circulating levels of hGH.
|Overexpression of Wnt-1 in thyrocytes enhances cellular growth but suppresses transcription of the thyroperoxidase gene via different signaling mechanisms. |
Kim, WB; Lewis, CJ; McCall, KD; Malgor, R; Kohn, AD; Moon, RT; Kohn, LD
The Journal of endocrinology 193 93-106 2007
Wnt binding to cell surface receptors can activate a 'canonical' pathway that increases cellular beta-catenin or a 'noncanonical' Ca(++) pathway which can increase protein kinase C (PKC) activity. Although components of both Wnt/beta-catenin-signaling pathways exist in thyrocytes, their biological role is largely unknown. In evaluating the biological role of Wnt signaling in differentiated FRTL-5 thyroid cells, we showed that TSH increased canonical Wnt-1 but, surprisingly, decreased the active form of beta-catenin. Transient overexpression of Wnt-1 or beta-catenin in FRTL-5 cells increased active beta-catenin (ABC), decreased thyroperoxidase (TPO) mRNA, and suppressed TPO-promoter activity. The target of beta-catenin suppressive action was a consensus T cell factor/lymphoid enhancing factor (TCF/LEF)-binding site 5'-A/T A/T CAAAG-3', -137 to -129 bp on the rat TPO promoter. beta-Catenin overexpression significantly increased complex formation between beta-catenin/TCF-1 and an oligonucleotide containing the TCF/LEF sequence, suggesting that the beta-catenin/TCF-1 complex acts as a transcriptional repressor of the TPO gene. Stable over-expression of Wnt-1 in FRTL-5 cells significantly increased the growth rate without increasing beta-catenin levels. Increased growth was blunted by a PKC inhibitor, staurosporin. Wnt-1 overexpression increased serine phosphorylation, without affecting tyrosine phosphorylation, of signal transducers and activators of transcription 3 (STAT3) protein. In addition, these final results suggest that TSH-induced increase in Wnt-1 levels in thyrocytes contributes to enhanced cellular growth via a PKC pathway that increases STAT3 serine phosphorylation and activation, whereas TSH-induced decrease in activation of beta-catenin simultaneously relieves transcriptional suppression of TPO. We hypothesize that Wnt signaling contributes to the ability of TSH to simultaneously increase cell growth and functional, thyroid-specific, gene expression.
|Novel role for amphiregulin in protection from liver injury. |
Berasain, C; García-Trevijano, ER; Castillo, J; Erroba, E; Santamaría, M; Lee, DC; Prieto, J; Avila, MA
The Journal of biological chemistry 280 19012-20 2005
Clinically, the Fas and Fas ligand system plays a central role in the development of hepatocyte apoptosis, a process contributing to a broad spectrum of liver diseases. Therefore, the development of therapies aimed at the inhibition of hepatocyte apoptosis is a major issue. Activation of the epidermal growth factor receptor has been shown to convey survival signals to the hepatocyte. To learn about the endogenous response of epidermal growth factor receptor ligands during Fas-mediated liver injury we investigated the expression of epidermal growth factor, transforming growth factor alpha, heparin-binding epidermal growth factor-like growth factor, betacellulin, epiregulin, and amphiregulin in the liver of mice challenged with Fas-agonist antibody. Amphiregulin expression, barely detectable in healthy liver, was significantly up-regulated. Amphiregulin administration abrogated Fas-mediated liver injury in mice and showed direct anti-apoptotic effects in primary hepatocytes. Amphiregulin activated the Akt and signal transducer and activator of transcription-3 survival pathways, and up-regulated Bcl-xL expression. Amphiregulin knock-out mice showed signs of chronic liver damage in the absence of any noxious treatment, and died faster than wild type mice in response to lethal doses of Fas-agonist antibody. In contrast, these mice were more resistant against sublethal liver damage, supporting the hypothesis that chronic liver injury can precondition hepatocytes inducing resistance to subsequent cell death. These results show that amphiregulin is a protective factor induced in response to liver damage and that it may be therapeutic in liver diseases.
|Retinoic acid inhibits leukemia inhibitory factor signaling pathways in mouse embryonic stem cells. |
Ann P Tighe, Lorraine J Gudas
Journal of cellular physiology 198 223-9 2004
Retinoic acid (RA) induces the differentiation of murine embryonic stem (ES) cells to cell types resembling those found in the early embryo. When cultured in the presence of leukemia inhibitory factor (LIF), ES cells are maintained in an undifferentiated (self-renewing) state. Addition of RA to the culture media overrides the self-renewing effects of LIF to induce ES cell differentiation. Therefore, we hypothesized that RA-induced differentiation of ES cells may be accomplished by antagonism of LIF-induced signaling pathways. We demonstrate that RA-induced differentiation of CCE ES cells is associated with (1) downregulation of the LIF receptor (LIFR); (2) decreased tyrosine phosphorylation of signal transducer and activator of transcription 3 protein (Stat3); and (3) increased activation of extracellular regulated kinase (Erk1/2). We conclude that RA induces CCE ES cell differentiation in the presence of LIF, in part, by disrupting signaling between the LIFR/gp130 receptor and nuclear targets that are required to prevent ES cell differentiation. Our data indicate that RA-induced inhibition of LIF signaling does not involve Erk1/2-dependent actions.
|IL-5 and granulocyte-macrophage colony-stimulating factor activate STAT3 and STAT5 and promote Pim-1 and cyclin D3 protein expression in human eosinophils. |
Barbara A Stout, Mary Ellen Bates, Lin Ying Liu, Natasha N Farrington, Paul J Bertics
Journal of immunology (Baltimore, Md. : 1950) 173 6409-17 2004
Allergic inflammation is characterized by elevated eosinophil numbers and by the increased production of the cytokines IL-5 and GM-CSF, which control several eosinophil functions, including the suppression of apoptosis. The JAK/STAT pathway is important for several functions in hemopoietic cells, including the suppression of apoptosis. We report in this study that STAT3, STAT5a, and STAT5b are expressed in human eosinophils and that their signaling pathways are active following IL-5 or GM-CSF treatment. However, in airway eosinophils, the phosphorylation of STAT5 by IL-5 is reduced, an event that may be related to the reduced expression of the IL-5Ralpha on airway eosinophils. Furthermore, IL-5 and GM-CSF induced the protein expression of cyclin D3 and the kinase Pim-1, both of which are regulated by STAT-dependent processes in some cell systems. Pim-1 is more abundantly expressed in airway eosinophils than in blood eosinophils. Because Pim-1 reportedly has a role in the modulation of apoptosis, these results suggest that Pim-1 action is linked to the suppression of eosinophil apoptosis by these cytokines. Although cyclin D3 is known to be critical for cell cycle progression, eosinophils are terminally differentiated cells that do not proceed through the cell cycle. Thus, this apparent cytokine regulation of cyclin D3 suggests that there is an alternative role(s) for cyclin D3 in eosinophil biology.
|The AG1478 tyrosine kinase inhibitor is an effective suppressor of leiomyoma cell growth. |
Shushan, A; Rojansky, N; Laufer, N; Klein, BY; Shlomai, Z; Levitzki, R; Hartzstark, Z; Ben-Bassat, H
Human reproduction (Oxford, England) 19 1957-67 2004
Uterine leiomyomas are the most common benign smooth muscle cell tumours in women. Formation of leiomyomas, still not completely understood, is viewed as a multistep process, with involvement of ovarian steroid hormones, cytokines and growth factors. Our study aimed to identify tyrosine kinase inhibitors as potential 'signal transduction therapeutics' for leiomyomas, underlying the effect of ovarian steroidal hormones.The selective epidermal growth factor (EGF) receptor blocker AG1478 was evaluated as a potential target, since EGF has been shown to mediate estrogen action and to play a crucial role in regulating leiomyoma growth. Paired cultures of leiomyoma and normal myometrium samples were established and the suppressive effect of AG1478 on the cells prior and subsequent to steroidal hormone treatment was examined: cell proliferation, recovery after treatment, cell cycle analysis and immunochemical analysis of relevant proteins.Leiomyoma cell growth is effectively blocked by AG1478 and is unaffected by the presence of physiological concentrations of progesterone and estradiol. AG1478 (10 microM) completely suppressed proliferation and the cells did not recover after cessation of treatment.The growth-arresting properties of AG1478, unaffected by ovarian steroidal hormones, identify it as a potential lead agent for the non-surgical management of uterine leiomyomas.
|Interferon consensus sequence-binding protein is constitutively expressed and differentially regulated in the ocular lens |
Li, W., et al
J Biol Chem, 274:9686-91 (1999) 1999
|Electrophoretic Mobility Shift Assay||10092656|
|The carboxyl-terminal domains of gp130-related cytokine receptors are necessary for suppressing embryonic stem cell differentiation. Involvement of STAT3 |
Ernst, M., et al
J Biol Chem, 274:9729-37 (1999) 1999
|Erythropoietin and Friend virus gp55 activate different JAK/STAT pathways through the erythropoietin receptor in erythroid cells |
Yamamura, Y., et al
Mol Cell Biol, 18:1172-80 (1998) 1998
|Immunoblotting (Western), Immunoprecipitation||9488432|
|Differentiation of oligodendroglial progenitors derived from cortical multipotent cells requires extrinsic signals including activation of gp130/LIFbeta receptors |
Marmur, R., et al
J Neurosci, 18:9800-11 (1998) 1998
|The box-1 region of the leukemia inhibitory factor receptor alpha-chain cytoplasmic domain is sufficient for hemopoietic cell proliferation and differentiation |
Zhang, Y., et al
J Biol Chem, 273:34370-83 (1998) 1998
|Electrophoretic Mobility Shift Assay||9852103|
|Critical cytoplasmic domains of human interleukin-9 receptor alpha chain in interleukin-9-mediated cell proliferation and signal transduction |
Zhu, Y. X., et al
J Biol Chem, 272:21334-40 (1997) 1997
|Urokinase receptor is associated with the components of the JAK1/STAT1 signaling pathway and leads to activation of this pathway upon receptor clustering in the human kidney epithelial tumor cell line TCL-598. |
Koshelnick, Y, et al.
J. Biol. Chem., 272: 28563-7 (1997) 1997
The urokinase-type plasminogen activator (uPA) binds to cells via a specific receptor attached to the plasma membrane by a glycosylphosphatidylinositol (GPI) anchor. Despite the lack of a transmembrane domain, the urokinase receptor (uPAR) is capable of transducing extracellular signals affecting growth, migration, and adhesion. Several Tyr kinases of the src family as well as beta1, beta2, and beta3 integrins were found to be associated with the uPAR. We found that in the human kidney epithelial line TCL-598, also components of the JAK1/STAT1 signal transduction pathway including gp130, are associated with uPAR as revealed by coimmunoprecipitation and are co-localized in caveolae. Upon clustering of uPA.uPAR complex by a monoclonal antibody, JAK1 associates with uPAR, which in turn leads to STAT1 phosphorylation, dimerization, specific binding to DNA, and gene activation. To prove the dependence of STAT1 activation on the uPAR, TCL-598 cells were treated with sense and antisense uPAR oligonucleotides. In antisense-treated cells in which uPAR expression was reduced to less then one third, activation of STAT1 by the clustering antibody was abolished while STAT1 activation by interferon-gamma was unaffected. Therefore, in this cell line, uPA.uPAR also utilizes the JAK1/STAT1 pathway for signaling, and gp130 might be the transmembrane adapter for this signal transduction pathway.
|Electrophoretic Mobility Shift Assay||9353320|
|Requirement of serine phosphorylation for formation of STAT-promoter complexes. |
Zhang, X, et al.
Science, 267: 1990-4 (1995) 1995
Members of the interleukin-6 family of cytokines bind to and activate receptors that contain a common subunit, gp130. This leads to the activation of Stat3 and Stat1, two cytoplasmic signal transducers and activators of transcription (STATs), by tyrosine phosphorylation. Serine phosphorylation of Stat3 was constitutive and was enhanced by signaling through gp130. In cells of lymphoid and neuronal origins, inhibition of serine phosphorylation prevented the formation of complexes of DNA with Stat3-Stat3 but not with Stat3-Stat1 or Stat1-Stat1 dimers. In vitro serine dephosphorylation of Stat3 also inhibited DNA binding of Stat3-Stat3. The requirement of serine phosphorylation for Stat3-Stat3.DNA complex formation was inversely correlated with the affinity of Stat3-Stat3 for the binding site. Thus, serine phosphorylation appears to enhance or to be required for the formation of stable Stat3-Stat3.DNA complexes.
|Molecular cloning of APRF, a novel IFN-stimulated gene factor 3 p91-related transcription factor involved in the gp130-mediated signaling pathway. |
Akira, S, et al.
Cell, 77: 63-71 (1994) 1994
|Stat3: a STAT family member activated by tyrosine phosphorylation in response to epidermal growth factor and interleukin-6 |
Zhong, Z, et al
Science, 264:95-8 (1994) 1994