WorldWideScience

Sample records for cell receptor signalling

  1. Lipid rafts in T cell receptor signalling (Review)

    OpenAIRE

    KABOURIDIS, PANAGIOTIS S.

    2006-01-01

    The molecular events and the protein components that are involved in signalling by the T cell receptor (TCR) for antigen have been extensively studied. Activation of signalling cascades following TCR stimulation depends on the phosphorylation of the receptor by the tyrosine kinase Lck, which localizes to the cytoplasmic face of the plasma membrane by virtue of its post-translational modification. However, the precise order of events during TCR phosphorylation at the plasma membrane, remains t...

  2. Erythropoietin regulates Treg cells in asthma through TGFβ receptor signaling.

    Science.gov (United States)

    Wan, Guoshi; Wei, Bing

    2015-01-01

    Asthma is a chronic inflammatory disorder of the airways, the development of which is suppressed by regulatory T cells (Treg). Erythropoietin (EPO) is originally defined as a hematopoietic growth factor. Recently, the anti-inflammatory effects of EPO in asthma have been acknowledged. However, the underlying mechanisms remain ill-defined. Here, we showed that EPO treatment significantly reduced the severity of an ovalbumin (OVA)-induced asthma in mice, seemingly through promoting Foxp3-mediated activation of Treg cells in OVA-treated mouse lung. The activation of Treg cells resulted from increases in transforming growth factor β1 (TGFβ1), which were mainly produced by M2 macrophages (M2M). In vitro, Co-culture with M2M increased Foxp3 levels in Treg cells and the Treg cell number, in a TGFβ receptor signaling dependent manner. Moreover, elimination of macrophages abolished the therapeutic effects of EPO in vivo. Together, our data suggest that EPO may increase M2M, which activate Treg cells through TGFβ receptor signaling to mitigate the severity of asthma. PMID:26807178

  3. Vitamin D Receptor Signaling and Pancreatic Cancer Cell EMT

    Science.gov (United States)

    Li, Zhiwei; Guo, Junli; Xie, Keping; Zheng, Shaojiang

    2016-01-01

    Pancreatic ductal adenocarcinoma remains one of the most lethal of human malignancies. Even in patients who undergo resection, long-term survival rates remain extremely low. A major contributor to the aggressiveness of pancreatic ductal adenocarcinoma is epithelial-to-mesenchymal transition (EMT), a physiologic process of morphological and genetic changes in carcinoma cells from an epithelial phenotype to a mesenchymal phenotype, which is the basis of the high metastatic potential of pancreatic cancer cells. EMT is triggered by various tumor microenvironmental factors, including cytokines, growth factors, and chemotherapeutic agents. This review highlights the growing evidence of the effect of EMT on pancreatic cancer progression, focusing on the interaction of EMT with other pathways central to cancer progression, especially vitamin D receptor signaling. Studies of the signaling pathways that lead to the inactivation of EMT programs during these disease processes are providing new insights into the plasticity of cellular phenotypes and possible therapeutic interventions. PMID:25506892

  4. DMPD: Signals and receptors involved in recruitment of inflammatory cells. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 7744810 Signals and receptors involved in recruitment of inflammatory cells. Ben-Ba...ruch A, Michiel DF, Oppenheim JJ. J Biol Chem. 1995 May 19;270(20):11703-6. (.png) (.svg) (.html) (.csml) Show Signal...s and receptors involved in recruitment of inflammatory cells. PubmedID 7744810 Title Signals and r

  5. Tumor necrosis factor receptor superfamily costimulation couples T cell receptor signal strength to thymic regulatory T cell differentiation

    OpenAIRE

    Mahmud, Shawn A.; Manlove, Luke S.; Schmitz, Heather M.; Xing, Yan; Wang, Yanyan; Owen, David L.; Schenkel, Jason M.; Boomer, Jonathan S; Jonathan M Green; Yagita, Hideo; Chi, Hongbo; Hogquist, Kristin A.; Farrar, Michael A.

    2014-01-01

    Regulatory T (Treg) cells express tumor necrosis factor receptor superfamily (TNFRSF) members, but their role in thymic Treg development is undefined. We demonstrate that Treg progenitors highly express the TNFRSF members GITR, OX40, and TNFR2. Expression of these receptors correlates directly with T cell receptor (TCR) signal strength, and requires CD28 and the kinase TAK1. Neutralizing TNFSF ligands markedly reduced Treg development. Conversely, TNFRSF agonists enhanced Treg differentiation...

  6. P2X and P2Y receptor signaling in red blood cells

    Science.gov (United States)

    Sluyter, Ronald

    2015-01-01

    Purinergic signaling involves the activation of cell surface P1 and P2 receptors by extracellular nucleosides and nucleotides such as adenosine and adenosine triphosphate (ATP), respectively. P2 receptors comprise P2X and P2Y receptors, and have well-established roles in leukocyte and platelet biology. Emerging evidence indicates important roles for these receptors in red blood cells. P2 receptor activation stimulates a number of signaling pathways in progenitor red blood cells resulting in microparticle release, reactive oxygen species formation, and apoptosis. Likewise, activation of P2 receptors in mature red blood cells stimulates signaling pathways mediating volume regulation, eicosanoid release, phosphatidylserine exposure, hemolysis, impaired ATP release, and susceptibility or resistance to infection. This review summarizes the distribution of P2 receptors in red blood cells, and outlines the functions of P2 receptor signaling in these cells and its implications in red blood cell biology. PMID:26579528

  7. Vitamin D receptor-retinoid X receptor heterodimer signaling regulates oligodendrocyte progenitor cell differentiation.

    Science.gov (United States)

    de la Fuente, Alerie Guzman; Errea, Oihana; van Wijngaarden, Peter; Gonzalez, Ginez A; Kerninon, Christophe; Jarjour, Andrew A; Lewis, Hilary J; Jones, Clare A; Nait-Oumesmar, Brahim; Zhao, Chao; Huang, Jeffrey K; ffrench-Constant, Charles; Franklin, Robin J M

    2015-12-01

    The mechanisms regulating differentiation of oligodendrocyte (OLG) progenitor cells (OPCs) into mature OLGs are key to understanding myelination and remyelination. Signaling via the retinoid X receptor γ (RXR-γ) has been shown to be a positive regulator of OPC differentiation. However, the nuclear receptor (NR) binding partner of RXR-γ has not been established. In this study we show that RXR-γ binds to several NRs in OPCs and OLGs, one of which is vitamin D receptor (VDR). Using pharmacological and knockdown approaches we show that RXR-VDR signaling induces OPC differentiation and that VDR agonist vitamin D enhances OPC differentiation. We also show expression of VDR in OLG lineage cells in multiple sclerosis. Our data reveal a role for vitamin D in the regenerative component of demyelinating disease and identify a new target for remyelination medicines. PMID:26644513

  8. Peroxisome Proliferator-Activated Receptor and Vitamin D Receptor Signaling Pathways in Cancer Cells

    International Nuclear Information System (INIS)

    Peroxisome proliferator-activated receptors (PPARs) are members of the superfamily of nuclear hormone receptors, which respond to specific ligands such as polyunsaturated fatty acids by altering gene expression. Three subtypes of this receptor have been discovered, each evolving to achieve different biological functions. Like other nuclear receptors, the transcriptional activity of PPARs is affected not only by ligand-stimulation, but also by cross-talk with other molecules. For example, both PPARs and the RXRs are ligand-activated transcription factors that coordinately regulate gene expression. In addition, PPARs and vitamin D receptor (VDR) signaling pathways regulate a multitude of genes that are of importance for cellular functions including cell proliferation and cell differentiation. Interaction of the PPARs and VDR signaling pathways has been shown at the level of molecular cross-regulation of their transcription factor. A variety of ligands influencing the PPARs and VDR signaling pathways have been shown to reveal chemopreventive potential by mediating tumor suppressive activities in human cancers. Use of these compounds may represent a potential novel strategy to prevent cancers. This review summarizes the roles of the PPARs and the VDR in pathogenesis and progression of cancer

  9. Peroxisome Proliferator-Activated Receptor and Vitamin D Receptor Signaling Pathways in Cancer Cells

    Directory of Open Access Journals (Sweden)

    Yasuko Kitagishi

    2013-10-01

    Full Text Available Peroxisome proliferator-activated receptors (PPARs are members of the superfamily of nuclear hormone receptors, which respond to specific ligands such as polyunsaturated fatty acids by altering gene expression. Three subtypes of this receptor have been discovered, each evolving to achieve different biological functions. Like other nuclear receptors, the transcriptional activity of PPARs is affected not only by ligand-stimulation, but also by cross-talk with other molecules. For example, both PPARs and the RXRs are ligand-activated transcription factors that coordinately regulate gene expression. In addition, PPARs and vitamin D receptor (VDR signaling pathways regulate a multitude of genes that are of importance for cellular functions including cell proliferation and cell differentiation. Interaction of the PPARs and VDR signaling pathways has been shown at the level of molecular cross-regulation of their transcription factor. A variety of ligands influencing the PPARs and VDR signaling pathways have been shown to reveal chemopreventive potential by mediating tumor suppressive activities in human cancers. Use of these compounds may represent a potential novel strategy to prevent cancers. This review summarizes the roles of the PPARs and the VDR in pathogenesis and progression of cancer.

  10. Peroxisome Proliferator-Activated Receptor and Vitamin D Receptor Signaling Pathways in Cancer Cells

    Energy Technology Data Exchange (ETDEWEB)

    Matsuda, Satoru, E-mail: smatsuda@cc.nara-wu.ac.jp; Kitagishi, Yasuko [Department of Food Science and Nutrition, Nara Women’s University, Kita-Uoya Nishimachi, Nara 630-8506 (Japan)

    2013-10-21

    Peroxisome proliferator-activated receptors (PPARs) are members of the superfamily of nuclear hormone receptors, which respond to specific ligands such as polyunsaturated fatty acids by altering gene expression. Three subtypes of this receptor have been discovered, each evolving to achieve different biological functions. Like other nuclear receptors, the transcriptional activity of PPARs is affected not only by ligand-stimulation, but also by cross-talk with other molecules. For example, both PPARs and the RXRs are ligand-activated transcription factors that coordinately regulate gene expression. In addition, PPARs and vitamin D receptor (VDR) signaling pathways regulate a multitude of genes that are of importance for cellular functions including cell proliferation and cell differentiation. Interaction of the PPARs and VDR signaling pathways has been shown at the level of molecular cross-regulation of their transcription factor. A variety of ligands influencing the PPARs and VDR signaling pathways have been shown to reveal chemopreventive potential by mediating tumor suppressive activities in human cancers. Use of these compounds may represent a potential novel strategy to prevent cancers. This review summarizes the roles of the PPARs and the VDR in pathogenesis and progression of cancer.

  11. Regulation of C3a Receptor Signaling in Human Mast Cells by G Protein Coupled Receptor Kinases

    OpenAIRE

    Qiang Guo; Hariharan Subramanian; Kshitij Gupta; Hydar Ali

    2011-01-01

    BACKGROUND: The complement component C3a activates human mast cells via its cell surface G protein coupled receptor (GPCR) C3aR. For most GPCRs, agonist-induced receptor phosphorylation leads to receptor desensitization, internalization as well as activation of downstream signaling pathways such as ERK1/2 phosphorylation. Previous studies in transfected COS cells overexpressing G protein coupled receptor kinases (GRKs) demonstrated that GRK2, GRK3, GRK5 and GRK6 participate in agonist-induced...

  12. GLYCINE AND GLYCINE RECEPTOR SIGNALING IN IMMUNE CELLS

    OpenAIRE

    Van den Eynden, Jimmy

    2010-01-01

    The central nervous system (CNS) is an integration center for signal processing, receiving signals from the different sensory systems and transmitting signals to the motor system. The main cells conducting signals are neurons, and for the largest part of the 20th century most attention of neuroscientist was focused on neurons. A role of glial cells, for a long time considered as passive connective tissue elements, in normal physiology and pathophysiology is now becoming increasingly appreciat...

  13. Vitamin D controls T cell antigen receptor signaling and activation of human T cells

    DEFF Research Database (Denmark)

    von Essen, Marina Rode; Kongsbak, Martin; Schjerling, Peter;

    2010-01-01

    Phospholipase C (PLC) isozymes are key signaling proteins downstream of many extracellular stimuli. Here we show that naive human T cells had very low expression of PLC-gamma1 and that this correlated with low T cell antigen receptor (TCR) responsiveness in naive T cells. However, TCR triggering...... led to an upregulation of approximately 75-fold in PLC-gamma1 expression, which correlated with greater TCR responsiveness. Induction of PLC-gamma1 was dependent on vitamin D and expression of the vitamin D receptor (VDR). Naive T cells did not express VDR, but VDR expression was induced by TCR...... signaling via the alternative mitogen-activated protein kinase p38 pathway. Thus, initial TCR signaling via p38 leads to successive induction of VDR and PLC-gamma1, which are required for subsequent classical TCR signaling and T cell activation....

  14. G Protein-Coupled Receptor Signaling in Stem Cells and Cancer

    OpenAIRE

    Jennifer R. Lynch; Jenny Yingzi Wang

    2016-01-01

    G protein-coupled receptors (GPCRs) are a large superfamily of cell-surface signaling proteins that bind extracellular ligands and transduce signals into cells via heterotrimeric G proteins. GPCRs are highly tractable drug targets. Aberrant expression of GPCRs and G proteins has been observed in various cancers and their importance in cancer stem cells has begun to be appreciated. We have recently reported essential roles for G protein-coupled receptor 84 (GPR84) and G protein subunit Gαq in ...

  15. High Cell Surface Death Receptor Expression Determines Type I Versus Type II Signaling*

    Science.gov (United States)

    Meng, Xue Wei; Peterson, Kevin L.; Dai, Haiming; Schneider, Paula; Lee, Sun-Hee; Zhang, Jin-San; Koenig, Alexander; Bronk, Steve; Billadeau, Daniel D.; Gores, Gregory J.; Kaufmann, Scott H.

    2011-01-01

    Previous studies have suggested that there are two signaling pathways leading from ligation of the Fas receptor to induction of apoptosis. Type I signaling involves Fas ligand-induced recruitment of large amounts of FADD (FAS-associated death domain protein) and procaspase 8, leading to direct activation of caspase 3, whereas type II signaling involves Bid-mediated mitochondrial perturbation to amplify a more modest death receptor-initiated signal. The biochemical basis for this dichotomy has previously been unclear. Here we show that type I cells have a longer half-life for Fas message and express higher amounts of cell surface Fas, explaining the increased recruitment of FADD and subsequent signaling. Moreover, we demonstrate that cells with type II Fas signaling (Jurkat or HCT-15) can signal through a type I pathway upon forced receptor overexpression and that shRNA-mediated Fas down-regulation converts cells with type I signaling (A498) to type II signaling. Importantly, the same cells can exhibit type I signaling for Fas and type II signaling for TRAIL (TNF-α-related apoptosis-inducing ligand), indicating that the choice of signaling pathway is related to the specific receptor, not some other cellular feature. Additional experiments revealed that up-regulation of cell surface death receptor 5 levels by treatment with 7-ethyl-10-hydroxy-camptothecin converted TRAIL signaling in HCT116 cells from type II to type I. Collectively, these results suggest that the type I/type II dichotomy reflects differences in cell surface death receptor expression. PMID:21865165

  16. Signaling through C5a receptor and C3a receptor diminishes function of murine natural regulatory T cells

    OpenAIRE

    Kwan, Wing-hong; Van der Touw, William; Paz-Artal, Estela; Li, Ming O.; Heeger, Peter S.

    2013-01-01

    Thymus-derived (natural) CD4+ FoxP3+ regulatory T cells (nT reg cells) are required for immune homeostasis and self-tolerance, but must be stringently controlled to permit expansion of protective immunity. Previous findings linking signals transmitted through T cell–expressed C5a receptor (C5aR) and C3a receptor (C3aR) to activation, differentiation, and expansion of conventional CD4+CD25− T cells (T conv cells), raised the possibility that C3aR/C5aR signaling on nT reg cells could physiologi...

  17. Is The CD200/CD200 Receptor Interaction More Than Just a Myeloid Cell Inhibitory Signal?

    OpenAIRE

    Minas, Konstantinos; Liversidge, Janet

    2006-01-01

    The membrane glycoprotein CD200, which has a widespread but defined distribution and a structurally similar receptor (CD200R) that transmits an inhibitory signal to cells of the hematopoetic lineage, especially myeloid cells, has been characterized. CD200R expression is restricted predominantly to cells of the myeloid lineage indicating that this ligand/receptor pair has a specific role in controlling myeloid cell function. In addition to CD200R, several related genes have been identified. Wh...

  18. Regulation of T cell receptor signaling by the actin cytoskeleton and poroelastic cytoplasm

    OpenAIRE

    Beemiller, Peter; Krummel, Matthew F.

    2013-01-01

    The actin cytoskeleton plays essential roles in modulating T-cell activation. Most models of T-cell receptor (TCR) triggering, signalosome assembl, y and immune synapse formation invoke actin-dependent mechanisms. As T cells are constitutively motile cells, TCR triggering and signaling occur against a cytoskeletal backdrop that is constantly remodeling. While the interplay between actin dynamics and TCR signaling have been the focus of research for many years, much of the work in T cells has ...

  19. Sodium/Calcium Exchangers Selectively Regulate Calcium Signaling in Mouse Taste Receptor Cells

    OpenAIRE

    Szebenyi, Steven A.; Laskowski, Agnieszka I.; Medler, Kathryn F.

    2010-01-01

    Taste cells use multiple signaling mechanisms to generate appropriate cellular responses to discrete taste stimuli. Some taste stimuli activate G protein coupled receptors (GPCRs) that cause calcium release from intracellular stores while other stimuli depolarize taste cells to cause calcium influx through voltage-gated calcium channels (VGCCs). While the signaling mechanisms that initiate calcium signals have been described in taste cells, the calcium clearance mechanisms (CCMs) that contrib...

  20. Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand–receptor binding and therefore impairs cancer cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Feng [Department of Gastroenterology, The Tenth People’s Hospital of Shanghai, Tongji University, Shanghai 200072 (China); Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Yang, Yong, E-mail: yyang@houstonmethodist.org [Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Department of Medicine, Weill Cornell Medical College, New York, NY 10065 (United States)

    2014-10-03

    Graphical abstract: - Highlights: • Quercetin inhibits insulin ligand–receptor interactions. • Quercetin reduces downstream insulin receptor signaling. • Quercetin blocks insulin induced glucose uptake. • Quercetin suppresses insulin stimulated cancer cell proliferation and tumor growth. - Abstract: Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand–receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocation of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers.

  1. Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand–receptor binding and therefore impairs cancer cell proliferation

    International Nuclear Information System (INIS)

    Graphical abstract: - Highlights: • Quercetin inhibits insulin ligand–receptor interactions. • Quercetin reduces downstream insulin receptor signaling. • Quercetin blocks insulin induced glucose uptake. • Quercetin suppresses insulin stimulated cancer cell proliferation and tumor growth. - Abstract: Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand–receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocation of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers

  2. Phosphorylation site dynamics of early T-cell receptor signaling

    DEFF Research Database (Denmark)

    Chylek, Lily A; Akimov, Vyacheslav; Dengjel, Jörn;

    2014-01-01

    systems-level understanding of how these components cooperate to control signaling dynamics, especially during the crucial first seconds of stimulation. Here, we used quantitative proteomics to characterize reshaping of the T-cell phosphoproteome in response to TCR/CD28 co-stimulation, and found that...... diverse dynamic patterns emerge within seconds. We detected phosphorylation dynamics as early as 5 s and observed widespread regulation of key TCR signaling proteins by 30 s. Development of a computational model pointed to the presence of novel regulatory mechanisms controlling phosphorylation of sites...

  3. Proteomic and phosphoproteomic analysis of signalling by adhesion and growth factor receptors in mammary epithelial cells

    OpenAIRE

    Paul, Nikki

    2014-01-01

    Cell adhesion and communication are essential for tissue morphogenesis and repair in healthy multicellular organisms. However, dysregulation of these processes can drive disease progression in conditions such as cancer. Selective cell adhesion to the extracellular matrix is mediated by integrins, a family of transmembrane receptors that compartmentalise signalling and organise the cytoskeleton. Adhesion receptors provide spatial cues to cells to allow them to respond to growth factor and cyto...

  4. Selective Accumulation of Raft-Associated Membrane Protein Lat in T Cell Receptor Signaling Assemblies

    OpenAIRE

    Harder, Thomas; Kuhn, Marina

    2000-01-01

    Activation of T cell antigen receptor (TCR) induces tyrosine phosphorylations that mediate the assembly of signaling protein complexes. Moreover, cholesterol-sphingolipid raft membrane domains have been implicated to play a role in TCR signal transduction. Here, we studied the assembly of TCR with signal transduction proteins and raft markers in plasma membrane subdomains of Jurkat T leukemic cells. We employed a novel method to immunoisolate plasma membrane subfragments that were highly conc...

  5. Targeting early B-cell receptor signaling induces apoptosis in leukemic mantle cell lymphoma

    Directory of Open Access Journals (Sweden)

    Boukhiar Mohand-Akli

    2013-02-01

    Full Text Available Abstract Background We previously showed that B-cell receptor (BCR signaling pathways are important for in vitro survival of mantle cell lymphoma (MCL cells. To further identify early BCR-activated signaling pathways involved in MCL cell survival, we focused our study on BCR-proximal kinases such as LYN whose dysregulations could contribute to the aggressive course of MCL. Methods Primary MCL cells were isolated from 14 leukemic patients. Early BCR-induced genes were identified by qRT-PCR array. The basal and BCR-induced phosphorylation of LYN and JNK were evaluated by immunoblottting. Cell survival signals were evaluated by apoptosis using flow cytometry. Results We showed that LYN was constitutively phosphorylated in MCL cell lines and in 9/10 leukemic MCL cases. Treatment with dasatinib or with a specific inhibitor of Src kinases such as PP2 suppressed constitutive LYN activation and increased in vitro spontaneous apoptosis of primary MCL cells. BCR engagement resulted in an increase of LYN phosphorylation leading to activation of c-JUN NH2-terminal kinase (JNK and over-expression of the early growth response gene-1 (EGR-1. Inhibition of JNK with SP600125 induced apoptosis and reduced level of basal and BCR-induced expression of EGR-1. Furthermore, decreasing EGR1 expression by siRNA reduced BCR-induced cell survival. Treatment with PP2 or with dasatinib suppressed BCR-induced LYN and JNK phosphorylation as well as EGR-1 upregulation and is associated with a decrease of cell survival in all cases analysed. Conclusions This study highlights the importance of BCR signaling in MCL cell survival and points out to the efficiency of kinase inhibitors in suppressing proximal BCR signaling events and in inducing apoptosis.

  6. Altered insulin receptor signalling and β-cell cycle dynamics in type 2 diabetes mellitus.

    Directory of Open Access Journals (Sweden)

    Franco Folli

    Full Text Available Insulin resistance, reduced β-cell mass, and hyperglucagonemia are consistent features in type 2 diabetes mellitus (T2DM. We used pancreas and islets from humans with T2DM to examine the regulation of insulin signaling and cell-cycle control of islet cells. We observed reduced β-cell mass and increased α-cell mass in the Type 2 diabetic pancreas. Confocal microscopy, real-time PCR and western blotting analyses revealed increased expression of PCNA and down-regulation of p27-Kip1 and altered expression of insulin receptors, insulin receptor substrate-2 and phosphorylated BAD. To investigate the mechanisms underlying these findings, we examined a mouse model of insulin resistance in β-cells--which also exhibits reduced β-cell mass, the β-cell-specific insulin receptor knockout (βIRKO. Freshly isolated islets and β-cell lines derived from βIRKO mice exhibited poor cell-cycle progression, nuclear restriction of FoxO1 and reduced expression of cell-cycle proteins favoring growth arrest. Re-expression of insulin receptors in βIRKO β-cells reversed the defects and promoted cell cycle progression and proliferation implying a role for insulin-signaling in β-cell growth. These data provide evidence that human β- and α-cells can enter the cell-cycle, but proliferation of β-cells in T2DM fails due to G1-to-S phase arrest secondary to defective insulin signaling. Activation of insulin signaling, FoxO1 and proteins in β-cell-cycle progression are attractive therapeutic targets to enhance β-cell regeneration in the treatment of T2DM.

  7. N-wasp is essential for the negative regulation of B cell receptor signaling.

    Directory of Open Access Journals (Sweden)

    Chaohong Liu

    2013-11-01

    Full Text Available Negative regulation of receptor signaling is essential for controlling cell activation and differentiation. In B-lymphocytes, the down-regulation of B-cell antigen receptor (BCR signaling is critical for suppressing the activation of self-reactive B cells; however, the mechanism underlying the negative regulation of signaling remains elusive. Using genetically manipulated mouse models and total internal reflection fluorescence microscopy, we demonstrate that neuronal Wiskott-Aldrich syndrome protein (N-WASP, which is coexpressed with WASP in all immune cells, is a critical negative regulator of B-cell signaling. B-cell-specific N-WASP gene deletion causes enhanced and prolonged BCR signaling and elevated levels of autoantibodies in the mouse serum. The increased signaling in N-WASP knockout B cells is concurrent with increased accumulation of F-actin at the B-cell surface, enhanced B-cell spreading on the antigen-presenting membrane, delayed B-cell contraction, inhibition in the merger of signaling active BCR microclusters into signaling inactive central clusters, and a blockage of BCR internalization. Upon BCR activation, WASP is activated first, followed by N-WASP in mouse and human primary B cells. The activation of N-WASP is suppressed by Bruton's tyrosine kinase-induced WASP activation, and is restored by the activation of SH2 domain-containing inositol 5-phosphatase that inhibits WASP activation. Our results reveal a new mechanism for the negative regulation of BCR signaling and broadly suggest an actin-mediated mechanism for signaling down-regulation.

  8. Role of Cbl-associated protein/ponsin in receptor tyrosine kinase signaling and cell adhesion

    Directory of Open Access Journals (Sweden)

    Ritva Tikkanen

    2012-10-01

    Full Text Available The Cbl-associated protein/ponsin (CAP is an adaptor protein that contains a so-called Sorbin homology (SoHo domain and three Src homology 3 (SH3 domains which are engaged in diverse protein-protein interactions. CAP has been shown to function in the regulation of the actin cytoskeleton and cell adhesion and to be involved in the differentiation of muscle cells and adipocytes. In addition, it participates in signaling pathways through several receptor tyrosine kinases such as insulin and neurotrophin receptors. In the last couple of years, several studies have shed light on the details of these processes and identified novel interaction partners of CAP. In this review, we summarize these recent findings and provide an overview on the function of CAP especially in cell adhesion and membrane receptor signaling.

  9. Square Cell Packing in the Drosophila Embryo through Spatiotemporally Regulated EGF Receptor Signaling.

    Science.gov (United States)

    Tamada, Masako; Zallen, Jennifer A

    2015-10-26

    Cells display dynamic and diverse morphologies during development, but the strategies by which differentiated tissues achieve precise shapes and patterns are not well understood. Here we identify a developmental program that generates a highly ordered square cell grid in the Drosophila embryo through sequential and spatially regulated cell alignment, oriented cell division, and apicobasal cell elongation. The basic leucine zipper transcriptional regulator Cnc is necessary and sufficient to produce a square cell grid in the presence of a midline signal provided by the EGF receptor ligand Spitz. Spitz orients cell divisions through a Pins/LGN-dependent spindle-positioning mechanism and controls cell shape and alignment through a transcriptional pathway that requires the Pointed ETS domain protein. These results identify a strategy for producing ordered square cell packing configurations in epithelia and reveal a molecular mechanism by which organized tissue structure is generated through spatiotemporally regulated responses to EGF receptor activation. PMID:26506305

  10. T Cell Receptor-Independent Basal Signaling via Erk and Abl Kinases Suppresses RAG Gene Expression

    Directory of Open Access Journals (Sweden)

    Roose Jeroen P

    2003-01-01

    Full Text Available Signal transduction pathways guided by cellular receptors commonly exhibit low-level constitutive signaling in a continuous, ligand-independent manner. The dynamic equilibrium of positive and negative regulators establishes such a tonic signal. Ligand-independent signaling by the precursors of mature antigen receptors regulates development of B and T lymphocytes. Here we describe a basal signal that controls gene expression profiles in the Jurkat T cell line and mouse thymocytes. Using DNA microarrays and Northern blots to analyze unstimulated cells, we demonstrate that expression of a cluster of genes, including RAG-1 and RAG-2, is repressed by constitutive signals requiring the adapter molecules LAT and SLP-76. This TCR-like pathway results in constitutive low-level activity of Erk and Abl kinases. Inhibition of Abl by the drug STI-571 or inhibition of signaling events upstream of Erk increases RAG-1 expression. Our data suggest that physiologic gene expression programs depend upon tonic activity of signaling pathways independent of receptor ligation.

  11. Regulation of T cell receptor signaling by the actin cytoskeleton and poroelastic cytoplasm

    Science.gov (United States)

    Beemiller, Peter; Krummel, Matthew F.

    2013-01-01

    Summary The actin cytoskeleton plays essential roles in modulating T-cell activation. Most models of T-cell receptor (TCR) triggering, signalosome assembl, y and immune synapse formation invoke actin-dependent mechanisms. As T cells are constitutively motile cells, TCR triggering and signaling occur against a cytoskeletal backdrop that is constantly remodeling. While the interplay between actin dynamics and TCR signaling have been the focus of research for many years, much of the work in T cells has considered actin largely for its ‘scaffolding’ function. We examine the roles of the actin cytoskeleton in TCR signaling and immune synapse formation with an emphasis on how poroelasticity, an ensemble feature of actin dynamics with the cytosol, relates to how T cells respond to stimulation. PMID:24117819

  12. Regulation of T-cell receptor signaling by the actin cytoskeleton and poroelastic cytoplasm.

    Science.gov (United States)

    Beemiller, Peter; Krummel, Matthew F

    2013-11-01

    The actin cytoskeleton plays essential roles in modulating T-cell activation. Most models of T-cell receptor (TCR) triggering signalosome assembly and immune synapse formation invoke actin-dependent mechanisms. As T cells are constitutively motile cells, TCR triggering and signaling occur against a cytoskeletal backdrop that is constantly remodeling. While the interplay between actin dynamics and TCR signaling have been the focus of research for many years, much of the work in T cells has considered actin largely for its 'scaffolding' function. We examine the roles of the actin cytoskeleton in TCR signaling and immune synapse formation with an emphasis on how poroelasticity, an ensemble feature of actin dynamics with the cytosol, relates to how T cells respond to stimulation. PMID:24117819

  13. Rationale for targeting the pre-B-cell receptor signaling pathway in acute lymphoblastic leukemia.

    Science.gov (United States)

    Müschen, Markus

    2015-06-11

    Inhibitors of B-cell receptor (BCR) and pre-BCR signaling were successfully introduced into patient care for various subtypes of mature B-cell lymphoma (e.g., ibrutinib, idelalisib). Acute lymphoblastic leukemia (ALL) typically originates from pre-B cells that critically depend on survival signals emanating from a functional pre-BCR. However, whether patients with ALL benefit from treatment with (pre-) BCR inhibitors has not been explored. Recent data suggest that the pre-BCR functions as tumor suppressor in the majority of cases of human ALL. However, a distinct subset of human ALL is selectively sensitive to pre-BCR antagonists. PMID:25878119

  14. A model for the biosynthesis and transport of plasma membrane-associated signaling receptors to the cell surface

    Directory of Open Access Journals (Sweden)

    Sorina Claudia Popescu

    2012-04-01

    Full Text Available Intracellular protein transport is emerging as critical in determining the outcome of receptor-activated signal transduction pathways. In plants, relatively little is known about the nature of the molecular components and mechanisms involved in coordinating receptor synthesis and transport to the cell surface. Recent advances in this field indicate that signaling pathways and intracellular transport machinery converge and coordinate to render receptors competent for signaling at their plasma membrane activity sites. The biogenesis and transport to the cell surface of signaling receptors appears to require both general trafficking and receptor-specific factors. Several molecular determinants, residing or associated with compartments of the secretory pathway and known to influence aspects in receptor biogenesis, are discussed and integrated into a predictive cooperative model for the functional expression of signaling receptors at the plasma membrane.

  15. TRPM5, a taste-signaling transient receptor potential ion-channel, is a ubiquitous signaling component in chemosensory cells

    Directory of Open Access Journals (Sweden)

    Hofmann Thomas

    2007-07-01

    Full Text Available Abstract Background A growing number of TRP channels have been identified as key players in the sensation of smell, temperature, mechanical forces and taste. TRPM5 is known to be abundantly expressed in taste receptor cells where it participates in sweet, amino acid and bitter perception. A role of TRPM5 in other sensory systems, however, has not been studied so far. Results Here, we systematically investigated the expression of TRPM5 in rat and mouse tissues. Apart from taste buds, where we found TRPM5 to be predominantly localized on the basolateral surface of taste receptor cells, TRPM5 immunoreactivity was seen in other chemosensory organs – the main olfactory epithelium and the vomeronasal organ. Most strikingly, we found solitary TRPM5-enriched epithelial cells in all parts of the respiratory and gastrointestinal tract. Based on their tissue distribution, the low cell density, morphological features and co-immunostaining with different epithelial markers, we identified these cells as brush cells (also known as tuft, fibrillovesicular, multivesicular or caveolated cells. In terms of morphological characteristics, brush cells resemble taste receptor cells, while their origin and biological role are still under intensive debate. Conclusion We consider TRPM5 to be an intrinsic signaling component of mammalian chemosensory organs, and provide evidence for brush cells being an important cellular correlate in the periphery.

  16. Charged MVB protein 5 is involved in T-cell receptor signaling.

    Science.gov (United States)

    Wi, Sae Mi; Min, Yoon; Lee, Ki-Young

    2016-01-01

    Charged multivesicular body protein 5 (CHMP5) has a key role in multivesicular body biogenesis and a critical role in the downregulation of signaling pathways through receptor degradation. However, the role of CHMP5 in T-cell receptor (TCR)-mediated signaling has not been previously investigated. In this study, we utilized a short hairpin RNA-based RNA interference approach to investigate the functional role of CHMP5. Upon TCR stimulation, CHMP5-knockdown (CHMP5(KD)) Jurkat T cells exhibited activation of TCR downstream signaling molecules, such as PKCθ and IKKαβ, and resulted in the activation of nuclear factor-κB and the marked upregulation of TCR-induced gene expression. Moreover, we found that activator protein-1 and nuclear factor of activated T-cells transcriptional factors were markedly activated in CHMP5(KD) Jurkat cells in response to TCR stimulation, which led to a significant increase in interleukin-2 secretion. Biochemical studies revealed that CHMP5 endogenously forms high-molecular-weight complexes, including TCR molecules, and specifically interacts with TCRβ. Interestingly, flow cytometry analysis also revealed that CHMP5(KD) Jurkat T cells exhibit upregulation of TCR expression on the cell surface compared with control Jurkat T cells. Taken together, these findings demonstrated that CHMP5 might be involved in the homeostatic regulation of TCR on the cell surface, presumably through TCR recycling or degradation. Thus CHMP5 is implicated in TCR-mediated signaling. PMID:26821576

  17. HIV-1 Nef: a multifaceted modulator of T cell receptor signaling

    Directory of Open Access Journals (Sweden)

    Abraham Libin

    2012-12-01

    Full Text Available Abstract Nef, an accessory protein of the Human Immunodeficiency Virus type 1 (HIV-1, is dispensable for viral replication in cell culture, but promotes virus replication and pathogenesis in the infected host. Acting as protein-interaction adaptor, HIV-1 Nef modulates numerous target cell activities including cell surface receptor expression, cytoskeletal remodeling, vesicular transport, and signal transduction. In infected T-lymphocytes, altering T-cell antigen receptor (TCR signaling has long been recognized as one key function of the viral protein. However, reported effects of Nef range from inhibition to activation of this cascade. Recent advances in the field begin to explain these seemingly contradictory observations and suggest that Nef alters intracellular trafficking of TCR proximal machinery to disrupt plasma membrane bound TCR signaling while at the same time, the viral protein induces localized signal transduction at the trans-Golgi network. This review summarizes these new findings on how HIV-1 Nef reprograms TCR signalling output from a broad response to selective activation of the RAS-Erk pathway. We also discuss the implications of these alterations in the context of HIV-1 infection and in light of current concepts of TCR signal transduction.

  18. Purinergic receptors and calcium signalling in human pancreatic duct cell lines

    DEFF Research Database (Denmark)

    Hansen, Mette R; Krabbe, Simon; Novak, Ivana

    2008-01-01

    expression of P2X4 and P2X7 receptors. Expression of P2Y2, P2X4 and P2X7 receptors was confirmed by immunocytochemistry. This fingerprint of P2 receptors in human pancreatic duct models forms the basis for studying effect of nucleotides on ion and fluid secretion, as well as on Ca(2+) and tissue homeostasis...... pancreatic duct cell lines PANC-1 and CFPAC-1. Expression of P2 receptors was examined using RT-PCR and immunocytochemistry. Both cell lines, and also Capan-1 cells, express RNA transcripts for the following receptors: P2Y1, P2Y2, P2Y4, P2Y6, P2Y11-14 and P2X1, P2X2, P2X4, P2X5, P2X6 and P2X7. Using Fura-2...... and single-cell imaging we tested effects of various nucleotide analogues on intracellular Ca(2+) signals in PANC-1 and CFPAC-1 cells. The cell lines responded to all nucleotides with the following efficiency: UTP >or= ATP = ATPgammaS > BzATP. ATP, UTP and ATPgammaS elicited oscillatory responses. Bz...

  19. Erythropoietin regulates Treg cells in asthma through TGFβ receptor signaling

    OpenAIRE

    Wan, Guoshi; Wei, Bing

    2015-01-01

    Asthma is a chronic inflammatory disorder of the airways, the development of which is suppressed by regulatory T cells (Treg). Erythropoietin (EPO) is originally defined as a hematopoietic growth factor. Recently, the anti-inflammatory effects of EPO in asthma have been acknowledged. However, the underlying mechanisms remain ill-defined. Here, we showed that EPO treatment significantly reduced the severity of an ovalbumin (OVA)-induced asthma in mice, seemingly through promoting Foxp3-mediate...

  20. Alternate estrogen receptors promote invasion of inflammatory breast cancer cells via non-genomic signaling.

    Directory of Open Access Journals (Sweden)

    Kazufumi Ohshiro

    Full Text Available Although Inflammatory Breast Cancer (IBC is a rare and an aggressive type of locally advanced breast cancer with a generally worst prognosis, little work has been done in identifying the status of non-genomic signaling in the invasiveness of IBC. The present study was performed to explore the status of non-genomic signaling as affected by various estrogenic and anti-estrogenic agents in IBC cell lines SUM149 and SUM190. We have identified the presence of estrogen receptor α (ERα variant, ERα36 in SUM149 and SUM190 cells. This variant as well as ERβ was present in a substantial concentration in IBC cells. The treatment with estradiol (E2, anti-estrogenic agents 4-hydroxytamoxifen and ICI 182780, ERβ specific ligand DPN and GPR30 agonist G1 led to a rapid activation of p-ERK1/2, suggesting the involvement of ERα36, ERβ and GPR30 in the non-genomic signaling pathway in these cells. We also found a substantial increase in the cell migration and invasiveness of SUM149 cells upon the treatment with these ligands. Both basal and ligand-induced migration and invasiveness of SUM149 cells were drastically reduced in the presence of MEK inhibitor U0126, implicating that the phosphorylation of ERK1/2 by MEK is involved in the observed motility and invasiveness of IBC cells. We also provide evidence for the upregulation of p-ERK1/2 through immunostaining in IBC patient samples. These findings suggest a role of non-genomic signaling through the activation of p-ERK1/2 in the hormonal dependence of IBC by a combination of estrogen receptors. These findings only explain the failure of traditional anti-estrogen therapies in ER-positive IBC which induces the non-genomic signaling, but also opens newer avenues for design of modified therapies targeting these estrogen receptors.

  1. Combinatory annotation of cell membrane receptors and signalling pathways of Bombyx mori prothoracic glands

    Science.gov (United States)

    Moulos, Panagiotis; Samiotaki, Martina; Panayotou, George; Dedos, Skarlatos G.

    2016-01-01

    The cells of prothoracic glands (PG) are the main site of synthesis and secretion of ecdysteroids, the biochemical products of cholesterol conversion to steroids that shape the morphogenic development of insects. Despite the availability of genome sequences from several insect species and the extensive knowledge of certain signalling pathways that underpin ecdysteroidogenesis, the spectrum of signalling molecules and ecdysteroidogenic cascades is still not fully comprehensive. To fill this gap and obtain the complete list of cell membrane receptors expressed in PG cells, we used combinatory bioinformatic, proteomic and transcriptomic analysis and quantitative PCR to annotate and determine the expression profiles of genes identified as putative cell membrane receptors of the model insect species, Bombyx mori, and subsequently enrich the repertoire of signalling pathways that are present in its PG cells. The genome annotation dataset we report here highlights modules and pathways that may be directly involved in ecdysteroidogenesis and aims to disseminate data and assist other researchers in the discovery of the role of such receptors and their ligands. PMID:27576083

  2. Single molecule analysis of B cell receptor motion during signaling activation

    Science.gov (United States)

    Rey Suarez, Ivan; Koo, Peter; Mochrie, Simon; Song, Wenxia; Upadhyaya, Arpita

    B cells are an essential part of the adaptive immune system. They patrol the body looking for signs of infection in the form of antigen on the surface of antigen presenting cells. The binding of the B cell receptor (BCR) to antigen induces signaling cascades that lead to B cell activation and eventual production of high affinity antibodies. During activation, BCR organize into signaling microclusters, which are platforms for signal amplification. The physical processes underlying receptor movement and aggregation are not well understood. Here we study the dynamics of single BCRs on activated murine primary B cells using TIRF imaging and single particle tracking. The tracks obtained are analyzed using perturbation expectation-maximization (pEM) a systems-level analysis that allows the identification of different short-time diffusive states from a set of single particle tracks. We identified five different diffusive states on wild type cells, which correspond to different molecular states of the BCR. By using actin polymerization inhibitors and mutant cells lacking important actin regulators we were able to identify the BCR molecule configuration associated with each diffusive state.

  3. Toll-like receptor signaling is functional in immune cells of the endangered Tasmanian devil.

    Science.gov (United States)

    Patchett, Amanda L; Latham, Roger; Brettingham-Moore, Kate H; Tovar, Cesar; Lyons, A Bruce; Woods, Gregory M

    2015-11-01

    Devil facial tumour disease (DFTD) is a fatally transmissible cancer that threatens the Tasmanian devil population. As Tasmanian devils do not produce an immune response against DFTD cells, an effective vaccine will require a strong adjuvant. Activation of innate immune system cells through toll-like receptors (TLRs) could provide this stimulation. It is unknown whether marsupials, including Tasmanian devils, express functional TLRs. We isolated RNA from peripheral blood mononuclear cells and, with PCR, detected transcripts for TLRs 2, 3, 4, 5, 6, 7, 8, 9, 10 and 13. Stimulation of the mononuclear cells with agonists to these TLRs increased the expression of downstream TLR signaling products (IL1α, IL6, IL12A and IFNβ). Our data provide the first evidence that TLR signaling is functional in the mononuclear cells of the Tasmanian devil. Future DFTD vaccination trials will incorporate TLR agonists to enhance the immune response against DFTD. PMID:26182986

  4. Ethanol affects NMDA receptor signaling at climbing fiber-Purkinje cell synapses in mice and impairs cerebellar LTD

    OpenAIRE

    He, Qionger; Titley, Heather; Grasselli, Giorgio; Piochon, Claire; Hansel, Christian

    2012-01-01

    Ethanol profoundly influences cerebellar circuit function and motor control. It has recently been demonstrated that functional N-methyl-d-aspartate (NMDA) receptors are postsynaptically expressed at climbing fiber (CF) to Purkinje cell synapses in the adult cerebellum. Using whole cell patch-clamp recordings from mouse cerebellar slices, we examined whether ethanol can affect NMDA receptor signaling in mature Purkinje cells. NMDA receptor-mediated currents were isolated by bath application of...

  5. Systems model of T cell receptor proximal signaling reveals emergent ultrasensitivity.

    Directory of Open Access Journals (Sweden)

    Himadri Mukhopadhyay

    Full Text Available Receptor phosphorylation is thought to be tightly regulated because phosphorylated receptors initiate signaling cascades leading to cellular activation. The T cell antigen receptor (TCR on the surface of T cells is phosphorylated by the kinase Lck and dephosphorylated by the phosphatase CD45 on multiple immunoreceptor tyrosine-based activation motifs (ITAMs. Intriguingly, Lck sequentially phosphorylates ITAMs and ZAP-70, a cytosolic kinase, binds to phosphorylated ITAMs with differential affinities. The purpose of multiple ITAMs, their sequential phosphorylation, and the differential ZAP-70 affinities are unknown. Here, we use a systems model to show that this signaling architecture produces emergent ultrasensitivity resulting in switch-like responses at the scale of individual TCRs. Importantly, this switch-like response is an emergent property, so that removal of multiple ITAMs, sequential phosphorylation, or differential affinities abolishes the switch. We propose that highly regulated TCR phosphorylation is achieved by an emergent switch-like response and use the systems model to design novel chimeric antigen receptors for therapy.

  6. Perturbed T cell IL7 receptor-signaling in chronic Chagas disease

    Science.gov (United States)

    Albareda, M. Cecilia; Perez-Mazliah, Damián; Natale, M. Ailén; Castro-Eiro, Melisa; Alvarez, María G.; Viotti, Rodolfo; Bertocchi, Graciela; Lococo, Bruno; Tarleton, Rick L; Laucella, Susana A.

    2015-01-01

    We have previously demonstrated that immune responses in subjects with chronic Trypanosoma cruzi infection display features common to other persistent infections with signs of T cell exhaustion. Alterations in cytokine receptor signal transduction have emerged as one of the cell-intrinsic mechanisms of T cell exhaustion. Herein, we performed an analysis of the expression of IL-7R components (CD127 and CD132) on CD4+ and CD8+ T cells, and evaluated IL-7-dependent signaling events in patients at different clinical stages of chronic chagasic heart disease. Subjects with no signs of cardiac disease showed a decrease in CD127+CD132+ cells and a reciprocal gain of CD127-CD132+ in CD8+ and CD4+ T cells compared to either patients exhibiting heart enlargement or uninfected controls. T. cruzi infection, in vitro, was able to stimulate the downregulation of CD127 and the upregulation of CD132 on T cells. IL-7-induced phosphorylation of STAT5 as well as Bcl-2 and CD25 expression were lower in T. cruzi-infected subjects compared with uninfected controls. The serum levels of IL-7 was also increased in chronic chagasic patients. The present study highlights perturbed IL-7/IL-7R T cell signaling through STAT5 as a potential mechanism of T cell exhaustion in chronic T. cruzi infection. PMID:25769928

  7. Prostaglandin Receptor Signaling in Disease

    Directory of Open Access Journals (Sweden)

    Toshiyuki Matsuoka

    2007-01-01

    Full Text Available Prostanoids, consisting of the prostaglandins (PGs and the thromboxanes (TXs, are a group of lipid mediators formed in response to various stimuli. They include PGD2, PGE2, PGF2α, PGI2, and TXA2. They are released outside of the cells immediately after synthesis, and exert their actions by binding to a G-protein coupled rhodopsin-type receptor on the surface of target cells. There are eight types of the prostanoid receptors conserved in mammals from mouse to human. They are the PGD receptor (DP, four subtypes of the PGE receptor (EP1, EP2, EP3, and EP4, the PGF receptor (FP, PGI receptor (IP, and TXA receptor (TP. Recently, mice deficient in each of these prostanoid receptors were generated and subjected to various experimental models of disease. These studies have revealed the roles of PG receptor signaling in various pathological conditions, and suggest that selective manipulation of the prostanoid receptors may be beneficial in treatment of the pathological conditions. Here we review these recent findings of roles of prostanoid receptor signaling and their therapeutic implications.

  8. Dopamine Receptor Signaling in MIN6 β-Cells Revealed by Fluorescence Fluctuation Spectroscopy.

    Science.gov (United States)

    Caldwell, Brittany; Ustione, Alessandro; Piston, David W

    2016-08-01

    Insulin secretion defects are central to the development of type II diabetes mellitus. Glucose stimulation of insulin secretion has been extensively studied, but its regulation by other stimuli such as incretins and neurotransmitters is not as well understood. We investigated the mechanisms underlying the inhibition of insulin secretion by dopamine, which is synthesized in pancreatic β-cells from circulating L-dopa. Previous research has shown that this inhibition is mediated primarily by activation of the dopamine receptor D3 subtype (DRD3), even though both DRD2 and DRD3 are expressed in β-cells. To understand this dichotomy, we investigated the dynamic interactions between the dopamine receptor subtypes and their G-proteins using two-color fluorescence fluctuation spectroscopy (FFS) of mouse MIN6 β-cells. We show that proper membrane localization of exogenous G-proteins depends on both the Gβ and Gγ subunits being overexpressed in the cell. Triple transfections of the dopamine receptor subtype and Gβ and Gγ subunits, each labeled with a different-colored fluorescent protein (FP), yielded plasma membrane expression of all three FPs and permitted an FFS evaluation of interactions between the dopamine receptors and the Gβγ complex. Upon dopamine stimulation, we measured a significant decrease in interactions between DRD3 and the Gβγ complex, which is consistent with receptor activation. In contrast, dopamine stimulation did not cause significant changes in the interactions between DRD2 and the Gβγ complex. These results demonstrate that two-color FFS is a powerful tool for measuring dynamic protein interactions in living cells, and show that preferential DRD3 signaling in β-cells occurs at the level of G-protein release. PMID:27508444

  9. OCA-B integrates B cell antigen receptor-, CD40L- and IL 4-mediated signals for the germinal center pathway of B cell development.

    OpenAIRE

    Qin, X F; Reichlin, A; Luo, Y.; Roeder, R. G.; Nussenzweig, M.C.

    1998-01-01

    Many of the key decisions in lymphocyte differentiation and activation are dependent on integration of antigen receptor and co-receptor signals. Although there is significant understanding of these receptors and their signaling pathways, little is known about the molecular requirements for signal integration at the level of activation of gene expression. Here we show that in primary B cells, expression of the B-cell specific transcription coactivator OCA-B (also known as OBF-1 or Bob-1) is re...

  10. Desipramine inhibits histamine H1 receptor-induced Ca2+ signaling in rat hypothalamic cells.

    Directory of Open Access Journals (Sweden)

    Ji-Ah Kang

    Full Text Available The hypothalamus in the brain is the main center for appetite control and integrates signals from adipose tissue and the gastrointestinal tract. Antidepressants are known to modulate the activities of hypothalamic neurons and affect food intake, but the cellular and molecular mechanisms by which antidepressants modulate hypothalamic function remain unclear. Here we have investigated how hypothalamic neurons respond to treatment with antidepressants, including desipramine and sibutramine. In primary cultured rat hypothalamic cells, desipramine markedly suppressed the elevation of intracellular Ca(2+ evoked by histamine H1 receptor activation. Desipramine also inhibited the histamine-induced Ca(2+ increase and the expression of corticotrophin-releasing hormone in hypothalamic GT1-1 cells. The effect of desipramine was not affected by pretreatment with prazosin or propranolol, excluding catecholamine reuptake activity of desipramine as an underlying mechanism. Sibutramine which is also an antidepressant but decreases food intake, had little effect on the histamine-induced Ca(2+ increase or AMP-activated protein kinase activity. Our results reveal that desipramine and sibutramine have different effects on histamine H1 receptor signaling in hypothalamic cells and suggest that distinct regulation of hypothalamic histamine signaling might underlie the differential regulation of food intake between antidepressants.

  11. CD28 and T cell antigen receptor signal transduction coordinately regulate interleukin 2 gene expression in response to superantigen stimulation

    OpenAIRE

    1992-01-01

    Activation of an immune response requires intercellular contact between T lymphocytes and antigen-presenting cells (APC). Interaction of the T cell antigen receptor (TCR) with antigen in the context of major histocompatibility molecules mediates signal transduction, but T cell activation appears to require the induction of a second costimulatory signal transduction pathway. Recent studies suggest that interaction of CD28 with B7 on APC might deliver such a costimulatory signal. To investigate...

  12. A Novel Nectin-mediated Cell Adhesion Apparatus That Is Implicated in Prolactin Receptor Signaling for Mammary Gland Development.

    Science.gov (United States)

    Kitayama, Midori; Mizutani, Kiyohito; Maruoka, Masahiro; Mandai, Kenji; Sakakibara, Shotaro; Ueda, Yuki; Komori, Takahide; Shimono, Yohei; Takai, Yoshimi

    2016-03-11

    Mammary gland development is induced by the actions of various hormones to form a structure consisting of collecting ducts and milk-secreting alveoli, which comprise two types of epithelial cells known as luminal and basal cells. These cells adhere to each other by cell adhesion apparatuses whose roles in hormone-dependent mammary gland development remain largely unknown. Here we identified a novel cell adhesion apparatus at the boundary between the luminal and basal cells in addition to desmosomes. This apparatus was formed by the trans-interaction between the cell adhesion molecules nectin-4 and nectin-1, which were expressed in the luminal and basal cells, respectively. Nectin-4 of this apparatus further cis-interacted with the prolactin receptor in the luminal cells to enhance the prolactin-induced prolactin receptor signaling for alveolar development with lactogenic differentiation. Thus, a novel nectin-mediated cell adhesion apparatus regulates the prolactin receptor signaling for mammary gland development. PMID:26757815

  13. Role of ERK/MAPK in endothelin receptor signaling in human aortic smooth muscle cells

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    Edvinsson Lars

    2009-07-01

    Full Text Available Abstract Background Endothelin-1 (ET-1 is a potent vasoactive peptide, which induces vasoconstriction and proliferation in vascular smooth muscle cells (VSMCs through activation of endothelin type A (ETA and type B (ETB receptors. The extracellular signal-regulated kinase 1 and 2 (ERK1/2 mitogen-activated protein kinases (MAPK are involved in ET-1-induced VSMC contraction and proliferation. This study was designed to investigate the ETA and ETB receptor intracellular signaling in human VSMCs and used phosphorylation (activation of ERK1/2 as a functional signal molecule for endothelin receptor activity. Results Subconfluent human VSMCs were stimulated by ET-1 at different concentrations (1 nM-1 μM. The activation of ERK1/2 was examined by immunofluorescence, Western blot and phosphoELISA using specific antibody against phosphorylated ERK1/2 protein. ET-1 induced a concentration- and time- dependent activation of ERK1/2 with a maximal effect at 10 min. It declined to baseline level at 30 min. The ET-1-induced activation of ERK1/2 was completely abolished by MEK1/2 inhibitors U0126 and SL327, and partially inhibited by the MEK1 inhibitor PD98059. A dual endothelin receptor antagonist bosentan or the ETA antagonist BQ123 blocked the ET-1 effect, while the ETB antagonist BQ788 had no significant effect. However, a selective ETB receptor agonist, Sarafotoxin 6c (S6c caused a time-dependent ERK1/2 activation with a maximal effect by less than 20% of the ET-1-induced activation of ERK1/2. Increase in bosentan concentration up to 10 μM further inhibited ET-1-induced activation of ERK1/2 and had a stronger inhibitory effect than BQ123 or the combined use of BQ123 and BQ788. To further explore ET-1 intracellular signaling, PKC inhibitors (staurosporin and GF109203X, PKC-delta inhibitor (rottlerin, PKA inhibitor (H-89, and phosphatidylinositol 3-kinase (PI3K inhibitor (wortmannin were applied. The inhibitors showed significant inhibitory effects on ET-1

  14. Modulation of B-cell receptor and microenvironment signaling by a guanine exchange factor in B-cell malignancies

    Institute of Scientific and Technical Information of China (English)

    Wei Liao; Sanjai Sharma

    2016-01-01

    Objective: Chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL) cells over-express a guanine exchange factor (GEF), Rasgrf-1. This GEF increases active Ras as it catalyzes the removal of GDP from Ras so that GTP can bind and activate Ras. This study aims to study the mechanism of action of Rasgrf-1 in B-cell malignancies. Methods: N-terminus truncated Rasgrf-1 variants have a higher GEF activity as compared to the full-length transcript therefore a MCL cell line with stable over-expression of truncated Rasgrf-1 was established. The B-cell receptor (BCR) and chemokine signaling pathways were compared in the Rasgrf-1 over-expressing and a control transfected cell line. Results: Cells over-expressing truncated form of Rasgrf-1 have a higher proliferative rate as compared to control transfected cells. BCR was activated by lower concentrations of anti-IgM antibody in Rasgrf-1 over-expressing cells as compared to control cells indicating that these cells are more sensitive to BCR signaling. BCR signaling also phosphorylates Rasgrf-1 that further increases its GEF function and amplifies BCR signaling. This activation of Rasgrf-1 in over-expressing cells resulted in a higher expression of phospho-ERK, AKT, BTK and PKC-alpha as compared to control cells. Besides BCR, Rasgrf-1 over-expressing cells were also more sensitive to microenvironment stimuli as determined by resistance to apoptosis, chemotaxis and ERK pathway activation. Conclusions: This GEF protein sensitizes B-cells to BCR and chemokine mediated signaling and also upregulates a number of other signaling pathways which promotes growth and survival of these cells.

  15. Signaling through urokinase and urokinase receptor in lung cancer cells requires interactions with beta1 integrins.

    Science.gov (United States)

    Tang, Chi-Hui; Hill, Marla L; Brumwell, Alexis N; Chapman, Harold A; Wei, Ying

    2008-11-15

    The urokinase receptor (uPAR) is upregulated upon tumor cell invasion and correlates with poor lung cancer survival. Although a cis-interaction with integrins has been ascribed to uPAR, whether this interaction alone is critical to urokinase (uPA)- and uPAR-dependent signaling and tumor promotion is unclear. Here we report the functional consequences of point mutations of uPAR (H249A-D262A) that eliminate beta1 integrin interactions but maintain uPA binding, vitronectin attachment and association with alphaV integrins, caveolin and epidermal growth factor receptor. Disruption of uPAR interactions with beta1 integrins recapitulated previously reported findings with beta1-integrin-derived peptides that attenuated matrix-dependent ERK activation, MMP expression and in vitro migration by human lung adenocarcinoma cell lines. The uPAR mutant cells acquired enhanced capacity to adhere to vitronectin via uPAR-alphaVbeta5-integrin, rather than through the uPAR-alpha3beta1-integrin complex and they were unable to initiate uPA signaling to activate ERK, Akt or Stat1. In an orthotopic lung cancer model, uPAR mutant cells exhibited reduced tumor size compared with cells expressing wild-type uPAR. Taken together, the results indicate that uPAR-beta1-integrin interactions are essential to signals induced by integrin matrix ligands or uPA that support lung cancer cell invasion in vitro and progression in vivo. PMID:18940913

  16. Role of ERK/MAPK in endothelin receptor signaling in human aortic smooth muscle cells

    DEFF Research Database (Denmark)

    Chen, Qing-wen; Edvinsson, Lars; Xu, Cang-Bao

    2009-01-01

    muscle cells (VSMCs) through activation of endothelin type A (ETA) and type B (ETB) receptors. The extracellular signal-regulated kinase 1 and 2 (ERK1/2) mitogen-activated protein kinases (MAPK) are involved in ET-1-induced VSMC contraction and proliferation. This study was designed to investigate...... showed significant inhibitory effects on ET-1-induced activation of ERK1/2. However, blockage of L-type Ca2+ channels or calcium/calmodulin-dependent protein kinase II, chelating extracellular Ca2+ or emptying internal Ca2+ stores, did not affect ET-1-induced activation of ERK1/2. CONCLUSION: The ETA...

  17. Reactive oxygen species differentially affect T cell receptor-signaling pathways.

    Science.gov (United States)

    Cemerski, Saso; Cantagrel, Alain; Van Meerwijk, Joost P M; Romagnoli, Paola

    2002-05-31

    Oxidative stress plays an important role in the induction of T lymphocyte hyporesponsiveness observed in several human pathologies including cancer, rheumatoid arthritis, leprosy, and AIDS. To investigate the molecular basis of oxidative stress-induced T cell hyporesponsiveness, we have developed an in vitro system in which T lymphocytes are rendered hyporesponsive by co-culture with oxygen radical-producing activated neutrophils. We have observed a direct correlation between the level of T cell hyporesponsiveness induced and the concentration of reactive oxygen species produced. Moreover, induction of T cell hyporesponsiveness is blocked by addition of N-acetyl cysteine, Mn(III)tetrakis(4-benzoic acid)porphyrin chloride, and catalase, confirming the critical role of oxidative stress in this system. The pattern of tyrosine-phosphorylated proteins was profoundly altered in hyporesponsive as compared with normal T cells. In hyporesponsive T cells, T cell receptor (TCR) ligation no longer induced phospholipase C-gamma1 activation and caused reduced Ca(2+) flux. In contrast, despite increased levels of ERK1/2 phosphorylation, TCR-dependent activation of mitogen-activated protein kinase ERK1/2 was unaltered in hyporesponsive T lymphocytes. A late TCR-signaling event such as caspase 3 activation was as well unaffected in hyporesponsive T lymphocytes. Our data indicate that TCR-signaling pathways are differentially affected by physiological levels of oxidative stress and would suggest that although "hyporesponsive" T cells have lost certain effector functions, they may have maintained or gained others. PMID:11916964

  18. Quantitative Single-Cell Analysis of Signaling Pathways Activated Immediately Downstream of Histamine Receptor Subtypes.

    Science.gov (United States)

    van Unen, Jakobus; Rashidfarrokhi, Ali; Hoogendoorn, Eelco; Postma, Marten; Gadella, Theodorus W J; Goedhart, Joachim

    2016-09-01

    Genetically encoded biosensors based on Förster resonance energy transfer (FRET) can visualize responses of individual cells in real time. Here, we evaluated whether FRET-based biosensors provide sufficient contrast and specificity to measure activity of G-protein-coupled receptors. The four histamine receptor subtypes (H1R, H2R, H3R, and H4R) respond to the ligand histamine by activating three canonical heterotrimeric G-protein-mediated signaling pathways with a reported high degree of specificity. Using FRET-based biosensors, we demonstrate that H1R activates Gαq. We also observed that H1R activates Gαi, albeit at a 10-fold lower potency. In addition to increasing cAMP levels, most likely via Gαs, we found that the H2R induces Gαq-mediated calcium release. The H3R and H4R activated Gαi with high specificity and a high potency. We demonstrate that a number of FRET sensors provide sufficient contrast to: 1) analyze the specificity of the histamine receptor subtypes for different heterotrimeric G-protein families with single-cell resolution, 2) probe for antagonist specificity, and 3) allow the measurement of single-cell concentration-response curves. PMID:27358232

  19. Enhanced airway smooth muscle cell thromboxane receptor signaling via activation of JNK MAPK and extracellular calcium influx

    DEFF Research Database (Denmark)

    Lei, Ying; Cao, Yongxiao; Zhang, Yaping;

    2011-01-01

    airway smooth muscle cells by using an organ culture model and a set of selective pharmacological inhibitors for mitogen-activated protein kinase (MAPK) and calcium signal pathways. Western-blot, immunohistochemistry, myograph and a selective TP receptor agonist U46619 were used for examining TP receptor...... signal proteins and function. Organ culture of rat bronchial segments for up to 48 h induces a time-dependently increased airway contractile response to U46619. This indicates that organ culture increases TP receptor signaling in the airway smooth muscle cells. The enhanced bronchial contraction was...... attenuated by the inhibition of c-Jun N-terminal kinase (JNK) MAPK activity, chelation of extracellular calcium and calcium channel blocker nifedipine, suggesting that JNK MAPK activity and elevated intracellular calcium level are required for the TP receptor signaling. In conclusion, airway smooth muscle...

  20. Tumor-specific T cells signal tumor destruction via the lymphotoxin β receptor

    Directory of Open Access Journals (Sweden)

    Fox Bernard A

    2007-03-01

    Full Text Available Abstract Background Previously, we reported that adoptively transferred perforin k/o (PKO, and IFN-γ k/o (GKO, or perforin/IFN-γ double k/o (PKO/GKO effector T cells mediated regression of B16BL6-D5 (D5 pulmonary metastases and showed that TNF receptor signaling played a critical role in mediating tumor regression. In this report we investigated the role of lymphotoxin-α (LT-α as a potential effector molecules of tumor-specific effector T cells. Methods Effector T cells were generated from tumor vaccine-draining lymph node (TVDLN of wt, GKO, LT-α deficient (LKO, or PKO/GKO mice and tested for their ability to mediate regression of D5 pulmonary metastases in the presence or absence of LT-βR-Fc fusion protein or anti-IFN-γ antibody. Chemokine production by D5 tumor cells was determined by ELISA, RT-PCR and Chemotaxis assays. Results Stimulated effector T cells from wt, GKO, or PKO/GKO mice expressed ligands for LT-β receptor (LT-βR. D5 tumor cells were found to constitutively express the LT-βR. Administration of LT-βR-Fc fusion protein completely abrogated the therapeutic efficacy of GKO or PKO/GKO but not wt effector T cells (p Conclusion The contribution of LT-α expression by effector T cells to anti-tumor activity in vivo was not discernable when wt effector T cells were studied. However, the contribution of LT-β R signaling was identified for GKO or PKO/GKO effector T cells. Since LT-α does not directly induce killing of D5 tumor cells in vitro, but does stimulate D5 tumor cells to secrete chemokines, these data suggest a model where LT-α expression by tumor-specific effector T cells interacts via cross-linking of the LT-βR on tumor cells to induce secretion of chemokines that are chemotactic for macrophages. While the contribution of macrophages to tumor elimination in our system requires additional study, this model provides a possible explanation for the infiltration of inate effector cells that is seen coincident with tumor

  1. Role of AKT/ PKB and 14-3-3 in the regulation of B cell receptor signaling and signalosome assembly

    OpenAIRE

    Mohammad, Dara

    2015-01-01

    AKT/PKB is an oncogenic serine/threonine kinase regulated via the PI3K pathways. 14-3-3s represent a large group of adaptor proteins that are known to interact with a plethora of signaling proteins and regulate diverse signal transduction pathways. The B-cell antigen receptor (BCR) activation and signalosome assembly are dynamic processes controlled by protein phosphorylation. The signaling events share their functions in controlling cell proliferation, differentiation, and/or apoptosis. ...

  2. Signaling mechanism for modulation by ATP of glycine receptors on rat retinal ganglion cells.

    Science.gov (United States)

    Zhang, Ping-Ping; Zhang, Gong; Zhou, Wei; Weng, Shi-Jun; Yang, Xiong-Li; Zhong, Yong-Mei

    2016-01-01

    ATP modulates voltage- and ligand-gated channels in the CNS via the activation of ionotropic P2X and metabotropic P2Y receptors. While P2Y receptors are expressed in retinal neurons, the function of these receptors in the retina is largely unknown. Using whole-cell patch-clamp techniques in rat retinal slice preparations, we demonstrated that ATP suppressed glycine receptor-mediated currents of OFF type ganglion cells (OFF-GCs) dose-dependently, and the effect was in part mediated by P2Y1 and P2Y11, but not by P2X. The ATP effect was abolished by intracellular dialysis of a Gq/11 protein inhibitor and phosphatidylinositol (PI)-phospholipase C (PLC) inhibitor, but not phosphatidylcholine (PC)-PLC inhibitor. The ATP effect was accompanied by an increase in [Ca(2+)]i through the IP3-sensitive pathway and was blocked by intracellular Ca(2+)-free solution. Furthermore, the ATP effect was eliminated in the presence of PKC inhibitors. Neither PKA nor PKG system was involved. These results suggest that the ATP-induced suppression may be mediated by a distinct Gq/11/PI-PLC/IP3/Ca(2+)/PKC signaling pathway, following the activation of P2Y1,11 and other P2Y subtypes. Consistently, ATP suppressed glycine receptor-mediated light-evoked inhibitory postsynaptic currents of OFF-GCs. These results suggest that ATP may modify the ON-to-OFF crossover inhibition, thus changing action potential patterns of OFF-GCs. PMID:27357477

  3. G-protein coupled receptor signaling architecture of mammalian immune cells.

    Directory of Open Access Journals (Sweden)

    Natalia Polouliakh

    Full Text Available A series of recent studies on large-scale networks of signaling and metabolic systems revealed that a certain network structure often called "bow-tie network" are observed. In signaling systems, bow-tie network takes a form with diverse and redundant inputs and outputs connected via a small numbers of core molecules. While arguments have been made that such network architecture enhances robustness and evolvability of biological systems, its functional role at a cellular level remains obscure. A hypothesis was proposed that such a network function as a stimuli-reaction classifier where dynamics of core molecules dictate downstream transcriptional activities, hence physiological responses against stimuli. In this study, we examined whether such hypothesis can be verified using experimental data from Alliance for Cellular Signaling (AfCS that comprehensively measured GPCR related ligands response for B-cell and macrophage. In a GPCR signaling system, cAMP and Ca2+ act as core molecules. Stimuli-response for 32 ligands to B-Cells and 23 ligands to macrophages has been measured. We found that ligands with correlated changes of cAMP and Ca2+ tend to cluster closely together within the hyperspaces of both cell types and they induced genes involved in the same cellular processes. It was found that ligands inducing cAMP synthesis activate genes involved in cell growth and proliferation; cAMP and Ca2+ molecules that increased together form a feedback loop and induce immune cells to migrate and adhere together. In contrast, ligands without a core molecules response are scattered throughout the hyperspace and do not share clusters. G-protein coupling receptors together with immune response specific receptors were found in cAMP and Ca2+ activated clusters. Analyses have been done on the original software applicable for discovering 'bow-tie' network architectures within the complex network of intracellular signaling where ab initio clustering has been

  4. Epidermal growth factor receptor signalling in human breast cancer cells operates parallel to estrogen receptor α signalling and results in tamoxifen insensitive proliferation

    International Nuclear Information System (INIS)

    Tamoxifen resistance is a major problem in the treatment of estrogen receptor (ER) α -positive breast cancer patients. Although the mechanisms behind tamoxifen resistance are still not completely understood, clinical data suggests that increased expression of receptor tyrosine kinases is involved. Here, we studied the estrogen and anti-estrogen sensitivity of human breast cancer MCF7 cells that have a moderate, retroviral-mediated, ectopic expression of epidermal growth factor receptor (MCF7-EGFR). Proliferation of MCF7-EGFR and parental cells was induced by 17β-estradiol (E2), epidermal growth factor (EGF) or a combination of these. Inhibition of proliferation under these conditions was investigated with 4-hydroxy-tamoxifen (TAM) or fulvestrant at 10-12 to 10-6 M. Cells were lysed at different time points to determine the phosphorylation status of EGFR, MAPK1/3, AKT and the expression of ERα. Knockdown of target genes was established using smartpool siRNAs. Transcriptomics analysis was done 6 hr after stimulation with growth factors using Affymetrix HG-U133 PM array plates. While proliferation of parental MCF7 cells could only be induced by E2, proliferation of MCF7-EGFR cells could be induced by either E2 or EGF. Treatment with TAM or fulvestrant did significantly inhibit proliferation of MCF7-EGFR cells stimulated with E2 alone. EGF treatment of E2/TAM treated cells led to a marked cell proliferation thereby overruling the anti-estrogen-mediated inhibition of cell proliferation. Under these conditions, TAM however did still inhibit ERα- mediated transcription. While siRNA-mediated knock-down of EGFR inhibited the EGF- driven proliferation under TAM/E2/EGF condition, knock down of ERα did not. The TAM resistant cell proliferation mediated by the conditional EGFR-signaling may be dependent on the PI3K/Akt pathway but not the MEK/MAPK pathway, since a MEK inhibitor (U0126), did not block the proliferation. Transcriptomic analysis under the various E2/TAM

  5. Diverse signaling systems activated by the sweet taste receptor in human GLP-1-secreting cells.

    Science.gov (United States)

    Ohtsu, Yoshiaki; Nakagawa, Yuko; Nagasawa, Masahiro; Takeda, Shigeki; Arakawa, Hirokazu; Kojima, Itaru

    2014-08-25

    Sweet taste receptor regulates GLP-1 secretion in enteroendocrine L-cells. We investigated the signaling system activated by this receptor using Hutu-80 cells. We stimulated them with sucralose, saccharin, acesulfame K and glycyrrhizin. These sweeteners stimulated GLP-1 secretion, which was attenuated by lactisole. All these sweeteners elevated cytoplasmic cyclic AMP ([cAMP]c) whereas only sucralose and saccharin induced a monophasic increase in cytoplasmic Ca(2+) ([Ca(2+)]c). Removal of extracellular calcium or sodium and addition of a Gq/11 inhibitor greatly reduced the [Ca(2+)]c responses to two sweeteners. In contrast, acesulfame K induced rapid and sustained reduction of [Ca(2+)]c. In addition, glycyrrhizin first reduced [Ca(2+)]c which was followed by an elevation of [Ca(2+)]c. Reductions of [Ca(2+)]c induced by acesulfame K and glycyrrhizin were attenuated by a calmodulin inhibitor or by knockdown of the plasma membrane calcium pump. These results indicate that various sweet molecules act as biased agonists and evoke strikingly different patterns of intracellular signals. PMID:25017733

  6. Piperlongumine inhibits atherosclerotic plaque formation and vascular smooth muscle cell proliferation by suppressing PDGF receptor signaling

    Energy Technology Data Exchange (ETDEWEB)

    Son, Dong Ju [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Kim, Soo Yeon [Division of Life Science, Korea Basic Science Institute, Daejeon (Korea, Republic of); Han, Seong Su [University of Iowa Carver College of Medicine, Department of Pathology, Iowa City, IA (United States); Kim, Chan Woo [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Kumar, Sandeep [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Park, Byeoung Soo [Nanotoxtech Co., Ansan (Korea, Republic of); Lee, Sung Eun [Division of Applied Biology and Chemistry, Kyungpook National University, Daegu (Korea, Republic of); Yun, Yeo Pyo [College of Pharmacy, Chungbuk National University, Cheongju (Korea, Republic of); Jo, Hanjoong, E-mail: hjo@emory.edu [Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA (United States); Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA (United States); Department of Bioinspired Science, Ehwa Womans University, Seoul (Korea, Republic of); Park, Young Hyun, E-mail: pyh012@sch.ac.kr [Department of Food Science and Nutrition, College of Natural Sciences, Soonchunhyang University, Asan (Korea, Republic of)

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer Anti-atherogenic effect of PL was examined using partial carotid ligation model in ApoE KO mice. Black-Right-Pointing-Pointer PL prevented atherosclerotic plaque development, VSMCs proliferation, and NF-{kappa}B activation. Black-Right-Pointing-Pointer Piperlongumine reduced vascular smooth muscle cell activation through PDGF-R{beta} and NF-{kappa}B-signaling. Black-Right-Pointing-Pointer PL may serve as a new therapeutic molecule for atherosclerosis treatment. -- Abstract: Piperlongumine (piplartine, PL) is an alkaloid found in the long pepper (Piper longum L.) and has well-documented anti-platelet aggregation, anti-inflammatory, and anti-cancer properties; however, the role of PL in prevention of atherosclerosis is unknown. We evaluated the anti-atherosclerotic potential of PL in an in vivo murine model of accelerated atherosclerosis and defined its mechanism of action in aortic vascular smooth muscle cells (VSMCs) in vitro. Local treatment with PL significantly reduced atherosclerotic plaque formation as well as proliferation and nuclear factor-kappa B (NF-{kappa}B) activation in an in vivo setting. PL treatment in VSMCs in vitro showed inhibition of migration and platelet-derived growth factor BB (PDGF-BB)-induced proliferation to the in vivo findings. We further identified that PL inhibited PDGF-BB-induced PDGF receptor beta activation and suppressed downstream signaling molecules such as phospholipase C{gamma}1, extracellular signal-regulated kinases 1 and 2 and Akt. Lastly, PL significantly attenuated activation of NF-{kappa}B-a downstream transcriptional regulator in PDGF receptor signaling, in response to PDGF-BB stimulation. In conclusion, our findings demonstrate a novel, therapeutic mechanism by which PL suppresses atherosclerosis plaque formation in vivo.

  7. Piperlongumine inhibits atherosclerotic plaque formation and vascular smooth muscle cell proliferation by suppressing PDGF receptor signaling

    International Nuclear Information System (INIS)

    Highlights: ► Anti-atherogenic effect of PL was examined using partial carotid ligation model in ApoE KO mice. ► PL prevented atherosclerotic plaque development, VSMCs proliferation, and NF-κB activation. ► Piperlongumine reduced vascular smooth muscle cell activation through PDGF-Rβ and NF-κB-signaling. ► PL may serve as a new therapeutic molecule for atherosclerosis treatment. -- Abstract: Piperlongumine (piplartine, PL) is an alkaloid found in the long pepper (Piper longum L.) and has well-documented anti-platelet aggregation, anti-inflammatory, and anti-cancer properties; however, the role of PL in prevention of atherosclerosis is unknown. We evaluated the anti-atherosclerotic potential of PL in an in vivo murine model of accelerated atherosclerosis and defined its mechanism of action in aortic vascular smooth muscle cells (VSMCs) in vitro. Local treatment with PL significantly reduced atherosclerotic plaque formation as well as proliferation and nuclear factor-kappa B (NF-κB) activation in an in vivo setting. PL treatment in VSMCs in vitro showed inhibition of migration and platelet-derived growth factor BB (PDGF-BB)-induced proliferation to the in vivo findings. We further identified that PL inhibited PDGF-BB-induced PDGF receptor beta activation and suppressed downstream signaling molecules such as phospholipase Cγ1, extracellular signal-regulated kinases 1 and 2 and Akt. Lastly, PL significantly attenuated activation of NF-κB—a downstream transcriptional regulator in PDGF receptor signaling, in response to PDGF-BB stimulation. In conclusion, our findings demonstrate a novel, therapeutic mechanism by which PL suppresses atherosclerosis plaque formation in vivo.

  8. Microenvironmental stiffness enhances glioma cell proliferation by stimulating epidermal growth factor receptor signaling.

    Directory of Open Access Journals (Sweden)

    Vaibhavi Umesh

    Full Text Available The aggressive and rapidly lethal brain tumor glioblastoma (GBM is associated with profound tissue stiffening and genomic lesions in key members of the epidermal growth factor receptor (EGFR pathway. Previous studies from our laboratory have shown that increasing microenvironmental stiffness in culture can strongly enhance glioma cell behaviors relevant to tumor progression, including proliferation, yet it has remained unclear whether stiffness and EGFR regulate proliferation through common or independent signaling mechanisms. Here we test the hypothesis that microenvironmental stiffness regulates cell cycle progression and proliferation in GBM tumor cells by altering EGFR-dependent signaling. We began by performing an unbiased reverse phase protein array screen, which revealed that stiffness modulates expression and phosphorylation of a broad range of signals relevant to proliferation, including members of the EGFR pathway. We subsequently found that culturing human GBM tumor cells on progressively stiffer culture substrates both dramatically increases proliferation and facilitates passage through the G1/S checkpoint of the cell cycle, consistent with an EGFR-dependent process. Western Blots showed that increasing microenvironmental stiffness enhances the expression and phosphorylation of EGFR and its downstream effector Akt. Pharmacological loss-of-function studies revealed that the stiffness-sensitivity of proliferation is strongly blunted by inhibition of EGFR, Akt, or PI3 kinase. Finally, we observed that stiffness strongly regulates EGFR clustering, with phosphorylated EGFR condensing into vinculin-positive focal adhesions on stiff substrates and dispersing as microenvironmental stiffness falls to physiological levels. Our findings collectively support a model in which tissue stiffening promotes GBM proliferation by spatially and biochemically amplifying EGFR signaling.

  9. Microenvironmental stiffness enhances glioma cell proliferation by stimulating epidermal growth factor receptor signaling.

    Science.gov (United States)

    Umesh, Vaibhavi; Rape, Andrew D; Ulrich, Theresa A; Kumar, Sanjay

    2014-01-01

    The aggressive and rapidly lethal brain tumor glioblastoma (GBM) is associated with profound tissue stiffening and genomic lesions in key members of the epidermal growth factor receptor (EGFR) pathway. Previous studies from our laboratory have shown that increasing microenvironmental stiffness in culture can strongly enhance glioma cell behaviors relevant to tumor progression, including proliferation, yet it has remained unclear whether stiffness and EGFR regulate proliferation through common or independent signaling mechanisms. Here we test the hypothesis that microenvironmental stiffness regulates cell cycle progression and proliferation in GBM tumor cells by altering EGFR-dependent signaling. We began by performing an unbiased reverse phase protein array screen, which revealed that stiffness modulates expression and phosphorylation of a broad range of signals relevant to proliferation, including members of the EGFR pathway. We subsequently found that culturing human GBM tumor cells on progressively stiffer culture substrates both dramatically increases proliferation and facilitates passage through the G1/S checkpoint of the cell cycle, consistent with an EGFR-dependent process. Western Blots showed that increasing microenvironmental stiffness enhances the expression and phosphorylation of EGFR and its downstream effector Akt. Pharmacological loss-of-function studies revealed that the stiffness-sensitivity of proliferation is strongly blunted by inhibition of EGFR, Akt, or PI3 kinase. Finally, we observed that stiffness strongly regulates EGFR clustering, with phosphorylated EGFR condensing into vinculin-positive focal adhesions on stiff substrates and dispersing as microenvironmental stiffness falls to physiological levels. Our findings collectively support a model in which tissue stiffening promotes GBM proliferation by spatially and biochemically amplifying EGFR signaling. PMID:25000176

  10. Elucidation of tonic and activated B-cell receptor signaling in Burkitt's lymphoma provides insights into regulation of cell survival.

    Science.gov (United States)

    Corso, Jasmin; Pan, Kuan-Ting; Walter, Roland; Doebele, Carmen; Mohr, Sebastian; Bohnenberger, Hanibal; Ströbel, Philipp; Lenz, Christof; Slabicki, Mikolaj; Hüllein, Jennifer; Comoglio, Federico; Rieger, Michael A; Zenz, Thorsten; Wienands, Jürgen; Engelke, Michael; Serve, Hubert; Urlaub, Henning; Oellerich, Thomas

    2016-05-17

    Burkitt's lymphoma (BL) is a highly proliferative B-cell neoplasm and is treated with intensive chemotherapy that, because of its toxicity, is often not suitable for the elderly or for patients with endemic BL in developing countries. BL cell survival relies on signals transduced by B-cell antigen receptors (BCRs). However, tonic as well as activated BCR signaling networks and their relevance for targeted therapies in BL remain elusive. We have systematically characterized and compared tonic and activated BCR signaling in BL by quantitative phosphoproteomics to identify novel BCR effectors and potential drug targets. We identified and quantified ∼16,000 phospho-sites in BL cells. Among these sites, 909 were related to tonic BCR signaling, whereas 984 phospho-sites were regulated upon BCR engagement. The majority of the identified BCR signaling effectors have not been described in the context of B cells or lymphomas yet. Most of these newly identified BCR effectors are predicted to be involved in the regulation of kinases, transcription, and cytoskeleton dynamics. Although tonic and activated BCR signaling shared a considerable number of effector proteins, we identified distinct phosphorylation events in tonic BCR signaling. We investigated the functional relevance of some newly identified BCR effectors and show that ACTN4 and ARFGEF2, which have been described as regulators of membrane-trafficking and cytoskeleton-related processes, respectively, are crucial for BL cell survival. Thus, this study provides a comprehensive dataset for tonic and activated BCR signaling and identifies effector proteins that may be relevant for BL cell survival and thus may help to develop new BL treatments. PMID:27155012

  11. Suppression of sustained and transient ON signals of amacrine cells by GABA is mediated by different receptor subtypes

    Institute of Scientific and Technical Information of China (English)

    张道启; 杨如; 杨雄里

    1999-01-01

    Intracellular recordings were made from amacrine cells in the isolated, superfused carp retina, and the effects of γ-aminobutyric acid (GABA) on sustained and transient ON signals of these cells were studied. Exogenous GABA application partially suppressed the sustained response of ON amacrine cells, which could be completely reversed by picrotoxin (PTX), a chloride channel blocker, and by bicuculline (BCC), a specific GABA_A receptor antagonist. On the other hand, suppression by GABA of the ON response which was predominantly driven by rod signals in a certain portion of transient ON-OFF amacrine cells was completely blocked by PTX, but not by BCC, indicating that GABA_C receptors may be involved in the effect. These results suggest that GABA_A and GABA_C receptors may be respectively involved in mediating the transmission of sustained and transient signals in the carp inner retina.

  12. Signalling mechanism for somatostatin receptor 5-mediated suppression of AMPA responses in rat retinal ganglion cells.

    Science.gov (United States)

    Deng, Qin-Qin; Sheng, Wen-Long; Zhang, Gong; Weng, Shi-Jun; Yang, Xiong-Li; Zhong, Yong-Mei

    2016-08-01

    Somatostatin (SRIF) is involved in a variety of physiological functions via the activation of five subtypes of specific receptors (sst1-5). Here, we investigated the effects of SRIF on AMPA receptor (AMPAR)-mediated currents (AMPA currents) in isolated rat retinal ganglion cells (GCs) using patch-clamp techniques. Immunofluorescence double labelling demonstrated the expression of sst5 in rat GCs. Consistent to this, whole cell AMPA currents of GCs were dose-dependently suppressed by SRIF, and the effect was reversed by the sst5 antagonist BIM-23056. Intracellular dialysis of GDP-β-S or pre-incubation with the Gi/o inhibitor pertussis toxin (PTX) abolished the SRIF effect. The SRIF effect was mimicked by the administration of either 8-Br-cAMP or forskolin, but was eliminated by the protein kinase A (PKA) antagonists H-89/KT5720/Rp-cAMP. Moreover, SRIF increased intracellular Ca(2+) levels and did not suppress the AMPA currents when GCs were infused with an intracellular Ca(2+)-free solution or in the presence of ryanodine receptor modulators caffeine/ryanodine. Furthermore, the SRIF effect was eliminated when the activity of calmodulin (CaM), calcineurin and protein phosphatase 1 (PP1) was blocked with W-7, FK-506 and okadaic acid, respectively. SRIF persisted to suppress the AMPA currents when cGMP-protein kinase G (PKG) and phosphatidylinositol (PI)-/phosphatidylcholine (PC)-phospholipase C (PLC) signalling pathways were blocked. In rat flat-mount retinas, SRIF suppressed AMPAR-mediated light-evoked excitatory postsynaptic currents (L-EPSCs) in GCs. We conclude that a distinct Gi/o/cAMP-PKA/ryanodine/Ca(2+)/CaM/calcineurin/PP1 signalling pathway comes into play due to the activation of sst5 to mediate the SRIF effect on GCs. PMID:26969240

  13. Genomics of signaling crosstalk of estrogen receptor alpha in breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Peter Dudek

    Full Text Available BACKGROUND: The estrogen receptor alpha (ERalpha is a ligand-regulated transcription factor. However, a wide variety of other extracellular signals can activate ERalpha in the absence of estrogen. The impact of these alternate modes of activation on gene expression profiles has not been characterized. METHODOLOGY/PRINCIPAL FINDINGS: We show that estrogen, growth factors and cAMP elicit surprisingly distinct ERalpha-dependent transcriptional responses in human MCF7 breast cancer cells. In response to growth factors and cAMP, ERalpha primarily activates and represses genes, respectively. The combined treatments with the anti-estrogen tamoxifen and cAMP or growth factors regulate yet other sets of genes. In many cases, tamoxifen is perverted to an agonist, potentially mimicking what is happening in certain tamoxifen-resistant breast tumors and emphasizing the importance of the cellular signaling environment. Using a computational analysis, we predicted that a Hox protein might be involved in mediating such combinatorial effects, and then confirmed it experimentally. Although both tamoxifen and cAMP block the proliferation of MCF7 cells, their combined application stimulates it, and this can be blocked with a dominant-negative Hox mutant. CONCLUSIONS/SIGNIFICANCE: The activating signal dictates both target gene selection and regulation by ERalpha, and this has consequences on global gene expression patterns that may be relevant to understanding the progression of ERalpha-dependent carcinomas.

  14. Protease activated receptor signaling is required for African trypanosome traversal of human brain microvascular endothelial cells.

    Directory of Open Access Journals (Sweden)

    Dennis J Grab

    Full Text Available BACKGROUND: Using human brain microvascular endothelial cells (HBMECs as an in vitro model for how African trypanosomes cross the human blood-brain barrier (BBB we recently reported that the parasites cross the BBB by generating calcium activation signals in HBMECs through the activity of parasite cysteine proteases, particularly cathepsin L (brucipain. In the current study, we examined the possible role of a class of protease stimulated HBMEC G protein coupled receptors (GPCRs known as protease activated receptors (PARs that might be implicated in calcium signaling by African trypanosomes. METHODOLOGY/PRINCIPAL FINDINGS: Using RNA interference (RNAi we found that in vitro PAR-2 gene (F2RL1 expression in HBMEC monolayers could be reduced by over 95%. We also found that the ability of Trypanosoma brucei rhodesiense to cross F2RL1-silenced HBMEC monolayers was reduced (39%-49% and that HBMECs silenced for F2RL1 maintained control levels of barrier function in the presence of the parasite. Consistent with the role of PAR-2, we found that HBMEC barrier function was also maintained after blockade of Galpha(q with Pasteurella multocida toxin (PMT. PAR-2 signaling has been shown in other systems to have neuroinflammatory and neuroprotective roles and our data implicate a role for proteases (i.e. brucipain and PAR-2 in African trypanosome/HBMEC interactions. Using gene-profiling methods to interrogate candidate HBMEC pathways specifically triggered by brucipain, several pathways that potentially link some pathophysiologic processes associated with CNS HAT were identified. CONCLUSIONS/SIGNIFICANCE: Together, the data support a role, in part, for GPCRs as molecular targets for parasite proteases that lead to the activation of Galpha(q-mediated calcium signaling. The consequence of these events is predicted to be increased permeability of the BBB to parasite transmigration and the initiation of neuroinflammation, events precursory to CNS disease.

  15. T Cell Receptor-Proximal Signals Are Sustained in Peripheral Microclusters and Terminated in the Central Supramolecular Activation Cluster

    OpenAIRE

    Varma, Rajat; Campi, Gabriele; Yokosuka, Tadashi; Saito, Takashi; Dustin, Michael L

    2006-01-01

    T cell receptor (TCR) signaling is initiated and sustained in microclusters; however, it’s not known whether signaling also occurs in the TCR-rich central supramolecular activation cluster (cSMAC). We showed that the cSMAC formed by fusion of microclusters contained more CD45 than microclusters and is a site enriched in lysobisphosphatidic acid, a lipid involved in sorting ubiquitinated membrane proteins for degradation. Calcium signaling via TCR was blocked within 2 min by anti-MHCp treatmen...

  16. G protein-coupled receptor 30 ligand G-1 increases aryl hydrocarbon receptor signalling by inhibition of tubulin assembly and cell cycle arrest in human MCF-7 cells.

    Science.gov (United States)

    Tarnow, Patrick; Tralau, Tewes; Luch, Andreas

    2016-08-01

    Regulatory crosstalk between the aryl hydrocarbon receptor (AHR) and oestrogen receptor α (ERα) is well established. Apart from the nuclear receptors ERα and ERβ, oestrogen signalling further involves an unrelated G protein-coupled receptor termed GPR30. In order to investigate potential regulatory crosstalk, this study investigated the influence of G-1 as one of the few GPR30-specific ligands on the AHR regulon in MCF-7 cells. As a well-characterised model system, these human mammary carcinoma cells co-express all three receptors (AHR, ERα and GPR30) and are thus ideally suited to study corresponding regulatory pathway interactions on transcript level. Indeed, treatment with micromolar concentrations of the GPR30-specific agonist G-1 resulted in up-regulation of AHR as well as the transcripts for cytochromes P450 1A1 and 1B1, two well-known targets of the AHR regulon. While this was partly attributable to G-1-mediated inhibition of tubulin assembly and subsequent cell cycle arrest in the G2/M phase, the effects nevertheless required functional AHR. However, G-1-induced up-regulation of CYP 1A1 was not mediated by GPR30, as G15 antagonist treatment as well as a knockdown of GPR30 and AHR failed to inhibit this effect. PMID:26475489

  17. Δ9-Tetrahydrocannabinol enhances MCF-7 cell proliferation via cannabinoid receptor-independent signaling

    International Nuclear Information System (INIS)

    We recently reported that Δ9-tetrahydrocannabinol (Δ9-THC) has the ability to stimulate the proliferation of human breast carcinoma MCF-7 cells. However, the mechanism of action remains to be clarified. The present study focused on the relationship between receptor expression and the effects of Δ9-THC on cell proliferation. RT-PCR analysis demonstrated that there was no detectable expression of CB receptors in MCF-7 cells. In accordance with this, no effects of cannabinoid 1/2 (CB1/2) receptor antagonists and pertussis toxin on cell proliferation were observed. Although MCF-7 cell proliferation is suggested to be suppressed by Δ9-THC in the presence of CB receptors, it was revealed that Δ9-THC could exert upregulation of living cells in the absence of the receptors. Interestingly, Δ9-THC upregulated human epithelial growth factor receptor type 2 (HER2) expression, which is known to be a predictive factor of human breast cancer and is able to stimulate cancer cells as well as MCF-7 cells. Actinomycin D-treatment interfered with the upregulation of HER2 and cell proliferation by cannabinoid. Taken together, these studies suggest that, in the absence of CB receptors, Δ9-THC can stimulate the proliferation of MCF-7 cells by modulating, at least in part, HER2 transcription

  18. Olfactory receptor signaling.

    Science.gov (United States)

    Antunes, Gabriela; Simoes de Souza, Fabio Marques

    2016-01-01

    The guanine nucleotide protein (G protein)-coupled receptors (GPCRs) superfamily represents the largest class of membrane protein in the human genome. More than a half of all GPCRs are dedicated to interact with odorants and are termed odorant-receptors (ORs). Linda Buck and Richard Axel, the Nobel Prize laureates in physiology or medicine in 2004, first cloned and characterized the gene family that encode ORs, establishing the foundations to the understanding of the molecular basis for odor recognition. In the last decades, a lot of progress has been done to unravel the functioning of the sense of smell. This chapter gives a general overview of the topic of olfactory receptor signaling and reviews recent advances in this field. PMID:26928542

  19. Delineation of the GPRC6A Receptor Signaling Pathways Using a Mammalian Cell Line Stably Expressing the Receptor

    DEFF Research Database (Denmark)

    Jacobsen, Stine Engesgaard; Nørskov-Lauritsen, Lenea; Thomsen, Alex Rojas Bie;

    2013-01-01

    of the stable CHO cell line with robust receptor responsiveness and optimization of the highly sensitive homogeneous time resolved fluorescence technology allow fast assessment of Gq activation without previous manipulations like cotransfection of mutated G proteins. This cell-based assay system for...... GPRC6A is thus useful in high-throughput screening for novel pharmacological tool compounds, which are necessary to unravel the physiologic function of the receptor....

  20. A Computational Study of the Effects of Syk Activity on B Cell Receptor Signaling Dynamics

    Directory of Open Access Journals (Sweden)

    Reginald L. McGee

    2015-02-01

    Full Text Available The kinase Syk is intricately involved in early signaling events in B cells and isrequired for proper response when antigens bind to B cell receptors (BCRs. Experimentsusing an analog-sensitive version of Syk (Syk-AQL have better elucidated its role, buthave not completely characterized its behavior. We present a computational model for BCRsignaling, using dynamical systems, which incorporates both wild-type Syk and Syk-AQL.Following the use of sensitivity analysis to identify significant reaction parameters, we screenfor parameter vectors that produced graded responses to BCR stimulation as is observedexperimentally. We demonstrate qualitative agreement between the model and dose responsedata for both mutant and wild-type kinases. Analysis of our model suggests that the level of NF-KB activation, which is reduced in Syk-AQL cells relative to wild-type, is more sensitiveto small reductions in kinase activity than Erkp activation, which is essentially unchanged.Since this profile of high Erkp and reduced NF-KB is consistent with anergy, this implies thatanergy is particularly sensitive to small changes in catalytic activity. Also, under a range offorward and reverse ligand binding rates, our model of Erkp and NF-KB activation displaysa dependence on a power law affinity: the ratio of the forward rate to a non-unit power of thereverse rate. This dependence implies that B cells may respond to certain details of bindingand unbinding rates for ligands rather than simple affinity alone.

  1. EGF-Receptor-Mediated Mammary Epithelial Cell Migration is Driven by Sustained ERK Signaling from Autocrine Stimulation

    Energy Technology Data Exchange (ETDEWEB)

    Joslin, Elizabeth J.; Opresko, Lee; Wells, Alan; Wiley, H. S.; Lauffenburger, Douglas A.

    2007-10-15

    Aberrant expression of epidermal growth factor (EGF) receptor family ligands, as well as the receptors themselves, has been implicated in various types of cancers. EGF family ligands are synthesized as membrane-anchored proteins requiring proteolytic release to form the mature soluble factor. Despite the pathophysiological importance of autocrine systems, how the rate of protease-mediated ligand release quantitatively influences receptor-mediated signaling and consequent cell behavior is poorly understood. Therefore, we explored the relationship between autocrine EGF release rates and receptor-mediated ERK activation and migration in human mammary epithelial cells. A quantitative spectrum of EGF release rates was achieved using a set of chimeric transmembrane EGF ligand precursors modulated by the addition of the metalloprotease inhibitor batimastat. We found that ERK activation increased with increasing ligand release rates despite concomitant EGF receptor downregulation. Cell migration speed depended linearly on the steady-state phospho-ERK level obtained from either autocrine or exogenous ligand, but was much greater at any given phospho-ERK level for autocrine compared to exogenous stimulation. In contrast, cell proliferation rates were relatively constant across the various treatment conditions. Thus, in these cells, ERK-mediated migration stimulated by EGF receptor signaling is most sensitively regulated by autocrine ligand control mechanisms.

  2. FGF-receptor signalling controls neural cell diversity in the zebrafish hindbrain by regulating olig2 and sox9.

    Science.gov (United States)

    Esain, Virginie; Postlethwait, John H; Charnay, Patrick; Ghislain, Julien

    2010-01-01

    The mechanisms underlying the generation of neural cell diversity are the subject of intense investigation, which has highlighted the involvement of different signalling molecules including Shh, BMP and Wnt. By contrast, relatively little is known about FGF in this process. In this report we identify an FGF-receptor-dependent pathway in zebrafish hindbrain neural progenitors that give rise to somatic motoneurons, oligodendrocyte progenitors and differentiating astroglia. Using a combination of chemical and genetic approaches to conditionally inactivate FGF-receptor signalling, we investigate the role of this pathway. We show that FGF-receptor signalling is not essential for the survival or maintenance of hindbrain neural progenitors but controls their fate by coordinately regulating key transcription factors. First, by cooperating with Shh, FGF-receptor signalling controls the expression of olig2, a patterning gene essential for the specification of somatic motoneurons and oligodendrocytes. Second, FGF-receptor signalling controls the development of both oligodendrocyte progenitors and astroglia through the regulation of sox9, a gliogenic transcription factor the function of which we show to be conserved in the zebrafish hindbrain. Overall, for the first time in vivo, our results reveal a mechanism of FGF in the control of neural cell diversity. PMID:20023158

  3. Identification of Cell Type-Specific Differences in Erythropoietin Receptor Signaling in Primary Erythroid and Lung Cancer Cells

    Science.gov (United States)

    Salopiata, Florian; Depner, Sofia; Wäsch, Marvin; Böhm, Martin E.; Mücke, Oliver; Plass, Christoph; Lehmann, Wolf D.; Kreutz, Clemens; Timmer, Jens; Klingmüller, Ursula

    2016-01-01

    Lung cancer, with its most prevalent form non-small-cell lung carcinoma (NSCLC), is one of the leading causes of cancer-related deaths worldwide, and is commonly treated with chemotherapeutic drugs such as cisplatin. Lung cancer patients frequently suffer from chemotherapy-induced anemia, which can be treated with erythropoietin (EPO). However, studies have indicated that EPO not only promotes erythropoiesis in hematopoietic cells, but may also enhance survival of NSCLC cells. Here, we verified that the NSCLC cell line H838 expresses functional erythropoietin receptors (EPOR) and that treatment with EPO reduces cisplatin-induced apoptosis. To pinpoint differences in EPO-induced survival signaling in erythroid progenitor cells (CFU-E, colony forming unit-erythroid) and H838 cells, we combined mathematical modeling with a method for feature selection, the L1 regularization. Utilizing an example model and simulated data, we demonstrated that this approach enables the accurate identification and quantification of cell type-specific parameters. We applied our strategy to quantitative time-resolved data of EPO-induced JAK/STAT signaling generated by quantitative immunoblotting, mass spectrometry and quantitative real-time PCR (qRT-PCR) in CFU-E and H838 cells as well as H838 cells overexpressing human EPOR (H838-HA-hEPOR). The established parsimonious mathematical model was able to simultaneously describe the data sets of CFU-E, H838 and H838-HA-hEPOR cells. Seven cell type-specific parameters were identified that included for example parameters for nuclear translocation of STAT5 and target gene induction. Cell type-specific differences in target gene induction were experimentally validated by qRT-PCR experiments. The systematic identification of pathway differences and sensitivities of EPOR signaling in CFU-E and H838 cells revealed potential targets for intervention to selectively inhibit EPO-induced signaling in the tumor cells but leave the responses in erythroid

  4. Identification of Cell Type-Specific Differences in Erythropoietin Receptor Signaling in Primary Erythroid and Lung Cancer Cells.

    Science.gov (United States)

    Merkle, Ruth; Steiert, Bernhard; Salopiata, Florian; Depner, Sofia; Raue, Andreas; Iwamoto, Nao; Schelker, Max; Hass, Helge; Wäsch, Marvin; Böhm, Martin E; Mücke, Oliver; Lipka, Daniel B; Plass, Christoph; Lehmann, Wolf D; Kreutz, Clemens; Timmer, Jens; Schilling, Marcel; Klingmüller, Ursula

    2016-08-01

    Lung cancer, with its most prevalent form non-small-cell lung carcinoma (NSCLC), is one of the leading causes of cancer-related deaths worldwide, and is commonly treated with chemotherapeutic drugs such as cisplatin. Lung cancer patients frequently suffer from chemotherapy-induced anemia, which can be treated with erythropoietin (EPO). However, studies have indicated that EPO not only promotes erythropoiesis in hematopoietic cells, but may also enhance survival of NSCLC cells. Here, we verified that the NSCLC cell line H838 expresses functional erythropoietin receptors (EPOR) and that treatment with EPO reduces cisplatin-induced apoptosis. To pinpoint differences in EPO-induced survival signaling in erythroid progenitor cells (CFU-E, colony forming unit-erythroid) and H838 cells, we combined mathematical modeling with a method for feature selection, the L1 regularization. Utilizing an example model and simulated data, we demonstrated that this approach enables the accurate identification and quantification of cell type-specific parameters. We applied our strategy to quantitative time-resolved data of EPO-induced JAK/STAT signaling generated by quantitative immunoblotting, mass spectrometry and quantitative real-time PCR (qRT-PCR) in CFU-E and H838 cells as well as H838 cells overexpressing human EPOR (H838-HA-hEPOR). The established parsimonious mathematical model was able to simultaneously describe the data sets of CFU-E, H838 and H838-HA-hEPOR cells. Seven cell type-specific parameters were identified that included for example parameters for nuclear translocation of STAT5 and target gene induction. Cell type-specific differences in target gene induction were experimentally validated by qRT-PCR experiments. The systematic identification of pathway differences and sensitivities of EPOR signaling in CFU-E and H838 cells revealed potential targets for intervention to selectively inhibit EPO-induced signaling in the tumor cells but leave the responses in erythroid

  5. Glycoprotein 130 receptor signaling mediates α-cell dysfunction in a rodent model of type 2 diabetes

    DEFF Research Database (Denmark)

    Chow, Samuel Z; Speck, Madeleine; Yoganathan, Piriya;

    2014-01-01

    knockout (αgp130KO) mice showed no differences in glycemic control, α-cell function, or α-cell mass. However, when subjected to streptozotocin plus high-fat diet to induce islet inflammation and pathophysiology modeling type 2 diabetes, αgp130KO mice had reduced fasting glycemia, improved glucose tolerance......, reduced fasting insulin, and improved α-cell function. Hyperinsulinemic-euglycemic clamps revealed no differences in insulin sensitivity. We conclude that in a setting of islet inflammation and pathophysiology modeling type 2 diabetes, activation of α-cell gp130 receptor signaling has deleterious effects...... on α-cell function, promoting hyperglycemia. Antagonism of α-cell gp130 receptor signaling may be useful for the treatment of type 2 diabetes....

  6. ERK controls epithelial cell death receptor signalling and cellular FLICE-like inhibitory protein (c-FLIP) in ulcerative colitis

    DEFF Research Database (Denmark)

    Seidelin, Jakob Benedict; Coskun, Mehmet; Vainer, Ben;

    2013-01-01

    Intestinal epithelial cell (IEC) death signalling through the Fas receptor is impaired in active ulcerative colitis (UC). This is possibly due to the activation of cytoprotective pathways resulting in limitation of the tissue injury secondary to inflammation. We hypothesized that inflammatory sig...

  7. The Multiple Faces of Prostaglandin E2 G-Protein Coupled Receptor Signaling during the Dendritic Cell Life Cycle

    Directory of Open Access Journals (Sweden)

    Alessandra Cambi

    2013-03-01

    Full Text Available Many processes regulating immune responses are initiated by G-protein coupled receptors (GPCRs and report biochemical changes in the microenvironment. Dendritic cells (DCs are the most potent antigen-presenting cells and crucial for the regulation of innate and adaptive immune responses. The lipid mediator Prostaglandin E2 (PGE2 via four GPCR subtypes (EP1-4 critically regulates DC generation, maturation and migration. The role of PGE2 signaling in DC biology was unraveled by the characterization of EP receptor subtype expression in DC progenitor cells and DCs, the identification of the signaling pathways initiated by these GPCR subtypes and the classification of DC responses to PGE2 at different stages of differentiation. Here, we review the advances in PGE2 signaling in DCs and describe the efforts still to be made to understand the spatio-temporal fine-tuning of PGE2 responses by DCs.

  8. Pathogenic role of B-cell receptor signaling and canonical NF-κB activation in mantle cell lymphoma.

    Science.gov (United States)

    Saba, Nakhle S; Liu, Delong; Herman, Sarah E M; Underbayev, Chingiz; Tian, Xin; Behrend, David; Weniger, Marc A; Skarzynski, Martin; Gyamfi, Jennifer; Fontan, Lorena; Melnick, Ari; Grant, Cliona; Roschewski, Mark; Navarro, Alba; Beà, Sílvia; Pittaluga, Stefania; Dunleavy, Kieron; Wilson, Wyndham H; Wiestner, Adrian

    2016-07-01

    To interrogate signaling pathways activated in mantle cell lymphoma (MCL) in vivo, we contrasted gene expression profiles of 55 tumor samples isolated from blood and lymph nodes from 43 previously untreated patients with active disease. In addition to lymph nodes, MCL often involves blood, bone marrow, and spleen and is incurable for most patients. Recently, the Bruton tyrosine kinase (BTK) inhibitor ibrutinib demonstrated important clinical activity in MCL. However, the role of specific signaling pathways in the lymphomagenesis of MCL and the biologic basis for ibrutinib sensitivity of these tumors are unknown. Here, we demonstrate activation of B-cell receptor (BCR) and canonical NF-κB signaling specifically in MCL cells in the lymph node. Quantification of BCR signaling strength, reflected in the expression of BCR regulated genes, identified a subset of patients with inferior survival after cytotoxic therapy. Tumor proliferation was highest in the lymph node and correlated with the degree of BCR activation. A subset of leukemic tumors showed active BCR and NF-κB signaling apparently independent of microenvironmental support. In one of these samples, we identified a novel somatic mutation in RELA (E39Q). This sample was resistant to ibrutinib-mediated inhibition of NF-κB and apoptosis. In addition, we identified germ line variants in genes encoding regulators of the BCR and NF-κB pathway previously implicated in lymphomagenesis. In conclusion, BCR signaling, activated in the lymph node microenvironment in vivo, appears to promote tumor proliferation and survival and may explain the sensitivity of this lymphoma to BTK inhibitors. PMID:27127301

  9. Progesterone receptors - animal models and cell signalling in breast cancer: Diverse activation pathways for the progesterone receptor: possible implications for breast biology and cancer

    International Nuclear Information System (INIS)

    Progesterone and estradiol, and their nuclear receptors, play essential roles in the physiology of the reproductive tract, the mammary gland and the nervous system. Estrogens have traditionally been considered associated with an increased risk of breast cancer. There is, however, compelling evidence that progesterone plays an important role in breast cell proliferation and cancer. Herein, we review the possible role of progestins and the progesterone receptor-associated signaling pathways in the development of breast cancer, as well as the therapeutic possibilities arising from our growing knowledge of the activation of the progesterone receptor by other proliferative mechanisms

  10. Multiple Transduction Pathways Mediate Thyrotropin Receptor Signaling in Preosteoblast-Like Cells.

    Science.gov (United States)

    Boutin, Alisa; Neumann, Susanne; Gershengorn, Marvin C

    2016-05-01

    It has been shown that the TSH receptor (TSHR) couples to a number of different signaling pathways, although the Gs-cAMP pathway has been considered primary. Here, we measured the effects of TSH on bone marker mRNA and protein expression in preosteoblast-like U2OS cells stably expressing TSHRs. We determined which signaling cascades are involved in the regulation of IL-11, osteopontin (OPN), and alkaline phosphatase (ALPL). We demonstrated that TSH-induced up-regulation of IL-11 is primarily mediated via the Gs pathway as IL-11 was up-regulated by forskolin (FSK), an adenylyl cyclase activator, and inhibited by protein kinase A inhibitor H-89 and by silencing of Gαs by small interfering RNA. OPN levels were not affected by FSK, but its up-regulation was inhibited by TSHR/Gi-uncoupling by pertussis toxin. Pertussis toxin decreased p38 MAPK kinase phosphorylation, and a p38 inhibitor and small interfering RNA knockdown of p38α inhibited OPN induction by TSH. Up-regulation of ALPL expression required high doses of TSH (EC50 = 395nM), whereas low doses (EC50 = 19nM) were inhibitory. FSK-stimulated cAMP production decreased basal ALPL expression, whereas protein kinase A inhibition by H-89 and silencing of Gαs increased basal levels of ALPL. Knockdown of Gαq/11 and a protein kinase C inhibitor decreased TSH-stimulated up-regulation of ALPL, whereas a protein kinase C activator increased ALPL levels. A MAPK inhibitor and silencing of ERK1/2 inhibited TSH-stimulated ALPL expression. We conclude that TSH regulates expression of different bone markers via distinct signaling pathways. PMID:26950201

  11. Effect of Eicosapentaenoic Acid on E-type Prostaglandin Synthesis and EP4 Receptor Signaling Human Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Gillian Hawcroft

    2010-08-01

    Full Text Available The ω-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA, in the free fatty acid (FFA form, has been demonstrated to reduce adenoma number and size in patients with familial adenomatous polyposis. However, the mechanistic basis of the antineoplastic activity of EPA in the colorectum remains unclear. We tested the hypothesis that EPAFFA negatively modulates synthesis of and signaling by prostaglandin (PG E2 in human colorectal cancer (CRC cells. EPA-FFA induced apoptosis of cyclooxygenase (COX-2-positive human HCA-7 CRC cells in vitro. EPA-FFA in cell culture medium was incorporated rapidly into phospholipid membranes of HCA-7 human CRC cells and acted as a substrate for COX-2, leading to reduced synthesis of PGE2 and generation of PGE3. Alone, PGE3 bound and activated the PGE2 EP4 receptor but with reduced affinity and efficacy compared with its “natural” ligand PGE2. However, in the presence of PGE2, PGE3 acted as an antagonist of EP4 receptor-dependent 3’,5’ cyclic adenosine monophosphate induction in naturally EP4 receptor-positive LoVo human CRC cells and of resistance to apoptosis in HT-29-EP4 human CRC cells overexpressing the EP4 receptor. We conclude that EPA-FFA drives a COX-2dependent “PGE2-to-PGE3 switch” in human CRC cells and that PGE3 acts as a partial agonistat the PGE2 EP4 receptor.

  12. Integration of Nuclear- and Extranuclear-Initiated Estrogen Receptor Signaling in Breast Cancer Cells

    Science.gov (United States)

    Madak Erdogan, Zeynep

    2009-01-01

    Estrogenic hormones exert their effects through binding to Estrogen Receptors (ERs), which work in concert with coregulators and extranuclear signaling pathways to control gene expression in normal as well as cancerous states, including breast tumors. In this thesis, we have used multiple genome-wide analysis tools to elucidate various ways that…

  13. Unraveling the Complexities of Androgen Receptor Signaling in Prostate Cancer Cells

    OpenAIRE

    Heemers, Hannelore V.; Tindall, Donald J.

    2009-01-01

    Androgen signaling is critical for proliferation of prostate cancer cells but cannot be fully inhibited by current androgen deprivation therapies. A study by Xu et al. in this issue of Cancer Cell provides insights into the complexities of androgen signaling in prostate cancer and suggests avenues to target a subset of androgen-sensitive genes.

  14. Adaptive and innate immune reactions regulating mast cell activation: from receptor-mediated signaling to responses

    DEFF Research Database (Denmark)

    Tkaczyk, Christine; Jensen, Bettina M; Iwaki, Shoko; Gilfillan, Alasdair M

    2006-01-01

    In this article, we have described studies that have demonstrated that mast cells can be activated as a consequence of adaptive and innate immune reactions and that these responses can be modified by ligands for other receptors expressed on the surface of mast cells. These various stimuli differe...

  15. Differential expression of Toll-like receptor (TLR) and B cell receptor (BCR) signaling molecules in primary diffuse large B-cell lymphoma of the central nervous system.

    Science.gov (United States)

    Akhter, Ariz; Masir, Noraidah; Elyamany, Ghaleb; Phang, Kean-Chang; Mahe, Etienne; Al-Zahrani, Ali Matar; Shabani-Rad, Meer-Taher; Stewart, Douglas Allan; Mansoor, Adnan

    2015-01-01

    Primary diffuse large B-cell lymphoma of the central nervous system (CNS DLBCL) is a distinct and aggressive lymphoma that is confined to CNS. Since, central nervous system is barrier-protected and immunologically silent; role of TLR/BCR signaling in pathogenesis and biology of CNS DLBCL is intriguing. Genomic mutations in key regulators of TLR/BCR signaling pathway (MYD88/CD79B/CARD11) have recently been reported in this disease. These observations raised possible implications in novel targeted therapies; however, expression pattern of molecules related to TLR/BCR pathways in this lymphoma remains unknown. We have analyzed the expression of 19 genes encoding TLR/BCR pathways and targets in CNS DLBCLs (n = 20) by Nanostring nCounter™ analysis and compared it with expression patterns in purified reactive B-lymphocytes and systemic diffuse large B cell lymphoma (DLBCL) (n = 20). Relative expression of TLR4, TLR5, TLR9, CD79B and BLNK was higher in CNS DLBCLs than in control B-lymphocytes; where as TLR7, MALT1, BCL10, CD79A and LYN was lower in CNS DLBCLs (P 1.5 fold change; P < 0.01). The B cell receptor molecules like BLNK and CD79B were also associated with higher expression of MYD88 dependent TLRs (TLR4/5/9). In conclusion, we have shown over expression of TLR/BCR related genes or their targets, where genomic mutations have commonly been identified in CNS DLBCL. We have also demonstrated that TLR over expression closely relate with up regulation of genes associated with BCR pathway like CD79B/BLNK and CARD11, which play an important role in NF-kB pathway activation. Our results provide an important insight into the possibility of TLR and/or B-cell receptor signaling molecules as possible therapeutic targets in CNS DLBCL. PMID:25391967

  16. Complementary signaling through flt3 and interleukin-7 receptor alpha is indispensable for fetal and adult B cell genesis

    DEFF Research Database (Denmark)

    Sitnicka, Ewa; Brakebusch, Cord; Martensson, Inga-Lill;

    2003-01-01

    FL-/- x IL-7Ralpha-/- BM that also lacks expression of the B cell commitment factor Pax5 and its direct target genes. Furthermore, in contrast to IL-7Ralpha-/- mice, FL-/- x IL-7Ralpha-/- mice also lack mature B cells and detectable committed B cell progenitors during fetal development. Thus......, signaling through the cytokine tyrosine kinase receptor flt3 and IL-7Ralpha are indispensable for fetal and adult B cell development.......Extensive studies of mice deficient in one or several cytokine receptors have failed to support an indispensable role of cytokines in development of multiple blood cell lineages. Whereas B1 B cells and Igs are sustained at normal levels throughout life of mice deficient in IL-7, IL-7Ralpha, common...

  17. Rice Bran Feruloylated Oligosaccharides Activate Dendritic Cells via Toll-Like Receptor 2 and 4 Signaling

    Directory of Open Access Journals (Sweden)

    Chi Chen Lin

    2014-04-01

    Full Text Available This work presents the effects of feruloylated oligosaccharides (FOs of rice bran on murine bone marrow-derived dendritic cells (BMDCs and the potential pathway through which the effects are mediated. We found that FOs induced phenotypic maturation of DCs, as shown by the increased expression of CD40, CD80/CD86 and MHC-I/II molecules. FOs efficiently induced maturation of DCs generated from C3H/HeN or C57BL/6 mice with normal toll-like receptor 4 (TLR-4 or TLR-2 but not DCs from mice with mutated TLR4 or TLR2. The mechanism of action of FOs may be mediated by increased phosphorylation of ERK, p38 and JNK mitogen-activated protein kinase (MAPKs and increased NF-kB activity, which are important signaling molecules downstream of TLR-4 and TLR-2. These data suggest that FOs induce DCs maturation through TLR-4 and/or TLR-2 and that FOs might have potential efficacy against tumor or virus infection or represent a candidate-adjuvant approach for application in immunotherapy and vaccination.

  18. Rice bran feruloylated oligosaccharides activate dendritic cells via Toll-like receptor 2 and 4 signaling.

    Science.gov (United States)

    Lin, Chi Chen; Chen, Hua Han; Chen, Yu Kuo; Chang, Hung Chia; Lin, Ping Yi; Pan, I-Hong; Chen, Der-Yuan; Chen, Chuan Mu; Lin, Su Yi

    2014-01-01

    This work presents the effects of feruloylated oligosaccharides (FOs) of rice bran on murine bone marrow-derived dendritic cells (BMDCs) and the potential pathway through which the effects are mediated. We found that FOs induced phenotypic maturation of DCs, as shown by the increased expression of CD40, CD80/CD86 and MHC-I/II molecules. FOs efficiently induced maturation of DCs generated from C3H/HeN or C57BL/6 mice with normal toll-like receptor 4 (TLR-4) or TLR-2 but not DCs from mice with mutated TLR4 or TLR2. The mechanism of action of FOs may be mediated by increased phosphorylation of ERK, p38 and JNK mitogen-activated protein kinase (MAPKs) and increased NF-kB activity, which are important signaling molecules downstream of TLR-4 and TLR-2. These data suggest that FOs induce DCs maturation through TLR-4 and/or TLR-2 and that FOs might have potential efficacy against tumor or virus infection or represent a candidate-adjuvant approach for application in immunotherapy and vaccination. PMID:24762969

  19. T cell homeostasis requires G protein-coupled receptor-mediated access to trophic signals that promote growth and inhibit chemotaxis

    OpenAIRE

    Cinalli, Ryan M.; Herman, Catherine E.; Lew, Brian O.; Wieman, Heather L.; Thompson, Craig B.; Rathmell, Jeffrey C.

    2005-01-01

    Signals that regulate T cell homeostasis are not fully understood. G protein-coupled receptors (GPCR), such as the chemokine receptors, may affect homeostasis by direct signaling or by guiding T cell migration to distinct location-restricted signals. Here, we show that blockade of Gαi-associated GPCR signaling by treatment with pertussis toxin led to T cell atrophy and shortened life-span in T cell-replete hosts and prevented T cell homeostatic growth and proliferation in T cell-deficient hos...

  20. Glucose-Sensing Receptor T1R3: A New Signaling Receptor Activated by Glucose in Pancreatic β-Cells.

    Science.gov (United States)

    Kojima, Itaru; Nakagawa, Yuko; Hamano, Kunihisa; Medina, Johan; Li, Longfei; Nagasawa, Masahiro

    2015-01-01

    Subunits of the sweet taste receptors T1R2 and T1R3 are expressed in pancreatic β-cells. Compared with T1R3, mRNA expression of T1R2 is considerably lower. At the protein level, expression of T1R2 is undetectable in β-cells. Accordingly, a major component of the sweet taste-sensing receptor in β-cells may be a homodimer of T1R3 rather than a heterodimer of T1R2/T1R3. Inhibition of this receptor by gurmarin or deletion of the T1R3 gene attenuates glucose-induced insulin secretion from β-cells. Hence the T1R3 homodimer functions as a glucose-sensing receptor (GSR) in pancreatic β-cells. When GSR is activated by the T1R3 agonist sucralose, elevation of intracellular ATP concentration ([ATP]i) is observed. Sucralose increases [ATP]i even in the absence of ambient glucose, indicating that sucralose increases [ATP]i not simply by activating glucokinase, a rate-limiting enzyme in the glycolytic pathway. In addition, sucralose augments elevation of [ATP]i induced by methylsuccinate, suggesting that sucralose activates mitochondrial metabolism. Nonmetabolizable 3-O-methylglucose also increases [ATP]i and knockdown of T1R3 attenuates elevation of [ATP]i induced by high concentration of glucose. Collectively, these results indicate that the T1R3 homodimer functions as a GSR; this receptor is involved in glucose-induced insulin secretion by activating glucose metabolism probably in mitochondria. PMID:25947913

  1. The hepatocyte growth factor (HGF)-MET receptor tyrosine kinase signaling pathway: Diverse roles in modulating immune cell functions.

    Science.gov (United States)

    Ilangumaran, Subburaj; Villalobos-Hernandez, Alberto; Bobbala, Diwakar; Ramanathan, Sheela

    2016-06-01

    Hepatocyte growth factor (HGF) signaling via the MET receptor is essential for embryonic development and tissue repair. On the other hand, deregulated MET signaling promotes tumor progression in diverse types of cancers. Even though oncogenic MET signaling remains the major research focus, the HGF-MET axis has also been implicated in diverse aspects of immune cell development and functions. In the presence of other hematopoietic growth factors, HGF promotes the development of erythroid, myeloid and lymphoid lineage cells and thrombocytes. In monocytes and macrophages responding to inflammatory stimuli, induction of autocrine HGF-MET signaling can contribute to tissue repair via stimulating anti-inflammatory cytokine production. HGF-MET signaling can also modulate adaptive immune response by facilitating the migration of Langerhans cells and dendritic cells to draining lymph nodes. However, MET signaling has also been shown to induce tolerogenic dendritic cells in mouse models of graft-versus-host disease and experimental autoimmune encephalomyelitis. HGF-MET axis is also implicated in promoting thymopoiesis and the survival and migration of B lymphocytes. Recent studies have shown that MET signaling induces cardiotropism in activated T lymphocytes. Further understanding of the HGF-MET axis in the immune system would allow its therapeutic manipulation to improve immune cell reconstitution, restore immune homeostasis and to treat immuno-inflammatory diseases. PMID:26822708

  2. Microenvironment interactions and B-cell receptor signaling in Chronic Lymphocytic Leukemia: Implications for disease pathogenesis and treatment.

    Science.gov (United States)

    Ten Hacken, Elisa; Burger, Jan A

    2016-03-01

    Chronic Lymphocytic Leukemia (CLL) is a malignancy of mature B lymphocytes which are highly dependent on interactions with the tissue microenvironment for their survival and proliferation. Critical components of the microenvironment are monocyte-derived nurselike cells (NLCs), mesenchymal stromal cells, T cells and NK cells, which communicate with CLL cells through a complex network of adhesion molecules, chemokine receptors, tumor necrosis factor (TNF) family members, and soluble factors. (Auto-) antigens and/or autonomous mechanisms activate the B-cell receptor (BCR) and its downstream signaling cascade in secondary lymphatic tissues, playing a central pathogenetic role in CLL. Novel small molecule inhibitors, including the Bruton's tyrosine kinase (BTK) inhibitor ibrutinib and the phosphoinositide-3-kinase delta (PI3Kδ) inhibitor idelalisib, target BCR signaling and have become the most successful new therapeutics in this disease. We here review the cellular and molecular characteristics of CLL cells, and discuss the cellular components and key pathways involved in the cross-talk with their microenvironment. We also highlight the relevant novel treatment strategies, focusing on immunomodulatory agents and BCR signaling inhibitors and how these treatments disrupt CLL-microenvironment interactions. This article is part of a Special Issue entitled: Tumor Microenvironment Regulation of Cancer Cell Survival, Metastasis, Inflammation, and Immune Surveillance edited by Peter Ruvolo and Gregg L. Semenza. PMID:26193078

  3. Biased Signaling of Protease-activated Receptors

    OpenAIRE

    PeishenZhao; NigelWilliamBunnett

    2014-01-01

    In addition to their role in protein degradation and digestion, proteases can also function as hormone-like signaling molecules that regulate vital patho-physiological processes, including inflammation, hemostasis, pain and repair mechanisms. Certain proteases can signal to cells by cleaving protease-activated receptors (PARs), a family of four G protein-coupled receptors. PARs are expressed by almost all cell types, control important physiological and disease-relevant processes, and are an e...

  4. Progesterone receptors - animal models and cell signaling in breast cancer: Expression and transcriptional activity of progesterone receptor A and progesterone receptor B in mammalian cells

    International Nuclear Information System (INIS)

    Progesterone is an essential regulator of normal female reproductive function. Its effects are mediated by two nuclear progesterone receptor (PR) proteins, PRA and PRB, which are identical except for an additional 164 amino acids at the N-terminal end of PRB. Transcriptional analyses of the two receptor forms have assigned strikingly distinct functional signatures to the two PRs, despite their apparent physical similarity. The basis of these differences is yet to be fully understood. Furthermore, these differences are strongly influenced by the cell type and the promoter used. We review the mammalian transcriptional studies of PRA and PRB, and compare them with what is known about their expression and function in target tissues

  5. Ethanol affects NMDA receptor signaling at climbing fiber-Purkinje cell synapses in mice and impairs cerebellar LTD.

    Science.gov (United States)

    He, Qionger; Titley, Heather; Grasselli, Giorgio; Piochon, Claire; Hansel, Christian

    2013-03-01

    Ethanol profoundly influences cerebellar circuit function and motor control. It has recently been demonstrated that functional N-methyl-(D)-aspartate (NMDA) receptors are postsynaptically expressed at climbing fiber (CF) to Purkinje cell synapses in the adult cerebellum. Using whole cell patch-clamp recordings from mouse cerebellar slices, we examined whether ethanol can affect NMDA receptor signaling in mature Purkinje cells. NMDA receptor-mediated currents were isolated by bath application of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoylbenzol[f]quinoxaline (NBQX). The remaining (D)-2-amino-5-phosphonovaleric acid ((D)-APV)-sensitive current was reduced by ethanol at concentrations as low as 10 mM. At a concentration of 50 mM ethanol, the blockade of (D)-APV-sensitive CF-excitatory postsynaptic currents was significantly stronger. Ethanol also altered the waveform of CF-evoked complex spikes by reducing the afterdepolarization. This effect was not seen when NMDA receptors were blocked by (D)-APV before ethanol wash-in. In contrast to CF synaptic transmission, parallel fiber (PF) synaptic inputs were not affected by ethanol. Finally, ethanol (10 mM) impaired long-term depression (LTD) at PF to Purkinje cell synapses as induced under control conditions by paired PF and CF activity. However, LTD induced by pairing PF stimulation with depolarizing voltage steps (substituting for CF activation) was not blocked by ethanol. These observations suggest that the sensitivity of cerebellar circuit function and plasticity to low concentrations of ethanol may be caused by an ethanol-mediated impairment of NMDA receptor signaling at CF synapses onto cerebellar Purkinje cells. PMID:23221414

  6. Histamine H4 Receptor mediates interleukin-8 and TNF-α release in human mast cells via multiple signaling pathways.

    Science.gov (United States)

    Chen, X-F; Zhang, Z; Dou, X; Li, J-J; Zhang, W; Yu, Y-Y; Yu, B; Yu, B

    2016-01-01

    Histamine, mainly produced by mast cells, is an important inflammatory mediator in immune response. Recently Histamine H4 Receptor (H4R) was also identified in mast cells, from which pro-inflammatory cytokines and chemokines are released. However, the mechanism of how H4R mediates these cytokines and chemokines release in mast cells was still unclear. To further explore the role of H4R in the immune inflammatory response in mast cells, we tested the release of inflammatory cytokine tumor necrosis factor-α (TNF-α), chemokine interleukin-8 (IL-8) and the relevant signaling pathways activated by H4R on LAD2 cells (a human mast cell line). We found that the release of IL-8 and TNF-α were blocked by inhibitors of PI3K, ERK and Ca2+-Calcineurin-NFAT signaling pathways, while the release of these cytokines and chemokines were enhanced by the inhibitor of P38 signaling pathway. However, inhibitors of the JNK and NF-κB signaling pathways had little effect on the expression of the pro-inflammatory mediators. Moreover, activation of the H4R could induce phosphorylation of ERK, p38 and AKT in mast cells. In conclusion, we found that H4R mediates the release of inflammatory cytokine TNF-α and chemokine IL-8 in human mast cells via PI3K, Ca2+-Calcineurin-NFAT and MAPKs signaling pathways. PMID:26828993

  7. Toddler: An Embryonic Signal That Promotes Cell Movement via Apelin Receptors

    OpenAIRE

    Pauli, Andrea; Norris, Megan L.; Valen, Eivind; Chew, Guo-Liang; Gagnon, James A.; Zimmerman, Steven; Mitchell, Andrew; Ma, Jiao; Dubrulle, Julien; Reyon, Deepak; Tsai, Shengdar Q.; Joung, J. Keith; Saghatelian, Alan; Schier, Alexander F.

    2014-01-01

    It has been assumed that most, if not all, signals regulating early development have been identified. Contrary to this expectation, we identified 28 candidate signaling proteins expressed during zebrafish embryogenesis, including Toddler, a short, conserved, and secreted peptide. Both absence and overproduction of Toddler reduce the movement of mesendodermal cells during zebrafish gastrulation. Local and ubiquitous production of Toddler promote cell movement, suggesting that Toddler is neithe...

  8. FGF-receptor signalling controls neural cell diversity in the zebrafish hindbrain by regulating olig2 and sox9

    OpenAIRE

    Esain, Virginie; Postlethwait, John H.; Charnay, Patrick; Ghislain, Julien

    2010-01-01

    The mechanisms underlying the generation of neural cell diversity are the subject of intense investigation, which has highlighted the involvement of different signalling molecules including Shh, BMP and Wnt. By contrast, relatively little is known about FGF in this process. In this report we identify an FGF-receptor-dependent pathway in zebrafish hindbrain neural progenitors that give rise to somatic motoneurons, oligodendrocyte progenitors and differentiating astroglia. Using a combination o...

  9. Orphan nuclear receptor TLX activates Wnt/β-catenin signalling to stimulate neural stem cell proliferation and self-renewal

    OpenAIRE

    Qu, Qiuhao; Sun, Guoqiang; Li, Wenwu; Yang, Su; Ye, Peng; Zhao, Chunnian; Yu, Ruth T.; Gage, Fred H; Evans, Ronald M; Shi, Yanhong

    2009-01-01

    The nuclear receptor TLX (also known as NR2E1) is essential for adult neural stem cell self-renewal; however, the molecular mechanisms involved remain elusive. Here we show that TLX activates the canonical Wnt/β-catenin pathway in adult mouse neural stem cells. Furthermore, we demonstrate that Wnt/β-catenin signalling is important in the proliferation and self-renewal of adult neural stem cells in the presence of epidermal growth factor and fibroblast growth factor. Wnt7a and active β-catenin...

  10. Notch Signaling Activation in Cervical Cancer Cells Induces Cell Growth Arrest with the Involvement of the Nuclear Receptor NR4A2

    Science.gov (United States)

    Sun, Lichun; Liu, Mingqiu; Sun, Guang-Chun; Yang, Xu; Qian, Qingqing; Feng, Shuyu; Mackey, L. Vienna; Coy, David H.

    2016-01-01

    Cervical cancer is a second leading cancer death in women world-wide, with most cases in less developed countries. Notch signaling is highly conserved with its involvement in many cancers. In the present study, we established stable cervical cell lines with Notch activation and inactivation and found that Notch activation played a suppressive role in cervical cancer cells. Meanwhile, the transient overexpression of the active intracellular domain of all four Notch receptors (ICN1, 2, 3, and 4) also induced the suppression of cervical cancer Hela cell growth. ICN1 also induced cell cycle arrest at phase G1. Notch1 signaling activation affected the expression of serial genes, especially the genes associated with cAMP signaling, with an increase of genes like THBS1, VCL, p63, c-Myc and SCG2, a decrease of genes like NR4A2, PCK2 and BCL-2. Particularly, The nuclear receptor NR4A2 was observed to induce cell proliferation via MTT assay and reduce cell apoptosis via FACS assay. Furthermore, NR4A2's activation could reverse ICN1-induced suppression of cell growth while erasing ICN1-induced increase of tumor suppressor p63. These findings support that Notch signaling mediates cervical cancer cell growth suppression with the involvement of nuclear receptor NR4A2. Notably, Notch/NR4A2/p63 signaling cascade possibly is a new signling pathway undisclosed. PMID:27471554

  11. δ-Opioid receptor-stimulated Akt signaling in neuroblastoma x glioma (NG108-15) hybrid cells involves receptor tyrosine kinase-mediated PI3K activation

    International Nuclear Information System (INIS)

    δ-Opioid receptor (DOR) agonists possess cytoprotective properties, an effect associated with activation of the 'pro-survival' kinase Akt. Here we delineate the signal transduction pathway by which opioids induce Akt activation in neuroblastoma x glioma (NG108-15) hybrid cells. Exposure of the cells to both [D-Pen2,5]enkephalin and etorphine resulted in a time- and dose-dependent increase in Akt activity, as measured by means of an activation-specific antibody recognizing phosphoserine-473. DOR-mediated Akt signaling is blocked by the opioid antagonist naloxone and involves inhibitory Gi/o proteins, because pre-treatment with pertussis toxin, but not over-expression of the Gq/11 scavengers EBP50 and GRK2-K220R, prevented this effect. Further studies with Wortmannin and LY294002 revealed that phophoinositol-3-kinase (PI3K) plays a central role in opioid-induced Akt activation. Opioids stimulate Akt activity through transactivation of receptor tyrosine kinases (RTK), because pre-treatment of the cells with inhibitors for neurotrophin receptor tyrosine kinases (AG879) and the insulin-like growth factor receptor IGF-1 (AG1024), but not over-expression of the Gβγ scavenger phosducin, abolished this effect. Activated Akt translocates to the nuclear membrane, where it promotes GSK3 phosphorylation and prevents caspase-3 cleavage, two key events mediating inhibition of cell apoptosis and enhancement of cell survival. Taken together, these results demonstrate that in NG108-15 hybrid cells DOR agonists possess cytoprotective properties mediated by activation of the RTK/PI3K/Akt signaling pathway.

  12. {delta}-Opioid receptor-stimulated Akt signaling in neuroblastoma x glioma (NG108-15) hybrid cells involves receptor tyrosine kinase-mediated PI3K activation

    Energy Technology Data Exchange (ETDEWEB)

    Heiss, Anika; Ammer, Hermann [Institute of Pharmacology, Toxicology and Pharmacy Ludwig-Maximilians-University of Munich Koeniginstrasse 16 80539 Muenchen Federal Republic of Germany (Germany); Eisinger, Daniela A., E-mail: eisinger@pharmtox.vetmed.uni-muenchen.de [Institute of Pharmacology, Toxicology and Pharmacy Ludwig-Maximilians-University of Munich Koeniginstrasse 16 80539 Muenchen Federal Republic of Germany (Germany)

    2009-07-15

    {delta}-Opioid receptor (DOR) agonists possess cytoprotective properties, an effect associated with activation of the 'pro-survival' kinase Akt. Here we delineate the signal transduction pathway by which opioids induce Akt activation in neuroblastoma x glioma (NG108-15) hybrid cells. Exposure of the cells to both [D-Pen{sup 2,5}]enkephalin and etorphine resulted in a time- and dose-dependent increase in Akt activity, as measured by means of an activation-specific antibody recognizing phosphoserine-473. DOR-mediated Akt signaling is blocked by the opioid antagonist naloxone and involves inhibitory G{sub i/o} proteins, because pre-treatment with pertussis toxin, but not over-expression of the G{sub q/11} scavengers EBP50 and GRK2-K220R, prevented this effect. Further studies with Wortmannin and LY294002 revealed that phophoinositol-3-kinase (PI3K) plays a central role in opioid-induced Akt activation. Opioids stimulate Akt activity through transactivation of receptor tyrosine kinases (RTK), because pre-treatment of the cells with inhibitors for neurotrophin receptor tyrosine kinases (AG879) and the insulin-like growth factor receptor IGF-1 (AG1024), but not over-expression of the G{beta}{gamma} scavenger phosducin, abolished this effect. Activated Akt translocates to the nuclear membrane, where it promotes GSK3 phosphorylation and prevents caspase-3 cleavage, two key events mediating inhibition of cell apoptosis and enhancement of cell survival. Taken together, these results demonstrate that in NG108-15 hybrid cells DOR agonists possess cytoprotective properties mediated by activation of the RTK/PI3K/Akt signaling pathway.

  13. GnRH receptor activation competes at a low level with growth signaling in stably transfected human breast cell lines

    International Nuclear Information System (INIS)

    Gonadotrophin releasing hormone (GnRH) analogs lower estrogen levels in pre-menopausal breast cancer patients. GnRH receptor (GnRH-R) activation also directly inhibits the growth of certain cells. The applicability of GnRH anti-proliferation to breast cancer was therefore analyzed. GnRH-R expression in 298 primary breast cancer samples was measured by quantitative immunofluorescence. Levels of functional GnRH-R in breast-derived cell lines were assessed using 125I-ligand binding and stimulation of 3H-inositol phosphate production. Elevated levels of GnRH-R were stably expressed in cells by transfection. Effects of receptor activation on in vitro cell growth were investigated in comparison with IGF-I and EGF receptor inhibition, and correlated with intracellular signaling using western blotting. GnRH-R immunoscoring was highest in hormone receptor (triple) negative and grade 3 breast tumors. However prior to transfection, functional endogenous GnRH-R were undetectable in four commonly studied breast cancer cell lines (MCF-7, ZR-75-1, T47D and MDA-MB-231). After transfection with GnRH-R, high levels of cell surface GnRH-R were detected in SVCT and MDA-MB-231 clones while low-moderate levels of GnRH-R occurred in MCF-7 clones and ZR-75-1 clones. MCF-7 sub-clones with high levels of GnRH-R were isolated following hygromycin phosphotransferase transfection. High level cell surface GnRH-R enabled induction of high levels of 3H-inositol phosphate and modest growth-inhibition in SVCT cells. In contrast, growth of MCF-7, ZR-75-1 or MDA-MB-231 clones was unaffected by GnRH-R activation. Cell growth was inhibited by IGF-I or EGF receptor inhibitors. IGF-I receptor inhibitor lowered levels of p-ERK1/2 in MCF-7 clones. Washout of IGF-I receptor inhibitor resulted in transient hyper-elevation of p-ERK1/2, but co-addition of GnRH-R agonist did not alter the dynamics of ERK1/2 re-phosphorylation. Breast cancers exhibit a range of GnRH-R immunostaining, with higher levels of

  14. The Wnt Frizzled Receptor MOM-5 Regulates the UNC-5 Netrin Receptor through Small GTPase-Dependent Signaling to Determine the Polarity of Migrating Cells.

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    Naomi Levy-Strumpf

    2015-08-01

    Full Text Available Wnt and Netrin signaling regulate diverse essential functions. Using a genetic approach combined with temporal gene expression analysis, we found a regulatory link between the Wnt receptor MOM-5/Frizzled and the UNC-6/Netrin receptor UNC-5. These two receptors play key roles in guiding cell and axon migrations, including the migration of the C. elegans Distal Tip Cells (DTCs. DTCs migrate post-embryonically in three sequential phases: in the first phase along the Antero-Posterior (A/P axis, in the second, along the Dorso-Ventral (D/V axis, and in the third, along the A/P axis. Loss of MOM-5/Frizzled function causes third phase A/P polarity reversals of the migrating DTCs. We show that an over-expression of UNC-5 causes similar DTC A/P polarity reversals and that unc-5 deficits markedly suppress the A/P polarity reversals caused by mutations in mom-5/frizzled. This implicates MOM-5/Frizzled as a negative regulator of unc-5. We provide further evidence that small GTPases mediate MOM-5's regulation of unc-5 such that one outcome of impaired function of small GTPases like CED-10/Rac and MIG-2/RhoG is an increase in unc-5 function. The work presented here demonstrates the existence of cross talk between components of the Netrin and Wnt signaling pathways and provides further insights into the way guidance signaling mechanisms are integrated to orchestrate directed cell migration.

  15. T cell receptor signaling pathway is overexpressed in CD4 + T cells from HAM/TSP individuals

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    Mariana Tomazini Pinto

    2015-12-01

    Full Text Available ABSTRACT Human T-lymphotropic virus type 1 (HTLV-1 is a human retrovirus related to the chronic neuroinflammatory disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP. CD4+ T cells activation appears to play a key role on HTLV-1 infection. Here we investigated the expression of genes associated to T cell activation CD3e molecule, epsilon (CD3?, lymphocyte-specific protein tyrosine kinase (LCK, vav 1 guanine nucleotide exchange factor (VAV1, and zeta-chain (TCR associated protein kinase 70 kDa (ZAP70 on T lymphocytes of HTLV-1-infected individuals and compared to healthy uninfected individuals (CT. We observed that CD3?, LCK, ZAP70, and VAV1 gene expression were increased in CD4+ T cells from HAM/TSP group compared to HTLV-1 asymptomatic patients (HAC. Moreover, ZAP70 and VAV1 were also upregulated in HAM/TSP compared to CT group. We detected a positive correlation among all these genes. We also observed that CD3?, LCK, and VAV1 genes had a positive correlation with the proviral load (PVL and Tax expression. These results suggest that PVL and Tax protein could drive CD3?, LCK, and VAV1 gene expression in CD4+ T cells, and these genes function on a synchronized way on the CD4+ T cell activation. The elucidation of the mechanisms underlying T cell receptor signaling pathway is of considerable interest and might lead to new insights into the mechanism of HAM/TSP.

  16. Roles for NHERF1 and NHERF2 on the regulation of C3a receptor signaling in human mast cells.

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    Hariharan Subramanian

    Full Text Available BACKGROUND: The anaphylatoxin C3a binds to the G protein coupled receptor (GPCR, C3aR and activates divergent signaling pathways to induce degranulation and cytokine production in human mast cells. Adapter proteins such as the Na(+/H(+ exchange regulatory factor (NHERF1 and NHERF2 have been implicated in regulating functions of certain GPCRs by binding to the class I PDZ (PSD-95/Dlg/Zo1 motifs present on their cytoplasmic tails. Although C3aR possesses a class I PDZ motif, the possibility that it interacts with NHERF proteins to modulate signaling in human mast cells has not been determined. METHODOLOGY/PRINCIPAL FINDINGS: Using reverse transcription PCR and Western blotting, we found that NHERF1 and NHERF2 are expressed in human mast cell lines (HMC-1, LAD2 and CD34(+-derived primary human mast cells. Surprisingly, however, C3aR did not associate with these adapter proteins. To assess the roles of NHERFs on signaling downstream of C3aR, we used lentiviral shRNA to stably knockdown the expression of these proteins in human mast cells. Silencing the expression of NHERF1 and NHERF2 had no effect on C3aR desensitization, agonist-induced receptor internalization, ERK/Akt phosphorylation or chemotaxis. However, loss of NHERF1 and NHERF2 resulted in significant inhibition of C3a-induced mast cell degranulation, NF-κB activation and chemokine production. CONCLUSION/SIGNIFICANCE: This study demonstrates that although C3aR possesses a class I PDZ motif, it does not associate with NHERF1 and NHERF2. Surprisingly, these proteins provide stimulatory signals for C3a-induced degranulation, NF-κB activation and chemokine generation in human mast cells. These findings reveal a new level of complexity for the functional regulation of C3aR by NHERFs in human mast cells.

  17. Distinct signalling pathways of murine histamine H1- and H4-receptors expressed at comparable levels in HEK293 cells.

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    Silke Beermann

    Full Text Available Histamine (HA is recognized by its target cells via four G-protein-coupled receptors, referred to as histamine H1-receptor (H1R, H2R, H3R, and H4R. Both H1R and H4R exert pro-inflammatory functions. However, their signal transduction pathways have never been analyzed in a directly comparable manner side by side. Moreover, the analysis of pharmacological properties of the murine orthologs, representing the main targets of pre-clinical research, is very important. Therefore, we engineered recombinant HEK293 cells expressing either mouse (mH1R or mH4R at similar levels and analyzed HA-induced signalling in these cells. HA induced intracellular calcium mobilization via both mH1R and mH4R, with the mH1R being much more effective. Whereas cAMP accumulation was potentiated via the mH1R, it was reduced via the mH4R. The regulation of both second messengers via the H4R, but not the H1R, was sensitive to pertussis toxin (PTX. The mitogen-activated protein kinases (MAPKs ERK 1/2 were massively activated downstream of both receptors and demonstrated a functional involvement in HA-induced EGR-1 gene expression. The p38 MAPK was moderately activated via both receptors as well, but was functionally involved in HA-induced EGR-1 gene expression only in H4R-expressing cells. Surprisingly, in this system p38 MAPK activity reduced the HA-induced gene expression. In summary, using this system which allows a direct comparison of mH1R- and mH4R-induced signalling, qualitative and quantitative differences on the levels of second messenger generation and also in terms of p38 MAPK function became evident.

  18. HIV-1 Tat Regulates Occludin and Aβ Transfer Receptor Expression in Brain Endothelial Cells via Rho/ROCK Signaling Pathway

    Science.gov (United States)

    Chen, Yanlan; Jiang, Wenlin; Wu, Xianghong; Ye, Biao; Zhou, Xiaoting

    2016-01-01

    HIV-1 transactivator protein (Tat) has been shown to play an important role in HIV-associated neurocognitive disorders. The aim of the present study was to evaluate the relationship between occludin and amyloid-beta (Aβ) transfer receptors in human cerebral microvascular endothelial cells (hCMEC/D3) in the context of HIV-1-related pathology. The protein expressions of occludin, receptor for advanced glycation end products (RAGE), and low-density lipoprotein receptor-related protein 1 (LRP1) in hCMEC/D3 cells were examined using western blotting and immunofluorescent staining. The mRNA levels of occludin, RAGE, and LRP1 were measured using quantitative real-time polymerase chain reaction. HIV-1 Tat at 1 µg/mL and the Rho inhibitor hydroxyfasudil (HF) at 30 µmol/L, with 24 h exposure, had no significant effect on hCMEC/D3 cell viability. Treatment with HIV-1 Tat protein decreased mRNA and protein levels of occludin and LRP1 and upregulated the expression of RAGE; however, these effects were attenuated by HF. These data suggest that the Rho/ROCK signaling pathway is involved in HIV-1 Tat-mediated changes in occludin, RAGE, and LRP1 in hCMEC/D3 cells. HF may have a beneficial influence by protecting the integrity of the blood-brain barrier and the expression of Aβ transfer receptors.

  19. Morphine Induces Desensitization of Insulin Receptor Signaling

    OpenAIRE

    Li, Yu; Eitan, Shoshana; Wu, Jiong; Evans, Christopher J.; Kieffer, Brigitte; Sun, Xiaojian; Polakiewicz, Roberto D.

    2003-01-01

    Morphine analgesia is mediated principally by the μ-opioid receptor (MOR). Since morphine and other opiates have been shown to influence glucose homeostasis, we investigated the hypothesis of direct cross talk between the MOR and the insulin receptor (IR) signaling cascades. We show that prolonged morphine exposure of cell lines expressing endogenous or transfected MOR, IR, and the insulin substrate 1 (IRS-1) protein specifically desensitizes IR signaling to Akt and ERK cascades. Morphine cau...

  20. ZAP-70, CTLA-4 and proximal T cell receptor signaling in cows infected with Mycobacterium avium subsp. paratuberculosis.

    Science.gov (United States)

    Leite, Fernando L; Eslabão, Livia B; Pesch, Bruce; Bannantine, John P; Reinhardt, Timothy A; Stabel, Judith R

    2015-09-15

    Paratuberculosis is a chronic intestinal disease of ruminant animals caused by Mycobacterium avium subsp. paratuberculosis (MAP). A hallmark of paratuberculosis is a transition from a cell-mediated Th1 type response to a humoral Th2 response with the progression of disease from a subclinical to clinical state. The objective of this study was to investigate the expression of two crucial molecules in T cell function, ZAP-70 (zeta-chain-associated protein of 70 kDa) and CTLA-4 (cytotoxic T-lymphocyte antigen-4), in cows naturally infected with MAP. Peripheral blood mononuclear cells (PBMCs) isolated from control non-infected cows (n=5), and cows in subclinical (n=6) and clinical stages of paratuberculosis (n=6) were cultured alone (medium only), and with concanavalin A, and a whole cell sonicate of MAP for 24, 72 and 144 h to measure the dynamic changes of ZAP-70 and CTLA-4 expression on CD4, CD8, and gamma delta (γδ) T cells. Flow cytometry was also performed to measure ZAP-70 phosphorylation to examine proximal T cell receptor signaling in animals of different disease status. The surface expression of CTLA-4 was increased in animals in subclinical stage of infection while levels of ZAP-70 were decreased in CD4+ T cells of both subclinical and clinical animals, indicating a change in T cell phenotype with disease state. Interestingly, proximal T cell receptor signaling was not altered in infected animals. This study demonstrated changes in crucial signaling molecules in animals infected with MAP, thereby elucidating T cell alterations associated with disease progression. PMID:26163934

  1. Conserved roles of fibroblast growth factor receptor 2 signaling in the regulation of inner cell mass development in bovine blastocysts.

    Science.gov (United States)

    Akizawa, Hiroki; Nagatomo, Hiroaki; Odagiri, Haruka; Kohri, Nanami; Yamauchi, Nobuhiko; Yanagawa, Yojiro; Nagano, Masashi; Takahashi, Masashi; Kawahara, Manabu

    2016-06-01

    A common process during preimplantation mammalian development is blastocyst formation, which utilizes signaling through fibroblast growth factor receptor 2 (FGFR2), yet the mechanisms through which FGFR2 signaling affect preimplantation development in bovine embryos remain incompletely understood. Here, we used RNA-interference to investigate the in vitro development, the frequency of blastomere apoptosis, and the mRNA expression of developmental marker genes in FGF receptor 2-knockdown (FGFR2-KD) bovine embryos. A reduction in FGFR2 mRNA did not affect preimplantation development or the frequency of apoptotic blastomeres, but did enhanced proliferation of the inner cell mass in blastocysts (P cattle. Mol. Reprod. Dev. 83: 516-525, 2016. © 2016 Wiley Periodicals, Inc. PMID:27060901

  2. PCA3 noncoding RNA is involved in the control of prostate-cancer cell survival and modulates androgen receptor signaling

    International Nuclear Information System (INIS)

    PCA3 is a non-coding RNA (ncRNA) that is highly expressed in prostate cancer (PCa) cells, but its functional role is unknown. To investigate its putative function in PCa biology, we used gene expression knockdown by small interference RNA, and also analyzed its involvement in androgen receptor (AR) signaling. LNCaP and PC3 cells were used as in vitro models for these functional assays, and three different siRNA sequences were specifically designed to target PCA3 exon 4. Transfected cells were analyzed by real-time qRT-PCR and cell growth, viability, and apoptosis assays. Associations between PCA3 and the androgen-receptor (AR) signaling pathway were investigated by treating LNCaP cells with 100 nM dihydrotestosterone (DHT) and with its antagonist (flutamide), and analyzing the expression of some AR-modulated genes (TMPRSS2, NDRG1, GREB1, PSA, AR, FGF8, CdK1, CdK2 and PMEPA1). PCA3 expression levels were investigated in different cell compartments by using differential centrifugation and qRT-PCR. LNCaP siPCA3-transfected cells significantly inhibited cell growth and viability, and increased the proportion of cells in the sub G0/G1 phase of the cell cycle and the percentage of pyknotic nuclei, compared to those transfected with scramble siRNA (siSCr)-transfected cells. DHT-treated LNCaP cells induced a significant upregulation of PCA3 expression, which was reversed by flutamide. In siPCA3/LNCaP-transfected cells, the expression of AR target genes was downregulated compared to siSCr-transfected cells. The siPCA3 transfection also counteracted DHT stimulatory effects on the AR signaling cascade, significantly downregulating expression of the AR target gene. Analysis of PCA3 expression in different cell compartments provided evidence that the main functional roles of PCA3 occur in the nuclei and microsomal cell fractions. Our findings suggest that the ncRNA PCA3 is involved in the control of PCa cell survival, in part through modulating AR signaling, which may raise new

  3. PCA3 noncoding RNA is involved in the control of prostate-cancer cell survival and modulates androgen receptor signaling

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    Ferreira Luciana Bueno

    2012-11-01

    Full Text Available Abstract Background PCA3 is a non-coding RNA (ncRNA that is highly expressed in prostate cancer (PCa cells, but its functional role is unknown. To investigate its putative function in PCa biology, we used gene expression knockdown by small interference RNA, and also analyzed its involvement in androgen receptor (AR signaling. Methods LNCaP and PC3 cells were used as in vitro models for these functional assays, and three different siRNA sequences were specifically designed to target PCA3 exon 4. Transfected cells were analyzed by real-time qRT-PCR and cell growth, viability, and apoptosis assays. Associations between PCA3 and the androgen-receptor (AR signaling pathway were investigated by treating LNCaP cells with 100 nM dihydrotestosterone (DHT and with its antagonist (flutamide, and analyzing the expression of some AR-modulated genes (TMPRSS2, NDRG1, GREB1, PSA, AR, FGF8, CdK1, CdK2 and PMEPA1. PCA3 expression levels were investigated in different cell compartments by using differential centrifugation and qRT-PCR. Results LNCaP siPCA3-transfected cells significantly inhibited cell growth and viability, and increased the proportion of cells in the sub G0/G1 phase of the cell cycle and the percentage of pyknotic nuclei, compared to those transfected with scramble siRNA (siSCr-transfected cells. DHT-treated LNCaP cells induced a significant upregulation of PCA3 expression, which was reversed by flutamide. In siPCA3/LNCaP-transfected cells, the expression of AR target genes was downregulated compared to siSCr-transfected cells. The siPCA3 transfection also counteracted DHT stimulatory effects on the AR signaling cascade, significantly downregulating expression of the AR target gene. Analysis of PCA3 expression in different cell compartments provided evidence that the main functional roles of PCA3 occur in the nuclei and microsomal cell fractions. Conclusions Our findings suggest that the ncRNA PCA3 is involved in the control of PCa cell survival

  4. Erythropoietin receptor signaling is membrane raft dependent.

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    Kathy L McGraw

    Full Text Available Upon erythropoietin (Epo engagement, Epo-receptor (R homodimerizes to activate JAK2 and Lyn, which phosphorylate STAT5. Although recent investigations have identified key negative regulators of Epo-R signaling, little is known about the role of membrane localization in controlling receptor signal fidelity. Here we show a critical role for membrane raft (MR microdomains in creation of discrete signaling platforms essential for Epo-R signaling. Treatment of UT7 cells with Epo induced MR assembly and coalescence. Confocal microscopy showed that raft aggregates significantly increased after Epo stimulation (mean, 4.3±1.4(SE vs. 25.6±3.2 aggregates/cell; p≤0.001, accompanied by a >3-fold increase in cluster size (p≤0.001. Raft fraction immunoblotting showed Epo-R translocation to MR after Epo stimulation and was confirmed by fluorescence microscopy in Epo stimulated UT7 cells and primary erythroid bursts. Receptor recruitment into MR was accompanied by incorporation of JAK2, Lyn, and STAT5 and their activated forms. Raft disruption by cholesterol depletion extinguished Epo induced Jak2, STAT5, Akt and MAPK phosphorylation in UT7 cells and erythroid progenitors. Furthermore, inhibition of the Rho GTPases Rac1 or RhoA blocked receptor recruitment into raft fractions, indicating a role for these GTPases in receptor trafficking. These data establish a critical role for MR in recruitment and assembly of Epo-R and signal intermediates into discrete membrane signaling units.

  5. Chronic hyperglycemia downregulates GLP-1 receptor signaling in pancreatic β-cells via protein kinase A

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    Sindhu Rajan

    2015-04-01

    Conclusions: These data show that glucose acting, at least in part, via PKA leads to the loss of the GLP-1R from the cell surface and an impairment of GLP-1R signaling, which may underlie the reduced clinical efficacy of GLP-1R based therapies in individuals with poorly controlled hyperglycemia.

  6. Estrogen stimulated migration and invasion of estrogen receptor-negative breast cancer cells involves an ezrin-dependent crosstalk between G protein-coupled receptor 30 and estrogen receptor beta signaling.

    Science.gov (United States)

    Zhou, Kewen; Sun, Peng; Zhang, Yaxing; You, Xinchao; Li, Ping; Wang, Tinghuai

    2016-07-01

    Estrogen mediates important cellular activities in estrogen receptor negative (ER-) breast cancer cells via membrane associated G protein-coupled receptor 30 (GPR30). However, the biological role and mechanism of estrogen action on cell motility and invasion in this aggressive kind of tumors remains poorly understood. We showed here that treatment with 17β-estradiol (E2) in ER-negative cancer cells resulted in ezrin-dependent cytoskeleton rearrangement and elicited a stimulatory effect on cell migration and invasion. Mechanistically, E2 induced ezrin activation was mediated by distinct mechanisms in different cell contexts. In SK-BR-3 cells with a high GPR30/ERβ ratio, silencing of GPR30 was able to abolish E2 induced ERK1/2, AKT phosphorylation and ezrin activation, whereas in MDA-MB-231 cells with low GPR30/ERβ ratio, E2 stimulated ezrin activation was mediated by the ERβ/PI3K/AKT signaling pathway. Importantly, we showed that activation of GPR30 signaling significantly prevents ERβ activation induced ezrin phosphorylation, cell migration and invasion, indicating an antagonist effect between GPR30 and ERβ signaling in MDA-MB-231 cells. These findings highlight the important interplay between different estrogen receptors in estrogen induced cell motility and invasiveness in ER-negative breast cancer cells. PMID:26850467

  7. Engagement of CD22 on B cells with the monoclonal antibody epratuzumab stimulates the phosphorylation of upstream inhibitory signals of the B cell receptor.

    Science.gov (United States)

    Lumb, Simon; Fleischer, Sarah J; Wiedemann, Annika; Daridon, Capucine; Maloney, Alison; Shock, Anthony; Dörner, Thomas

    2016-06-01

    The binding of antigen to the B cell receptor (BCR) results in a cascade of signalling events that ultimately drive B cell activation. Uncontrolled B cell activation is regulated by negative feedback loops that involve inhibitory co-receptors such as CD22 and CD32B that exert their functions following phosphorylation of immunoreceptor tyrosine-based inhibition motifs (ITIMs). The CD22-targeted antibody epratuzumab has previously been shown to inhibit BCR-driven signalling events, but its effects on ITIM phosphorylation of CD22 and CD32B have not been properly evaluated. The present study therefore employed both immunoprecipitation and flow cytometry approaches to elucidate the effects of epratuzumab on direct phosphorylation of key tyrosine (Tyr) residues on both these proteins, using both transformed B cell lines and primary human B cells. Epratuzumab induced the phosphorylation of Tyr(822) on CD22 and enhanced its co-localisation with SHP-1. Additionally, in spite of high basal phosphorylation of other key ITIMs on CD22, in primary human B cells epratuzumab also enhanced phosphorylation of Tyr(807), a residue involved in the recruitment of Grb2. Such initiation events could explain the effects of epratuzumab on downstream signalling in B cells. Finally, we were able to demonstrate that epratuzumab stimulated the phosphorylation of Tyr(292) on the low affinity inhibitory Fc receptor CD32B which would further attenuate BCR-induced signalling. Together, these data demonstrate that engagement of CD22 with epratuzumab leads to the direct phosphorylation of key upstream inhibitory receptors of BCR signalling and may help to explain how this antibody modulates B cell function. PMID:27125377

  8. Caveolin-1 Regulates the P2Y2 Receptor Signaling in Human 1321N1 Astrocytoma Cells.

    Science.gov (United States)

    Martinez, Namyr A; Ayala, Alondra M; Martinez, Magdiel; Martinez-Rivera, Freddyson J; Miranda, Jorge D; Silva, Walter I

    2016-06-01

    Damage to the CNS can cause a differential spatio-temporal release of multiple factors, such as nucleotides, ATP and UTP. The latter interact with neuronal and glial nucleotide receptors. The P2Y2 nucleotide receptor (P2Y2R) has gained prominence as a modulator of gliotic responses after CNS injury. Still, the molecular mechanisms underlying these responses in glia are not fully understood. Membrane-raft microdomains, such as caveolae, and their constituent caveolins, modulate receptor signaling in astrocytes; yet, their role in P2Y2R signaling has not been adequately explored. Hence, this study evaluated the role of caveolin-1 (Cav-1) in modulating P2Y2R subcellular distribution and signaling in human 1321N1 astrocytoma cells. Recombinant hP2Y2R expressed in 1321N1 cells and Cav-1 were found to co-fractionate in light-density membrane-raft fractions, co-localize via confocal microscopy, and co-immunoprecipitate. Raft localization was dependent on ATP stimulation and Cav-1 expression. This hP2Y2R/Cav-1 distribution and interaction was confirmed with various cell model systems differing in the expression of both P2Y2R and Cav-1, and shRNA knockdown of Cav-1 expression. Furthermore, shRNA knockdown of Cav-1 expression decreased nucleotide-induced increases in the intracellular Ca(2+) concentration in 1321N1 and C6 glioma cells without altering TRAP-6 and carbachol Ca(2+) responses. In addition, Cav-1 shRNA knockdown also decreased AKT phosphorylation and altered the kinetics of ERK1/2 activation in 1321N1 cells. Our findings strongly suggest that P2Y2R interaction with Cav-1 in membrane-raft caveolae of 1321N1 cells modulates receptor coupling to its downstream signaling machinery. Thus, P2Y2R/Cav-1 interactions represent a novel target for controlling P2Y2R function after CNS injury. PMID:27129210

  9. p38 signaling and receptor recycling events in a microfluidic endothelial cell adhesion assay.

    Directory of Open Access Journals (Sweden)

    Dwayne A L Vickers

    Full Text Available Adhesion-based microfluidic cell separation has proven to be very useful in applications ranging from cancer diagnostics to tissue engineering. This process involves functionalizing microchannel surfaces with a capture molecule. High specificity and purity capture can be achieved using this method. Despite these advances, little is known about the mechanisms that govern cell capture within these devices and their relationships to basic process parameters such as fluid shear stress and the presence of soluble factors. This work examines how the adhesion of human endothelial cells (ECs is influenced by a soluble tetrapeptide, Arg-Glu-Asp-Val (REDV and fluidic shear stress. The ability of these ECs to bind within microchannels coated with REDV is shown to be governed by shear- and soluble-factor mediated changes in p38 mitogen-activated protein kinase expression together with recycling of adhesion receptors from the endosome.

  10. Nuclear Receptor Signaling Atlas (NURSA)

    Data.gov (United States)

    U.S. Department of Health & Human Services — The Nuclear Receptor Signaling Atlas (NURSA) is designed to foster the development of a comprehensive understanding of the structure, function, and role in disease...

  11. Sub-lethal irradiation of human colorectal tumor cells imparts enhanced and sustained susceptibility to multiple death receptor signaling pathways.

    Directory of Open Access Journals (Sweden)

    Victoria Ifeadi

    Full Text Available BACKGROUND: Death receptors (DR of the TNF family function as anti-tumor immune effector molecules. Tumor cells, however, often exhibit DR-signaling resistance. Previous studies indicate that radiation can modify gene expression within tumor cells and increase tumor cell sensitivity to immune attack. The aim of this study is to investigate the synergistic effect of sub-lethal doses of ionizing radiation in sensitizing colorectal carcinoma cells to death receptor-mediated apoptosis. METHODOLOGY/PRINCIPAL FINDINGS: The ability of radiation to modulate the expression of multiple death receptors (Fas/CD95, TRAILR1/DR4, TRAILR2/DR5, TNF-R1 and LTβR was examined in colorectal tumor cells. The functional significance of sub-lethal doses of radiation in enhancing tumor cell susceptibility to DR-induced apoptosis was determined by in vitro functional sensitivity assays. The longevity of these changes and the underlying molecular mechanism of irradiation in sensitizing diverse colorectal carcinoma cells to death receptor-mediated apoptosis were also examined. We found that radiation increased surface expression of Fas, DR4 and DR5 but not LTβR or TNF-R1 in these cells. Increased expression of DRs was observed 2 days post-irradiation and remained elevated 7-days post irradiation. Sub-lethal tumor cell irradiation alone exhibited minimal cell death, but effectively sensitized three of three colorectal carcinoma cells to both TRAIL and Fas-induced apoptosis, but not LTβR-induced death. Furthermore, radiation-enhanced Fas and TRAIL-induced cell death lasted as long as 5-days post-irradiation. Specific analysis of intracellular sensitizers to apoptosis indicated that while radiation did reduce Bcl-X(L and c-FLIP protein expression, this reduction did not correlate with the radiation-enhanced sensitivity to Fas and/or TRAIL mediated apoptosis among the three cell types. CONCLUSIONS/SIGNIFICANCE: Irradiation of tumor cells can overcome Fas and TRAIL

  12. Neuropeptide Y1 receptor inhibits cell growth through inactivating mitogen-activated protein kinase signal pathway in human hepatocellular carcinoma.

    Science.gov (United States)

    Lv, Xiufang; Zhao, Fengbo; Huo, Xisong; Tang, Weidong; Hu, Baoying; Gong, Xiu; Yang, Juan; Shen, Qiujin; Qin, Wenxin

    2016-07-01

    Hepatocellular carcinoma (HCC) is one of the most common cancers, and its incidence is increasing worldwide. Neuropeptide Y (NPY) broadly expressed in the central and peripheral nervous system. It participates in multiple physiological and pathological processes through specific receptors. Evidences are accumulating that NPY is involved in development and progression in neuro- or endocrine-related cancers. However, little is known about the potential roles and underlying mechanisms of NPY receptors in HCC. In this study, we analyzed the expression of NPY receptors by real-time polymerase chain reaction, Western blot, and immunohistochemical staining. Correlation between NPY1R levels and clinicopathological characteristics, and survival of HCC patients were explored, respectively. Cell proliferation was researched by CCK-8 in vitro, and tumor growth was studied by nude mice xenografts in vivo. We found that mRNA and protein level of NPY receptor Y1 subtype (NPY1R) significantly decreased in HCC tissues. Low expression of NPY1R closely correlated with poor prognosis in HCC patients. Proliferation of HCC cells was significantly inhibited by recombinant NPY protein in vitro. This inhibitory effect could be blocked by selected NPY1R antagonist BIBP3226. Furthermore, overexpression of NPY1R could significantly inhibit HCC cell proliferation. Knockdown of NPY1R promoted cell multiplication in vitro and increased tumorigenicity and tumor growth in vivo. NPY1R was found to participate in the inhibition of cell proliferation via inactivating mitogen-activated protein kinase signal pathway in HCC cells. Collectively, NPY1R plays an inhibitory role in tumor growth and may be a promising therapeutic target for HCC. PMID:27262566

  13. Rice Bran Feruloylated Oligosaccharides Activate Dendritic Cells via Toll-Like Receptor 2 and 4 Signaling

    OpenAIRE

    Chi Chen Lin; Hua Han Chen; Yu Kuo Chen; Hung Chia Chang; Ping Yi Lin; I-Hong Pan; Der-Yuan Chen; Chuan Mu Chen; Su Yi Lin

    2014-01-01

    This work presents the effects of feruloylated oligosaccharides (FOs) of rice bran on murine bone marrow-derived dendritic cells (BMDCs) and the potential pathway through which the effects are mediated. We found that FOs induced phenotypic maturation of DCs, as shown by the increased expression of CD40, CD80/CD86 and MHC-I/II molecules. FOs efficiently induced maturation of DCs generated from C3H/HeN or C57BL/6 mice with normal toll-like receptor 4 (TLR-4) or TLR-2 but not DCs from mice with ...

  14. Activated factor X signaling via protease-activated receptor 2 suppresses pro-inflammatory cytokine production from LPS-stimulated myeloid cells.

    LENUS (Irish Health Repository)

    Gleeson, Eimear M

    2013-07-19

    Vitamin K-dependent proteases generated in response to vascular injury and infection enable fibrin clot formation, but also trigger distinct immuno-regulatory signaling pathways on myeloid cells. Factor Xa, a protease crucial for blood coagulation, also induces protease-activated receptor-dependent cell signaling. Factor Xa can bind both monocytes and macrophages, but whether factor Xa-dependent signaling stimulates or suppresses myeloid cell cytokine production in response to Toll-like receptor activation is not known. In this study, exposure to factor Xa significantly impaired pro-inflammatory cytokine production from lipopolysaccharide-treated peripheral blood mononuclear cells, THP-1 monocytic cells and murine macrophages. Furthermore, factor Xa inhibited nuclear factor-kappa B activation in THP-1 reporter cells, requiring phosphatidylinositide 3-kinase activity for its anti-inflammatory effect. Active-site blockade, γ-carboxyglutamic acid domain truncation and a peptide mimic of the factor Xa inter-epidermal growth factor-like region prevented factor Xa inhibition of lipopolysaccharide-induced tumour necrosis factor-α release. In addition, factor Xa anti-inflammatory activity was markedly attenuated by the presence of an antagonist of protease-activated receptor 2, but not protease-activated receptor 1. The key role of protease-activated receptor 2 in eliciting factor Xa-dependent anti-inflammatory signaling on macrophages was further underscored by the inability of factor Xa to mediate inhibition of tumour necrosis factor-α and interleukin-6 release from murine bone marrow-derived protease-activated receptor 2-deficient macrophages. We also show for the first time that, in addition to protease-activated receptor 2, factor Xa requires a receptor-associated protein-sensitive low-density lipoprotein receptor to inhibit lipopolysaccharide-induced cytokine production. Collectively, this study supports a novel function for factor Xa as an endogenous, receptor

  15. Complexity of Receptor Tyrosine Kinase Signal Processing

    OpenAIRE

    Volinsky, Natalia; Kholodenko, Boris N.

    2013-01-01

    Our knowledge of molecular mechanisms of receptor tyrosine kinase (RTK) signaling advances with ever-increasing pace. Yet our understanding of how the spatiotemporal dynamics of RTK signaling control specific cellular outcomes has lagged behind. Systems-centered experimental and computational approaches can help reveal how overlapping networks of signal transducers downstream of RTKs orchestrate specific cell-fate decisions. We discuss how RTK network regulatory structures, which involve the ...

  16. Muscarinic Receptor Signaling in Colon Cancer

    Directory of Open Access Journals (Sweden)

    Jean-Pierre Raufman

    2011-03-01

    Full Text Available According to the adenoma-carcinoma sequence, colon cancer results from accumulating somatic gene mutations; environmental growth factors accelerate and augment this process. For example, diets rich in meat and fat increase fecal bile acids and colon cancer risk. In rodent cancer models, increased fecal bile acids promote colon dysplasia. Conversely, in rodents and in persons with inflammatory bowel disease, low-dose ursodeoxycholic acid treatment alters fecal bile acid composition and attenuates colon neoplasia. In the course of elucidating the mechanism underlying these actions, we discovered that bile acids interact functionally with intestinal muscarinic receptors. The present communication reviews muscarinic receptor expression in normal and neoplastic colon epithelium, the role of autocrine signaling following synthesis and release of acetylcholine from colon cancer cells, post-muscarinic receptor signaling including the role of transactivation of epidermal growth factor receptors and activation of the ERK and PI3K/AKT signaling pathways, the structural biology and metabolism of bile acids and evidence for functional interaction of bile acids with muscarinic receptors on human colon cancer cells. In murine colon cancer models, deficiency of subtype 3 muscarinic receptors attenuates intestinal neoplasia; a proof-of-concept supporting muscarinic receptor signaling as a therapeutic target for colon cancer.

  17. Muscarinic Receptor Signaling in Colon Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Rosenvinge, Erik C. von, E-mail: evonrose@medicine.umaryland.edu; Raufman, Jean-Pierre [University of Maryland School of Medicine, Division of Gastroenterology & Hepatology, 22 S. Greene Street, N3W62, Baltimore, MD 21201 (United States); Department of Veterans Affairs, VA Maryland Health Care System, 10 North Greene Street, Baltimore, MD 21201 (United States)

    2011-03-02

    According to the adenoma-carcinoma sequence, colon cancer results from accumulating somatic gene mutations; environmental growth factors accelerate and augment this process. For example, diets rich in meat and fat increase fecal bile acids and colon cancer risk. In rodent cancer models, increased fecal bile acids promote colon dysplasia. Conversely, in rodents and in persons with inflammatory bowel disease, low-dose ursodeoxycholic acid treatment alters fecal bile acid composition and attenuates colon neoplasia. In the course of elucidating the mechanism underlying these actions, we discovered that bile acids interact functionally with intestinal muscarinic receptors. The present communication reviews muscarinic receptor expression in normal and neoplastic colon epithelium, the role of autocrine signaling following synthesis and release of acetylcholine from colon cancer cells, post-muscarinic receptor signaling including the role of transactivation of epidermal growth factor receptors and activation of the ERK and PI3K/AKT signaling pathways, the structural biology and metabolism of bile acids and evidence for functional interaction of bile acids with muscarinic receptors on human colon cancer cells. In murine colon cancer models, deficiency of subtype 3 muscarinic receptors attenuates intestinal neoplasia; a proof-of-concept supporting muscarinic receptor signaling as a therapeutic target for colon cancer.

  18. Muscarinic Receptor Signaling in Colon Cancer

    International Nuclear Information System (INIS)

    According to the adenoma-carcinoma sequence, colon cancer results from accumulating somatic gene mutations; environmental growth factors accelerate and augment this process. For example, diets rich in meat and fat increase fecal bile acids and colon cancer risk. In rodent cancer models, increased fecal bile acids promote colon dysplasia. Conversely, in rodents and in persons with inflammatory bowel disease, low-dose ursodeoxycholic acid treatment alters fecal bile acid composition and attenuates colon neoplasia. In the course of elucidating the mechanism underlying these actions, we discovered that bile acids interact functionally with intestinal muscarinic receptors. The present communication reviews muscarinic receptor expression in normal and neoplastic colon epithelium, the role of autocrine signaling following synthesis and release of acetylcholine from colon cancer cells, post-muscarinic receptor signaling including the role of transactivation of epidermal growth factor receptors and activation of the ERK and PI3K/AKT signaling pathways, the structural biology and metabolism of bile acids and evidence for functional interaction of bile acids with muscarinic receptors on human colon cancer cells. In murine colon cancer models, deficiency of subtype 3 muscarinic receptors attenuates intestinal neoplasia; a proof-of-concept supporting muscarinic receptor signaling as a therapeutic target for colon cancer

  19. Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase

    International Nuclear Information System (INIS)

    T-cell protein tyrosine phosphatase (TCPTP/TC45) is a ubiquitously expressed intra-cellular non-receptor protein tyrosine phosphatase involved in the negative regulation of several cancer relevant cellular signalling pathways. We have previously shown that interaction between the α-cytoplasmic tail of α1β1 integrin and TCPTP activates TCPTP by disrupting an inhibitory intra-molecular bond in TCPTP. Thus, inhibition of the regulatory interaction in TCPTP is a desirable strategy for TCPTP activation and attenuation of oncogenic RTK signalling. However, this is challenging with low molecular weight compounds. We developed a high-throughput compatible assay to analyse activity of recombinant TCPTP in vitro. Using this assay we have screened 64280 small molecules to identify novel agonists for TCPTP. Dose-dependent response to TCPTP agonist was performed using the in vitro assay. Inhibition effects and specificity of TCPTP agonists were evaluated using TCPTP expressing and null mouse embryonic fibroblasts. Western blot analysis was used to evaluate attenuation of PDGFRβ and EGFR phosphorylation. Inhibition of VEGF signalling was analysed with VEGF-induced endothelial cell sprouting assays. From the screen we identified six TCPTP agonists. Two compounds competed with α1-cytoplasmic domain for binding to TCPTP, suggesting that they activate TCPTP similar to α1-cyt by disrupting the intra-molecular bond in TCPTP. Importantly, one of the compounds (spermidine) displayed specificity towards TCPTP in cells, since TCPTP -/- cells were 43-fold more resistant to the compound than TCPTP expressing cells. This compound attenuates PDGFRβ and VEGFR2 signalling in cells in a TCPTP-dependent manner and functions as a negative regulator of EGFR phosphorylation in cancer cells. In this study we showed that small molecules mimicking TCPTP-α1 interaction can be used as TCPTP agonists. These data provide the first proof-of-concept description of the use of high-throughput screening

  20. 2B4 (CD244) signaling via chimeric receptors costimulates tumor-antigen specific proliferation and in vitro expansion of human T cells.

    Science.gov (United States)

    Altvater, Bianca; Landmeier, Silke; Pscherer, Sibylle; Temme, Jaane; Juergens, Heribert; Pule, Martin; Rossig, Claudia

    2009-12-01

    Regulatory NK cell receptors can contribute to antigen-specific adaptive immune responses by modulating T cell receptor (TCR)-induced T cell activation. We investigated the potential of the NK cell receptor 2B4 (CD244) to enhance tumor antigen-induced activation of human T cells. 2B4 is a member of the CD2 receptor subfamily with both activating and inhibitory functions in NK cells. In T cells, its expression is positively associated with the acquisition of a cytolytic effector memory phenotype. Recombinant chimeric receptors that link extracellular single-chain Fv fragments specific for the tumor-associated surface antigens CD19 and G(D2) to the signaling domains of human 2B4 and/or TCRzeta were expressed in non-specifically activated peripheral blood T cells by retroviral gene transfer. While 2B4 signaling alone failed to induce T cell effector functions or proliferation, it significantly augmented the antigen-specific activation responses induced by TCRzeta. 2B4 costimulation did not affect the predominant effector memory phenotype of expanding T cells, nor did it increase the proportion of T cells with regulatory phenotype (CD4+CD25(hi)FoxP3+). These data support a costimulatory role for 2B4 in human T cell subpopulations. As an amplifier of TCR-mediated signals, 2B4 may provide a powerful new tool for immunotherapy of cancer, promoting sustained activation and proliferation of gene-modified antitumor T cells. PMID:19360406

  1. Taurolithocholic acid promotes intrahepatic cholangiocarcinoma cell growth via muscarinic acetylcholine receptor and EGFR/ERK1/2 signaling pathway

    Science.gov (United States)

    AMONYINGCHAROEN, SUMET; SURIYO, TAWIT; THIANTANAWAT, APINYA; WATCHARASIT, PIYAJIT; SATAYAVIVAD, JUTAMAAD

    2015-01-01

    Cholangiocarcinoma (CCA) is a malignant cancer of the biliary tract and its occurrence is associated with chronic cholestasis which causes an elevation of bile acids in the liver and bile duct. The present study aimed to investigate the role and mechanistic effect of bile acids on the CCA cell growth. Intrahepatic CCA cell lines, RMCCA-1 and HuCCA-1, were treated with bile acids and their metabolites to determine the growth promoting effect. Cell viability, cell cycle analysis, EdU incorporation assays were conducted. Intracellular signaling proteins were detected by western immunoblotting. Among eleven forms of bile acids and their metabolites, only taurolithocholic acid (TLCA) concentration dependently (1–40 μM) increased the cell viability of RMCCA-1, but not HuCCA-1 cells. The cell cycle analysis showed induction of cells in the S phase and the EdU incorporation assay revealed induction of DNA synthesis in the TLCA-treated RMCCA-1 cells. Moreover, TLCA increased the phosphorylation of EGFR, ERK 1/2 and also increased the expression of cyclin D1 in RMCCA-1 cells. Furthermore, TLCA-induced RMCCA-1 cell growth could be inhibited by atropine, a non-selective muscarinic acetylcholine receptor (mAChR) antagonist, AG 1478, a specific EGFR inhibitor, or U 0126, a specific MEK 1/2 inhibitor. These results suggest that TLCA induces CCA cell growth via mAChR and EGFR/EKR1/2 signaling pathway. Moreover, the functional presence of cholinergic system plays a certain role in TLCA-induced CCA cell growth. PMID:25815516

  2. Farnesoid X receptor signal is involved in deoxycholic acid-induced intestinal metaplasia of normal human gastric epithelial cells.

    Science.gov (United States)

    Li, Shu; Chen, Xin; Zhou, Lu; Wang, Bang-Mao

    2015-11-01

    The farnesoid X receptor (FXR) signaling pathway is known to be involved in the metabolism of bile acid, glucose and lipid. In the present study, we demonstrated that 400 µmol/l deoxycholic acid (DCA) stimulation promotes the proliferation of normal human gastric epithelial cells (GES-1). In addition, DCA activated FXR and increased the expression of intestinal metaplasia genes, including caudal-related homeobox transcription factor 2 (Cdx2) and mucin 2 (MUC2). The treatment of FXR agonist GW4064/antagonist guggulsterone (Gug.) significantly increased/decreased the expression levels of FXR, Cdx2 and MUC2 protein in DCA-induced GES-1 cells. GW4064/Gug. also enhanced/reduced the nuclear factor-κB (NF-κB) activity and binding of the Cdx2 promoter region and NF-κB, the most common subunit p50 protein. Taken together, the results indicated that DCA is capable of modulating the expression of Cdx2 and the downstream MUC2 via the nuclear receptor FXR-NF-κB activity in normal gastric epithelial cells. FXR signaling pathway may therefore be involved in the intestinal metaplasia of human gastric mucosa. PMID:26324224

  3. T cell receptor-proximal signals are sustained in peripheral microclusters and terminated in the central supramolecular activation cluster.

    Science.gov (United States)

    Varma, Rajat; Campi, Gabriele; Yokosuka, Tadashi; Saito, Takashi; Dustin, Michael L

    2006-07-01

    T cell receptor (TCR) signaling is initiated and sustained in microclusters; however, it's not known whether signaling also occurs in the TCR-rich central supramolecular activation cluster (cSMAC). We showed that the cSMAC formed by fusion of microclusters contained more CD45 than microclusters and is a site enriched in lysobisphosphatidic acid, a lipid involved in sorting ubiquitinated membrane proteins for degradation. Calcium signaling via TCR was blocked within 2 min by anti-MHCp treatment and 1 min by latrunculin-A treatment. TCR-MHCp interactions in the cSMAC survived these perturbations for 10 min and hence were not sufficient to sustain signaling. TCR microclusters were also resistant to disruption by anti-MHCp and latrunculin-A treatments. We propose that TCR signaling is sustained by stabilized microclusters and is terminated in the cSMAC, a structure from which TCR are sorted for degradation. Our studies reveal a role for F-actin in TCR signaling beyond microcluster formation. PMID:16860761

  4. Role of protein kinase C and epidermal growth factor receptor signalling in growth stimulation by neurotensin in colon carcinoma cells

    International Nuclear Information System (INIS)

    Neurotensin has been found to promote colon carcinogenesis in rats and mice, and proliferation of human colon carcinoma cell lines, but the mechanisms involved are not clear. We have examined signalling pathways activated by neurotensin in colorectal and pancreatic carcinoma cells. Colon carcinoma cell lines HCT116 and HT29 and pancreatic adenocarcinoma cell line Panc-1 were cultured and stimulated with neurotensin or epidermal growth factor (EGF). DNA synthesis was determined by incorporation of radiolabelled thymidine into DNA. Levels and phosphorylation of proteins in signalling pathways were assessed by Western blotting. Neurotensin stimulated the phosphorylation of both extracellular signal-regulated kinase (ERK) and Akt in all three cell lines, but apparently did so through different pathways. In Panc-1 cells, neurotensin-induced phosphorylation of ERK, but not Akt, was dependent on protein kinase C (PKC), whereas an inhibitor of the β-isoform of phosphoinositide 3-kinase (PI3K), TGX221, abolished neurotensin-induced Akt phosphorylation in these cells, and there was no evidence of EGF receptor (EGFR) transactivation. In HT29 cells, in contrast, the EGFR tyrosine kinase inhibitor gefitinib blocked neurotensin-stimulated phosphorylation of both ERK and Akt, indicating transactivation of EGFR, independently of PKC. In HCT116 cells, neurotensin induced both a PKC-dependent phosphorylation of ERK and a metalloproteinase-mediated transactivation of EGFR that was associated with a gefitinib-sensitive phosphorylation of the downstream adaptor protein Shc. The activation of Akt was also inhibited by gefitinib, but only partly, suggesting a mechanism in addition to EGFR transactivation. Inhibition of PKC blocked neurotensin-induced DNA synthesis in HCT116 cells. While acting predominantly through PKC in Panc-1 cells and via EGFR transactivation in HT29 cells, neurotensin used both these pathways in HCT116 cells. In these cells, neurotensin-induced activation of ERK

  5. Role of protein kinase C and epidermal growth factor receptor signalling in growth stimulation by neurotensin in colon carcinoma cells

    Directory of Open Access Journals (Sweden)

    Dajani Olav

    2011-10-01

    Full Text Available Abstract Background Neurotensin has been found to promote colon carcinogenesis in rats and mice, and proliferation of human colon carcinoma cell lines, but the mechanisms involved are not clear. We have examined signalling pathways activated by neurotensin in colorectal and pancreatic carcinoma cells. Methods Colon carcinoma cell lines HCT116 and HT29 and pancreatic adenocarcinoma cell line Panc-1 were cultured and stimulated with neurotensin or epidermal growth factor (EGF. DNA synthesis was determined by incorporation of radiolabelled thymidine into DNA. Levels and phosphorylation of proteins in signalling pathways were assessed by Western blotting. Results Neurotensin stimulated the phosphorylation of both extracellular signal-regulated kinase (ERK and Akt in all three cell lines, but apparently did so through different pathways. In Panc-1 cells, neurotensin-induced phosphorylation of ERK, but not Akt, was dependent on protein kinase C (PKC, whereas an inhibitor of the β-isoform of phosphoinositide 3-kinase (PI3K, TGX221, abolished neurotensin-induced Akt phosphorylation in these cells, and there was no evidence of EGF receptor (EGFR transactivation. In HT29 cells, in contrast, the EGFR tyrosine kinase inhibitor gefitinib blocked neurotensin-stimulated phosphorylation of both ERK and Akt, indicating transactivation of EGFR, independently of PKC. In HCT116 cells, neurotensin induced both a PKC-dependent phosphorylation of ERK and a metalloproteinase-mediated transactivation of EGFR that was associated with a gefitinib-sensitive phosphorylation of the downstream adaptor protein Shc. The activation of Akt was also inhibited by gefitinib, but only partly, suggesting a mechanism in addition to EGFR transactivation. Inhibition of PKC blocked neurotensin-induced DNA synthesis in HCT116 cells. Conclusions While acting predominantly through PKC in Panc-1 cells and via EGFR transactivation in HT29 cells, neurotensin used both these pathways in HCT116

  6. PGF2α-F-prostanoid receptor signalling via ADAMTS1 modulates epithelial cell invasion and endothelial cell function in endometrial cancer

    International Nuclear Information System (INIS)

    An increase in cancer cell invasion and microvascular density is associated with a poorer prognosis for patients with endometrial cancer. In endometrial adenocarcinoma F-prostanoid (FP) receptor expression is elevated, along with its ligand prostaglandin (PG)F2α, where it regulates expression and secretion of a host of growth factors and chemokines involved in tumorigenesis. This study investigates the expression, regulation and role of a disintegrin and metalloproteinase with thrombospondin repeat 1 (ADAMTS1) in endometrial adenocarcinoma cells by PGF2α via the FP receptor. Human endometrium and adenocarcinoma tissues were obtained in accordance with Lothian Research Ethics Committee guidance with informed patient consent. Expression of ADAMTS1 mRNA and protein in tissues was determined by quantitative RT-PCR analysis and immunohistochemistry. Signal transduction pathways regulating ADAMTS1 expression in Ishikawa cells stably expressing the FP receptor to levels seen in endometrial cancer (FPS cells) were determined by quantitative RT-PCR analysis. In vitro invasion and proliferation assays were performed with FPS cells and human umbilical vein endothelial cells (HUVECs) using conditioned medium (CM) from PGF2α-treated FPS cells from which ADAMTS1 was immunoneutralised and/or recombinant ADAMTS1. The role of endothelial ADAMTS1 in endothelial cell proliferation was confirmed with RNA interference. The data in this study were analysed by T-test or ANOVA. ADAMTS1 mRNA and protein expression is elevated in endometrial adenocarcinoma tissues compared with normal proliferative phase endometrium and is localised to the glandular and vascular cells. Using FPS cells, we show that PGF2α-FP signalling upregulates ADAMTS1 expression via a calmodulin-NFAT-dependent pathway and this promotes epithelial cell invasion through ECM and inhibits endothelial cell proliferation. Furthermore, we show that CM from FPS cells regulates endothelial cell ADAMTS1 expression in a rapid

  7. Infection mobilizes hematopoietic stem cells through cooperative NOD-like receptor and Toll-like receptor signaling.

    Science.gov (United States)

    Burberry, Aaron; Zeng, Melody Y; Ding, Lei; Wicks, Ian; Inohara, Naohiro; Morrison, Sean J; Núñez, Gabriel

    2014-06-11

    Adult hematopoietic stem cells (HSCs) are maintained in specialized niches within the bone marrow under steady-state conditions and mobilize for extramedullary hematopoiesis during periods of stress such as bacterial infections. However, the underlying mechanisms are unclear. We show that systemic infection of mice with Escherichia coli, commonly associated with bacteremia in humans, mobilizes functional HSCs to the spleen. Accumulation of splenic HSCs (CD150+CD48-Lin(-/low)Sca1+cKit+) was diminished in TLR4-deficient and RIPK2-deficient mice, implicating TLRs and cytosolic NOD1/NOD2 signaling in the process. Accordingly, dual stimulation of NOD1 and TLR4 in radio-resistant cells alone was sufficient to mobilize HSCs, while TLR4 expression on HSCs was dispensable. Mechanistically, TLR4 and NOD1 synergistically induced granulocyte colony-stimulating factor (G-CSF), which was required for extramedullary HSC accumulation. Mobilized HSCs and progenitor cells gave rise to neutrophils and monocytes and contributed to limiting secondary infection. PMID:24882704

  8. Flagellin-induced tolerance of the Toll-like receptor 5 signaling pathway in polarized intestinal epithelial cells.

    Science.gov (United States)

    Sun, Jun; Fegan, Pamela E; Desai, Anjali S; Madara, James L; Hobert, Michael E

    2007-03-01

    Salmonella typhimurium is a gram-negative enteric pathogen that invades the mucosal epithelium and is associated with diarrheal illness in humans. Flagellin from S. typhimurium and other gram-negative bacteria has been shown to be the predominant proinflammatory mediator through activation of the basolateral Toll-like receptor 5 (TLR5). Recent evidence has shown that prior exposure can render immune cells tolerant to subsequent challenges by TLR ligands. Accordingly, we examined whether prior exposure to purified flagellin would render human intestinal epithelial cells insensitive to future contact. We found that flagellin-induced tolerance is common to polarized epithelial cells and prevents further activation of proinflammatory signaling cascades by both purified flagellin and Salmonella bacteria but does not affect TNF-alpha stimulation of the same pathways. Flagellin tolerance is a rapid process that does not require protein synthesis, and that occurs within 1 to 2 h of flagellin exposure. Prolonged flagellin exposure blocks activation of the NF-kappaB, MAPK, and phosphoinositol 3-kinase signaling pathways and results in the internalization of a fraction of the basolateral TLR5 without affecting the polarity or total expression of TLR5. After removal of flagellin, cells require more than 24 h to fully recover their ability to mount a normal proinflammatory response. We have found that activation of phosphoinositol 3-kinase and Akt by flagellin has a small damping effect in the early stages of flagellin signaling but is not responsible for tolerance. Our study indicates that inhibition of TLR5-associated IL-1 receptor-associated kinase-4 activity occurs during the development of flagellin tolerance and is likely to be the cause of tolerance. PMID:17138965

  9. Camptothecin disrupts androgen receptor signaling and suppresses prostate cancer cell growth

    International Nuclear Information System (INIS)

    The androgen receptor (AR) is the main therapeutic target for treatment of metastatic prostate cancers. The present study demonstrates that the topoisomerase I inhibitor camptothecin selectively inhibits androgen-responsive growth of prostate cancer cells. Camptothecin strikingly inhibited mutated and wild-type AR protein expression in LNCaP and PC-3/AR cells. This inhibition coincided with decreased androgen-mediated AR phosphorylation at Ser81 and reduced androgen-mediated AR transcriptional activity in a dose-dependent manner. Additionally, camptothecin disrupted the association between AR and heat shock protein 90 and impeded binding of the synthetic androgen [3H]R1881 to AR in LNCaP cells. Camptothecin also blocked androgen-induced AR nuclear translocation, leading to downregulation of the AR target gene PSA. In addition to decreasing the intracellular and secreted prostate-specific antigen (PSA) levels, camptothecin markedly inhibited androgen-stimulated PSA promoter activity. Collectively, our data reveal that camptothecin not only serves as a traditional genotoxic agent but, by virtue of its ability to target and disrupt AR, may also be a novel candidate for the treatment of prostate cancer.

  10. PLK1 Signaling in Breast Cancer Cells Cooperates with Estrogen Receptor-Dependent Gene Transcription

    Directory of Open Access Journals (Sweden)

    Michael Wierer

    2013-06-01

    Full Text Available Polo-like kinase 1 (PLK1 is a key regulator of cell division and is overexpressed in many types of human cancers. Compared to its well-characterized role in mitosis, little is known about PLK1 functions in interphase. Here, we report that PLK1 mediates estrogen receptor (ER-regulated gene transcription in human breast cancer cells. PLK1 interacts with ER and is recruited to ER cis-elements on chromatin. PLK1-coactivated genes included classical ER target genes such as Ps2, Wisp2, and Serpina3 and were enriched in developmental and tumor-suppressive functions. Performing large-scale phosphoproteomics of estradiol-treated MCF7 cells in the presence or absence of the specific PLK1 inhibitor BI2536, we identified several PLK1 end targets involved in transcription, including the histone H3K4 trimethylase MLL2, the function of which on ER target genes was impaired by PLK1 inhibition. Our results propose a mechanism for the tumor-suppressive role of PLK1 in mammals as an interphase transcriptional regulator.

  11. Camptothecin disrupts androgen receptor signaling and suppresses prostate cancer cell growth

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Shicheng, E-mail: liusc59@yahoo.co.jp [Research and Development Department, Nipro Patch Co., Ltd., 8-1, Minamisakae-cho, Kasukabe, Saitama 344-0057 (Japan); Yuan, Yiming [Department of Geriatrics, West China Hospital, Sichuan University, Chengdu 610041 (China); Okumura, Yutaka; Shinkai, Norihiro; Yamauchi, Hitoshi [Research and Development Department, Nipro Patch Co., Ltd., 8-1, Minamisakae-cho, Kasukabe, Saitama 344-0057 (Japan)

    2010-04-02

    The androgen receptor (AR) is the main therapeutic target for treatment of metastatic prostate cancers. The present study demonstrates that the topoisomerase I inhibitor camptothecin selectively inhibits androgen-responsive growth of prostate cancer cells. Camptothecin strikingly inhibited mutated and wild-type AR protein expression in LNCaP and PC-3/AR cells. This inhibition coincided with decreased androgen-mediated AR phosphorylation at Ser{sup 81} and reduced androgen-mediated AR transcriptional activity in a dose-dependent manner. Additionally, camptothecin disrupted the association between AR and heat shock protein 90 and impeded binding of the synthetic androgen [{sup 3}H]R1881 to AR in LNCaP cells. Camptothecin also blocked androgen-induced AR nuclear translocation, leading to downregulation of the AR target gene PSA. In addition to decreasing the intracellular and secreted prostate-specific antigen (PSA) levels, camptothecin markedly inhibited androgen-stimulated PSA promoter activity. Collectively, our data reveal that camptothecin not only serves as a traditional genotoxic agent but, by virtue of its ability to target and disrupt AR, may also be a novel candidate for the treatment of prostate cancer.

  12. Heterologous transmembrane signaling by a human insulin receptor-v-ros hybrid in Chinese hamster ovary cells

    International Nuclear Information System (INIS)

    A hybrid receptor molecule composed of the extracellular ligand-binding domain of the human insulin receptor and the transmembrane and cytoplasmic (protein-tyrosine kinase) domains of the chicken sarcoma virus UR2 transforming protein p68/sup gag-ros/ has been constructed and expressed in Chinese hamster ovary (CHO) cells. The hybrid is processed normally into α and hybrid β subunits, is expressed on the cell surface at high levels, and binds insulin with near-wild-type affinity. Furthermore, insulin stimulates the phosphorylation on tyrosine resides of the hybrid β-subunit in vivo and the phosphorylation of an exogeneous substrate [poly(Glu,Tyr)] in vitro. Thus the hybrid is capable of heterologous transmembrane signaling. However, the hybrid mediates neither the insulin-activated uptake of 2-deoxyglucose nor the incorporation of [3H]thymidine into DNA, suggesting that the physiological response(s) mediated by ligand-activated protein-tyrosine kinases may utilize distinct intracellular mechanisms for postreceptor signaling

  13. Heterologous transmembrane signaling by a human insulin receptor-v-ros hybrid in Chinese hamster ovary cells

    Energy Technology Data Exchange (ETDEWEB)

    Ellis, L.; Morgan, D.O.; Jong, S.M.; Wang, L.H.; Roth, R.A.; Rutter, W.J.

    1987-08-01

    A hybrid receptor molecule composed of the extracellular ligand-binding domain of the human insulin receptor and the transmembrane and cytoplasmic (protein-tyrosine kinase) domains of the chicken sarcoma virus UR2 transforming protein p68/sup gag-ros/ has been constructed and expressed in Chinese hamster ovary (CHO) cells. The hybrid is processed normally into ..cap alpha.. and hybrid ..beta.. subunits, is expressed on the cell surface at high levels, and binds insulin with near-wild-type affinity. Furthermore, insulin stimulates the phosphorylation on tyrosine resides of the hybrid ..beta..-subunit in vivo and the phosphorylation of an exogeneous substrate (poly(Glu,Tyr)) in vitro. Thus the hybrid is capable of heterologous transmembrane signaling. However, the hybrid mediates neither the insulin-activated uptake of 2-deoxyglucose nor the incorporation of (/sup 3/H)thymidine into DNA, suggesting that the physiological response(s) mediated by ligand-activated protein-tyrosine kinases may utilize distinct intracellular mechanisms for postreceptor signaling

  14. Antagonism of Dopamine Receptor 2 Long Affects Cannabinoid Receptor 1 Signaling in a Cell Culture Model of Striatal Medium Spiny Projection Neurons.

    Science.gov (United States)

    Bagher, Amina M; Laprairie, Robert B; Kelly, Melanie E M; Denovan-Wright, Eileen M

    2016-06-01

    Activation of dopamine receptor 2 long (D2L) switches the signaling of type 1 cannabinoid receptor (CB1) from Gαi to Gαs, a process thought to be mediated through CB1-D2L heteromerization. Given the clinical importance of D2 antagonists, the goal of this study was to determine if D2 antagonists could modulate CB1 signaling. Interactions between CB1 and D2L, Gαi, Gαs, and β-arrestin1 were studied using bioluminescence resonance energy transfer 2 (BRET(2)) in STHdh(Q7/Q7) cells. CB1-dependent extracellular regulated kinase (ERK)1/2, CREB phosphorylation, and CB1 internalization following cotreatment of CB1 agonist and D2 antagonist were quantified. Preassembled CB1-Gαi complexes were detected by BRET(2) Arachidonyl-2'-chloroethylamide (ACEA), a selective CB1 agonist, caused a rapid and transient increase in BRET efficiency (BRETEff) between Gαi-Rluc and CB1-green fluorescent protein 2 (GFP(2)), and a Gαi-dependent increase in ERK phosphorylation. Physical interactions between CB1 and D2L were observed using BRET(2) Cotreatment of STHdh(Q7/Q7) cells with ACEA and haloperidol, a D2 antagonist, inhibited BRETEff signals between Gαi-Rluc and CB1-GFP(2) and reduced the EMax and pEC50 of ACEA-mediated Gαi-dependent ERK phosphorylation. ACEA and haloperidol cotreatments produced a delayed and sustained increase in BRETEff between Gαs-Rluc and CB1-GFP(2) and increased the EMax and pEC50 of ACEA-induced Gαs-dependent cAMP response element-binding protein phosphorylation. In cells expressing CB1 and D2L treated with ACEA, binding of haloperidol to D2 receptors switched CB1 coupling from Gαi to Gαs In addition, haloperidol treatment reduced ACEA-induced β-arrestin1 recruitment to CB1 and CB1 internalization. D2 antagonists allosterically modulate cannabinoid-induced CB1 coupling, signaling, and β-arrestin1 recruitment through binding to CB1-D2L heteromers. These findings indicate that D2 antagonism, like D2 agonists, change agonist-mediated CB1 coupling and

  15. RAG-mediated DNA double-strand breaks activate a cell type-specific checkpoint to inhibit pre-B cell receptor signals.

    Science.gov (United States)

    Bednarski, Jeffrey J; Pandey, Ruchi; Schulte, Emily; White, Lynn S; Chen, Bo-Ruei; Sandoval, Gabriel J; Kohyama, Masako; Haldar, Malay; Nickless, Andrew; Trott, Amanda; Cheng, Genhong; Murphy, Kenneth M; Bassing, Craig H; Payton, Jacqueline E; Sleckman, Barry P

    2016-02-01

    DNA double-strand breaks (DSBs) activate a canonical DNA damage response, including highly conserved cell cycle checkpoint pathways that prevent cells with DSBs from progressing through the cell cycle. In developing B cells, pre-B cell receptor (pre-BCR) signals initiate immunoglobulin light (Igl) chain gene assembly, leading to RAG-mediated DNA DSBs. The pre-BCR also promotes cell cycle entry, which could cause aberrant DSB repair and genome instability in pre-B cells. Here, we show that RAG DSBs inhibit pre-BCR signals through the ATM- and NF-κB2-dependent induction of SPIC, a hematopoietic-specific transcriptional repressor. SPIC inhibits expression of the SYK tyrosine kinase and BLNK adaptor, resulting in suppression of pre-BCR signaling. This regulatory circuit prevents the pre-BCR from inducing additional Igl chain gene rearrangements and driving pre-B cells with RAG DSBs into cycle. We propose that pre-B cells toggle between pre-BCR signals and a RAG DSB-dependent checkpoint to maintain genome stability while iteratively assembling Igl chain genes. PMID:26834154

  16. Hypoxia attenuates inflammatory mediators production induced by Acanthamoeba via Toll-like receptor 4 signaling in human corneal epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Hong [Department of Ophthalmology, Qilu Hospital, Shandong University, 107, Wenhua Xi Road, Jinan 250012 (China); The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Public Health, Qilu Hospital, Shandong University, 107, Wenhua Xi Road, Jinan 250012 (China); Wu, Xinyi, E-mail: xywu8868@163.com [Department of Ophthalmology, Qilu Hospital, Shandong University, 107, Wenhua Xi Road, Jinan 250012 (China)

    2012-04-13

    Highlights: Black-Right-Pointing-Pointer Hypoxia attenuates Acanthamoeba-induced the production of IL-8 and IFN-{beta}. Black-Right-Pointing-Pointer Hypoxia inhibits TLR4 expression in a time-dependent manner in HCECs. Black-Right-Pointing-Pointer Hypoxia inhibits Acanthamoeba-induced the activation of NF-{kappa}B and ERK1/2 in HCECs. Black-Right-Pointing-Pointer Hypoxia decreases Acanthamoeba-induced inflammatory response via TLR4 signaling. Black-Right-Pointing-Pointer LPS-induced the secretion of IL-6 and IL-8 is abated by hypoxia via TLR4 signaling. -- Abstract: Acanthamoeba keratitis (AK) is a vision-threatening corneal infection that is intimately associated with contact lens use which leads to hypoxic conditions on the corneal surface. However, the effect of hypoxia on the Acanthamoeba-induced host inflammatory response of corneal epithelial cells has not been studied. In the present study, we investigated the effect of hypoxia on the Acanthamoeba-induced production of inflammatory mediators interleukin-8 (IL-8) and interferon-{beta} (IFN-{beta}) in human corneal epithelial cells and then evaluated its effects on the Toll-like receptor 4 (TLR4) signaling, including TLR4 and myeloid differentiation primary response gene (88) (MyD88) expression as well as the activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-{kappa}B) and extracellular signal-regulated kinases 1/2 (ERK1/2). We then studied the effect of hypoxia on a TLR4-specific inflammatory response triggered by the TLR4 ligand lipopolysaccharide (LPS). Our data showed that hypoxia significantly decreased the production of IL-8 and IFN-{beta}. Furthermore, hypoxia attenuated Acanthamoeba-triggered TLR4 expression as well as the activation of NF-{kappa}B and ERK1/2, indicating that hypoxia abated Acanthamoeba-induced inflammatory responses by affecting TLR4 signaling. Hypoxia also inhibited LPS-induced IL-6 and IL-8 secretion, myeloid differentiation primary response gene (88

  17. Ca2+ is involved in muscarine-acetylcholine-receptor-mediated acetylcholine signal transduction in guard cells of Vicia faba L.

    Institute of Scientific and Technical Information of China (English)

    MENG Fanxia; MIAO Long; ZHANG Shuqiu; LOU Chenghou

    2004-01-01

    Acetylcholine (ACh) is an important neurochemical transmitter in animals; it also exists in plants and plays a significant role in various kinds of physiological functions in plants. ACh has been known to induce the stomatal opening. By monitoring the changes of cytosolic Ca2+ with fluorescent probe Fluo-3 AM under the confocal microscopy,we found that exogenous ACh increased cytosolic Ca2+ concentration of guard cells of Vicia faba L. Muscarine, an agonist of muscarine acetylcholine receptor (mAChR), could do so as well. In contrast, atropine, the antagonist of mAChR abolished the ability of ACh to increase Ca2+ in guard cells.This mechanism is similar to mAChR in animals. When EGTA was used to chelate Ca2+ or ruthenium red to block Ca2+ released from vacuole respectively, the results showed that the increased cytosolic Ca2+ mainly come from intracellular Ca2+ store. The evidence supports that Ca2+ is involved in guard-cell response to ACh and that Ca2+ signal is coupled to mAChRs in ACh signal transduction in guard cells.

  18. Crosstalk between Wnt/β-catenin and estrogen receptor signaling synergistically promotes osteogenic differentiation of mesenchymal progenitor cells.

    Directory of Open Access Journals (Sweden)

    Yanhong Gao

    Full Text Available Osteogenic differentiation from mesenchymal progenitor cells (MPCs are initiated and regulated by a cascade of signaling events. Either Wnt/β-catenin or estrogen signaling pathway has been shown to play an important role in regulating skeletal development and maintaining adult tissue homeostasis. Here, we investigate the potential crosstalk and synergy of these two signaling pathways in regulating osteogenic differentiation of MPCs. We find that the activation of estrogen receptor (ER signaling by estradiol (E2 or exogenously expressed ERα in MPCs synergistically enhances Wnt3A-induced early and late osteogenic markers, as well as matrix mineralization. The E2 or ERα-mediated synergy can be effectively blocked by ERα antagonist tamoxifen. E2 stimulation can enhance endochondral ossification of Wnt3A-transduced mouse fetal limb explants. Furthermore, exogenously expressed ERα significantly enhances the maturity and mineralization of Wnt3A-induced subcutaneous and intramuscular ectopic bone formation. Mechanistically, we demonstrate that E2 does not exert any detectable effect on β-catenin/Tcf reporter activity. However, ERα expression is up-regulated within the first 48h in AdWnt3A-transduced MPCs, whereas ERβ expression is significantly inhibited within 24h. Moreover, the key enzyme for the biosynthesis of estrogens aromatase is modulated by Wnt3A in a biphasic manner, up-regulated at 24h but reduced after 48h. Our results demonstrate that, while ER signaling acts synergistically with Wnt3A in promoting osteogenic differentiation, Wnt3A may crosstalk with ER signaling by up-regulating ERα expression and down-regulating ERβ expression in MPCs. Thus, the signaling crosstalk and synergy between these two pathways should be further explored as a potential therapeutic approach to combating bone and skeletal disorders, such as fracture healing and osteoporosis.

  19. Phosphoproteomic analysis of adhesion receptor signalling

    OpenAIRE

    Robertson, Joseph

    2014-01-01

    The binding of integrin adhesion receptors to their extracellular matrix (ECM) ligands activates intracellular signalling pathways that control diverse and fundamental aspects of cell behaviour. While it is clear that protein kinases and phosphatases play an integral role in such adhesion-mediated signalling, current knowledge of the phosphorylation events regulated downstream of integrin ligation is limited and prohibits a systems-level understanding of the molecular mechanisms through which...

  20. Muscarinic receptor signaling and colon cancer progression

    Institute of Scientific and Technical Information of China (English)

    Guofeng Xie; Jean-Pierre Raufman

    2016-01-01

    Due to the lack of effective treatments, advanced colorectal cancer (CRC) remains a leading cause of cancer death in the United States. Emerging evidence supports the observation that muscarinic receptor (MR) signaling plays a critical role in growth and progression of CRC. MR activation by acetylcholine and bile acids results in transactivation of epidermal growth factor receptors (EGFR) and post-EGFR signal transduction that enhances cell proliferation, migration, and invasion. Here, the authors review recent progress in understanding the molecular mechanisms underlying MR-mediated CRC progression and its therapeutic implications.

  1. An additive interaction between the NFκB and estrogen receptor signalling pathways in human endometrial epithelial cells

    Science.gov (United States)

    King, A.E.; Collins, F.; Klonisch, T.; Sallenave, J.-M.; Critchley, H.O.D.; Saunders, P.T.K.

    2010-01-01

    BACKGROUND Human embryo implantation is regulated by estradiol (E2), progesterone and locally produced mediators including interleukin-1β (IL-1β). Interactions between the estrogen receptor (ER) and NF kappa B (NFκB) signalling pathways have been reported in other systems but have not been detailed in human endometrium. METHODS AND RESULTS Real-time PCR showed that mRNA for the p65 and p105 NFκB subunits is maximally expressed in endometrium from the putative implantation window. Both subunits are localized in the endometrial epithelium throughout the menstrual cycle. Reporter assays for estrogen response element (ERE) activity were used to examine functional interactions between ER and NFκB in telomerase immortalized endometrial epithelial cells (TERT-EEC). E2 and IL-1β treatment of TERT-EECs enhances ERE activity by a NFκB and ER dependent mechanism; this effect could be mediated by ERα or ERβ. E2 and IL-1β also positively interact to increase endogenous gene expression of prostaglandin E synthase and c-myc. This is a gene-dependent action as there is no additive effect on cyclin D1 or progesterone receptor expression. CONCLUSION In summary, we have established that NFκB signalling proteins are expressed in normal endometrium and report that IL-1β can enhance the actions of E2 in a cell line derived from healthy endometrium. This mechanism may allow IL-1β, possibly from the developing embryo, to modulate the function of the endometrial epithelium to promote successful implantation, for example by regulating prostaglandin production. Aberrations in the interaction between the ER and NFκB signalling pathways may have a negative impact on implantation contributing to pathologies such as early pregnancy loss and infertility. PMID:19955102

  2. A therapeutic anti-CD4 monoclonal antibody inhibits T cell receptor signal transduction in mouse autoimmune cardiomyopathy

    Institute of Scientific and Technical Information of China (English)

    WANG Zhao-hui; LIAO Yu-hua; YUAN Jing; ZHANG Li; WANG Min; ZHANG Jing-hui; LIU Zhong-ping; DONG Ji-hua

    2007-01-01

    Background T cell immune abnormalities in patients with dilated cardiomyopathy (DCM) has been intensively studied over the past 10 years. Our previous study has suggested that immunization of mice with the peptides derived from human adenine nucleotide translocator (ANT) result in the production of autoantibodies against the ANT and histopathological changes similar to those in human DCM. The ANT peptides can induce autoimmune cardiomyopathy like DCM in Balb/c mice. In this study we aimed to focus on the molecular mechanism of T cells in the autoimmune cardiomyopathy mouse model by detecting the expression of the two T cell signaling molecules.Methods The ANT peptides were used to cause autoimmune cardiomyopathy in Balb/c mice. Anti-L3T4 or rat anti-mouse IgG was administered to the mice (n=6 in each group) simultaneously immunized with ANT. ELISA analysis was used to detect autoantibodies against the ANT peptides and the percentages of interferon-Y and interleukin-4 producing cells among splenic CD4+ lymphocytes was determined by using flow cytometry analysis. The expression of CD45 in spleen T cells was determined by immunohistochemistry and the mRNAs of T cell signaling molecules were detected by real-time PCR.Results Treatment of ANT immunized Balb/c mice with anti-CD4 mAb caused a reduction in the gene expression of P56lck and Zap-70 and a lower level of CD45 expression by spleen T cells. Aiso, a reverse of the Th1/Th2 ratio that results in the reduced production of antibodies against ANT was found in the anti-CD4 monoclonal antibodies (mAb)group. Whereas irrelevant antibody (rat anti-mouse IgG) did not suppress T cell signaling molecules nor inhibit CD45 expression, and control-antibody mice did not show any significant differences compared with the DCM group.Conclusion The results show that anti-CD4 mAb is a powerful inhibitor of the early initiating events of T cell receptor(TCR) signal transduction in mouse autoimmune dilated cardiomyopathy.

  3. Lipid rafts are required for signal transduction by angiotensin II receptor type 1 in neonatal glomerular mesangial cells

    International Nuclear Information System (INIS)

    Angiotensin II (ANG-II) receptors (AGTRs) contribute to renal physiology and pathophysiology, but the underlying mechanisms that regulate AGTR function in glomerular mesangium are poorly understood. Here, we show that AGTR1 is the functional AGTR subtype expressed in neonatal pig glomerular mesangial cells (GMCs). Cyclodextrin (CDX)-mediated cholesterol depletion attenuated cell surface AGTR1 protein expression and ANG-II-induced intracellular Ca2+ ([Ca2+]i) elevation in the cells. The COOH-terminus of porcine AGTR1 contains a caveolin (CAV)-binding motif. However, neonatal GMCs express CAV-1, but not CAV-2 and CAV-3. Colocalization and in situ proximity ligation assay detected an association between endogenous AGTR1 and CAV-1 in the cells. A synthetic peptide corresponding to the CAV-1 scaffolding domain (CSD) sequence also reduced ANG-II-induced [Ca2+]i elevation in the cells. Real-time imaging of cell growth revealed that ANG-II stimulates neonatal GMC proliferation. ANG-II-induced GMC growth was attenuated by EMD 66684, an AGTR1 antagonist; BAPTA, a [Ca2+]i chelator; KN-93, a Ca2+/calmodulin-dependent protein kinase II inhibitor; CDX; and a CSD peptide, but not PD 123319, a selective AGTR2 antagonist. Collectively, our data demonstrate [Ca2+]i-dependent proliferative effect of ANG-II and highlight a critical role for lipid raft microdomains in AGTR1-mediated signal transduction in neonatal GMCs. - Highlights: • AGTR1 is the functional AGTR subtype expressed in neonatal mesangial cells. • Endogenous AGTR1 associates with CAV-1 in neonatal mesangial cells. • Lipid raft disruption attenuates cell surface AGTR1 protein expression. • Lipid raft disruption reduces ANG-II-induced [Ca2+]i elevation in neonatal mesangial cells. • Lipid raft disruption inhibits ANG-II-induced neonatal mesangial cell growth

  4. Orexin A Affects INS-1 Rat Insulinoma Cell Proliferation via Orexin Receptor 1 and the AKT Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Li Chen

    2013-01-01

    Full Text Available Our aim is to investigate the role of the AKT/PKB (protein kinase B signaling pathway acting via orexin receptor 1 (OX1R and the effects of orexin A (OXA on cell proliferation in the insulin-secreting beta-cell line (INS-1 cells. Rat INS-1 cells were exposed to different concentrations of OXA in vitro and treated with OX1R antagonist (SB334867, PI3K antagonist (wortmannin, AKT antagonist (PF-04691502, or negative control. INS-1 amount of cell proliferation, viability and apoptosis, insulin secretion, OX1R protein expression, caspase-3 activity, and AKT protein levels were determined. We report that OXA (10-10 to 10-6 M stimulates INS-1 cell proliferation and viability, reduces the proapoptotic activity of caspase-3 to protect against apoptotic cell death, and increases insulin secretion. Additionally, AKT phosphorylation was stimulated by OXA (10-10 to 10-6 M. However, the OX1R antagonist SB334867 (10-6 M, the PI3K antagonist wortmannin (10-8 M, the AKT antagonist PF-04691502 (10-6 M, or the combination of both abolished the effects of OXA to a certain extent. These results suggest that the upregulation of OXA-OX1R mediated by AKT activation may inhibit cell apoptosis and promote cell proliferation in INS-1 cells. This finding provides functional evidence of the biological actions of OXA in rat insulinoma cells.

  5. Orexin A Affects INS-1 Rat Insulinoma Cell Proliferation via Orexin Receptor 1 and the AKT Signaling Pathway

    Science.gov (United States)

    Chen, Li; Zhao, Yuyan; Zheng, Delu; Ju, Shujing; Shen, Yang; Guo, Lei

    2013-01-01

    Our aim is to investigate the role of the AKT/PKB (protein kinase B) signaling pathway acting via orexin receptor 1 (OX1R) and the effects of orexin A (OXA) on cell proliferation in the insulin-secreting beta-cell line (INS-1 cells). Rat INS-1 cells were exposed to different concentrations of OXA in vitro and treated with OX1R antagonist (SB334867), PI3K antagonist (wortmannin), AKT antagonist (PF-04691502), or negative control. INS-1 amount of cell proliferation, viability and apoptosis, insulin secretion, OX1R protein expression, caspase-3 activity, and AKT protein levels were determined. We report that OXA (10−10 to 10−6 M) stimulates INS-1 cell proliferation and viability, reduces the proapoptotic activity of caspase-3 to protect against apoptotic cell death, and increases insulin secretion. Additionally, AKT phosphorylation was stimulated by OXA (10−10 to 10−6 M). However, the OX1R antagonist SB334867 (10−6 M), the PI3K antagonist wortmannin (10−8 M), the AKT antagonist PF-04691502 (10−6 M), or the combination of both abolished the effects of OXA to a certain extent. These results suggest that the upregulation of OXA-OX1R mediated by AKT activation may inhibit cell apoptosis and promote cell proliferation in INS-1 cells. This finding provides functional evidence of the biological actions of OXA in rat insulinoma cells. PMID:24382962

  6. Lipid rafts are required for signal transduction by angiotensin II receptor type 1 in neonatal glomerular mesangial cells

    Energy Technology Data Exchange (ETDEWEB)

    Adebiyi, Adebowale, E-mail: aadebiyi@uthsc.edu; Soni, Hitesh; John, Theresa A.; Yang, Fen

    2014-05-15

    Angiotensin II (ANG-II) receptors (AGTRs) contribute to renal physiology and pathophysiology, but the underlying mechanisms that regulate AGTR function in glomerular mesangium are poorly understood. Here, we show that AGTR1 is the functional AGTR subtype expressed in neonatal pig glomerular mesangial cells (GMCs). Cyclodextrin (CDX)-mediated cholesterol depletion attenuated cell surface AGTR1 protein expression and ANG-II-induced intracellular Ca{sup 2+} ([Ca{sup 2+}]{sub i}) elevation in the cells. The COOH-terminus of porcine AGTR1 contains a caveolin (CAV)-binding motif. However, neonatal GMCs express CAV-1, but not CAV-2 and CAV-3. Colocalization and in situ proximity ligation assay detected an association between endogenous AGTR1 and CAV-1 in the cells. A synthetic peptide corresponding to the CAV-1 scaffolding domain (CSD) sequence also reduced ANG-II-induced [Ca{sup 2+}]{sub i} elevation in the cells. Real-time imaging of cell growth revealed that ANG-II stimulates neonatal GMC proliferation. ANG-II-induced GMC growth was attenuated by EMD 66684, an AGTR1 antagonist; BAPTA, a [Ca{sup 2+}]{sub i} chelator; KN-93, a Ca{sup 2+}/calmodulin-dependent protein kinase II inhibitor; CDX; and a CSD peptide, but not PD 123319, a selective AGTR2 antagonist. Collectively, our data demonstrate [Ca{sup 2+}]{sub i}-dependent proliferative effect of ANG-II and highlight a critical role for lipid raft microdomains in AGTR1-mediated signal transduction in neonatal GMCs. - Highlights: • AGTR1 is the functional AGTR subtype expressed in neonatal mesangial cells. • Endogenous AGTR1 associates with CAV-1 in neonatal mesangial cells. • Lipid raft disruption attenuates cell surface AGTR1 protein expression. • Lipid raft disruption reduces ANG-II-induced [Ca{sup 2+}]{sub i} elevation in neonatal mesangial cells. • Lipid raft disruption inhibits ANG-II-induced neonatal mesangial cell growth.

  7. The EP4 receptor antagonist, L-161,982, blocks prostaglandin E2-induced signal transduction and cell proliferation in HCA-7 colon cancer cells

    International Nuclear Information System (INIS)

    Accumulating evidence indicates that elevated levels of prostaglandin E2 (PGE2) can increase intestinal epithelial cell proliferation, and thus play a role in colorectal tumorigenesis. PGE2 exerts its effects through four G-protein-coupled PGE receptor (EP) subtypes, named the EP1, EP2, EP3, and EP4. Increased phosphorylation of extracellular regulated kinases (ERK1/2) is required for PGE2 to stimulate cell proliferation of human colon cancer cells. However, the EP receptor(s) that are involved in this process remain unknown. We provide evidence that L-161,982, a selective EP4 receptor antagonist, completely blocks PGE2-induced ERK phosphorylation and cell proliferation of HCA-7 cells. In order to identify downstream target genes of ERK1/2 signaling, we found that PGE2 induces expression of early growth response gene-1 (EGR-1) downstream of ERK1/2 and regulates its expression at the level of transcription. PGE2 treatment induces phosphorylation of cyclic AMP response element binding protein (CREB) at Ser133 residue and CRE-mediated luciferase activity in HCA-7 cells. Studies with dominant-negative CREB mutant (ACREB) provide clear evidence for the involvement of CREB in PGE2 driven egr-1 transcription in HCA-7 cells. In conclusion, this study reveals that egr-1 is a target gene of PGE2 in HCA-7 cells and is regulated via the newly identified EP4/ERK/CREB pathway. Finally our results support the notion that antagonizing EP4 receptors may provide a novel therapeutic approach to the treatment of colon cancer

  8. Functional characterization of FLT3 receptor signaling deregulation in acute myeloid leukemia by single cell network profiling (SCNP.

    Directory of Open Access Journals (Sweden)

    David B Rosen

    Full Text Available BACKGROUND: Molecular characterization of the FMS-like tyrosine kinase 3 receptor (FLT3 in cytogenetically normal acute myeloid leukemia (AML has recently been incorporated into clinical guidelines based on correlations between FLT3 internal tandem duplications (FLT3-ITD and decreased disease-free and overall survival. These mutations result in constitutive activation of FLT3, and FLT3 inhibitors are currently undergoing trials in AML patients selected on FLT3 molecular status. However, the transient and partial responses observed suggest that FLT3 mutational status alone does not provide complete information on FLT3 biological activity at the individual patient level. Examination of variation in cellular responsiveness to signaling modulation may be more informative. METHODOLOGY/PRINCIPAL FINDINGS: Using single cell network profiling (SCNP, cells were treated with extracellular modulators and their functional responses were quantified by multiparametric flow cytometry. Intracellular signaling responses were compared between healthy bone marrow myeloblasts (BMMb and AML leukemic blasts characterized as FLT3 wild type (FLT3-WT or FLT3-ITD. Compared to healthy BMMb, FLT3-WT leukemic blasts demonstrated a wide range of signaling responses to FLT3 ligand (FLT3L, including elevated and sustained PI3K and Ras/Raf/Erk signaling. Distinct signaling and apoptosis profiles were observed in FLT3-WT and FLT3-ITD AML samples, with more uniform signaling observed in FLT3-ITD AML samples. Specifically, increased basal p-Stat5 levels, decreased FLT3L induced activation of the PI3K and Ras/Raf/Erk pathways, decreased IL-27 induced activation of the Jak/Stat pathway, and heightened apoptotic responses to agents inducing DNA damage were observed in FLT3-ITD AML samples. Preliminary analysis correlating these findings with clinical outcomes suggests that classification of patient samples based on signaling profiles may more accurately reflect FLT3 signaling

  9. Complexity of Receptor Tyrosine Kinase Signal Processing

    Science.gov (United States)

    Volinsky, Natalia; Kholodenko, Boris N.

    2013-01-01

    Our knowledge of molecular mechanisms of receptor tyrosine kinase (RTK) signaling advances with ever-increasing pace. Yet our understanding of how the spatiotemporal dynamics of RTK signaling control specific cellular outcomes has lagged behind. Systems-centered experimental and computational approaches can help reveal how overlapping networks of signal transducers downstream of RTKs orchestrate specific cell-fate decisions. We discuss how RTK network regulatory structures, which involve the immediate posttranslational and delayed transcriptional controls by multiple feed forward and feedback loops together with pathway cross talk, adapt cells to the combinatorial variety of external cues and conditions. This intricate network circuitry endows cells with emerging capabilities for RTK signal processing and decoding. We illustrate how mathematical modeling facilitates our understanding of RTK network behaviors by unraveling specific systems properties, including bistability, oscillations, excitable responses, and generation of intricate landscapes of signaling activities. PMID:23906711

  10. Receptor for Advanced Glycation End-Products Signaling Interferes with the Vascular Smooth Muscle Cell Contractile Phenotype and Function.

    Directory of Open Access Journals (Sweden)

    Elie Simard

    Full Text Available Increased blood glucose concentrations promote reactions between glucose and proteins to form advanced glycation end-products (AGE. Circulating AGE in the blood plasma can activate the receptor for advanced end-products (RAGE, which is present on both endothelial and vascular smooth muscle cells (VSMC. RAGE exhibits a complex signaling that involves small G-proteins and mitogen activated protein kinases (MAPK, which lead to increased nuclear factor kappa B (NF-κB activity. While RAGE signaling has been previously addressed in endothelial cells, little is known regarding its impact on the function of VSMC. Therefore, we hypothesized that RAGE signaling leads to alterations in the mechanical and functional properties of VSMC, which could contribute to complications associated with diabetes. We demonstrated that RAGE is expressed and functional in the A7r5 VSMC model, and its activation by AGE significantly increased NF-κB activity, which is known to interfere with the contractile phenotype of VSMC. The protein levels of the contraction-related transcription factor myocardin were also decreased by RAGE activation with a concomitant decrease in the mRNA and protein levels of transgelin (SM-22α, a regulator of VSMC contraction. Interestingly, we demonstrated that RAGE activation increased the overall cell rigidity, an effect that can be related to an increase in myosin activity. Finally, although RAGE stimulation amplified calcium signaling and slightly myosin activity in VSMC challenged with vasopressin, their contractile capacity was negatively affected. Overall, RAGE activation in VSMC could represent a keystone in the development of vascular diseases associated with diabetes by interfering with the contractile phenotype of VSMC through the modification of their mechanical and functional properties.

  11. Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase

    Directory of Open Access Journals (Sweden)

    Tähtinen Siri

    2010-01-01

    Full Text Available Abstract Background T-cell protein tyrosine phosphatase (TCPTP/TC45 is a ubiquitously expressed intra-cellular non-receptor protein tyrosine phosphatase involved in the negative regulation of several cancer relevant cellular signalling pathways. We have previously shown that interaction between the α-cytoplasmic tail of α1β1 integrin and TCPTP activates TCPTP by disrupting an inhibitory intra-molecular bond in TCPTP. Thus, inhibition of the regulatory interaction in TCPTP is a desirable strategy for TCPTP activation and attenuation of oncogenic RTK signalling. However, this is challenging with low molecular weight compounds. Methods We developed a high-throughput compatible assay to analyse activity of recombinant TCPTP in vitro. Using this assay we have screened 64280 small molecules to identify novel agonists for TCPTP. Dose-dependent response to TCPTP agonist was performed using the in vitro assay. Inhibition effects and specificity of TCPTP agonists were evaluated using TCPTP expressing and null mouse embryonic fibroblasts. Western blot analysis was used to evaluate attenuation of PDGFRβ and EGFR phosphorylation. Inhibition of VEGF signalling was analysed with VEGF-induced endothelial cell sprouting assays. Results From the screen we identified six TCPTP agonists. Two compounds competed with α1-cytoplasmic domain for binding to TCPTP, suggesting that they activate TCPTP similar to α1-cyt by disrupting the intra-molecular bond in TCPTP. Importantly, one of the compounds (spermidine displayed specificity towards TCPTP in cells, since TCPTP -/- cells were 43-fold more resistant to the compound than TCPTP expressing cells. This compound attenuates PDGFRβ and VEGFR2 signalling in cells in a TCPTP-dependent manner and functions as a negative regulator of EGFR phosphorylation in cancer cells. Conclusions In this study we showed that small molecules mimicking TCPTP-α1 interaction can be used as TCPTP agonists. These data provide the first

  12. Research Resource: Real-Time Analysis of Somatostatin and Dopamine Receptor Signaling in Pituitary Cells Using a Fluorescence-Based Membrane Potential Assay.

    Science.gov (United States)

    Günther, Thomas; Culler, Michael; Schulz, Stefan

    2016-04-01

    Stable somatostatin analogues and dopamine receptor agonists are the mainstay for the pharmacological treatment of functional pituitary adenomas; however, only a few cellular assays have been developed to detect receptor activation of novel compounds without disrupting cells to obtain the second messenger content. Here, we adapted a novel fluorescence-based membrane potential assay to characterize receptor signaling in a time-dependent manner. This minimally invasive technique provides a robust and reliable read-out for ligand-induced receptor activation in permanent and primary pituitary cells. The mouse corticotropic cell line AtT-20 endogenously expresses both the somatostatin receptors 2 (sst2) and 5 (sst5). Exposure of wild-type AtT-20 cells to the sst2- and sst5-selective agonists BIM-23120 and BIM-23268, respectively, promoted a pertussis toxin- and tertiapin-Q-sensitive reduction in fluorescent signal intensity, which is indicative of activation of G protein-coupled inwardly rectifying potassium (GIRK) channels. After heterologous expression, sst1, sst3, and sst4 receptors also coupled to GIRK channels in AtT-20 cells. Similar activation of GIRK channels by dopamine required overexpression of dopamine D2 receptors (D2Rs). Interestingly, the presence of D2Rs in AtT-20 cells strongly facilitated GIRK channel activation elicited by the sst2-D2 chimeric ligand BIM-23A760, suggesting a synergistic action of sst2 and D2Rs. Furthermore, stable somatostatin analogues produced strong responses in primary pituitary cultures from wild-type mice; however, in cultures from sst2 receptor-deficient mice, only pasireotide and somatoprim, but not octreotide, induced a reduction in fluorescent signal intensity, suggesting that octreotide mediates its pharmacological action primarily via the sst2 receptor. PMID:26967369

  13. Direct Melanoma Cell Contact Induces Stromal Cell Autocrine Prostaglandin E2-EP4 Receptor Signaling That Drives Tumor Growth, Angiogenesis, and Metastasis.

    Science.gov (United States)

    Inada, Masaki; Takita, Morichika; Yokoyama, Satoshi; Watanabe, Kenta; Tominari, Tsukasa; Matsumoto, Chiho; Hirata, Michiko; Maru, Yoshiro; Maruyama, Takayuki; Sugimoto, Yukihiko; Narumiya, Shuh; Uematsu, Satoshi; Akira, Shizuo; Murphy, Gillian; Nagase, Hideaki; Miyaura, Chisato

    2015-12-11

    The stromal cells associated with tumors such as melanoma are significant determinants of tumor growth and metastasis. Using membrane-bound prostaglandin E synthase 1 (mPges1(-/-)) mice, we show that prostaglandin E2 (PGE2) production by host tissues is critical for B16 melanoma growth, angiogenesis, and metastasis to both bone and soft tissues. Concomitant studies in vitro showed that PGE2 production by fibroblasts is regulated by direct interaction with B16 cells. Autocrine activity of PGE2 further regulates the production of angiogenic factors by fibroblasts, which are key to the vascularization of both primary and metastatic tumor growth. Similarly, cell-cell interactions between B16 cells and host osteoblasts modulate mPGES-1 activity and PGE2 production by the osteoblasts. PGE2, in turn, acts to stimulate receptor activator of NF-κB ligand expression, leading to osteoclast differentiation and bone erosion. Using eicosanoid receptor antagonists, we show that PGE2 acts on osteoblasts and fibroblasts in the tumor microenvironment through the EP4 receptor. Metastatic tumor growth and vascularization in soft tissues was abrogated by an EP4 receptor antagonist. EP4-null Ptger4(-/-) mice do not support B16 melanoma growth. In vitro, an EP4 receptor antagonist modulated PGE2 effects on fibroblast production of angiogenic factors. Our data show that B16 melanoma cells directly influence host stromal cells to generate PGE2 signals governing neoangiogenesis and metastatic growth in bone via osteoclast erosive activity as well as angiogenesis in soft tissue tumors. PMID:26475855

  14. Signaling through three chemokine receptors triggers the migration of transplanted neural precursor cells in a model of multiple sclerosis.

    Science.gov (United States)

    Cohen, Mikhal E; Fainstein, Nina; Lavon, Iris; Ben-Hur, Tamir

    2014-09-01

    Multiple sclerosis (MS) is a multifocal disease, and precursor cells need to migrate into the multiple lesions in order to exert their therapeutic effects. Therefore, cell migration is a crucial element in regenerative processes in MS, dictating the route of delivery, when cell transplantation is considered. We have previously shown that inflammation triggers migration of multi-potential neural precursor cells (NPCs) into the white matter of experimental autoimmune encephalomyelitis (EAE) rodents, a widely used model of MS. Here we investigated the molecular basis of this attraction. NPCs were grown from E13 embryonic mouse brains and transplanted into the lateral cerebral ventricles of EAE mice. Transplanted NPC migration was directed by three tissue-derived chemokines. Stromal cell-derived factor-1α, monocyte chemo-attractant protein-1 and hepatocyte growth factor were expressed in the EAE brain and specifically in microglia and astrocytes. Their cognate receptors, CXCR4, CCR2 or c-Met were constitutively expressed on NPCs. Selective blockage of CXCR4, CCR2 or c-Met partially inhibited NPC migration in EAE brains. Blocking all three receptors had an additive effect and resulted in profound inhibition of NPC migration, as compared to extensive migration of control NPCs. The inflammation-triggered NPC migration into white matter tracts was dependent on a motile NPC phenotype. Specifically, depriving NPCs from epidermal growth factor (EGF) prevented the induction of glial commitment and a motile phenotype (as indicated by an in vitro motility assay), hampering their response to neuroinflammation. In conclusion, signaling via three chemokine systems accounts for most of the inflammation-induced, tissue-derived attraction of transplanted NPCs into white matter tracts during EAE. PMID:25086214

  15. Interleukin-2 Receptor Signaling in Regulatory T Cell Development and Homeostasis

    OpenAIRE

    Burchill, Matthew A.; Yang, Jianying; Vang, Kieng B.; Farrar, Michael A.

    2007-01-01

    Interleukin-2 (IL2) was initially identified from supernatants of activated lymphocytes over 30 years ago [1]. In the ensuing 15 years, the cDNAs for both IL2 and the three chains of the interleukin-2 receptor (IL2R) were cloned [2–7]. Subsequently, many of the downstream biochemical pathways activated by the IL2 receptor complex were identified [8] and the structure of IL2 bound to this tripartite receptor complex was solved [9]. Thus, we now have a very good understanding of how each chain ...

  16. Epidermal growth factor receptor signalling in human breast cancer cells operates parallel to estrogen receptor α signalling and results in tamoxifen insensitive proliferation.

    NARCIS (Netherlands)

    Moerkens, M.; Zhang, Y.; Wester, L.; Water, van de B.; Meerman, J.H.N.

    2014-01-01

    BACKGROUND Tamoxifen resistance is a major problem in the treatment of estrogen receptor (ER) α -positive breast cancer patients. Although the mechanisms behind tamoxifen resistance are still not completely understood, clinical data suggests that increased expression of receptor tyrosine kinases is

  17. Receptor downregulation and desensitization enhance the information processing ability of signalling receptors

    OpenAIRE

    Resat Haluk; Wiley H Steven; Shankaran Harish

    2007-01-01

    Abstract Background In addition to initiating signaling events, the activation of cell surface receptors also triggers regulatory processes that restrict the duration of signaling. Acute attenuation of signaling can be accomplished either via ligand-induced internalization of receptors (endocytic downregulation) or via ligand-induced receptor desensitization. These phenomena have traditionally been viewed in the context of adaptation wherein the receptor system enters a refractory state in th...

  18. Opioid-receptor (OR) signaling cascades in rat cerebral cortex and model cell lines: the role of plasma membrane structure.

    Science.gov (United States)

    Ujčíková, H; Brejchová, J; Vošahlíková, M; Kagan, D; Dlouhá, K; Sýkora, J; Merta, L; Drastichová, Z; Novotný, J; Ostašov, P; Roubalová, L; Parenti, M; Hof, M; Svoboda, P

    2014-01-01

    Large number of extracellular signals is received by plasma membrane receptors which, upon activation, transduce information into the target cell interior via trimeric G-proteins (GPCRs) and induce activation or inhibition of adenylyl cyclase enzyme activity (AC). Receptors for opioid drugs such as morphine (micro-OR, delta-OR and kappa-OR) belong to rhodopsin family of GPCRs. Our recent results indicated a specific up-regulation of AC I (8-fold) and AC II (2.5-fold) in plasma membranes (PM) isolated from rat brain cortex exposed to increasing doses of morphine (10-50 mg/kg) for 10 days. Increase of ACI and ACII represented the specific effect as the amount of ACIII-ACIX, prototypical PM marker Na, K-ATPase and trimeric G-protein alpha and beta subunits was unchanged. The up-regulation of ACI and ACII faded away after 20 days since the last dose of morphine. Proteomic analysis of these PM indicated that the brain cortex of morphine-treated animals cannot be regarded as being adapted to this drug because significant up-regulation of proteins functionally related to oxidative stress and alteration of brain energy metabolism occurred. The number of delta-OR was increased 2-fold and their sensitivity to monovalent cations was altered. Characterization of delta-OR-G-protein coupling in model HEK293 cell line indicated high ability of lithium to support affinity of delta-OR response to agonist stimulation. Our studies of PM structure and function in context with desensitization of GPCRs action were extended by data indicating participation of cholesterol-enriched membrane domains in agonist-specific internalization of delta-OR. In HEK293 cells stably expressing delta-OR-G(i)1alpha fusion protein, depletion of PM cholesterol was associated with the decrease in affinity of G-protein response to agonist stimulation, whereas maximum response was unchanged. Hydrophobic interior of isolated PM became more "fluid", chaotically organized and accessible to water molecules

  19. Syndecans, signaling, and cell adhesion

    DEFF Research Database (Denmark)

    Couchman, J R; Woods, A

    1996-01-01

    structures within the heparan sulfate chains, leaving the roles of chondroitin sulfate chains and extracellular portion of the core proteins to be elucidated. Evidence that syndecans are a class of receptor involved in cell adhesion is mounting, and their small cytoplasmic domains may link with the...... transmembrane signaling from matrix to cytoskeleton, as proposed for other classes of adhesion receptors....

  20. The phenomenon of acquired resistance to metformin in breast cancer cells: The interaction of growth pathways and estrogen receptor signaling.

    Science.gov (United States)

    Scherbakov, Alexander M; Sorokin, Danila V; Tatarskiy, Victor V; Prokhorov, Nikolay S; Semina, Svetlana E; Berstein, Lev M; Krasil'nikov, Mikhail A

    2016-04-01

    Metformin, a biguanide antidiabetic drug, is used to decrease hyperglycemia in patients with type 2 diabetes. Recently, the epidemiological studies revealed the potential of metformin as an anti-tumor drug for several types of cancer, including breast cancer. Anti-tumor metformin action was found to be mediated, at least in part, via activation of adenosine monophosphate-activated protein kinase (AMPK)-intracellular energy sensor, which inhibits the mammalian target of rapamycin (mTOR) and some other signaling pathways. Nevertheless, some patients can be non-sensitive or resistant to metformin action. Here we analyzed the mechanism of the formation of metformin-resistant phenotype in breast cancer cells and its role in estrogen receptor (ER) regulation. The experiments were performed on the ER-positive MCF-7 breast cancer cells and metformin-resistant MCF-7 subline (MCF-7/M) developed due to long-term metformin treatment. The transcriptional activity of NF-κB and ER was measured by the luciferase reporter gene analysis. The protein expression was determined by immunoblotting (Snail1, (phospho)AMPK, (phospho)IκBα, (phospho)mTOR, cyclin D1, (phospho)Akt and ERα) and immunohistochemical analysis (E-cadherin). We have found that: 1) metformin treatment of MCF-7 cells is accompanied with the stimulation of AMPK and inhibition of growth-related proteins including IκBα, NF-κB, cyclin D1 and ERα; 2) long-term metformin treatment lead to the appearance and progression of cross-resistance to metformin and tamoxifen; the resistant cells are characterized with the unaffected AMPK activity, but the irreversible ER suppression and constitutive activation of Akt/Snail1 signaling; 3) Akt/Snail1 signaling is involved into progression of metformin resistance. The results presented may be considered as the first evidence of the progression of cross-resistance to metformin and tamoxifen in breast cancer cells. Importantly, the acquired resistance to both drugs is based on the

  1. Metformin-mediated growth inhibition involves suppression of the IGF-I receptor signalling pathway in human pancreatic cancer cells

    International Nuclear Information System (INIS)

    Epidemiological studies have shown direct associations between type 2 diabetes and obesity, both conditions associated with hyperglycaemia and hyperinsulinemia, and the risk of pancreatic cancer. Up to 80% of pancreatic cancer patients present with either new-onset type 2 diabetes or impaired glucose tolerance at the time of diagnosis. Recent population studies indicate that the incidence of pancreatic cancer is reduced among diabetics taking metformin. In this study, the effects of exposure of pancreatic cancer cells to high glucose levels on their growth and response to metformin were investigated. The human pancreatic cancer cell lines AsPC-1, BxPC-3, PANC-1 and MIAPaCa-2 were grown in normal (5 mM) or high (25 mM) glucose conditions, with or without metformin. The influence by metformin on proliferation, apoptosis and the AMPK and IGF-IR signalling pathways were evaluated in vitro. Metformin significantly reduced the proliferation of pancreatic cancer cells under normal glucose conditions. Hyperglycaemia however, protected against the metformin-induced growth inhibition. The anti-proliferative actions of metformin were associated with an activation of AMP-activated protein kinase AMPKThr172 together with an inhibition of the insulin/insulin-like growth factor-I (IGF-I) receptor activation and downstream signalling mediators IRS-1 and phosphorylated Akt. Furthermore, exposure to metformin during normal glucose conditions led to increased apoptosis as measured by poly(ADP-ribose) polymerase (PARP) cleavage. In contrast, exposure to high glucose levels promoted a more robust IGF-I response and Akt activation which correlated to stimulated AMPKSer485 phosphorylation and impaired AMPKThr172 phosphorylation, resulting in reduced anti-proliferative and apoptotic effects by metformin. Our results indicate that metformin has direct anti-tumour activities in pancreatic cancer cells involving AMPKThr172 activation and suppression of the insulin/IGF signalling pathways

  2. The inhibitory input to mouse cerebellar Purkinje cells is reciprocally modulated by Bergmann glial P2Y1 and AMPA receptor signaling.

    Science.gov (United States)

    Rudolph, Ramona; Jahn, Hannah M; Courjaret, Raphael; Messemer, Nanette; Kirchhoff, Frank; Deitmer, Joachim W

    2016-07-01

    Synaptic transmission has been shown to be modulated by glial functions, but the modes of specific glial action may vary in different neural circuits. We have tested the hypothesis, if Bergmann GLIA (BG) are involved in shaping neuronal communication in the mouse cerebellar cortex, using acutely isolated cerebellar slices of wild-type (WT) and of glia-specific receptor knockout mice. Activation of P2Y1 receptors by ADP (100 µM) or glutamatergic receptors by AMPA (0.3 µM) resulted in a robust, reversible and repeatable rise of evoked inhibitory input in Purkinje cells by 80% and 150%, respectively. The ADP-induced response was suppressed by prior application of AMPA, and the AMPA-induced response was suppressed by prior application of ADP. Genetic deletion or pharmacological blockade of either receptor restored the response to the other receptor agonist. Both ADP and AMPA responses were sensitive to Rose Bengal, which blocks vesicular glutamate uptake, and to the NMDA receptor antagonist D-AP5. Our results provide strong evidence that activation of both ADP and AMPA receptors, located on BGs, results in the release of glutamate, which in turn activates inhibitory interneurons via NMDA-type glutamate receptors. This infers that BG cells, by means of metabotropic signaling via their AMPA and P2Y1 receptors, which mutually suppress each other, would interdependently contribute to the fine-tuning of Purkinje cell activity in the cerebellar cortex. GLIA 2016. GLIA 2016;64:1265-1280. PMID:27144942

  3. Membrane binding mode of intrinsically disordered cytoplasmic domains of T cell receptor signaling subunits depends on lipid composition

    International Nuclear Information System (INIS)

    Intrinsically disordered cytoplasmic domains of T cell receptor (TCR) signaling subunits including ζcyt and CD3εcyt all contain one or more copies of an immunoreceptor tyrosine-based activation motif (ITAM), tyrosine residues of which are phosphorylated upon receptor triggering. Membrane binding-induced helical folding of ζcyt and CD3εcyt ITAMs is thought to control TCR activation. However, the question whether or not lipid binding of ζcyt and CD3εcyt is necessarily accompanied by a folding transition of ITAMs remains open. In this study, we investigate whether the membrane binding mechanisms of ζcyt and CD3εcyt depend on the membrane model used. Circular dichroic and fluorescence data indicate that binding of ζcyt and CD3εcyt to detergent micelles and unstable vesicles is accompanied by a disorder-to-order transition, whereas upon binding to stable vesicles these proteins remain unfolded. Using electron microscopy and dynamic light scattering, we show that upon protein binding, unstable vesicles fuse and rupture. In contrast, stable vesicles remain intact under these conditions. This suggests different membrane binding modes for ζcyt and CD3εcyt depending on the bilayer stability: (1) coupled binding and folding, and (2) binding without folding. These findings explain the long-standing puzzle in the literature and highlight the importance of the choice of an appropriate membrane model for protein-lipid interactions studies.

  4. Hepatitis C virus E2 protein promotes human hepatoma cell proliferation through the MAPK/ERK signaling pathway via cellular receptors

    International Nuclear Information System (INIS)

    Dysregulation of mitogen-activated protein kinase (MAPK) signaling pathways by various viruses has been shown to be responsible for viral pathogenicity. The molecular mechanism by which hepatitis C virus (HCV) infection caused human liver diseases has been investigated on the basis of abnormal intracellular signal events. Current data are very limited involved in transmembrane signal transduction triggered by HCV E2 protein. Here we explored regulation of the MAPK/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway by E2 expressed in Chinese hamster oval cells. In human hepatoma Huh-7 cells, E2 specifically activated the MAPK/ERK pathway including downstream transcription factor ATF-2 and greatly promoted cell proliferation. CD81 and low density lipoprotein receptor (LDLR) on the cell surface mediated binding of E2 to Huh-7 cells. The MAPK/ERK activation and cell proliferation driven by E2 were suppressed by blockage of CD81 as well as LDLR. Furthermore, pretreatment with an upstream kinase MEK1/2 inhibitor U0126 also impaired the MAPK/ERK activation and cell proliferation induced by E2. Our results suggest that the MAPK/ERK signaling pathway triggered by HCV E2 via its receptors maintains survival and growth of target cells

  5. Corazonin receptor signaling in ecdysis initiation.

    Science.gov (United States)

    Kim, Young-Joon; Spalovská-Valachová, Ivana; Cho, Kook-Ho; Zitnanova, Inka; Park, Yoonseong; Adams, Michael E; Zitnan, Dusan

    2004-04-27

    Corazonin is a highly conserved neuropeptide hormone of wide-spread occurrence in insects yet is associated with no universally recognized function. After discovery of the corazonin receptor in Drosophila, we identified its ortholog in the moth, Manduca sexta, as a prelude to physiological studies. The corazonin receptor cDNA in M. sexta encodes a protein of 436 amino acids with seven putative transmembrane domains and shares common ancestry with its Drosophila counterpart. The receptor exhibits high sensitivity and selectivity for corazonin when expressed in Xenopus oocytes (EC(50) approximately 200 pM) or Chinese hamster ovary cells (EC(50) approximately 75 pM). Northern blot analysis locates the receptor in peripheral endocrine Inka cells, the source of preecdysis- and ecdysis-triggering hormones. Injection of corazonin into pharate larvae elicits release of these peptides from Inka cells, which induce precocious preecdysis and ecdysis behaviors. In vitro exposure of isolated Inka cells to corazonin (25-100 pM) induces preecdysis- and ecdysis-triggering hormone secretion. Using corazonin receptor as a biosensor, we show that corazonin concentrations in the hemolymph 20 min before natural preecdysis onset range from 20 to 80 pM and then decline over the next 30-40 min. These findings support the role of corazonin signaling in initiation of the ecdysis behavioral sequence. We propose a model for peptide-mediated interactions between Inka cells and the CNS underlying this process in insect development. PMID:15096620

  6. Inactivation of TGFβ receptor II signalling in pancreatic epithelial cells promotes acinar cell proliferation, acinar-to-ductal metaplasia and fibrosis during pancreatitis.

    Science.gov (United States)

    Grabliauskaite, Kamile; Saponara, Enrica; Reding, Theresia; Bombardo, Marta; Seleznik, Gitta M; Malagola, Ermanno; Zabel, Anja; Faso, Carmen; Sonda, Sabrina; Graf, Rolf

    2016-02-01

    Determining signalling pathways that regulate pancreatic regeneration following pancreatitis is critical for implementing therapeutic interventions. In this study we elucidated the molecular mechanisms underlying the effects of transforming growth factor-β (TGFβ) in pancreatic epithelial cells during tissue regeneration. To this end, we conditionally inactivated TGFβ receptor II (TGFβ-RII) using a Cre-LoxP system under the control of pancreas transcription factor 1a (PTF1a) promoter, specific for the pancreatic epithelium, and evaluated the molecular and cellular changes in a mouse model of cerulein-induced pancreatitis. We show that TGFβ-RII signalling does not mediate the initial acinar cell damage observed at the onset of pancreatitis. However, TGFβ-RII signalling not only restricts acinar cell replication during the regenerative phase of the disease but also limits ADM formation in vivo and in vitro in a cell-autonomous manner. Analyses of molecular mechanisms underlying the observed phenotype revealed that TGFβ-RII signalling stimulates the expression of cyclin-dependent kinase inhibitors and intersects with the EGFR signalling axis. Finally, TGFβ-RII ablation in epithelial cells resulted in increased infiltration of inflammatory cells in the early phases of pancreatitis and increased activation of pancreatic stellate cells in the later stages of pancreatitis, thus highlighting a TGFβ-based crosstalk between epithelial and stromal cells regulating the development of pancreatic inflammation and fibrosis. Collectively, our data not only contribute to clarifying the cellular processes governing pancreatic tissue regeneration, but also emphasize the conserved role of TGFβ as a tumour suppressor, both in the regenerative process following pancreatitis and in the initial phases of pancreatic cancer. PMID:26510396

  7. HER2/ErbB2 Receptor Signaling in Rat and Human Prolactinoma Cells: Strategy for Targeted Prolactinoma Therapy

    OpenAIRE

    Fukuoka, Hidenori; Cooper, Odelia; MIZUTANI, JUN; Tong, Yunguang; Ren, Song-Guang; Bannykh, Serguei; Melmed, Shlomo

    2010-01-01

    Dopamine agonist resistance or intolerance is encountered in approximately 20% of prolactinoma patients. Because human epidermal growth factor receptor 2 (HER2)/ErbB2 is overexpressed in prolactinomas and ErbB receptor ligands regulate prolactin (PRL) gene expression, we tested the role of HER2/ErbB2 in prolactinoma hormone regulation and adenoma cell proliferation to assess the rationale for targeting this receptor for prolactinoma therapy. As we showed prolactinoma HER2 overexpression, we g...

  8. INSULIN ANALOGUES: ANALYSIS OF PROLIFERATIVE POTENCY AND CHARACTERIZATION OF RECEPTORS AND SIGNALLING PATHWAYS ACTIVATED IN HUMAN MAMMARY EPITHELIAL CELLS

    OpenAIRE

    Shukla, Ashish

    2009-01-01

    Insulin analogues have been developed with the aim to provide better glycaemic control to diabetic patients. They are generated by modifying the insulin backbone which, however, may alter relevant biochemical characteristics such as the affinity to insulin receptor and type I insulin-like growth factor receptor (IGF-IR), and the insulin receptor dissociation rate. As a result insulin analogues may exhibit stronger mitogenic potency than regular insulin. Normal mammary epithelial cells show hi...

  9. Long chain polyunsaturated fatty acids alter oxytocin signaling and receptor density in cultured pregnant human myometrial smooth muscle cells.

    Science.gov (United States)

    Kim, Paul Y; Zhong, Miao; Kim, Yoon-Sun; Sanborn, Barbara M; Allen, Kenneth G D

    2012-01-01

    Epidemiological studies and interventional clinical trials indicate that consumption of long chain n-3 polyunsaturated fatty acids (LC n-3 PUFA) such as docosahexaenoic acid (DHA) lengthen gestational duration. Although the mechanisms are not well understood, prostaglandins (PG) of the 2-series are known to play a role in the initiation and progress of labor. In animal studies, modest DHA provision has been shown to reduce placental and uterine PGE(2) and PGF(2α), matrix metalloproteinase (MMP)-2 and MMP-9 expression, and placental collagenase activity. However, modulation of PG biosynthesis may not account for all the effects of LC n-3 PUFAs in labor. We investigated one potential PG-independent mechanism of LC PUFA action using cultured pregnant human myometrial smooth muscle cells. Our goal was to characterize the effect of LC PUFA treatment on oxytocin signaling, a potent uterotonic hormone involved in labor. The addition of 10 µM-100 µM DHA or arachidonic acid (AA) to the culture media for 48 h resulted in dose dependent enrichment of these fatty acids in membrane lipid. DHA and AA significantly inhibited phosphatidylinositol turnover and [Ca(2+)](i) mobilization with oxytocin stimulation compared to bovine serum albumin control and equimolar oleic acid. DHA and AA significantly reduced oxytocin receptor membrane concentration without altering binding affinity or rate of receptor internalization. These findings demonstrate a role for LC n-3 PUFAs in regulation of oxytocin signaling and provide new insight into additional mechanisms pertaining to reports of dietary fish and fish oil consumption prolonging gestation. PMID:22848573

  10. Distinct roles for aryl hydrocarbon receptor nuclear translocator and ah receptor in estrogen-mediated signaling in human cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Mark P Labrecque

    Full Text Available The activated AHR/ARNT complex (AHRC regulates the expression of target genes upon exposure to environmental contaminants such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD. Importantly, evidence has shown that TCDD represses estrogen receptor (ER target gene activation through the AHRC. Our data indicates that AHR and ARNT act independently from each other at non-dioxin response element sites. Therefore, we sought to determine the specific functions of AHR and ARNT in estrogen-dependent signaling in human MCF7 breast cancer and human ECC-1 endometrial carcinoma cells. Knockdown of AHR with siRNA abrogates dioxin-inducible repression of estrogen-dependent gene transcription. Intriguingly, knockdown of ARNT does not effect TCDD-mediated repression of estrogen-regulated transcription, suggesting that AHR represses ER function independently of ARNT. This theory is supported by the ability of the selective AHR modulator 3',4'-dimethoxy-α-naphthoflavone (DiMNF to repress estrogen-inducible transcription. Furthermore, basal and estrogen-activated transcription of the genes encoding cathepsin-D and pS2 are down-regulated in MCF7 cells but up-regulated in ECC-1 cells in response to loss of ARNT. These responses are mirrored at the protein level with cathepsin-D. Furthermore, knock-down of ARNT led to opposite but corresponding changes in estrogen-stimulated proliferation in both MCF7 and ECC-1 cells. We have obtained experimental evidence demonstrating a dioxin-dependent repressor function for AHR and a dioxin-independent co-activator/co-repressor function for ARNT in estrogen signalling. These results provide us with further insight into the mechanisms of transcription factor crosstalk and putative therapeutic targets in estrogen-positive cancers.

  11. Toll-Like Receptor 4 (TLR4) of Retinal Pigment Epithelial Cells Participates in Transmembrane Signaling in Response to Photoreceptor Outer Segments

    OpenAIRE

    Kindzelskii, Andrei L.; Elner, Victor M.; Elner, Susan G.; Yang, Dongli; Hughes, Bret A.; Petty, Howard R.

    2004-01-01

    Retinal pigment epithelial (RPE) cells mediate the recognition and clearance of effete photoreceptor outer segments (POS), a process central to the maintenance of normal vision. Given the emerging importance of Toll-like receptors (TLRs) in transmembrane signaling in response to invading pathogens as well as endogenous substances, we hypothesized that TLRs are associated with RPE cell management of POS. TLR4 clusters on human RPE cells in response to human, but not bovine, POS. However, TLR4 ...

  12. Epidermal growth factor-receptor and radiation-induced signal transduction in human mammary and squamous carcinoma cells

    International Nuclear Information System (INIS)

    Purpose/Objective: The growth regulatory gene epidermal growth factor-receptor (EGF-R) and transforming growth factor-α (TGF-α) have been identified as radiation late response genes because of their permanent up-regulation after repeated radiation exposures. The steady-state mRNA levels of the same genes are up-regulated after single radiation exposures in the dose range of 2 - 5 Gy. This identifies EGF-R and TGF-α as genes critical in radiation responses and provides a unique opportunity to study the signal transduction cascade between immediate early and late responses. This study examines the role of protein tyrosine kinase receptors and immediate down-stream events in radiation-induced signal transduction. Material and Methods: For the analyses of EGF-R and TGF-α mRNA induction and EGF-R autophosphorylation MCF-7 mammary and A431 squamous carcinoma cells were grown in full medium without refeeding to 90% confluence within 4 days. EGF-R tyrosine phosphorylation (YP) after EGF exposures up to 60 min was compared to radiation-induced increases in EGF-R YP levels using YP-specific monoclonal antibody (Ab-5) for immune precipitation and immunoblotting with secondary antibodies. The phosphorylation pattern of EGF-R after radiation and EGF exposures was examined after metabolic labeling with 32P-ATP, immune precipitation and HPLC tryptic peptide mapping. The activation of phospholipase-C (PL-C), generating the secondary messengers (IP3) and diacylglycerol, was monitored by quantifying EGF- or radiation-induced IP3 production with a radio-immune assay kit. Results: Under the experimental conditions used, EGF induced dose-dependent 5- to 15-fold increases in autophosphorylation of EGF-R within 5 min of EGF exposure. The increased EGF-R YP levels were short-lived and reversed to less than base line levels within 60 min, most likely due to EGF-R turnover in the plasma membrane. Similar to EGF, single radiation exposures in the 1 to 4 Gy dose range induced an about 3

  13. Extra-nuclear signaling of progesterone receptor to breast cancer cell movement and invasion through the actin cytoskeleton.

    Directory of Open Access Journals (Sweden)

    Xiao-Dong Fu

    Full Text Available Progesterone plays a role in breast cancer development and progression but the effects on breast cancer cell movement or invasion have not been fully explored. In this study, we investigate the actions of natural progesterone and of the synthetic progestin medroxyprogesterone acetate (MPA on actin cytoskeleton remodeling and on breast cancer cell movement and invasion. In particular, we characterize the nongenomic signaling cascades implicated in these actions. T47-D breast cancer cells display enhanced horizontal migration and invasion of three-dimensional matrices in the presence of both progestins. Exposure to the hormones triggers a rapid remodeling of the actin cytoskeleton and the formation of membrane ruffles required for cell movement, which are dependent on the rapid phosphorylation of the actin-regulatory protein moesin. The extra-cellular small GTPase RhoA/Rho-associated kinase (ROCK-2 cascade plays central role in progesterone- and MPA-induced moesin activation, cell migration and invasion. In the presence of progesterone, progesterone receptor A (PRA interacts with the G protein G alpha(13, while MPA drives PR to interact with tyrosine kinase c-Src and to activate phosphatidylinositol-3 kinase, leading to the activation of RhoA/ROCK-2. In conclusion, our findings manifest that progesterone and MPA promote breast cancer cell movement via rapid actin cytoskeleton remodeling, which are mediated by moesin activation. These events are triggered by RhoA/ROCK-2 cascade through partially differing pathways by the two compounds. These results provide original mechanistic explanations for the effects of progestins on breast cancer progression and highlight potential targets to treat endocrine-sensitive breast cancers.

  14. Antigen receptor signaling: integration of protein tyrosine kinase functions.

    Science.gov (United States)

    Tamir, I; Cambier, J C

    1998-09-17

    Antigen receptors on T and B cells function to transduce signals leading to a variety of biologic responses minimally including antigen receptor editing, apoptotic death, developmental progression, cell activation, proliferation and survival. The response to antigen depends upon antigen affinity and valence, involvement of coreceptors in signaling and differentiative stage of the responding cell. The requirement that these receptors integrate signals that drive an array of responses may explain their evolved structural complexity. Antigen receptors are composed of multiple subunits compartmentalized to provide antigen recognition and signal transduction function. In lieu of on-board enzymatic activity these receptors rely on associated Protein Tyrosine Kinases (PTKs) for their signaling function. By aggregating the receptors, and hence their appended PTKs, antigens induce PTK transphosphorylation, activating them to phosphorylate the receptor within conserved motifs termed Immunoreceptor Tyrosine-based Activation Motifs (ITAMs) found in transducer subunits. The tyrosyl phosphorylated ITAMs then interact with Src Homology 2 (SH2) domains within the PTKs leading to their further activation. As receptor phosphorylation is amplified, other effectors, such as Shc, dock by virtue of SH2 binding, and serve, in-turn, as substrates for these PTKs. This sequence of events not only provides a signal amplification mechanism by combining multiple consecutive steps with positive feedback, but also allows for signal diversification by differential recruitment of effectors that provide access to distinct parallel downstream signaling pathways. The subject of antigen receptor signaling has been recently reviewed in depth (DeFranco, 1997; Kurosaki, 1997). Here we discuss the biochemical basis of antigen receptor signal transduction, using the B cell receptor (BCR) as a paradigm, with specific emphasis on the involved PTKs. We review several specific mechanisms by which responses

  15. G-Protein Coupled Receptor Signaling Architecture of Mammalian Immune Cells

    OpenAIRE

    Natalia Polouliakh; Richard Nock; Frank Nielsen; Hiroaki Kitano

    2009-01-01

    A series of recent studies on large-scale networks of signaling and metabolic systems revealed that a certain network structure often called "bow-tie network" are observed. In signaling systems, bow-tie network takes a form with diverse and redundant inputs and outputs connected via a small numbers of core molecules. While arguments have been made that such network architecture enhances robustness and evolvability of biological systems, its functional role at a cellular level remains obscure....

  16. Sweet Taste Receptor Signaling Network: Possible Implication for Cognitive Functioning

    Directory of Open Access Journals (Sweden)

    Menizibeya O. Welcome

    2015-01-01

    Full Text Available Sweet taste receptors are transmembrane protein network specialized in the transmission of information from special “sweet” molecules into the intracellular domain. These receptors can sense the taste of a range of molecules and transmit the information downstream to several acceptors, modulate cell specific functions and metabolism, and mediate cell-to-cell coupling through paracrine mechanism. Recent reports indicate that sweet taste receptors are widely distributed in the body and serves specific function relative to their localization. Due to their pleiotropic signaling properties and multisubstrate ligand affinity, sweet taste receptors are able to cooperatively bind multiple substances and mediate signaling by other receptors. Based on increasing evidence about the role of these receptors in the initiation and control of absorption and metabolism, and the pivotal role of metabolic (glucose regulation in the central nervous system functioning, we propose a possible implication of sweet taste receptor signaling in modulating cognitive functioning.

  17. Infection Mobilizes Hematopoietic Stem Cells through Cooperative NOD-like Receptor and Toll-like Receptor Signaling

    OpenAIRE

    Burberry, Aaron; Zeng, Melody Y.; Ding, Lei; Wicks, Ian; Inohara, Naohiro; Morrison, Sean J; Núñez, Gabriel

    2014-01-01

    Adult hematopoietic stem cells (HSCs) are maintained in specialized niches within the bone marrow under steady-state conditions and mobilized for extramedullary hematopoiesis during periods of stress such as bacterial infections. However, the underlying mechanisms are unclear. We show that systemic infection of mice with Escherichia coli, commonly associated with bacteremia in humans, mobilizes functional HSCs to the spleen. Accumulation of splenic HSCs (CD150+CD48-Lin−/lowScal1+cKit+) was di...

  18. Interleukin-7 Receptor Signaling Network: An Integrated Systems Perspective

    Institute of Scientific and Technical Information of China (English)

    Megan J. Palmer; Vinay S. Mahajan; Lily C. Trajman; Darrell J. Irvine; Douglas A.Lauffenburger; Jianzhu Chen

    2008-01-01

    Interleukin-7 (IL-7) is an essential cytokine for the development and homeostatic maintenance of T and B lymphocytes. Binding of IL-7 to its cognate receptor, the IL-7 receptor (IL-7R), activates multiple pathways that regulate lymphocyte survival, glucose uptake, proliferation and differentiation. There has been much interest in understanding how IL-7 receptor signaling is modulated at multiple interconnected network levels. This review examines how the strength of the signal through the IL-7 receptor is modulated in T and B cells, including the use of shared receptor components, signaling crosstaik, shared interaction domains, feedback loops, integrated gene regulation, muitimerization and ligand competition. We discuss how these network control mechanisms could integrate to govern the properties of IL-7R signaling in lymphocytes in health and disease. Analysis of IL-7receptor signaling at a network level in a systematic manner will allow for a comprehensive approach to understanding the impact of multiple signaling pathways on lymphocyte biology.

  19. Microtubule-Associated Protein EB3 Regulates IP3 Receptor Clustering and Ca2+ Signaling in Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Melissa Geyer

    2015-07-01

    Full Text Available The mechanisms by which the microtubule cytoskeleton regulates the permeability of endothelial barrier are not well understood. Here, we demonstrate that microtubule-associated end-binding protein 3 (EB3, a core component of the microtubule plus-end protein complex, binds to inositol 1,4,5-trisphosphate receptors (IP3Rs through an S/TxIP EB-binding motif. In endothelial cells, α-thrombin, a pro-inflammatory mediator that stimulates phospholipase Cβ, increases the cytosolic Ca2+ concentration and elicits clustering of IP3R3s. These responses, and the resulting Ca2+-dependent phosphorylation of myosin light chain, are prevented by depletion of either EB3 or mutation of the TxIP motif of IP3R3 responsible for mediating its binding to EB3. We also show that selective EB3 gene deletion in endothelial cells of mice abrogates α-thrombin-induced increase in endothelial permeability. We conclude that the EB3-mediated interaction of IP3Rs with microtubules controls the assembly of IP3Rs into effective Ca2+ signaling clusters, which thereby regulate microtubule-dependent endothelial permeability.

  20. Multimodal function of the sweet taste receptor expressed in pancreatic β-cells: generation of diverse patterns of intracellular signals by sweet agonists.

    Science.gov (United States)

    Nakagawa, Yuko; Nagasawa, Masahiro; Mogami, Hideo; Lohse, Martin; Ninomiya, Yuzo; Kojima, Itaru

    2013-01-01

    The sweet taste receptor is expressed in the taste bud and is activated by numerous sweet molecules with diverse chemical structures. It is, however, not known whether these sweet agonists induce a similar cellular response in target cells. Using MIN6 cells, a pancreatic β-cell line expressing endogenous sweet taste receptor, we addressed this question by monitoring changes in cytoplasmic Ca2+ ([Ca2+]i) and cAMP ([cAMP]i) induced by four sweet taste receptor agonists. Glycyrrhizin evoked sustained elevation of [Ca2+]i but [cAMP]i was not affected. Conversely, an artificial sweetener saccharin induced sustained elevation of [cAMP]i but did not increase [Ca2+]i. In contrast, sucralose and acesulfame K induced rapid and sustained increases in both [Ca2+]i and [cAMP]i. Although the latter two sweeteners increased [Ca2+]i and [cAMP]i, their actions were not identical: [Ca2+]i response to sucralose but not acesulfame K was inhibited by gurmarin, an antagonist of the sweet taste receptor which blocks the gustducin-dependent pathway. In addition, [Ca2+]i response to acesulfame K but not to sucralose was resistant to a Gq inhibitor. These results indicate that four types of sweeteners activate the sweet taste receptor differently and generate distinct patterns of intracellular signals. The sweet taste receptor has amazing multimodal functions producing multiple patterns of intracellular signals. PMID:23933592

  1. Complementary Signaling through flt3 and Interleukin-7 Receptor α Is Indispensable for Fetal and Adult B Cell Genesis

    OpenAIRE

    Sitnicka, Ewa; Brakebusch, Cord; Martensson, Inga-Lill; Svensson, Marcus; Agace, William W.; Sigvardsson, Mikael; Buza-Vidas, Natalija; Bryder, David; M.Cilio, Corrado; Ahlenius, Henrik; Maraskovsky, Eugene; Peschon, Jacques J.; Jacobsen, Sten Eirik W.

    2003-01-01

    Extensive studies of mice deficient in one or several cytokine receptors have failed to support an indispensable role of cytokines in development of multiple blood cell lineages. Whereas B1 B cells and Igs are sustained at normal levels throughout life of mice deficient in IL-7, IL-7Rα, common cytokine receptor gamma chain, or flt3 ligand (FL), we report here that adult mice double deficient in IL-7Rα and FL completely lack visible LNs, conventional IgM+ B cells, IgA+ plasma cells, and B1 cel...

  2. Toll-Like Receptor 3 and Suppressor of Cytokine Signaling Proteins Regulate CXCR4 and CXCR7 Expression in Bone Marrow-Derived Human Multipotent Stromal Cells

    OpenAIRE

    Tomchuck, Suzanne L.; Henkle, Sarah L.; Coffelt, Seth B.; Betancourt, Aline M.

    2012-01-01

    Background The use of bone marrow-derived human multipotent stromal cells (hMSC) in cell-based therapies has dramatically increased in recent years, as researchers have exploited the ability of these cells to migrate to sites of tissue injury, inflammation, and tumors. Our group established that hMSC respond to “danger” signals – by-products of damaged, infected or inflamed tissues – via activation of Toll-like receptors (TLRs). However, little is known regarding downstream signaling mediated...

  3. Cell surface-bound TIMP3 induces apoptosis in mesenchymal Cal78 cells through ligand-independent activation of death receptor signaling and blockade of survival pathways.

    Directory of Open Access Journals (Sweden)

    Christina Koers-Wunrau

    Full Text Available BACKGROUND: The matrix metalloproteinases (MMPs and their endogenous regulators, the tissue inhibitor of metalloproteinases (TIMPs 1-4 are responsible for the physiological remodeling of the extracellular matrix (ECM. Among all TIMPs, TIMP3 appears to play a unique role since TIMP3 is a secreted protein and, unlike the other TIMP family members, is tightly bound to the ECM. Moreover TIMP3 has been shown to be able to induce apoptotic cell death. As little is known about the underlying mechanisms, we set out to investigate the pro-apoptotic effect of TIMP3 in human mesenchymal cells. METHODOLOGY/PRINCIPAL FINDINGS: Lentiviral overexpression of TIMP3 in mesenchymal cells led to a strong dose-dependent induction of ligand-independent apoptosis as reflected by a five-fold increase in caspase 3 and 7 activity compared to control (pLenti6/V5-GW/lacZ or uninfected cells, whereas exogenous TIMP3 failed to induce apoptosis. Concordantly, increased cleavage of death substrate PARP and the caspases 3 and 7 was observed in TIMP3 overexpressing cultures. Notably, activation of caspase-8 but not caspase-9 was observed in TIMP3-overexpressing cells, indicating a death receptor-dependent mechanism. Moreover, overexpression of TIMP3 led to a further induction of apoptosis after stimulation with TNF-alpha, FasL and TRAIL. Most interestingly, TIMP3-overexpression was associated with a decrease in phosphorylation of cRaf, extracellular signal-regulated protein kinase (Erk1/2, ribosomal S6 kinase (RSK1 and Akt and serum deprivation of TIMP3-overexpressing cells resulted in a distinct enhancement of apoptosis, pointing to an impaired signaling of serum-derived survival factors. Finally, heparinase treatment of heparan sulfate proteoglycans led to the release of TIMP3 from the surface of overexpressing cells and to a significant decrease in apoptosis indicating that the binding of TIMP3 is necessary for apoptosis induction. CONCLUSION: The results demonstrate that

  4. B cell antigen receptor-induced activation of an IRAK4-dependent signaling pathway revealed by a MALT1-IRAK4 double knockout mouse model

    Directory of Open Access Journals (Sweden)

    Dufner Almut

    2011-03-01

    Full Text Available Abstract Background The B cell antigen receptor (BCR and pathogen recognition receptors, such as Toll-like receptor 4 (TLR4, act in concert to control adaptive B cell responses. However, little is known about the signaling pathways that integrate BCR activation with intrinsic TLR4 stimulation. Antigen receptors initialize activation of the inducible transcription factor nuclear factor-κB (NF-κB via recruitment of the membrane-associated guanylate kinase caspase recruitment domain protein 11 (CARD11, the adapter molecule B cell CLL/lymphoma 10 (BCL10, and the "paracaspase" mucosa-associated lymphoid tissue lymphoma translocation gene 1 (MALT1 into lipid rafts. Upon BCR triggering, this activation strictly depends on BCL10, but not on MALT1, leading to the hypothesis that a MALT1-independent NF-κB activation pathway contributes to BCR-induced NF-κB activation downstream of BCL10. The identity of this pathway has remained elusive. Results Using genetic and biochemical approaches, we demonstrate that the IRAK4- and IRAK1-dependent TLR signaling branch is activated upon BCR triggering to induce partial NF-κB activation. BCR-induced MALT1-independent IκB degradation and B cell proliferation were inhibited in MALT1/IRAK4 double knockout B cells. Moreover, IRAK1 was recruited into lipid rafts upon BCR stimulation and activated following transient recruitment of IRAK4. Conclusion We propose that the observed crosstalk between BCR and TLR signaling components may contribute to the discrimination of signals that emanate from single and dual receptor engagement to control adaptive B cell responses.

  5. Progesterone Signaling Mediated Through Progesterone Receptor Membrane Component-1 in Ovarian Cells with Special Emphasis on Ovarian Cancer

    OpenAIRE

    Peluso, John J.

    2011-01-01

    Various ovarian cell types including granulosa cells and ovarian surface epithelial cells express the progesterone (P4) binding protein, Progesterone Receptor Membrane Component-1 (PGRMC1). PGRMC1 is also expressed in ovarian tumors. PGRMC1 plays an essential role in promoting the survival of both normal and cancerous ovarian cell in vitro. Given the clinical significance of factors that regulate the viability of ovarian cancer, this review will focus on the role of PGRMC1 in ovarian cancer, ...

  6. Autocrine Bone Morphogenetic Protein-9 signals via Activin Receptor Like Kinase-2/Smad1/Smad4 to promote ovarian cancer cell proliferation

    OpenAIRE

    Herrera, Blanca; van Dinther, Maarten; ten Dijke, Peter; Inman, Gareth J.

    2009-01-01

    Bone morphogenetic proteins (BMPs) act as central regulators of ovarian physiology and may be involved in ovarian cancer development. In an effort to understand these processes we characterized TGFβ/BMP receptor and Smad expression in immortalised ovarian surface epithelial cells (IOSE) and a panel of ovarian cancer cell lines. These studies prompted us to evaluate the potential role of BMP9 signalling in ovarian cancer. Using siRNA, ligand trap, inhibitor and ligand stimulation approaches we...

  7. Rabies Virus Ocular Disease: T-Cell-Dependent Protection Is under the Control of Signaling by the p55 Tumor Necrosis Factor Alpha Receptor, p55TNFR

    OpenAIRE

    Camelo, Serge; Castellanos, Jaime; Lafage, Mireille; Lafon, Monique

    2001-01-01

    Following brain infection, the Challenge Virus Standard strain of rabies virus infects the retina. Rabies virus ocular infection induces the infiltration of neutrophils and predominantly T cells into the eye. The role of tumor necrosis factor alpha (TNF-α)-lymphotoxin signaling in the control of rabies virus ocular infection and inflammatory cell infiltration was assessed using mice lacking the p55 TNF-α receptor (p55TNFR−/− mice). The incidence of ocular disease and the intensity of retinal ...

  8. Membrane Trafficking of Death Receptors: Implications on Signalling

    Directory of Open Access Journals (Sweden)

    Wulf Schneider-Brachert

    2013-07-01

    Full Text Available Death receptors were initially recognised as potent inducers of apoptotic cell death and soon ambitious attempts were made to exploit selective ignition of controlled cellular suicide as therapeutic strategy in malignant diseases. However, the complexity of death receptor signalling has increased substantially during recent years. Beyond activation of the apoptotic cascade, involvement in a variety of cellular processes including inflammation, proliferation and immune response was recognised. Mechanistically, these findings raised the question how multipurpose receptors can ensure selective activation of a particular pathway. A growing body of evidence points to an elegant spatiotemporal regulation of composition and assembly of the receptor-associated signalling complex. Upon ligand binding, receptor recruitment in specialized membrane compartments, formation of receptor-ligand clusters and internalisation processes constitute key regulatory elements. In this review, we will summarise the current concepts of death receptor trafficking and its implications on receptor-associated signalling events.

  9. Attenuation of IgE receptor signalling in mast cells as molecular basis for the antiallergic action of glucocorticoids

    Energy Technology Data Exchange (ETDEWEB)

    Sancono, A.

    2003-04-01

    Glucocorticoids suppress the expression of Fc{epsilon}RI alpha chain gene by inhibiting its promoter activity. This effect requires de novo protein synthesis and regulatory elements at the Fc{epsilon}RI {alpha}-chain promoter that bind the transcription factors GATA-1, Elf-1, Pu.1 and YY1. The downregulation of the Fc{epsilon}RI {alpha}-chain gene expression correlates with a reduced surface expression of the Fc{epsilon}RI. Furthermore, glucocorticoid treatment promotes inactivation of the lck/yes-related novel (Lyn) Src-like tyrosine kinase, responsible of phosphorylation and activation of the Fc{epsilon}RI, via enhanced phosphorylation of the regulatory tyrosine residue of Lyn. The reduction of surface expressed IgE receptor and the inactivation of Lyn would suppress the signal transduction events originating from the Fc{epsilon}RI and ending up with the activation of the downstream targets ERK1/2. In addition, it has been shown here that glucocorticoids inhibit IgE receptor signalling by enhancing the expression of the MAPK phosphatase MKP-1, which in turn inhibits the activation of ERK1/2. The glucocorticoid-mediated enhancement in MKP-1 expression occurs at the MKP-1 promoter level. This regulation requires the glucocorticoid receptor (GR) dimerisation function and the presence of discrete elements on the promoter proximal sequence, suggesting that it is a direct action of the GR. A crucial role of MKP-1 in glucocorticoid-mediated repression of ERK1/2 phosphorylation was demonstrated in bone marrow derived mast cells from MKP-1-deficient mice. ERK1/2 in these cells are activated by IgE receptor cross-linking or by Stem Cell Factor through the c-kit receptor, and they were no longer inhibited by glucocorticoids, while this was the case in cells from wild-type mice. However, ERK1/2 activity in other cell types such as thymocytes and splenocytes of MKP-1 deficient mice, could still be inhibited by glucocorticoids. These results dmeonstrate that repression of ERK1

  10. Modulation of TLR3/TLR4 inflammatory signaling by the GABAB receptor agonist baclofen in glia and immune cells: relevance to therapeutic effects in multiple sclerosis

    Directory of Open Access Journals (Sweden)

    Tadhg eCrowley

    2015-07-01

    Full Text Available The GABAB receptor agonist, baclofen, is used to treat muscle tightness and cramping caused by spasticity in a number of disorders including Multiple Sclerosis (MS, but its precise mechanism of action is unknown. Neuroinflammation drives the central pathology in MS and is mediated by both immunoreactive glial cells and invading lymphocytes. Furthermore, a body of data indicates that the Toll-like receptor (TLR family of innate immune receptors is implicated in MS progression. In the present study we investigated whether modulation of GABAB receptors using baclofen can exert anti-inflammatory effects by targeting TLR3 and(or TLR4-induced inflammatory signaling in murine glial cells and human peripheral blood mononuclear cells (PBMCs isolated from healthy control individuals and patients with the relapse-remitting (RR form of MS. TLR3 and TLR4 stimulation promoted the nuclear sequestration of NF-κB and pro-inflammatory cytokine expression in murine glia, while TLR4, but not TLR3, promoted pro-inflammatory cytokine expression in PBMCs isolated from both healthy donors and RR-MS patients. Importantly, this effect was exacerbated in RR-MS patient immune cells. We present further evidence that baclofen dose-dependently attenuated TLR3- and TLR4-induced inflammatory signaling in primary glial cells. Pre-exposure of PBMCs isolated from healthy donors to baclofen attenuated TLR4-induced TNF-α expression, but did not affect TLR4-induced TNF-α expression in RR-MS patient PBMCs. Interestingly, mRNA expression of the GABAB receptor was reduced in PBMCs from RR-MS donors when compared to healthy controls, an effect that might contribute to the differential sensitivity to baclofen seen in healthy and RR-MS patient cells. Overall these findings indicate that baclofen differentially regulates TLR3 and TLR4 signaling in glia and immune cells, and offers insight on the role of baclofen in the treatment of neuroinflammatory disease states including MS.

  11. Piperlongumine inhibits atherosclerotic plaque formation and vascular smooth muscle cell proliferation by suppressing PDGF receptor signaling

    OpenAIRE

    Son, Dong Ju; Kim, Soo Yeon; Han, Seong Su; Kim, Chan Woo; Kumar, Sandeep; Park, Byeoung Soo; Lee, Sung Eun; Yun, Yeo Pyo; Jo, Hanjoong; Park, Young Hyun

    2012-01-01

    Piperlongumine (piplartine, PL) is an alkaloid found in the long pepper (Piper longum L.) and has well-documented anti-platelet aggregation, anti-inflammatory, and anti-cancer properties; however, the role of PL in prevention of atherosclerosis is unknown. We evaluated the anti-atherosclerotic potential of PL in an in vivo murine model of accelerated atherosclerosis and defined its mechanism of action in aortic vascular smooth muscle cells (VSMCs) in vitro. Local treatment with PL significant...

  12. Neuronal merlin influences ERBB2 receptor expression on Schwann cells through neuregulin 1 type III signalling

    OpenAIRE

    Schulz, Alexander; Kyselyova, Anna; Baader, Stephan L.; Jung, Marie Juliane; Zoch, Ansgar; Mautner, Victor-Felix; Hagel, Christian; Morrison, Helen

    2013-01-01

    Axonal surface proteins encompass a group of heterogeneous molecules, which exert a variety of different functions in the highly interdependent relationship between axons and Schwann cells. We recently revealed that the tumour suppressor protein merlin, mutated in the hereditary tumour syndrome neurofibromatosis type 2, impacts significantly on axon structure maintenance in the peripheral nervous system. We now report on a role of neuronal merlin in the regulation of the axonal surface protei...

  13. The Balance of Cell Surface and Soluble Type III TGF-β Receptor Regulates BMP Signaling in Normal and Cancerous Mammary Epithelial Cells

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    Catherine E. Gatza

    2014-06-01

    Full Text Available Bone morphogenetic proteins (BMPs are members of the TGF-β superfamily that are over-expressed in breast cancer, with context dependent effects on breast cancer pathogenesis. The type III TGF-β receptor (TβRIII mediates BMP signaling. While TβRIII expression is lost during breast cancer progression, the role of TβRIII in regulating BMP signaling in normal mammary epithelium and breast cancer cells has not been examined. Restoring TβRIII expression in a 4T1 murine syngeneic model of breast cancer suppressed Smad1/5/8 phosphorylation and inhibited the expression of the BMP transcriptional targets, Id1 and Smad6, in vivo. Similarly, restoring TβRIII expression in human breast cancer cell lines or treatment with sTβRIII inhibited BMP-induced Smad1/5/8 phosphorylation and BMP-stimulated migration and invasion. In normal mammary epithelial cells, shRNA-mediated silencing of TβRIII, TβRIII over-expression, or treatment with sTβRIII inhibited BMP-mediated phosphorylation of Smad1/5/8 and BMP induced migration. Inhibition of TβRIII shedding through treatment with TAPI-2 or expression of a non-shedding TβRIII mutant rescued TβRIII mediated inhibition of BMP induced Smad1/5/8 phosphorylation and BMP induced migration and/or invasion in both in normal mammary epithelial cells and breast cancer cells. Conversely, expression of a TβRIII mutant, which exhibited increased shedding, significantly reduced BMP-mediated Smad1/5/8 phosphorylation, migration, and invasion. These data demonstrate that TβRIII regulates BMP-mediated signaling and biological effects, primarily through the ligand sequestration effects of sTβRIII in normal and cancerous mammary epithelial cells and suggest that the ratio of membrane bound versus sTβRIII plays an important role in mediating these effects.

  14. Human Myeloblastic Leukemia Cells (HL-60) Express a Membrane Receptor for Estrogen that Signals and Modulates Retinoic Acid-induced Cell Differentiation

    OpenAIRE

    Kauss, M. Ariel; Reiterer, Gudrun; Rodica P Bunaciu; Yen, Andrew

    2008-01-01

    Estrogen receptors are historically perceived as nuclear ligand activated transcription factors. An estrogen receptor has now been found localized to the plasma membrane of human myeloblastic leukemia cells (HL-60). Its expression occurs throughout the cell cycle, progressively increasing as cells mature from G1 to S to G2/M. To ascertain that the receptor functioned, the effect of ligands, including a non-internalizable estradiol-BSA conjugate and tamoxifen, an antagonist of nuclear estrogen...

  15. Progesterone receptors - animal models and cell signaling in breast cancer: Role of steroid receptor coactivators and corepressors of progesterone receptors in breast cancer

    International Nuclear Information System (INIS)

    Progesterone, an ovarian steroid hormone, plays a key role in the development and function of the mammary gland, as it also does in the uterus and the ovary. The action of progesterone is mediated through its intracellular cognate receptor, the progesterone receptor (PR), which functions as a transcription factor that regulates gene expression. As with other nuclear receptors, coregulators (coactivators and corepressors) recruited by the liganded or unliganded PR, either to enhance or to suppress transcription activity, modulate the function of the PR. Mutation or aberrant expression of the coregulators might thus affect the normal function of the PR and hence disrupt the normal development of the mammary gland, which may lead to breast cancer

  16. Retinoids arrest breast cancer cell proliferation: retinoic acid selectively reduces the duration of receptor tyrosine kinase signaling

    OpenAIRE

    Tighe, Ann P.; Talmage, David A

    2004-01-01

    Retinoic acid (RA) induces cell cycle arrest of hormone-dependent human breast cancer (HBC) cells. Previously, we demonstrated that RA-induced growth arrest of T-47D HBC cells required the activity of the RA-induced protein kinase, protein kinase Cα (PKCα) [J. Cell Physiol. 172 (1997) 306]. Here, we demonstrate that RA treatment of T-47D cells interfered with growth factor signaling to downstream, cytoplasmic and nuclear targets. RA treatment did not inhibit epidermal growth factor (EGF) rece...

  17. Insulin signaling, lifespan and stress resistance are modulated by metabotropic GABA receptors on insulin producing cells in the brain of Drosophila.

    Science.gov (United States)

    Enell, Lina E; Kapan, Neval; Söderberg, Jeannette A E; Kahsai, Lily; Nässel, Dick R

    2010-01-01

    Insulin-like peptides (ILPs) regulate growth, reproduction, metabolic homeostasis, life span and stress resistance in worms, flies and mammals. A set of insulin producing cells (IPCs) in the Drosophila brain that express three ILPs (DILP2, 3 and 5) have been the main focus of interest in hormonal DILP signaling. Little is, however, known about factors that regulate DILP production and release by these IPCs. Here we show that the IPCs express the metabotropic GABA(B) receptor (GBR), but not the ionotropic GABA(A) receptor subunit RDL. Diminishing the GBR expression on these cells by targeted RNA interference abbreviates life span, decreases metabolic stress resistance and alters carbohydrate and lipid metabolism at stress, but not growth in Drosophila. A direct effect of diminishing GBR on IPCs is an increase in DILP immunofluorescence in these cells, an effect that is accentuated at starvation. Knockdown of irk3, possibly part of a G protein-activated inwardly rectifying K(+) channel that may link to GBRs, phenocopies GBR knockdown in starvation experiments. Our experiments suggest that the GBR is involved in inhibitory control of DILP production and release in adult flies at metabolic stress and that this receptor mediates a GABA signal from brain interneurons that may convey nutritional signals. This is the first demonstration of a neurotransmitter that inhibits insulin signaling in its regulation of metabolism, stress and life span in an invertebrate brain. PMID:21209905

  18. Raloxifene induces cell death and inhibits proliferation through multiple signaling pathways in prostate cancer cells expressing different levels of estrogen receptor α and β.

    Science.gov (United States)

    Rossi, V; Bellastella, G; De Rosa, C; Abbondanza, C; Visconti, D; Maione, L; Chieffi, P; Della Ragione, F; Prezioso, D; De Bellis, A; Bellastella, A; Sinisi, A A

    2011-05-01

    Raloxifene (RAL), a selective estrogen receptor (ER) modulator (SERM) seems to induce apoptosis in both androgen-dependent and -independent prostate cell (PC) lines via activation of ERβ and an antagonistic effect on ERα. In this study, we evaluated the effects of RAL on epithelial PC growth using the two following in vitro models: the androgen-dependent cell line EPN which expressed both ERs; and a stabilized epithelial cell line derived from a prostate cancer specimen (CPEC), which expressed low levels of ERβ and lacked ERα. In EPN cells, there was an increase in the pre-G1 apoptotic peak and a reduction in the S phase of the cell cycle with G0/G1 arrest after E2 or RAL treatment; bcl-2 mRNA and Bcl-2 protein levels were significantly reduced, while activated caspase-3 and Par-4 levels increased significantly after either E2 or RAL treatment; in addition, c-myc transcript was inhibited after 10(-6)  M RAL treatment. A dose-dependent increase of metallothionein II gene RNA level was also induced by RAL in EPN. In CPEC, there was only a weak apoptotic peak associated with caspase-3 activation and Par-4 increase after either E2 or RAL treatment; while c-myc transcript level increased. RAL induced a rapid but transient phosphorylation of ERK 1/2 in EPN cells but generated a sustained effect in CPEC. These findings suggest that RAL effects on PC growth control in vitro are cell-specific, depending on ERβ or ERβ/ERα relative expression levels. Moreover, this study demonstrated that RAL affected both transcriptional regulation and non-genomic signals, which resulted in the modulation of multiple signaling pathways of apoptosis and of cell cycle progression. PMID:20945400

  19. Characterisation of Signalling by the Endogenous GPER1 (GPR30 Receptor in an Embryonic Mouse Hippocampal Cell Line (mHippoE-18.

    Directory of Open Access Journals (Sweden)

    Nicholas J Evans

    Full Text Available Estrogen can modulate neuronal development and signalling by both genomic and non-genomic pathways. Many of its rapid, non-genomic effects on nervous tissue have been suggested to be mediated via the activation of the estrogen sensitive G-protein coupled receptor (GPER1 or GPR30. There has been much controversy over the cellular location, signalling properties and endogenous activators of GPER1. Here we describe the pharmacology and signalling properties of GPER1 in an immortalized embryonic hippocampal cell line, mHippoE-18. This cell line does not suffer from the inherent problems associated with the study of this receptor in native tissue or the problems associated with heterologously expression in clonal cell lines. In mHippoE-18 cells, 17β-Estradiol can mediate a dose-dependent rapid potentiation of forskolin-stimulated cyclic AMP levels but does not appear to activate the ERK1/2 pathway. The effect of 17β-Estradiol can be mimicked by the GPER1 agonist, G1, and also by tamoxifen and ICI 182,780 which activate GPER1 in a variety of other preparations. The response is not mimicked by the application of the classical estrogen receptor agonists, PPT, (an ERα agonist or DPN, (an ERβ agonist, further suggesting that this effect of 17β-Estradiol is mediated through the activation of GPER1. However, after exposure of the cells to the GPER1 specific antagonists, G15 and G36, the stimulatory effects of the above agonists are replaced by dose-dependent inhibitions of forskolin-stimulated cyclic AMP levels. This inhibitory effect is mimicked by aldosterone in a dose-dependent way even in the absence of the GPER1 antagonists. The results are discussed in terms of possible "Biased Antagonism" whereby the antagonists change the conformation of the receptor resulting in changes in the agonist induced coupling of the receptor to different second messenger pathways.

  20. A cleavable signal peptide enhances cell surface delivery and heterodimerization of Cerulean-tagged angiotensin II AT1 and bradykinin B2 receptor

    International Nuclear Information System (INIS)

    Highlights: → A new FRET-based method detects AT1/B2 receptor heterodimerization. → First time application of AT1-Cerulean as a FRET donor. → Method relies on signal peptide-enhanced cell surface delivery of AT1-Cerulean. → A high FRET efficiency revealed efficient heterodimerization of AT1/B2R proteins. → AT1/B2R heterodimers were functionally coupled to desensitization mechanisms. -- Abstract: Heterodimerization of the angiotensin II AT1 receptor with the receptor for the vasodepressor bradykinin, B2R, is known to sensitize the AT1-stimulated response of hypertensive individuals in vivo. To analyze features of that prototypic receptor heterodimer in vitro, we established a new method that uses fluorescence resonance energy transfer (FRET) and applies for the first time AT1-Cerulean as a FRET donor. The Cerulean variant of the green fluorescent protein as donor fluorophore was fused to the C-terminus of AT1, and the enhanced yellow fluorescent protein (EYFP) as acceptor fluorophore was fused to B2R. In contrast to AT1-EGFP, the AT1-Cerulean fusion protein was retained intracellularly. To facilitate cell surface delivery of AT1-Cerulean, a cleavable signal sequence was fused to the receptor's amino terminus. The plasma membrane-localized AT1-Cerulean resembled the native AT1 receptor regarding ligand binding and receptor activation. A high FRET efficiency of 24.7% between membrane-localized AT1-Cerulean and B2R-EYFP was observed with intact, non-stimulated cells. Confocal FRET microscopy further revealed that the AT1/B2 receptor heterodimer was functionally coupled to receptor desensitization mechanisms because activation of the AT1-Cerulean/B2R-EYFP heterodimer with a single agonist triggered the co-internalization of AT1/B2R. Receptor co-internalization was sensitive to inhibition of G protein-coupled receptor kinases, GRKs, as evidenced by a GRK-specific peptide inhibitor. In agreement with efficient AT1/B2R heterodimerization, confocal FRET imaging of

  1. A cleavable signal peptide enhances cell surface delivery and heterodimerization of Cerulean-tagged angiotensin II AT1 and bradykinin B2 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Quitterer, Ursula, E-mail: ursula.quitterer@pharma.ethz.ch [Molecular Pharmacology Unit, Swiss Federal Institute of Technology and University of Zurich, Zurich (Switzerland); Pohl, Armin; Langer, Andreas; Koller, Samuel; AbdAlla, Said [Molecular Pharmacology Unit, Swiss Federal Institute of Technology and University of Zurich, Zurich (Switzerland)

    2011-06-10

    Highlights: {yields} A new FRET-based method detects AT1/B2 receptor heterodimerization. {yields} First time application of AT1-Cerulean as a FRET donor. {yields} Method relies on signal peptide-enhanced cell surface delivery of AT1-Cerulean. {yields} A high FRET efficiency revealed efficient heterodimerization of AT1/B2R proteins. {yields} AT1/B2R heterodimers were functionally coupled to desensitization mechanisms. -- Abstract: Heterodimerization of the angiotensin II AT1 receptor with the receptor for the vasodepressor bradykinin, B2R, is known to sensitize the AT1-stimulated response of hypertensive individuals in vivo. To analyze features of that prototypic receptor heterodimer in vitro, we established a new method that uses fluorescence resonance energy transfer (FRET) and applies for the first time AT1-Cerulean as a FRET donor. The Cerulean variant of the green fluorescent protein as donor fluorophore was fused to the C-terminus of AT1, and the enhanced yellow fluorescent protein (EYFP) as acceptor fluorophore was fused to B2R. In contrast to AT1-EGFP, the AT1-Cerulean fusion protein was retained intracellularly. To facilitate cell surface delivery of AT1-Cerulean, a cleavable signal sequence was fused to the receptor's amino terminus. The plasma membrane-localized AT1-Cerulean resembled the native AT1 receptor regarding ligand binding and receptor activation. A high FRET efficiency of 24.7% between membrane-localized AT1-Cerulean and B2R-EYFP was observed with intact, non-stimulated cells. Confocal FRET microscopy further revealed that the AT1/B2 receptor heterodimer was functionally coupled to receptor desensitization mechanisms because activation of the AT1-Cerulean/B2R-EYFP heterodimer with a single agonist triggered the co-internalization of AT1/B2R. Receptor co-internalization was sensitive to inhibition of G protein-coupled receptor kinases, GRKs, as evidenced by a GRK-specific peptide inhibitor. In agreement with efficient AT1/B2R

  2. Integrated analysis reveals that STAT3 is central to the crosstalk between HER/ErbB receptor signaling pathways in human mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Gong, Chunhong; Zhang, Yi; Shankaran, Harish; Resat, Haluk

    2015-01-01

    Human epidermal growth factor receptors (HER, also known as ErbB) drive cellular proliferation, pro-survival and stress responses by activating several downstream kinases, in particular ERK, p38, JNK (SAPK), the PI3K/AKT, as well as various transcriptional regulators such as STAT3. When co-expressed, first three members of HER family (HER1-3) can form homo- and hetero-dimers. Based on the considerable evidence which suggest that every receptor dimer activates intracellular signaling pathways differentially, we hypothesized that the HER dimerization pattern is a better predictor of downstream signaling than the total receptor activation levels. We validated our hypothesis using a combination of model-based analysis to quantify the HER dimerization patterns and multi-factorial experiments where HER dimerization patterns and signaling crosstalk were rationally perturbed. We have measured the activation of HER1-3 receptors and of the sentinel signaling proteins ERK, AKT, p38, JNK, STAT3 as a function of time in a panel of human mammary epithelial (HME) cells expressing different levels of HER1-3 stimulated with various ligand combinations. Our analysis using multiple ways of clustering the activation data has confirmed that the HER receptor dimer is a better predictor of the signaling through p38, ERK and AKT pathways than the total HER receptor expression and activation levels. Targeted inhibition studies to identify the causal effects allowed us to obtain a consensus regulatory interaction model, which revealed that STAT3 occupies a central role in the crosstalk between the studied pathways.

  3. Rotavirus activates lymphocytes from non-obese diabetic mice by triggering toll-like receptor 7 signaling and interferon production in plasmacytoid dendritic cells.

    Directory of Open Access Journals (Sweden)

    Jessica A Pane

    2014-03-01

    Full Text Available It has been proposed that rotavirus infection promotes the progression of genetically-predisposed children to type 1 diabetes, a chronic autoimmune disease marked by infiltration of activated lymphocytes into pancreatic islets. Non-obese diabetic (NOD mice provide a model for the human disease. Infection of adult NOD mice with rhesus monkey rotavirus (RRV accelerates diabetes onset, without evidence of pancreatic infection. Rather, RRV spreads to the pancreatic and mesenteric lymph nodes where its association with antigen-presenting cells, including dendritic cells, induces cellular maturation. RRV infection increases levels of the class I major histocompatibility complex on B cells and proinflammatory cytokine expression by T cells at these sites. In autoimmunity-resistant mice and human mononuclear cells from blood, rotavirus-exposed plasmacytoid dendritic cells contribute to bystander polyclonal B cell activation through type I interferon expression. Here we tested the hypothesis that rotavirus induces bystander activation of lymphocytes from NOD mice by provoking dendritic cell activation and proinflammatory cytokine secretion. NOD mouse splenocytes were stimulated with rotavirus and assessed for activation by flow cytometry. This stimulation activated antigen-presenting cells and B cells independently of virus strain and replicative ability. Instead, activation depended on virus dose and was prevented by blockade of virus decapsidation, inhibition of endosomal acidification and interference with signaling through Toll-like receptor 7 and the type I interferon receptor. Plasmacytoid dendritic cells were more efficiently activated than conventional dendritic cells by RRV, and contributed to the activation of B and T cells, including islet-autoreactive CD8+ T cells. Thus, a double-stranded RNA virus can induce Toll-like receptor 7 signaling, resulting in lymphocyte activation. Our findings suggest that bystander activation mediated by type I

  4. Biased and g protein-independent signaling of chemokine receptors

    DEFF Research Database (Denmark)

    Steen, Anne; Larsen, Olav; Thiele, Stefanie;

    2014-01-01

    -switches based on recently published 7TM crystals and molecular dynamics studies. All three forms of biased signaling are abundant in the chemokine system. This challenges our understanding of "classic" redundancy inevitably ascribed to this system, where multiple chemokines bind to the same receptor and where a......Biased signaling or functional selectivity occurs when a 7TM-receptor preferentially activates one of several available pathways. It can be divided into three distinct forms: ligand bias, receptor bias, and tissue or cell bias, where it is mediated by different ligands (on the same receptor...... absolute, i.e., full versus no activation. Here we discuss biased signaling in the chemokine system, including the structural basis for biased signaling in chemokine receptors, as well as in class A 7TM receptors in general. This includes overall helical movements and the contributions of micro...

  5. A Natural Variant of the T Cell Receptor-Signaling Molecule Vav1 Reduces Both Effector T Cell Functions and Susceptibility to Neuroinflammation

    Science.gov (United States)

    Kassem, Sahar; Bernard, Isabelle; Dejean, Anne S.; Liblau, Roland; Fournié, Gilbert J.; Colacios, Céline

    2016-01-01

    The guanine nucleotide exchange factor Vav1 is essential for transducing T cell antigen receptor signals and therefore plays an important role in T cell development and activation. Our previous genetic studies identified a locus on rat chromosome 9 that controls the susceptibility to neuroinflammation and contains a non-synonymous polymorphism in the major candidate gene Vav1. To formally demonstrate the causal implication of this polymorphism, we generated a knock-in mouse bearing this polymorphism (Vav1R63W). Using this model, we show that Vav1R63W mice display reduced susceptibility to experimental autoimmune encephalomyelitis (EAE) induced by MOG35-55 peptide immunization. This is associated with a lower production of effector cytokines (IFN-γ, IL-17 and GM-CSF) by autoreactive CD4 T cells. Despite increased proportion of Foxp3+ regulatory T cells in Vav1R63W mice, we show that this lowered cytokine production is intrinsic to effector CD4 T cells and that Treg depletion has no impact on EAE development. Finally, we provide a mechanism for the above phenotype by showing that the Vav1R63W variant has normal enzymatic activity but reduced adaptor functions. Together, these data highlight the importance of Vav1 adaptor functions in the production of inflammatory cytokines by effector T cells and in the susceptibility to neuroinflammation. PMID:27438086

  6. Tissue kallikrein induces SH-SY5Y cell proliferation via epidermal growth factor receptor and extracellular signal-regulated kinase1/2 pathway

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Zhengyu [Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai 200040 (China); Department of Neurology, Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai 200437 (China); Yang, Qi; Cui, Mei; Liu, Yanping [Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai 200040 (China); Wang, Tao; Zhao, Hong [Department of Neurology, Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai 200437 (China); Dong, Qiang, E-mail: qiang_dong163@163.com [Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai 200040 (China)

    2014-03-28

    Highlights: • TK promotes EGFR phosphorylation in SH-SY5Y cells. • TK activates ERK1/2 and p38 phosphorylation in SH-SY5Y cells. • TK mediates SH-SY5Y cell proliferation via EGFR and ERK1/2 pathway. - Abstract: Tissue kallikrein (TK) is well known to take most of its biological functions through bradykinin receptors. In the present study, we found a novel signaling pathway mediated by TK through epidermal growth factor receptor (EGFR) in human SH-SY5Y cells. We discovered that TK facilitated the activation of EGFR, extracellular signal-regulated kinase (ERK) 1/2 and p38 cascade. Interestingly, not p38 but ERK1/2 phosphorylation was severely compromised in cells depleted of EGFR. Nevertheless, impairment of signaling of ERK1/2 seemed not to be restricted to EGFR phosphorylation. We also observed that TK stimulation could induce SH-SY5Y cell proliferation, which was reduced by EGFR down-regulation or ERK1/2 inhibitor. Overall, our findings provided convincing evidence that TK could mediate cell proliferation via EGFR and ERK1/2 pathway in vitro.

  7. Tissue kallikrein induces SH-SY5Y cell proliferation via epidermal growth factor receptor and extracellular signal-regulated kinase1/2 pathway

    International Nuclear Information System (INIS)

    Highlights: • TK promotes EGFR phosphorylation in SH-SY5Y cells. • TK activates ERK1/2 and p38 phosphorylation in SH-SY5Y cells. • TK mediates SH-SY5Y cell proliferation via EGFR and ERK1/2 pathway. - Abstract: Tissue kallikrein (TK) is well known to take most of its biological functions through bradykinin receptors. In the present study, we found a novel signaling pathway mediated by TK through epidermal growth factor receptor (EGFR) in human SH-SY5Y cells. We discovered that TK facilitated the activation of EGFR, extracellular signal-regulated kinase (ERK) 1/2 and p38 cascade. Interestingly, not p38 but ERK1/2 phosphorylation was severely compromised in cells depleted of EGFR. Nevertheless, impairment of signaling of ERK1/2 seemed not to be restricted to EGFR phosphorylation. We also observed that TK stimulation could induce SH-SY5Y cell proliferation, which was reduced by EGFR down-regulation or ERK1/2 inhibitor. Overall, our findings provided convincing evidence that TK could mediate cell proliferation via EGFR and ERK1/2 pathway in vitro

  8. Non-canonical kinase signaling by the death ligand TRAIL in cancer cells : discord in the death receptor family

    NARCIS (Netherlands)

    Azijli, K.; Weyhenmeyer, B.; Peters, G. J.; de Jong, S.; Kruyt, F. A. E.

    2013-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-based therapy is currently evaluated in clinical studies as a tumor cell selective pro-apoptotic approach. However, besides activating canonical caspase-dependent apoptosis by binding to TRAIL-specific death receptors, the TRAIL ligand

  9. De Novo Transcriptome Analysis Shows That SAV-3 Infection Upregulates Pattern Recognition Receptors of the Endosomal Toll-Like and RIG-I-Like Receptor Signaling Pathways in Macrophage/Dendritic Like TO-Cells.

    Science.gov (United States)

    Xu, Cheng; Evensen, Øystein; Mweemba Munang'andu, Hetron

    2016-01-01

    A fundamental step in cellular defense mechanisms is the recognition of "danger signals" made of conserved pathogen associated molecular patterns (PAMPs) expressed by invading pathogens, by host cell germ line coded pattern recognition receptors (PRRs). In this study, we used RNA-seq and the Kyoto encyclopedia of genes and genomes (KEGG) to identify PRRs together with the network pathway of differentially expressed genes (DEGs) that recognize salmonid alphavirus subtype 3 (SAV-3) infection in macrophage/dendritic like TO-cells derived from Atlantic salmon (Salmo salar L) headkidney leukocytes. Our findings show that recognition of SAV-3 in TO-cells was restricted to endosomal Toll-like receptors (TLRs) 3 and 8 together with RIG-I-like receptors (RLRs) and not the nucleotide-binding oligomerization domain-like receptors NOD-like receptor (NLRs) genes. Among the RLRs, upregulated genes included the retinoic acid inducible gene I (RIG-I), melanoma differentiation association 5 (MDA5) and laboratory of genetics and physiology 2 (LGP2). The study points to possible involvement of the tripartite motif containing 25 (TRIM25) and mitochondrial antiviral signaling protein (MAVS) in modulating RIG-I signaling being the first report that links these genes to the RLR pathway in SAV-3 infection in TO-cells. Downstream signaling suggests that both the TLR and RLR pathways use interferon (IFN) regulatory factors (IRFs) 3 and 7 to produce IFN-a2. The validity of RNA-seq data generated in this study was confirmed by quantitative real time qRT-PCR showing that genes up- or downregulated by RNA-seq were also up- or downregulated by RT-PCR. Overall, this study shows that de novo transcriptome assembly identify key receptors of the TLR and RLR sensors engaged in host pathogen interaction at cellular level. We envisage that data presented here can open a road map for future intervention strategies in SAV infection of salmon. PMID:27110808

  10. HBx induced AFP receptor expressed to activate PI3K/AKT signal to promote expression of Src in liver cells and hepatoma cells

    International Nuclear Information System (INIS)

    Hepatitis B virus (HBV)-X protein(HBx) is a transactivator of host several cellular genes including alpha-fetoprotein(AFP) and AFP receptor(AFPR) which contributes to HBV-associated tumor development. The expression of AFP/AFPR are correlated with hepatocellular carcinoma(HCC)-initial cells. But the role of AFP and AFPR in promoting occurrence of HBV-related HCC were still unclear. A total of 71 clinical patients’ liver specimens, normal human liver cells L-02 and HCC cell lines, PLC/PRF/5 were selected for analyzing the effects of HBx on expression of AFP, AFPR and Src. The expression of goal proteins were detected by Immunohistochemical stained and Western blotting; HBx-expressed vectors were constructed and transfected into L-02 cells, laser confocal microscopy was applied to observe expression and location of AFP, AFPR and Src in the normal liver cells and HCC cells, soft agar colony formation assay was used to observe colonies formed of the cells. We confirmed HBx gives preference to promote the expression of AFP and AFPR; HBx priors to up-regulate the expression of AFPR and AFP in L-02 cells and in normal liver specimens; AFPR signal been able to stimulate Src expression. The results also indicated that phosphatidylinositol 3-kinase(PI3K) inhibitors Ly294002 and GDC0941 effectively suppress AFPR mediated up-regulation expression of Src in AFPR positive HCC lines. HBx priors to drive the expression of AFP and AFPR to promote expression of Src in normal liver cells and hepatoma cells; AFP and AFPR maybe play pivotal role in HBV-related hepatocarcinogenesis; Targeting AFPR is an available therapeutic strategy of HCC. The online version of this article (doi:10.1186/s12885-015-1384-9) contains supplementary material, which is available to authorized users

  11. ABA Signaling in Guard Cells Entails a Dynamic Protein-Protein Interaction Relay from the PYL-RCAR Family Receptors to Ion Channels

    Institute of Scientific and Technical Information of China (English)

    Sung Chul Lee; Chae Woo Lim; Wenzhi Lan; Kai He; Sheng Luan

    2013-01-01

    Plant hormone abscisic acid (ABA) serves as an integrator of environmental stresses such as drought to trigger stomatal closure by regulating specific ion channels in guard cells.We previously reported that SLACl,an outward anion channel required for stomatal closure,was regulated via reversible protein phosphorylation events involving ABA signaling components,including protein phosphatase 2C members and a SnRK2-type kinase (OST1).In this study,we reconstituted the ABA signaling pathway as a protein-protein interaction relay from the PYL/RCAR-type receptors,to the PP2C-SnRK2 phosphatase-kinase pairs,to the ion channel SLACl.The ABA receptors interacted with and inhibited PP2C phosphatase activity against the SnRK2-type kinase,releasing active SnRK2 kinase to phosphorylate,and activate the SLACl channel,leading to reduced guard cell turgor and stomatal closure.Both yeast two-hybrid and bimolecular fluorescence complementation assays were used to verify the interactions among the components in the pathway.These biochemical assays demonstrated activity modifications of phosphatases and kinases by their interaction partners.The SLACl channel activity was used as an endpoint readout for the strength of the signaling pathway,depending on the presence of different combinations of signaling components.Further study using transgenic plants overexpressing one of the ABA receptors demonstrated that changing the relative level of interacting partners would change ABA sensitivity.

  12. Fermented milk containing Lactobacillus GG alleviated DSS-induced colitis in mice and activated epidermal growth factor receptor and Akt signaling in intestinal epithelial cells

    Directory of Open Access Journals (Sweden)

    Kazutoyo Yoda

    2012-06-01

    Full Text Available Lactobacillus rhamnosus GG was assessed for its ability to alleviate DSS-induced colitis in mice and activate epidermal growth factor receptor and Akt signaling in intestinal epithelial cells. In this study mice were treated with DSS to induce colitis and they were given Lactobacillus GG fermented milk to assess the effect of probiotic on colitis. Lactobacillus GG fermented milk significantly reduced the colitis associated changes suggesting a protective effect against DSS induced colitis.

  13. Fermented milk containing Lactobacillus GG alleviated DSS-induced colitis in mice and activated epidermal growth factor receptor and Akt signaling in intestinal epithelial cells

    OpenAIRE

    Yoda, Kazutoyo; He, Fang; Miyazawa, Kenji; Hiramatsu, Masaru

    2012-01-01

    Lactobacillus rhamnosus GG was assessed for its ability to alleviate DSS-induced colitis in mice and activate epidermal growth factor receptor and Akt signaling in intestinal epithelial cells. In this study mice were treated with DSS to induce colitis and they were given Lactobacillus GG fermented milk to assess the effect of probiotic on colitis. Lactobacillus GG fermented milk significantly reduced the colitis associated changes suggesting a protective effect against DSS induced colitis.Key...

  14. Breast cancer cells can switch between estrogen receptor alpha and ErbB signaling and combined treatment against both signaling pathways postpones development of resistance

    DEFF Research Database (Denmark)

    Sonne-Hansen, Katrine; Norrie, Ida C; Emdal, Kristina Bennet; Benjaminsen, Rikke Vicki; Frogne, Thomas; Christiansen, Ib J; Kirkegaard, Tove; Lykkesfeldt, Anne E

    2010-01-01

    The majority of breast cancers are estrogen responsive, but upon progression of disease other growth promoting pathways are activated, e.g., the ErbB receptor system. The present study focuses on resistance to the pure estrogen antagonist fulvestrant and strategies to treat resistant cells or eve...... with pan-ErbB inhibition may postpone or even prevent development of treatment resistance....

  15. Nicotine stimulates urokinase-type plasminogen activator receptor expression and cell invasiveness through mitogen-activated protein kinase and reactive oxygen species signaling in ECV304 endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Khoi, Pham Ngoc; Park, Jung Sun; Kim, Nam Ho; Jung, Young Do, E-mail: ydjung@chonnam.ac.kr

    2012-03-01

    Urokinase-type plasminogen activator receptor (uPAR) expression is elevated during inflammation, tissue remodeling and in many human cancers. This study investigated the effect of nicotine, a major alkaloid in tobacco, on uPAR expression and cell invasiveness in ECV304 endothelial cells. Nicotine stimulated uPAR expression in a dose-dependent manner and activated extracellular signal-regulated kinases-1/2 (Erk-1/2), c-Jun amino-terminal kinase (JNK) and p38 mitogen activated protein kinase (MAPK). Specific inhibitors of MEK-1 (PD98059) and JNK (SP600125) inhibited the nicotine-induced uPAR expression, while the p38 MAPK inhibitor SB203580 did not. Expression vectors encoding dominant negative MEK-1 (pMCL-K97M) and JNK (TAM67) also prevented nicotine-induced uPAR promoter activity. The intracellular hydrogen peroxide (H{sub 2}O{sub 2}) content was increased by nicotine treatment. The antioxidant N-acetylcysteine prevented nicotine-activated production of reactive oxygen species (ROS) and uPAR expression. Furthermore, exogenous H{sub 2}O{sub 2} increased uPAR mRNA expression. Deleted and site-directed mutagenesis demonstrated the involvement of the binding sites of transcription factor nuclear factor-kappaB (NF-κB) and activator protein (AP)-1 in the nicotine-induced uPAR expression. Studies with expression vectors encoding mutated NF-κB signaling molecules and AP-1 decoy confirmed that NF-κB and AP-1 were essential for the nicotine-stimulated uPAR expression. MAPK (Erk-1/2 and JNK) and ROS functioned as upstream signaling molecules in the activation of AP-1 and NF-κB, respectively. In addition, ECV304 endothelial cells treated with nicotine displayed markedly enhanced invasiveness, which was partially abrogated by uPAR neutralizing antibodies. The data indicate that nicotine induces uPAR expression via the MAPK/AP-1 and ROS/NF-κB signaling pathways and, in turn, stimulates invasiveness in human ECV304 endothelial cells. -- Highlights: ► Endothelial cells

  16. Nicotine stimulates urokinase-type plasminogen activator receptor expression and cell invasiveness through mitogen-activated protein kinase and reactive oxygen species signaling in ECV304 endothelial cells

    International Nuclear Information System (INIS)

    Urokinase-type plasminogen activator receptor (uPAR) expression is elevated during inflammation, tissue remodeling and in many human cancers. This study investigated the effect of nicotine, a major alkaloid in tobacco, on uPAR expression and cell invasiveness in ECV304 endothelial cells. Nicotine stimulated uPAR expression in a dose-dependent manner and activated extracellular signal-regulated kinases-1/2 (Erk-1/2), c-Jun amino-terminal kinase (JNK) and p38 mitogen activated protein kinase (MAPK). Specific inhibitors of MEK-1 (PD98059) and JNK (SP600125) inhibited the nicotine-induced uPAR expression, while the p38 MAPK inhibitor SB203580 did not. Expression vectors encoding dominant negative MEK-1 (pMCL-K97M) and JNK (TAM67) also prevented nicotine-induced uPAR promoter activity. The intracellular hydrogen peroxide (H2O2) content was increased by nicotine treatment. The antioxidant N-acetylcysteine prevented nicotine-activated production of reactive oxygen species (ROS) and uPAR expression. Furthermore, exogenous H2O2 increased uPAR mRNA expression. Deleted and site-directed mutagenesis demonstrated the involvement of the binding sites of transcription factor nuclear factor-kappaB (NF-κB) and activator protein (AP)-1 in the nicotine-induced uPAR expression. Studies with expression vectors encoding mutated NF-κB signaling molecules and AP-1 decoy confirmed that NF-κB and AP-1 were essential for the nicotine-stimulated uPAR expression. MAPK (Erk-1/2 and JNK) and ROS functioned as upstream signaling molecules in the activation of AP-1 and NF-κB, respectively. In addition, ECV304 endothelial cells treated with nicotine displayed markedly enhanced invasiveness, which was partially abrogated by uPAR neutralizing antibodies. The data indicate that nicotine induces uPAR expression via the MAPK/AP-1 and ROS/NF-κB signaling pathways and, in turn, stimulates invasiveness in human ECV304 endothelial cells. -- Highlights: ► Endothelial cells treated with nicotine

  17. Netrin-1 induces the migration of Schwann cells via p38 MAPK and PI3K-Akt signaling pathway mediated by the UNC5B receptor

    International Nuclear Information System (INIS)

    Schwann cells (SCs) play an essentially supportive role in the regeneration of injured peripheral nerve system (PNS). As Netrin-1 is crucial for the normal development of nervous system (NS) and can direct the process of damaged PNS regeneration, our study was designed to determine the role of Netrin-1 in RSC96 Schwann cells (an immortalized rat Schwann cell line) proliferation and migration. Our studies demonstrated that Netrin-1 had no effect on RSC96 cells proliferation, while significantly promoted RSC96 cells migration. The Netrin-1-induced RSC96 cells migration was significantly attenuated by inhibition of p38 and PI3K through pretreatment with SB203580 and LY294002 respectively, but not inhibition of MEK1/2 and JNK by U0126-EtOH and SP600125 individually. Treatment with Netrin-1 enhanced the phosphorylation of p38 and Akt. QRT-PCR indicated that Netrin-1 and only its receptors Unc5a, Unc5b and Neogenin were expressed in RSC96 cells, among which Unc5b expressed the most. And UNC5B protein was significantly increased after stimulated by Netrin-1. In conclusion, we show here that Netrin-1-enhanced SCs migration is mediated by activating p38 MAPK and PI3K-Akt signal cascades via receptor UNC5B, which suggests that Netrin-1 could serve as a new therapeutic strategy and has potential application value for PNS regeneration. - Highlights: • Netrin-1 attracts RSC96 Schwann cells migration in a dose dependent manner. • Netrin-1 induced Schwann cells migration is p38 and PI3K-Akt signaling dependent. • UNC5B may be dominant receptor mediating Netrin-1′ effect on RSC96 cells motility. • Netrin-1 may promote peripheral nerve repair by enhancing Schwann cells motility

  18. Netrin-1 induces the migration of Schwann cells via p38 MAPK and PI3K-Akt signaling pathway mediated by the UNC5B receptor

    Energy Technology Data Exchange (ETDEWEB)

    Lv, Jianwei [General Hospital of Tianjin Medical University, No. 154, Anshan Road, Heping District, Tianjin 300052 (China); Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China); Sun, Xiaolei; Ma, Jianxiong [Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China); Ma, Xinlong, E-mail: gengxiao502@163.com [General Hospital of Tianjin Medical University, No. 154, Anshan Road, Heping District, Tianjin 300052 (China); Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China); Zhang, Yang; Li, Fengbo; Li, Yanjun; Zhao, Zhihu [Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050 (China)

    2015-08-14

    Schwann cells (SCs) play an essentially supportive role in the regeneration of injured peripheral nerve system (PNS). As Netrin-1 is crucial for the normal development of nervous system (NS) and can direct the process of damaged PNS regeneration, our study was designed to determine the role of Netrin-1 in RSC96 Schwann cells (an immortalized rat Schwann cell line) proliferation and migration. Our studies demonstrated that Netrin-1 had no effect on RSC96 cells proliferation, while significantly promoted RSC96 cells migration. The Netrin-1-induced RSC96 cells migration was significantly attenuated by inhibition of p38 and PI3K through pretreatment with SB203580 and LY294002 respectively, but not inhibition of MEK1/2 and JNK by U0126-EtOH and SP600125 individually. Treatment with Netrin-1 enhanced the phosphorylation of p38 and Akt. QRT-PCR indicated that Netrin-1 and only its receptors Unc5a, Unc5b and Neogenin were expressed in RSC96 cells, among which Unc5b expressed the most. And UNC5B protein was significantly increased after stimulated by Netrin-1. In conclusion, we show here that Netrin-1-enhanced SCs migration is mediated by activating p38 MAPK and PI3K-Akt signal cascades via receptor UNC5B, which suggests that Netrin-1 could serve as a new therapeutic strategy and has potential application value for PNS regeneration. - Highlights: • Netrin-1 attracts RSC96 Schwann cells migration in a dose dependent manner. • Netrin-1 induced Schwann cells migration is p38 and PI3K-Akt signaling dependent. • UNC5B may be dominant receptor mediating Netrin-1′ effect on RSC96 cells motility. • Netrin-1 may promote peripheral nerve repair by enhancing Schwann cells motility.

  19. G protein-coupled receptor 30 mediates estrogen-induced proliferation of primordial germ cells via EGFR/Akt/β-catenin signaling pathway.

    Science.gov (United States)

    Ge, Chutian; Yu, Minli; Zhang, Caiqiao

    2012-07-01

    In vertebrates, estrogens are required for the normal development and function of postnatal gonads. However, it remains unclear whether estrogens are able to modulate development of the fetal germ cells. Here, we show that, unexpectedly, chicken primordial germ cells (PGC) lacking estrogen receptor α/β still proliferate in response to 17β-estradiol (E(2)). This is due to the capacity of G protein-coupled receptor 30 (GPR30), existing on PGC, to directly bind E(2). Knockdown experiments suggest that GPR30 is required for E(2)-stimulated PGC proliferation. Furthermore, this estrogen-induced activation of GPR30 is revealed to occur through the Gβγ-subunit protein-dependent and through the matrix metalloproteinase-dependent transactivation of the epidermal growth factor receptor. Epidermal growth factor receptor activation results in a series of intracellular events, including activation of the phosphatidylinositol 3-kinase/serine-threonine kinase/β-catenin pathway, which are followed by the induction of c-fos, c-myc, cyclin D1/E, and B-cell lymphoma 2 expression, and the inhibition of B-cell lymphoma 2-associated X protein expression and caspase3/9 activity. This eventually leads to decreased apoptosis and increased PGC proliferation. Collectively, these findings offer novel insights into the dynamic mechanism of estrogen action on PGC proliferation and suggest that E(2)/GPR30 signaling might play an important role in regulating fetal germ cell development, particularly at the stage before sexual differentiation. PMID:22635679

  20. Cell signaling review series

    Institute of Scientific and Technical Information of China (English)

    Aiming Lin; Zhenggang Liu

    2008-01-01

    @@ Signal transduction is pivotal for many, if not all, fundamental cellular functions including proliferation, differentiation, transformation and programmed cell death. Deregulation of cell signaling may result in certain types of cancers and other human diseases.

  1. Involvement of M3 Cholinergic Receptor Signal Transduction Pathway in Regulation of the Expression of Chemokine MOB-1, MCP-1 Genes in Pancreatic Acinar Cells

    Institute of Scientific and Technical Information of China (English)

    郑海; 陈道达; 张景輝; 田原

    2004-01-01

    Whether M3 cholinergic receptor signal transduction pathway is involved in regulation of the activation of NF-κB and the expression of chemokine MOB-1, MCP-1genes in pancreatic acinar cells was investigated. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, atropine and PDTC in vitro. The MOB-1 and MCP-1 mRNA expression was detected by using RT-PCR. The activation of NF-κB was monitored by using electrophoretic mobility shift assay.The results showed that as compared with control group, M3 cholinergic receptor agonist (103mol/L, 104-4ol/L carbachol) could induce a concentration-dependent and time-dependent increase in the expression of MOB-1, MCP-1 mRNA in pancreatic acinar cells. After treatment with 10 -3mol/L carbachol for 2 h, the expression of MOB-1, MCP-1 mRNA was strongest. The activity of NF-κB in pancreatic acinar cells was significantly increased (P<0.01) after treated with M3 cholinergic receptor agonist (10-3 mol/L carbachol) in vitro for 30 min. Either M3 cholinergic receptor antagonist (10-5 mol/L atropine) or NF-κB inhibitor (10-2 mol/L PDTC) could obviously inhibit the activation of NF-κB and the chemokine MOB-1, MCP-1 mRNA expression induced by carbachol (P <0.05). This inhibitory effect was significantly increased by atropine plus PDTC (P<0.01). The results of these studies indicated that M3 cholinergic receptor signal transduction pathway was likely involved in regulation of the expression of chemokine MOB-1 and MCP-1genes in pancreatic acinar cells in vitro through the activation of NF-κB.

  2. Interdependent epidermal growth factor receptor signalling and trafficking.

    Science.gov (United States)

    Jones, Sylwia; Rappoport, Joshua Z

    2014-06-01

    Epidermal growth factor (EGF) receptor (EGFR) signalling regulates diverse cellular functions, promoting cell proliferation, differentiation, migration, cell growth and survival. EGFR signalling is critical during embryogenesis, in particular in epithelial development, and disruption of the EGFR gene results in epithelial immaturity and perinatal death. EGFR signalling also functions during wound healing responses through accelerating wound re-epithelialisation, inducing cell migration, proliferation and angiogenesis. Upregulation of EGFR signalling is often observed in carcinomas and has been shown to promote uncontrolled cell proliferation and metastasis. Therefore aberrant EGFR signalling is a common target for anticancer therapies. Various reports indicate that EGFR signalling primarily occurs at the plasma membrane and EGFR degradation following endocytosis greatly attenuates signalling. Other studies argue that EGFR internalisation is essential for complete activation of downstream signalling cascades and that endosomes can serve as signalling platforms. The aim of this review is to discuss current understanding of intersection between EGFR signalling and trafficking. PMID:24681003

  3. Leucine leucine-37 uses formyl peptide receptor-like 1 to activate signal transduction pathways, stimulate oncogenic gene expression, and enhance the invasiveness of ovarian cancer cells.

    Science.gov (United States)

    Coffelt, Seth B; Tomchuck, Suzanne L; Zwezdaryk, Kevin J; Danka, Elizabeth S; Scandurro, Aline B

    2009-06-01

    Emerging evidence suggests that the antimicrobial peptide, leucine leucine-37 (LL-37), could play a role in the progression of solid tumors. LL-37 is expressed as the COOH terminus of human cationic antimicrobial protein-18 (hCAP-18) in ovarian, breast, and lung cancers. Previous studies have shown that the addition of LL-37 to various cancer cell lines in vitro stimulates proliferation, migration, and invasion. Similarly, overexpression of hCAP-18/LL-37 in vivo accelerates tumor growth. However, the receptor or receptors through which these processes are mediated have not been thoroughly examined. In the present study, expression of formyl peptide receptor-like 1 (FPRL1) was confirmed on ovarian cancer cells. Proliferation assays indicated that LL-37 does not signal through a G protein-coupled receptor, such as FPRL1, to promote cancer cell growth. By contrast, FPRL1 was required for LL-37-induced invasion through Matrigel. The peptide stimulated mitogen-activated protein kinase and Janus-activated kinase/signal transducers and activators of transcription signaling cascades and led to the significant activation of several transcription factors, through both FPRL1-dependent and FPRL1-independent pathways. Likewise, expression of some LL-37-stimulated genes was attenuated by the inhibition of FPRL1. Increased expression of CXCL10, EGF, and PDGF-BB as well as other soluble factors was confirmed from conditioned medium of LL-37-treated cells. Taken together, these data suggest that LL-37 potentiates a more aggressive behavior from ovarian cancer cells through its interaction with FPRL1. PMID:19491199

  4. The insulin receptor as a transmitter of a mitogenic signal in Chinese hamster ovary CHO-K1 cells

    International Nuclear Information System (INIS)

    Insulin is the only hormone required for continued growth of Chinese hamster ovary CHO-K1 cells in the defined medium M-F12. When CHO-K1 cells are incubated in M-F12 without insulin for 48-72 hr, the cells accumulate in G1. In response to physiological concentrations of insulin an 18-fold increase in rate of DNA synthesis occurs due to cells entering S phase after an 8- to 10-hr lag; cell division begins after 24 hr. The inhibitory effect of actinomycin D and 5,6-dichlorobenzimidazole riboside indicates that RNA synthesis is required for progression to S phase. CHO-K1 cells possess insulin receptors, and the insulin effect results from insulin binding to its own receptor: (i) binding occurs at physiological insulin concentrations with a half-maximal stimulation at ∼ 14 ng/ml. (ii) At insulin concentrations used, insulin-like growth factor I and II (IGF-I and IGF-II) have little or no effect. (iii) Scatchard analysis of 125I-labeled insulin binding shows the curvilinear response typical of insulin. (iv) The Kd for the so-called high-affinity binding site and the Ke are characteristic of the insulin receptor. (v) At the minimal insulin concentrations that stimulate growth, IGF-I and IGF-II compete poorly with insulin for insulin binding, insulin competes poorly with IGF-I for IGF-I binding, and affinity labeling with 125I-labeled insulin identifies as polypeptide typical of the α subunit of the insulin receptor

  5. Estrogen Regulates MAPK-Related Genes through Genomic and Nongenomic Interactions between IGF-I Receptor Tyrosine Kinase and Estrogen Receptor-Alpha Signaling Pathways in Human Uterine Leiomyoma Cells.

    Science.gov (United States)

    Yu, Linda; Moore, Alicia B; Castro, Lysandra; Gao, Xiaohua; Huynh, Hoang-Long C; Klippel, Michelle; Flagler, Norris D; Lu, Yi; Kissling, Grace E; Dixon, Darlene

    2012-01-01

    Estrogen and growth factors play a major role in uterine leiomyoma (UtLM) growth possibly through interactions of receptor tyrosine kinases (RTKs) and estrogen receptor-alpha (ERα) signaling. We determined the genomic and nongenomic effects of 17β-estradiol (E(2)) on IGF-IR/MAPKp44/42 signaling and gene expression in human UtLM cells with intact or silenced IGF-IR. Analysis by RT(2) Profiler PCR-array showed genes involved in IGF-IR/MAPK signaling were upregulated in UtLM cells by E(2) including cyclin D kinases, MAPKs, and MAPK kinases; RTK signaling mediator, GRB2; transcriptional factors ELK1 and E2F1; CCNB2 involved in cell cycle progression, proliferation, and survival; and COL1A1 associated with collagen synthesis. Silencing (si)IGF-IR attenuated the above effects and resulted in upregulation of different genes, such as transcriptional factor ETS2; the tyrosine kinase receptor, EGFR; and DLK1 involved in fibrosis. E(2) rapidly activated IGF-IR/MAPKp44/42 signaling nongenomically and induced phosphorylation of ERα at ser118 in cells with a functional IGF-IR versus those without. E(2) also upregulated IGF-I gene and protein expression through a prolonged genomic event. These results suggest a pivotal role of IGF-IR and possibly other RTKs in mediating genomic and nongenomic hormone receptor interactions and signaling in fibroids and provide novel genes and targets for future intervention and prevention strategies. PMID:23094148

  6. Tyrosine kinase signalling in breast cancer: Epidermal growth factor receptor - convergence point for signal integration and diversification

    International Nuclear Information System (INIS)

    Cross-communication between different signalling systems is critical for the integration of multiple and changing environmental influences on individual cells. The epidermal growth factor receptor (EGFR) has been identified as a key element in the complex signalling network that is utilized by various classes of cell-surface receptors. This nonclassical mode of signalling system cross-talk, in distinction to receptor activation induced by cognate ligands, has been termed 'signal transactivation'. With the EGFR as the convergence point and distribution focus, this scenario may involve signals emitted by other members of the tyrosine kinase family, cytokine receptors, ion channels, G-protein-coupled receptors and integrins

  7. Proliferative signaling initiated in ACTH receptors

    Directory of Open Access Journals (Sweden)

    C.F.P. Lotfi

    2000-10-01

    Full Text Available This article reviews recent results of studies aiming to elucidate modes of integrating signals initiated in ACTH receptors and FGF2 receptors, within the network system of signal transduction found in Y1 adrenocortical cells. These modes of signal integration should be central to the mechanisms underlying the regulation of the G0->G1->S transition in the adrenal cell cycle. FGF2 elicits a strong mitogenic response in G0/G1-arrested Y1 adrenocortical cells, that includes a rapid and transient activation of extracellular signal-regulated kinases-mitogen-activated protein kinases (ERK-MAPK (2 to 10 min, b transcription activation of c-fos, c-jun and c-myc genes (10 to 30 min, c induction of c-Fos and c-Myc proteins by 1 h and cyclin D1 protein by 5 h, and d onset of DNA synthesis stimulation within 8 h. ACTH, itself a weak mitogen, interacts with FGF2 in a complex manner, blocking the FGF2 mitogenic response during the early and middle G1 phase, keeping ERK-MAPK activation and c-Fos and cyclin D1 induction at maximal levels, but post-transcriptionally inhibiting c-Myc expression. c-Fos and c-Jun proteins are mediators in both the strong and the weak mitogenic responses respectively triggered by FGF2 and ACTH. Induction of c-Fos and stimulation of DNA synthesis by ACTH are independent of PKA and are inhibited by the PKC inhibitor GF109203X. In addition, ACTH is a poor activator of ERK-MAPK, but c-Fos induction and DNA synthesis stimulation by ACTH are strongly inhibited by the inhibitor of MEK1 PD98059.

  8. Progesterone receptors – animal models and cell signaling in breast cancer: The role of oestrogen and progesterone receptors in human mammary development and tumorigenesis

    International Nuclear Information System (INIS)

    A relatively small number of cells in the normal human mammary gland express receptors for oestrogen and progesterone (ER and PR), and there is almost complete dissociation between steroid receptor expression and proliferation. Increased expression of the ER alpha (ERα) and loss of the inverse relationship between receptor expression and proliferation occur at the very earliest stages of tumorigenesis, implying that dysregulation of ERα expression contributes to breast tumour formation. There is evidence also for alterations in the ratio between the two PR isoforms in premalignant breast lesions. Elucidation of the factors mediating the effects of oestradiol and progesterone on development of the normal breast and of the mechanisms by which expression of the ERα and the PR isoforms is controlled could identify new targets for breast cancer prevention and improved prediction of breast cancer risk

  9. LPA receptor signaling: pharmacology, physiology, and pathophysiology

    OpenAIRE

    Yung, Yun C.; Stoddard, Nicole C.; Chun, Jerold

    2014-01-01

    Lysophosphatidic acid (LPA) is a small ubiquitous lipid found in vertebrate and nonvertebrate organisms that mediates diverse biological actions and demonstrates medicinal relevance. LPA’s functional roles are driven by extracellular signaling through at least six 7-transmembrane G protein-coupled receptors. These receptors are named LPA1–6 and signal through numerous effector pathways activated by heterotrimeric G proteins, including Gi/o, G12/13, Gq, and Gs. LPA receptor-mediated effects ha...

  10. Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling

    International Nuclear Information System (INIS)

    Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2 and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. - Highlights: • PAUF confers resistance against oncolytic parvovirus H-1 infection. • PAUF enhances the expression of IFNAR in Panc-1 cells. • Increased activation of Tyk2 or Stat1 by PAUF provides resistance to parvovirus H-1-mediated apoptosis. • Constitutive inhibition of PAUF enhances parvovirus H-1-mediated oncolysis of Bxpc3 pancreatic cancer cells

  11. Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling

    Energy Technology Data Exchange (ETDEWEB)

    Kaowinn, Sirichat; Cho, Il-Rae; Moon, Jeong; Jun, Seung Won; Kim, Chang Seok [BK21+, Department of Cogno-Mechatronics Engineering, Pusan National University, Busan 609-736 (Korea, Republic of); Kang, Ho Young [Department of Microbiology, Pusan National University, Busan 609-736 (Korea, Republic of); Kim, Manbok [Department of Medical Science, Dankook University College of Medicine, Cheonan 330-714 (Korea, Republic of); Koh, Sang Seok [Department of Biological Sciences, Dong-A University, Busan 604-714 (Korea, Republic of); Chung, Young-Hwa, E-mail: younghc@pusan.ac.kr [BK21+, Department of Cogno-Mechatronics Engineering, Pusan National University, Busan 609-736 (Korea, Republic of)

    2015-04-03

    Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2 and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. - Highlights: • PAUF confers resistance against oncolytic parvovirus H-1 infection. • PAUF enhances the expression of IFNAR in Panc-1 cells. • Increased activation of Tyk2 or Stat1 by PAUF provides resistance to parvovirus H-1-mediated apoptosis. • Constitutive inhibition of PAUF enhances parvovirus H-1-mediated oncolysis of Bxpc3 pancreatic cancer cells.

  12. Regulator of G protein signaling 8 inhibits protease-activated receptor 1/Gi/o signaling by forming a distinct G protein-dependent complex in live cells.

    Science.gov (United States)

    Lee, Jinyong; Ghil, Sungho

    2016-05-01

    Activation of seven-transmembrane-domain-possessing G protein-coupled receptors (GPCRs) by extracellular stimuli elicits intracellular responses. One class of GPCRs-protease-activated receptors (PARs)-is activated by endogenous proteases, such as thrombin and trypsin. Members of the regulator of G protein signaling (RGS) family stimulate GTP hydrolysis of G protein alpha (Gα) subunits, thereby inhibiting GPCR/Gα-mediated signaling. We previously reported that RGS2 and RGS4 inhibit PAR1/Gα-mediated signaling by interacting with PAR1 in a Gα-dependent manner. Here, employing the bioluminescence resonance energy transfer (BRET) technique, we identified RGS8 as a novel PAR1-interacting protein. Very little BRET activity was observed between PAR1-Venus (PAR1-Ven) and RGS8-Luciferase (RGS8-Luc) in the absence of Gα. However, in the presence of Gαo, BRET activity was specifically and significantly increased. This interaction was confirmed by biochemical and immunofluorescence assays. Notably, RGS8 inhibited PAR1/Gαi/o-mediated adenylyl cyclase and ERK activation, and prevented Gαo-induced neurite outgrowth and activation of Necdin protein, a downstream target of Gαo. Our findings suggest a novel function of RGS8 and reveal cellular mechanisms by which RGS8 mediates PAR1 inhibition. PMID:26829215

  13. An additive interaction between the NFκB and estrogen receptor signalling pathways in human endometrial epithelial cells

    OpenAIRE

    King, A E; Collins, F; Klonisch, T; Sallenave, J-M; Critchley, H.O.D.; Saunders, P T K

    2009-01-01

    Human embryo implantation is regulated by estradiol (E2), progesterone and locally produced mediators including interleukin-1 beta (IL-1 beta). Interactions between the estrogen receptor (ER) and NF kappa B (NF kappa B) signalling pathways have been reported in other systems but have not been detailed in human endometrium.Real-time PCR showed that mRNA for the p65 and p105 NF kappa B subunits is maximally expressed in endometrium from the putative implantation window. Both subunits are locali...

  14. Redox-dependent regulation of epidermal growth factor receptor signaling.

    Science.gov (United States)

    Heppner, David E; van der Vliet, Albert

    2016-08-01

    Tyrosine phosphorylation-dependent cell signaling represents a unique feature of multicellular organisms, and is important in regulation of cell differentiation and specialized cell functions. Multicellular organisms also contain a diverse family of NADPH oxidases (NOXs) that have been closely linked with tyrosine kinase-based cell signaling and regulate tyrosine phosphorylation via reversible oxidation of cysteine residues that are highly conserved within many proteins involved in this signaling pathway. An example of redox-regulated tyrosine kinase signaling involves the epidermal growth factor receptor (EGFR), a widely studied receptor system with diverse functions in normal cell biology as well as pathologies associated with oxidative stress such as cancer. The purpose of this Graphical Redox Review is to highlight recently emerged concepts with respect to NOX-dependent regulation of this important signaling pathway. PMID:26722841

  15. Squamosamide derivative FLZ protects retinal pigment epithelium cells from oxidative stress through activation of epidermal growth factor receptor (EGFR)-AKT signaling.

    Science.gov (United States)

    Cheng, Li-Bo; Chen, Chun-Ming; Zhong, Hong; Zhu, Li-Juan

    2014-01-01

    Reactive oxygen species (ROS)-mediated retinal pigment epithelium (RPE) cell apoptosis is attributed to age-related macular degeneration (AMD) pathogenesis. FLZ, a novel synthetic squamosamide derivative from a Chinese herb, Annona glabra, has displayed significant cyto-protective activity. In the current study, we explored the pro-survival effect of FLZ in oxidative stressed-RPE cells and studied the underlying signaling mechanisms. Our results showed that FLZ attenuated hydrogen peroxide (H2O2)-induced viability decrease and apoptosis in the RPE cell line (ARPE-19 cells) and in primary mouse RPE cells. Western blotting results showed that FLZ activated AKT signaling in RPE cells. The AKT-specific inhibitor, MK-2206, the phosphoinositide 3-kinase (PI3K)/AKT pan inhibitor, wortmannin, and AKT1-shRNA (short hairpin RNA) depletion almost abolished FLZ-mediated pro-survival/anti-apoptosis activity. We discovered that epidermal growth factor receptor (EGFR) trans-activation mediated FLZ-induced AKT activation and the pro-survival effect in RPE cells, and the anti-apoptosis effect of FLZ against H2O2 was inhibited by the EGFR inhibitor, PD153035, or by EGFR shRNA-knockdown. In conclusion, FLZ protects RPE cells from oxidative stress through activation of EGFR-AKT signaling, and our results suggest that FLZ might have therapeutic values for AMD. PMID:25329617

  16. Thyrotropin Receptor Stimulates Internalization-Independent Persistent Phosphoinositide Signaling

    OpenAIRE

    Boutin, Alisa; Allen, Michael D.; Geras-Raaka, Elizabeth; Huang, Wenwei; Neumann, Susanne; Gershengorn, Marvin C.

    2011-01-01

    The thyrotropin [thyroid-stimulating hormone (TSH)] receptor (TSHR) is known to acutely and persistently stimulate cAMP signaling and at higher TSH concentrations to acutely stimulate phosphoinositide signaling. We measured persistent signaling by stimulating TSHR-expressing human embryonic kidney-EM293 cells with TSH and measuring cAMP or inositol monophosphate (IP1) production, a measure of phosphoinositide signaling, 60 min or longer after TSH removal. In contrast to persistent cAMP produc...

  17. Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling.

    Science.gov (United States)

    Kaowinn, Sirichat; Cho, Il-Rae; Moon, Jeong; Jun, Seung Won; Kim, Chang Seok; Kang, Ho Young; Kim, Manbok; Koh, Sang Seok; Chung, Young-Hwa

    2015-04-01

    Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2 and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. PMID:25727013

  18. Hyaluronan suppresses prostate tumor cell proliferation through diminished expression of N-cadherin and aberrant growth factor receptor signaling

    Energy Technology Data Exchange (ETDEWEB)

    Bharadwaj, Alamelu G.; Goodrich, Nathaniel P.; McAtee, Caitlin O.; Haferbier, Katie [Department of Biochemistry, University of Nebraska, Lincoln, NE 68588 (United States); Oakley, Gregory G.; Wahl, James K. [Department of Oral Biology, University of Nebraska College of Dentistry, Lincoln, NE 68588 (United States); Simpson, Melanie A., E-mail: msimpson2@unl.edu [Department of Biochemistry, University of Nebraska, Lincoln, NE 68588 (United States); Eppley Cancer Center, University of Nebraska Medical Center, Omaha, NE 68198 (United States)

    2011-05-01

    Hyaluronan (HA) production has been functionally implicated in prostate tumorigenesis and metastasis. We previously used prostate tumor cells overexpressing the HA synthesizing enzyme HAS3 or the clinically relevant hyaluronidase Hyal1 to show that excess HA production suppresses tumor growth, while HA turnover accelerates spontaneous metastasis from the prostate. Here, we examined pathways responsible for effects of HAS3 and Hyal1 on tumor cell phenotype. Detailed characterization of cell cycle progression revealed that expression of Hyal1 accelerated cell cycle re-entry following synchronization, whereas HAS3 alone delayed entry. Hyal1 expressing cells exhibited a significant reduction in their ability to sustain ERK phosphorylation upon stimulation by growth factors, and in their expression of the cyclin-dependent kinase inhibitor p21. In contrast, HAS3 expressing cells showed prolonged ERK phosphorylation and increased expression of both p21 and p27, in asynchronous and synchronized cultures. Changes in cell cycle regulatory proteins were accompanied by HA-induced suppression of N-cadherin, while E-cadherin expression and {beta}-catenin expression and distribution remained unchanged. Our results are consistent with a model in which excess HA synthesis suppresses cell proliferation by promoting homotypic E-cadherin mediated cell-cell adhesion, consequently signaling to elevate cell cycle inhibitor expression and suppress G1- to S-phase transition.

  19. Hyaluronan suppresses prostate tumor cell proliferation through diminished expression of N-cadherin and aberrant growth factor receptor signaling

    International Nuclear Information System (INIS)

    Hyaluronan (HA) production has been functionally implicated in prostate tumorigenesis and metastasis. We previously used prostate tumor cells overexpressing the HA synthesizing enzyme HAS3 or the clinically relevant hyaluronidase Hyal1 to show that excess HA production suppresses tumor growth, while HA turnover accelerates spontaneous metastasis from the prostate. Here, we examined pathways responsible for effects of HAS3 and Hyal1 on tumor cell phenotype. Detailed characterization of cell cycle progression revealed that expression of Hyal1 accelerated cell cycle re-entry following synchronization, whereas HAS3 alone delayed entry. Hyal1 expressing cells exhibited a significant reduction in their ability to sustain ERK phosphorylation upon stimulation by growth factors, and in their expression of the cyclin-dependent kinase inhibitor p21. In contrast, HAS3 expressing cells showed prolonged ERK phosphorylation and increased expression of both p21 and p27, in asynchronous and synchronized cultures. Changes in cell cycle regulatory proteins were accompanied by HA-induced suppression of N-cadherin, while E-cadherin expression and β-catenin expression and distribution remained unchanged. Our results are consistent with a model in which excess HA synthesis suppresses cell proliferation by promoting homotypic E-cadherin mediated cell-cell adhesion, consequently signaling to elevate cell cycle inhibitor expression and suppress G1- to S-phase transition.

  20. Impaired Ca(2+ signaling in β-cells lacking leptin receptors by Cre-loxP recombination.

    Directory of Open Access Journals (Sweden)

    Eva Tudurí

    Full Text Available Obesity is a major risk factor for diabetes and is typically associated with hyperleptinemia and a state of leptin resistance. The impact of chronically elevated leptin levels on the function of insulin-secreting β-cells has not been elucidated. We previously generated mice lacking leptin signaling in β-cells by using the Cre-loxP strategy and showed that these animals develop increased body weight and adiposity, hyperinsulinemia, impaired glucose-stimulated insulin secretion and insulin resistance. Here, we performed several in vitro studies and observed that β-cells lacking leptin signaling in this model are capable of properly metabolizing glucose, but show impaired intracellular Ca(2+ oscillations and lack of synchrony within the islets in response to glucose, display reduced response to tolbutamide and exhibit morphological abnormalities including increased autophagy. Defects in intracellular Ca(2+ signaling were observed even in neonatal islets, ruling out the possible contribution of obesity to the β-cell irregularities observed in adults. In parallel, we also detected a disrupted intracellular Ca(2+ pattern in response to glucose and tolbutamide in control islets from adult transgenic mice expressing Cre recombinase under the rat insulin promoter, despite these animals being glucose tolerant and secreting normal levels of insulin in response to glucose. This unexpected observation impeded us from discerning the consequences of impaired leptin signaling as opposed to long-term Cre expression in the function of insulin-secreting cells. These findings highlight the need to generate improved Cre-driver mouse models or new tools to induce Cre recombination in β-cells.

  1. Nexus of signaling and endocytosis in oncogenesis driven by non-small cell lung cancer-associated epidermal growth factor receptor mutants.

    Science.gov (United States)

    Chung, Byung Min; Tom, Eric; Zutshi, Neha; Bielecki, Timothy Alan; Band, Vimla; Band, Hamid

    2014-12-10

    Epidermal growth factor receptor (EGFR) controls a wide range of cellular processes, and aberrant EGFR signaling as a result of receptor overexpression and/or mutation occurs in many types of cancer. Tumor cells in non-small cell lung cancer (NSCLC) patients that harbor EGFR kinase domain mutations exhibit oncogene addiction to mutant EGFR, which confers high sensitivity to tyrosine kinase inhibitors (TKIs). As patients invariably develop resistance to TKIs, it is important to delineate the cell biological basis of mutant EGFR-induced cellular transformation since components of these pathways can serve as alternate therapeutic targets to preempt or overcome resistance. NSCLC-associated EGFR mutants are constitutively-active and induce ligand-independent transformation in nonmalignant cell lines. Emerging data suggest that a number of factors are critical for the mutant EGFR-dependent tumorigenicity, and bypassing the effects of TKIs on these pathways promotes drug resistance. For example, activation of downstream pathways such as Akt, Erk, STAT3 and Src is critical for mutant EGFR-mediated biological processes. It is now well-established that the potency and spatiotemporal features of cellular signaling by receptor tyrosine kinases such as EGFR, as well as the specific pathways activated, is determined by the nature of endocytic traffic pathways through which the active receptors traverse. Recent evidence indicates that NSCLC-associated mutant EGFRs exhibit altered endocytic trafficking and they exhibit reduced Cbl ubiquitin ligase-mediated lysosomal downregulation. More recent work has shown that mutant EGFRs undergo ligand-independent traffic into the endocytic recycling compartment, a behavior that plays a key role in Src pathway activation and oncogenesis. These studies are beginning to delineate the close nexus between signaling and endocytic traffic of EGFR mutants as a key driver of oncogenic processes. Therefore, in this review, we will discuss the links

  2. HOXA13 exerts a beneficial effect in albumin-induced epithelial-mesenchymal transition via the glucocorticoid receptor signaling pathway in human renal tubular epithelial cells.

    Science.gov (United States)

    Peng, Li; He, Qingnan; Li, Xiaoyan; Shuai, Lanjun; Chen, Haixia; Li, Yongzhen; Yi, Zhuwen

    2016-07-01

    Previous studies have suggested that albumin-induced renal tubular epithelial cell injury contributes to renal interstitial fibrosis. Epithelial-mesenchymal transition (EMT) is known to be a key mechanism in the pathogenesis and progression of renal interstitial fibrosis. Homeobox protein HOX‑A13 (HOXA13) is a nuclear transcriptional factor that has been reported to be involved in renal fibrosis. However, the mechanism underlying the effect of HOXA13 in human serum albumin (HSA)‑induced EMT in HKC renal tubular epithelial cells remains to be elucidated. Thus, the aim of the present study was to investigate the role of HOXA13 in HSA‑induced EMT in HKC cells and the potential mechanism of the glucocorticoid receptor (GR) signaling pathway. The protein and mRNA expression levels of HOXA13, cytokeratin, and vimentin were determined by western blot analysis and reverse transcription‑quantitative polymerase chain reaction in HKC cells, which were co‑incubated with HSA at different concentrations or for different time periods. The results demonstrated that HOXA13 mRNA and protein expression decreased in a dose‑ and time‑dependent manner when induced by HSA in HCK cells. The liposomal transfection experiment suggested that overexpression of HOXA13 activated the GR signal, which inhibits HSA-induced EMT. HOXA13 is involved in HSA‑induced EMT in HKC cells and upregulation of HOXA13 exerts a beneficial effect in EMT, which may be associated with the GR signaling pathway. PMID:27176855

  3. Cocaine inhibits dopamine D2 receptor signaling via sigma-1-D2 receptor heteromers.

    Directory of Open Access Journals (Sweden)

    Gemma Navarro

    Full Text Available Under normal conditions the brain maintains a delicate balance between inputs of reward seeking controlled by neurons containing the D1-like family of dopamine receptors and inputs of aversion coming from neurons containing the D2-like family of dopamine receptors. Cocaine is able to subvert these balanced inputs by altering the cell signaling of these two pathways such that D1 reward seeking pathway dominates. Here, we provide an explanation at the cellular and biochemical level how cocaine may achieve this. Exploring the effect of cocaine on dopamine D2 receptors function, we present evidence of σ1 receptor molecular and functional interaction with dopamine D2 receptors. Using biophysical, biochemical, and cell biology approaches, we discovered that D2 receptors (the long isoform of the D2 receptor can complex with σ1 receptors, a result that is specific to D2 receptors, as D3 and D4 receptors did not form heteromers. We demonstrate that the σ1-D2 receptor heteromers consist of higher order oligomers, are found in mouse striatum and that cocaine, by binding to σ1 -D2 receptor heteromers, inhibits downstream signaling in both cultured cells and in mouse striatum. In contrast, in striatum from σ1 knockout animals these complexes are not found and this inhibition is not seen. Taken together, these data illuminate the mechanism by which the initial exposure to cocaine can inhibit signaling via D2 receptor containing neurons, destabilizing the delicate signaling balance influencing drug seeking that emanates from the D1 and D2 receptor containing neurons in the brain.

  4. Dopamine D2-like receptor signaling suppresses human osteoclastogenesis.

    Science.gov (United States)

    Hanami, Kentaro; Nakano, Kazuhisa; Saito, Kazuyoshi; Okada, Yosuke; Yamaoka, Kunihiro; Kubo, Satoshi; Kondo, Masahiro; Tanaka, Yoshiya

    2013-09-01

    Dopamine, a major neurotransmitter, transmits signals via five different seven-transmembrane G protein-coupled receptors termed D1 to D5. Although the relevance of neuroendocrine system to bone metabolism has been emerging, the precise effects of dopaminergic signaling upon osteoclastogenesis remain unknown. Here, we demonstrate that human monocyte-derived osteoclast precursor cells express all dopamine-receptor subtypes. Dopamine and dopamine D2-like receptor agonists such as pramipexole and quinpirole reduced the formation of TRAP-positive multi-nucleated cells, cathepsin K mRNA expression, and pit formation area in vitro. These inhibitory effects were reversed by pre-treatment with a D2-like receptor antagonist haloperidol or a Gαi inhibitor pertussis toxin, but not with the D1-like receptor antagonist SCH-23390. Dopamine and dopamine D2-like receptor agonists, but not a D1-like receptor agonist, suppressed intracellular cAMP concentration as well as RANKL-meditated induction of c-Fos and NFATc1 mRNA expression in human osteoclast precursor cells. Finally, the dopamine D2-like receptor agonist suppressed LPS-induced osteoclast formation in murine bone marrow culture ex vivo. These findings indicate that dopaminergic signaling plays an important role in bone homeostasis via direct effects upon osteoclast differentiation and further suggest that the clinical use of neuroleptics is likely to affect bone mass. PMID:23631878

  5. Green tea polyphenol epigallocatechin-3-gallate inhibits TLR4 signaling through the 67-kDa laminin receptor on lipopolysaccharide-stimulated dendritic cells

    Energy Technology Data Exchange (ETDEWEB)

    Byun, Eui-Baek [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); Choi, Han-Gyu [Department of Microbiology and Research Institute for Medical Sciences, College of Medicine, Chungnam National University, Daejeon 301-747 (Korea, Republic of); Sung, Nak-Yun [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); Byun, Eui-Hong, E-mail: ehbyun80@gmail.com [Department of Microbiology and Research Institute for Medical Sciences, College of Medicine, Chungnam National University, Daejeon 301-747 (Korea, Republic of)

    2012-10-05

    Highlights: Black-Right-Pointing-Pointer Expressions of CD80, CD86, and MHC class I/II were inhibited by EGCG via 67LR. Black-Right-Pointing-Pointer EGCG-treated DCs inhibited LPS-induced pro-inflammatory cytokines via 67LR. Black-Right-Pointing-Pointer EGCG-treated DCs inhibited MAPKs activation and NF-{kappa}B p65 translocation via 67LR. Black-Right-Pointing-Pointer EGCG elevated the expression of the Tollip protein through 67LR in DCs. -- Abstract: Epigallocatechin-3-gallate (EGCG), a major active polyphenol of green tea, has been shown to down-regulate inflammatory responses in dendritic cells (DCs); however, the underlying mechanism has not been understood. Recently, we identified the 67-kDa laminin receptor (67LR) as a cell-surface EGCG receptor. In this study, we showed the molecular basis for the down-regulation of toll-like receptor 4 (TLR4) signal transduction by EGCG in DCs. The expressions of CD80, CD86, and MHC class I and II, which are molecules essential for antigen presentation by DCs, were inhibited by EGCG via 67LR. In addition, EGCG-treated DCs inhibited lipopolysaccharide (LPS)-induced production of pro-inflammatory cytokines (tumor necrosis factor [TNF]-{alpha}, interleukin [IL]-1{beta}, and IL-6) and activation of mitogen-activated protein kinases (MAPKs), e.g., extracellular signal-regulated kinase 1/2 (ERK1/2), p38, c-Jun N-terminal kinase (JNK), and nuclear factor {kappa}B (NF-{kappa}B) p65 translocation through 67LR. Interestingly, we also found that EGCG markedly elevated the expression of the Tollip protein, a negative regulator of TLR signaling, through 67LR. These novel findings provide new insight into the understanding of negative regulatory mechanisms of the TLR4 signaling pathway and consequent inflammatory responses that are implicated in the development and progression of many chronic diseases.

  6. Green tea polyphenol epigallocatechin-3-gallate inhibits TLR4 signaling through the 67-kDa laminin receptor on lipopolysaccharide-stimulated dendritic cells

    International Nuclear Information System (INIS)

    Highlights: ► Expressions of CD80, CD86, and MHC class I/II were inhibited by EGCG via 67LR. ► EGCG-treated DCs inhibited LPS-induced pro-inflammatory cytokines via 67LR. ► EGCG-treated DCs inhibited MAPKs activation and NF-κB p65 translocation via 67LR. ► EGCG elevated the expression of the Tollip protein through 67LR in DCs. -- Abstract: Epigallocatechin-3-gallate (EGCG), a major active polyphenol of green tea, has been shown to down-regulate inflammatory responses in dendritic cells (DCs); however, the underlying mechanism has not been understood. Recently, we identified the 67-kDa laminin receptor (67LR) as a cell-surface EGCG receptor. In this study, we showed the molecular basis for the down-regulation of toll-like receptor 4 (TLR4) signal transduction by EGCG in DCs. The expressions of CD80, CD86, and MHC class I and II, which are molecules essential for antigen presentation by DCs, were inhibited by EGCG via 67LR. In addition, EGCG-treated DCs inhibited lipopolysaccharide (LPS)-induced production of pro-inflammatory cytokines (tumor necrosis factor [TNF]-α, interleukin [IL]-1β, and IL-6) and activation of mitogen-activated protein kinases (MAPKs), e.g., extracellular signal-regulated kinase 1/2 (ERK1/2), p38, c-Jun N-terminal kinase (JNK), and nuclear factor κB (NF-κB) p65 translocation through 67LR. Interestingly, we also found that EGCG markedly elevated the expression of the Tollip protein, a negative regulator of TLR signaling, through 67LR. These novel findings provide new insight into the understanding of negative regulatory mechanisms of the TLR4 signaling pathway and consequent inflammatory responses that are implicated in the development and progression of many chronic diseases.

  7. Dependence of Wilms tumor cells on signaling through insulin-like growth factor 1 in an orthotopic xenograft model targetable by specific receptor inhibition

    DEFF Research Database (Denmark)

    Bielen, Aleksandra; Box, Gary; Perryman, Lara; Bjerke, Lynn; Popov, Sergey; Jamin, Yann; Jury, Alexa; Valenti, Melanie; Brandon, Alexis de Haven; Martins, Vanessa; Romanet, Vincent; Jeay, Sebastien; Raynaud, Florence I; Hofmann, Francesco; Robinson, Simon P; Eccles, Suzanne A; Jones, Chris

    2012-01-01

    We have previously demonstrated an increased DNA copy number and expression of IGF1R to be associated with poor outcome in Wilms tumors. We have now tested whether inhibiting this receptor may be a useful therapeutic strategy by using a panel of Wilms tumor cell lines. Both genetic and...... pharmacological targeting resulted in inhibition of downstream signaling through PI3 and MAP kinases, G(1) cell cycle arrest, and cell death, with drug efficacy dependent on the levels of phosphorylated IGF1R. These effects were further associated with specific gene expression signatures reflecting pathway...... inhibition, and conferred synergistic chemosensitisation to doxorubicin and topotecan. In the in vivo setting, s.c. xenografts of WiT49 cells resembled malignant rhabdoid tumors rather than Wilms tumors. Treatment with an IGF1R inhibitor (NVP-AEW541) showed no discernable antitumor activity and no downstream...

  8. A species-specific activation of Toll-like receptor signaling in bovine and sheep bronchial epithelial cells triggered by Mycobacterial infections.

    Science.gov (United States)

    Ma, Yan; Han, Fei; Liang, Jinping; Yang, Jiali; Shi, Juan; Xue, Jing; Yang, Li; Li, Yong; Luo, Meihui; Wang, Yujiong; Wei, Jun; Liu, Xiaoming

    2016-03-01

    Pulmonary tuberculosis caused by a Mycobacterium infection remains a major public health problem in most part of the world, in part owing to the transmission of its pathogens between hosts including human, domestic and wild animals. To date, molecular mechanisms of the pathogenesis of TB are still incompletely understood. In addition to alveolar macrophages, airway epithelial cells have also been recently recognized as main targets for Mycobacteria infections. In an effort to understand the pathogen-host interaction between Mycobacteria and airway epithelial cells in domestic animals, in present study, we investigated the Toll-like receptor (TLR) signaling in bovine and sheep airway epithelial cells in response to an infection of Mycobacterium tuberculosis avirulent H37Ra stain or Mycobacterium bovis BCG vaccine strain, using primary air-liquid interface (ALI) bronchial epithelial culture models. Our results revealed a host and pathogen species-specific TLR-mediated recognition of pathogen-associated molecular patterns (PAMPs), induction and activation of TLR signaling pathways, and substantial induction of inflammatory response in bronchial epithelial cells in response to Mycobacteria infections between these two species. Interestingly, the activation TLR signaling in bovine bronchial epithelial cells induced by Mycobacteria infection was mainly through a myeloid differentiation factor 88 (MyD88)-independent TLR signaling pathway, while both MyD88-dependent and independent TLR signaling cascades could be induced in sheep epithelial cells. Equally noteworthy, a BCG infection was able to induce both MyD88-dependent and independent signaling in sheep and bovine airway epithelial cells, but more robust inflammatory responses were induced in sheep epithelial cells relative to the bovines; whereas an H37Ra infection displayed an ability to mainly trigger a MyD88-independent TLR signaling cascade in these two host species, and induce a more extent expression of

  9. A selective estrogen receptor modulator inhibits TNF-alpha-induced apoptosis by activating ERK1/2 signaling pathway in vascular endothelial cells.

    Science.gov (United States)

    Yu, Jing; Eto, Masato; Akishita, Masahiro; Okabe, Tetsuro; Ouchi, Yasuyoshi

    2009-07-01

    Tumor necrosis factor (TNF-alpha) is a pleiotropic cytokine exerting both inflammatory and cell death activity and is thought to play a role in the pathogenesis of atherosclerosis. The present study was designed to examine whether the raloxifene analogue, LY117018 could inhibit TNF-alpha-induced apoptosis in vascular endothelial cells and to clarify the involved mechanisms. Apoptosis of endothelial cells was determined by DNA fragmentation assay and the activation of caspase-3. LY117018 significantly inhibited TNF-alpha-induced caspase-3 activation and cell DNA fragmentation levels in bovine carotid artery endothelial cells. The inhibitory effect of LY117018 was abolished by an estrogen receptor antagonist ICI 182,780. p38 MAPK, JNK, ERK1/2 and Akt have been shown to act as apoptotic or anti-apoptotic signals. TNF-alpha stimulated the phosphorylation levels of p38 MAPK, JNK, ERK1/2 and Akt in vascular endothelial cells. TNF-alpha-induced apoptosis was significantly decreased by SB203580, a p38 MAPK inhibitor or SP600125, a JNK inhibitor, but was enhanced by an ERK1/2 pathway inhibitor, PD98059 or a PI3-kinase/Akt pathway inhibitor, wortmannin. The anti-apoptotic effect of LY117018 was abrogated only by PD98059 but was not affected by the inhibitors for p38 MAPK, JNK, or Akt. LY117018 stimulated the further increase in phosphorylation of ERK1/2 in TNF-alpha treated endothelial cells but it did not affect phosphorylation levels of p38 MAPK, JNK or Akt. These results suggest that LY 110718 prevents caspase-3 dependent apoptosis induced by TNF-alpha in vascular endothelial cells through activation of the estrogen receptors and the ERK1/2 signaling pathway. PMID:19275968

  10. IL-27 receptor signalling restricts the formation of pathogenic, terminally differentiated Th1 cells during malaria infection by repressing IL-12 dependent signals.

    Directory of Open Access Journals (Sweden)

    Ana Villegas-Mendez

    Full Text Available The IL-27R, WSX-1, is required to limit IFN-γ production by effector CD4⁺ T cells in a number of different inflammatory conditions but the molecular basis of WSX-1-mediated regulation of Th1 responses in vivo during infection has not been investigated in detail. In this study we demonstrate that WSX-1 signalling suppresses the development of pathogenic, terminally differentiated (KLRG-1⁺ Th1 cells during malaria infection and establishes a restrictive threshold to constrain the emergent Th1 response. Importantly, we show that WSX-1 regulates cell-intrinsic responsiveness to IL-12 and IL-2, but the fate of the effector CD4⁺ T cell pool during malaria infection is controlled primarily through IL-12 dependent signals. Finally, we show that WSX-1 regulates Th1 cell terminal differentiation during malaria infection through IL-10 and Foxp3 independent mechanisms; the kinetics and magnitude of the Th1 response, and the degree of Th1 cell terminal differentiation, were comparable in WT, IL-10R1⁻/⁻ and IL-10⁻/⁻ mice and the numbers and phenotype of Foxp3⁺ cells were largely unaltered in WSX-1⁻/⁻ mice during infection. As expected, depletion of Foxp3⁺ cells did not enhance Th1 cell polarisation or terminal differentiation during malaria infection. Our results significantly expand our understanding of how IL-27 regulates Th1 responses in vivo during inflammatory conditions and establishes WSX-1 as a critical and non-redundant regulator of the emergent Th1 effector response during malaria infection.

  11. Retinoic acid receptor-dependent, cell-autonomous, endogenous retinoic acid signaling and its target genes in mouse collecting duct cells.

    Directory of Open Access Journals (Sweden)

    Yuen Fei Wong

    Full Text Available BACKGROUND: Vitamin A is necessary for kidney development and has also been linked to regulation of solute and water homeostasis and to protection against kidney stone disease, infection, inflammation, and scarring. Most functions of vitamin A are mediated by its main active form, all-trans retinoic acid (tRA, which binds retinoic acid receptors (RARs to modulate gene expression. We and others have recently reported that renal tRA/RAR activity is confined to the ureteric bud (UB and collecting duct (CD cell lineage, suggesting that endogenous tRA/RARs primarily act through regulating gene expression in these cells in embryonic and adult kidney, respectively. METHODOLOGY/PRINCIPAL FINDINGS: To explore target genes of endogenous tRA/RARs, we employed the mIMCD-3 mouse inner medullary CD cell line, which is a model of CD principal cells and exhibits constitutive tRA/RAR activity as CD principal cells do in vivo. Combining antagonism of RARs, inhibition of tRA synthesis, exposure to exogenous tRA, and gene expression profiling techniques, we have identified 125 genes as candidate targets and validated 20 genes that were highly regulated (Dhrs3, Sprr1a, and Ppbp were the top three. Endogenous tRA/RARs were more important in maintaining, rather than suppressing, constitutive gene expression. Although many identified genes were expressed in UBs and/or CDs, their exact functions in this cell lineage are still poorly defined. Nevertheless, gene ontology analysis suggests that these genes are involved in kidney development, renal functioning, and regulation of tRA signaling. CONCLUSIONS/SIGNIFICANCE: A rigorous approach to defining target genes for endogenous tRA/RARs has been established. At the pan-genomic level, genes regulated by endogenous tRA/RARs in a CD cell line have been catalogued for the first time. Such a catalogue will guide further studies on molecular mediators of endogenous tRA/RARs during kidney development and in relation to renal

  12. Activation of Protease-Activated Receptor 2-Mediated Signaling by Mast Cell Tryptase Modulates Cytokine Production in Primary Cultured Astrocytes

    Directory of Open Access Journals (Sweden)

    Xiaoning Zeng

    2013-01-01

    Full Text Available Protease-activated receptor 2 (PAR-2, which is abundantly expressed in astrocytes, is known to play major roles in brain inflammation. However, the influence of the natural agonist of PAR-2, tryptase, on proinflammatory mediator releasedfrom astrocytes remains uninvestigated. In the present study, we found that tryptase at lower concentrations modestly reduced intracellular ROS production but significantly increased IL-6 and TNF-α secretion at higher concentrations without affecting astrocytic viability and proliferation. The actions of tryptase were alleviated by specific PAR-2 antagonist FSLLRY-NH2 (FS, indicating that the actions of tryptase were via PAR-2. PI3K/AKT inhibitor LY294002 reversed the effect of tryptase on IL-6 production, whereas inhibitors specific for p38, JNK, and ERK1/2 abolished the effect of tryptase on TNF-α production, suggesting that different signaling pathways are involved. Moreover, tryptase-induced activation of MAPKs and AKT was eliminated by FS, implicating that PAR-2 is responsible for transmitting tryptase biosignals to MAPKs and AKT. Tryptase provoked also expression of TGF-β and CNTF in astrocytes. The present findings suggest for the first time that tryptase can regulate the release of cytokines from astrocytes via PAR-2-MAPKs or PAR-2-PI3K/AKT signaling pathways, which reveals PAR-2 as a new target actively participating in the regulation of astrocytic functions.

  13. Signal transduction by growth factor receptors: signaling in an instant

    DEFF Research Database (Denmark)

    Dengjel, Joern; Akimov, Vyacheslav; Blagoev, Blagoy; Andersen, Jens S

    2007-01-01

    mass spectrometry-based proteomics has allowed exciting views on the very early events in signal transduction. Activation profiles of regulated phosphorylation sites on epidermal growth factor receptor and downstream signal transducers showed different kinetics within the first ten seconds of...

  14. Molecular mechanisms of glucocorticoid receptor signaling

    Directory of Open Access Journals (Sweden)

    Marta Labeur

    2010-10-01

    Full Text Available This review highlights the most recent findings on the molecular mechanisms of the glucocorticoid receptor (GR. Most effects of glucocorticoids are mediated by the intracellular GR which is present in almost every tissue and controls transcriptional activation via direct and indirect mechanisms. Nevertheless the glucocorticoid responses are tissue -and gene- specific. GR associates selectively with corticosteroid ligands produced in the adrenal gland in response to changes of humoral homeostasis. Ligand interaction with GR promotes either GR binding to genomic glucocorticoid response elements, in turn modulating gene transcription, or interaction of GR monomers with other transcription factors activated by other signalling pathways leading to transrepression. The GR regulates a broad spectrum of physiological functions, including cell differentiation, metabolism and inflammatory responses. Thus, disruption or dysregulation of GR function will result in severe impairments in the maintenance of homeostasis and the control of adaptation to stress.

  15. Prolactin receptor and signal transduction to milk protein genes

    Energy Technology Data Exchange (ETDEWEB)

    Djiane, J.; Daniel, N.; Bignon, C. [Unite d`Endocrinologie Moleculaire, Jouy en Josas (France)] [and others

    1994-06-01

    After cloning of the mammary gland prolactin (PRL) receptor cDNA, a functional assay was established using co-transfection of PRL receptor cDNA together with a milk protein promoter/chloramphenicol acetyl transferase (CAT) construct in Chinese hamster ovary (CHO) cells. Different mutants of the PRL receptor were tested in this CAT assay to delimit the domains in the receptor necessary for signal transduction to milk protein genes. In CHO cells stably transfected with PRL receptor cDNA, high numbers of PRL receptor are expressed. By metabolic labeling and immunoprecipitation, expressed PRL receptor was identified as a single species of 100 kDa. Using these cells, we analyzed the effects of PRL on intracellular free Ca{sup ++} concentration. PRL stimulates Ca{sup ++} entry and induces secondary Ca{sup ++} mobilization. The entry of Ca{sup ++} is a result of an increase in K{sup +} conductance that hyperpolarizes the membranes. We have also analyzed tyrosine phosphorylation induced by PRL. In CHO cells stably transfected with PRL receptor cDNA, PRL induced a very rapid and transient tyrosine phosphorylation of a 100-kDa protein which is most probably the PRL receptor. The same finding was obtained in mammary membranes after PRL injection to lactating rabbits. Whereas tyrosine kinase inhibitors genistein and lavendustin were without effect, PRL stimulation of milk protein gene promoters was partially inhibited by 2 {mu}M herbimycin in CHO cells co-transfected with PRL receptor cDNA and the {Beta} lactoglobulin CAT construct. Taken together these observations indicate that the cytoplasmic domain of the PRL receptor interacts with one or several tyrosine kinases, which may represent early postreceptor events necessary for PRL signal transduction to milk protein genes. 14 refs., 4 figs.

  16. Steroid signaling activation and intracellular localization of sex steroid receptors

    OpenAIRE

    Giraldi, Tiziana; Giovannelli, Pia; Di Donato, Marzia; Castoria, Gabriella; Migliaccio, Antimo; Auricchio, Ferdinando

    2010-01-01

    In addition to stimulating gene transcription, sex steroids trigger rapid, non-genomic responses in the extra-nuclear compartment of target cells. These events take place within seconds or minutes after hormone administration and do not require transcriptional activity of sex steroid receptors. Depending on cell systems, activation of extra-nuclear signaling pathways by sex steroids fosters cell cycle progression, prevents apoptosis, leads to epigenetic modifications and increases cell migrat...

  17. Selective Aryl Hydrocarbon Receptor Modulator 3,3'-Diindolylmethane Impairs AhR and ARNT Signaling and Protects Mouse Neuronal Cells Against Hypoxia.

    Science.gov (United States)

    Rzemieniec, J; Litwa, E; Wnuk, A; Lason, W; Krzeptowski, W; Kajta, M

    2016-10-01

    The neuroprotective potential of 3,3'-diindolylmethane (DIM), which is a selective aryl hydrocarbon receptor modulator, has recently been shown in cellular and animal models of Parkinson's disease and lipopolysaccharide-induced inflammation. However, there are no data concerning the protective capacity and mechanisms of DIM action in neuronal cells exposed to hypoxia. The aim of the present study was to investigate the neuroprotective potential of DIM against the hypoxia-induced damage in mouse hippocampal cells in primary cultures, with a particular focus on DIM interactions with the aryl hydrocarbon receptor (AhR), its nuclear translocator ARNT, and estrogen receptor β (ERβ). In the present study, 18 h of hypoxia induced apoptotic processes, in terms of the mitochondrial membrane potential, activation of caspase-3, and fragmentation of cell nuclei. These effects were accompanied by substantial lactate dehydrogenase release and neuronal cell death. The results of the present study demonstrated strong neuroprotective and anti-apoptotic actions of DIM in hippocampal cells exposed to hypoxia. In addition, DIM decreased the Ahr and Arnt mRNA expression and stimulated Erβ mRNA expression level. DIM-induced mRNA alterations were mirrored by changes in protein levels, except for ERβ, as detected by ELISA, Western blotting, and immunofluorescence labeling. We also demonstrated that DIM decreased the expression of AhR-regulated CYP1A1. Using specific siRNAs, we provided evidence that impairment of AhR and ARNT, but not ERβ plays a key role in the neuroprotective action of DIM against hypoxia-induced cell damage. This study may have implication for identifying new agents that could protect neurons against hypoxia by targeting AhR/ARNT signaling. PMID:26476840

  18. Intermediate- and high-affinity interleukin-2 receptors expressed in an IL-4-dependent T-cell line induce different signals.

    Science.gov (United States)

    Rebollo, A; Silva, A

    1993-01-01

    In order to understand the respective roles of IL-2R alpha and IL-2R beta subunits in transmission of the interleukin-2 (IL-2)-mediated growth signals, we have established two IL-4-dependent murine T-cell clones stably expressing the human IL-2R beta chain and three clones stably expressing the human IL-2R alpha chain. Whereas parental LD8 cells (which express only the murine IL-2R beta chain) do not proliferate in response to IL-2, cell lines stably expressing human IL-2R beta or the chimeric IL-2R alpha beta complex proliferate in response to IL-2. Stably transfected cells expressing the chimeric high-affinity receptor (human IL-2R alpha and murine IL-2R beta) expressed de novo endogenous murine IL-2R alpha when cultured in the presence of IL-2 but not IL-4. Both chimeric and endogenous receptors are functional in response to IL-2, since only addition of both anti-human and anti-murine IL-2R alpha monoclonal antibodies (mAb) inhibited IL-2-induced proliferation. Taken together, our results strongly suggest that human and murine IL-2R beta molecules are different since interaction of IL-2 with human p70 IL-2R is sufficient for transduction of proliferative signals in the absence of p55 IL-2R or, alternatively, that over-expression of the IL-2R beta chain renders cells responsive to IL-2. In addition, IL-2 stimulation of T cells through different forms of IL-2R results in the induction of distinct cellular responses. PMID:8262551

  19. The ETS domain transcription factor ELK1 directs a critical component of growth signaling by the androgen receptor in prostate cancer cells.

    Science.gov (United States)

    Patki, Mugdha; Chari, Venkatesh; Sivakumaran, Suneethi; Gonit, Mesfin; Trumbly, Robert; Ratnam, Manohar

    2013-04-19

    The androgen receptor (AR) is essential for diverse aspects of prostate development and function. Molecular mechanisms by which prostate cancer (PC) cells redirect AR signaling to genes that primarily support growth are unclear. A systematic search for critical AR-tethering proteins led to ELK1, an ETS transcription factor of the ternary complex factor subfamily. Although genetically redundant, ELK1 was obligatory for AR-dependent growth and clonogenic survival in both hormone-dependent PC and castration-recurrent PC cells but not for AR-negative cell growth. AR required ELK1 to up-regulate a major subset of its target genes that was strongly and primarily enriched for cell growth functions. AR functioned as a coactivator of ELK1 by association through its A/B domain, bypassing the classical mechanism of ELK1 activation by phosphorylation and without inducing ternary complex target genes. The ELK1-AR synergy per se was ligand-independent, although it required ligand for nuclear localization of AR as targeting the AR A/B domain to the nucleus recapitulated the action of hormone; accordingly, Casodex was a poor antagonist of the synergy. ELK3, the closest substitute for ELK1 in structure/function and genome recognition, did not interact with AR. ELK1 thus directs selective and sustained gene induction that is a substantial and critical component of growth signaling by AR in PC cells. The ELK1-AR interaction offers a functionally tumor-selective drug target. PMID:23426362

  20. T cell receptor (TCR-transgenic CD8 lymphocytes rendered insensitive to transforming growth factor beta (TGFβ signaling mediate superior tumor regression in an animal model of adoptive cell therapy

    Directory of Open Access Journals (Sweden)

    Quatromoni Jon G

    2012-06-01

    Full Text Available Abstract Tumor antigen-reactive T cells must enter into an immunosuppressive tumor microenvironment, continue to produce cytokine and deliver apoptotic death signals to affect tumor regression. Many tumors produce transforming growth factor beta (TGFβ, which inhibits T cell activation, proliferation and cytotoxicity. In a murine model of adoptive cell therapy, we demonstrate that transgenic Pmel-1 CD8 T cells, rendered insensitive to TGFβ by transduction with a TGFβ dominant negative receptor II (DN, were more effective in mediating regression of established B16 melanoma. Smaller numbers of DN Pmel-1 T cells effectively mediated tumor regression and retained the ability to produce interferon-γ in the tumor microenvironment. These results support efforts to incorporate this DN receptor in clinical trials of adoptive cell therapy for cancer.

  1. Dynamics of Protein Expression Reveals Primary Targets and Secondary Messengers of Estrogen Receptor Alpha Signaling in MCF-7 Breast Cancer Cells.

    Science.gov (United States)

    Drabovich, Andrei P; Pavlou, Maria P; Schiza, Christina; Diamandis, Eleftherios P

    2016-06-01

    Estrogen receptor alpha (ERα)-mediated proliferation of breast cancer cells is facilitated through expression of multiple primary target genes, products of which induce a secondary response to stimulation. To differentiate between the primary and secondary target proteins of ERα signaling, we measured dynamics of protein expression induced by 17β-estradiol in MCF-7 breast cancer cells. Measurement of the global proteomic effects of estradiol by stable isotope labeling by amino acids in cell culture (SILAC) resulted in identification of 103 estrogen-regulated proteins, with only 40 of the corresponding genes having estrogen response elements. Selected reaction monitoring (SRM) assays were used to validate the differential expression of 19 proteins and measure the dynamics of their expression within 72 h after estradiol stimulation, and in the absence or presence of 4-hydroxytamoxifen, to confirm ERα-mediated signaling. Dynamics of protein expression unambiguously revealed early and delayed response proteins and well correlated with presence or absence of estrogen response elements in the corresponding genes. Finally, we quantified dynamics of protein expression in a rarely studied network of transcription factors with a negative feedback loop (ERα-EGR3-NAB2). Because NAB2 protein is a repressor of EGR3-induced transcription, siRNA-mediated silencing of NAB2 resulted in the enhanced expression of the EGR3-induced protein ITGA2. To conclude, we provided a high-quality proteomic resource to supplement genomic and transcriptomic studies of ERα signaling. PMID:27067054

  2. Small molecule receptor tyrosine kinase inhibitor of platelet-derived growth factor signaling (SU9518) modifies radiation response in fibroblasts and endothelial cells

    International Nuclear Information System (INIS)

    Several small receptor tyrosine kinase inhibitors (RTKI) have entered clinical cancer trials alone and in combination with radiotherapy or chemotherapy. The inhibitory spectrum of these compounds is often not restricted to a single target. For example Imatinib/Gleevec (primarily a bcr/abl kinase inhibitor) or SU11248 (mainly a VEGFR inhibitor) are also potent inhibitors of PDGFR and other kinases. We showed previously that PDGF signaling inhibition attenuates radiation-induced lung fibrosis in a mouse model. Here we investigate effects of SU9518, a PDGFR inhibitor combined with ionizing radiation in human primary fibroblasts and endothelial cells in vitro, with a view on utilizing RTKI for antifibrotic therapy. Protein levels of PDGFR-α/-β and phosphorylated PDGFR in fibroblasts were analyzed using western and immunocytochemistry assays. Functional proliferation and clonogenic assays were performed (i) to assess PDGFR-mediated survival and proliferation in fibroblasts and endothelial cells after SU9518 (small molecule inhibitor of PDGF receptor tyrosine kinase); (ii) to test the potency und selectivity of the PDGF RTK inhibitor after stimulation with PDGF isoforms (-AB, -AA, -BB) and VEGF+bFGF. In order to simulate in vivo conditions and to understand the role of radiation-induced paracrine PDGF secretion, co-culture models consisting of fibroblasts and endothelial cells were employed. In fibroblasts, radiation markedly activated PDGF signaling as detected by enhanced PDGFR phosphorylation which was potently inhibited by SU9518. In fibroblast clonogenic assay, SU9518 reduced PDGF stimulated fibroblast survival by 57%. Likewise, SU9518 potently inhibited fibroblast and endothelial cell proliferation. In the co-culture model, radiation of endothelial cells and fibroblast cells substantially stimulated proliferation of non irradiated fibroblasts and vice versa. Importantly, the RTK inhibitor significantly inhibited this paracrine radiation-induced fibroblast and

  3. The interaction of signal transduction pathways in FRTL5 thyroid follicular cells: Studies with stable expression of beta 2-adrenergic receptors

    International Nuclear Information System (INIS)

    Multiple signal transduction pathways interact in FRTL5 cells to promote thyroid follicular cell differentiated function and cell proliferation. In these cells, TSH is a tissue-specific mitogen that promotes DNA synthesis primarily through activation of adenylate cyclase. To further test the role of adenylate cyclase in regulating cell growth and differentiated function we have introduced into FRTL5 the human beta 2-adrenergic receptor (BAR) complementary DNA and have studied the ability of isoproterenol, alone and in combination with insulin-like growth factor I (IGF-I), to stimulate cAMP accumulation, iodide transport, [3H]thymidine incorporation into DNA, and cell growth. Wild-type FRTL5 were infected with a PLJ retroviral construct containing the BAR in either a sense (FRTL BAR) or antisense (FRTL RBAR) orientation, and cell populations were selected on the basis of resistance to the antibiotic geneticin. FRTL BAR expressed approximately 1.3 x 10(5) high affinity binding sites per cell for the beta 2-specific ligand, CGP-12177, while neither FRTL5 wild-type nor RBAR cells demonstrated any specific binding. FRTL BAR had significantly higher levels of intracellular cAMP, [3H]thymidine incorporation, and iodide uptake in the absence of added isoproterenol than FRTL RBAR or wild-type cells. In FRTL BAR, but not RBAR cells, isoproterenol stimulated a dose-dependent accumulation of cAMP, iodide uptake, [3H]thymidine incorporation, and cell growth. FRTL BAR and RBAR cells were equally responsive to TSH and to IGF-I. Isoproterenol enhanced the ability of IGF-I to stimulate [3H]thymidine incorporation in BAR but not RBAR cells. Isoproterenol partially inhibited the ability of TSH to stimulate cAMP generation and DNA synthesis

  4. Induction of Proinflammatory Responses in Macrophages by the Glycosylphosphatidylinositols (GPIs) of Plasmodium falciparum: CELL SIGNALING RECEPTORS, GPI STRUCTURAL REQUIREMENT, AND REGULATION OF GPI ACTIVITY*

    Science.gov (United States)

    Krishnegowda, Gowdahalli; Hajjar, Adeline M.; Zhu, Jianzhong; Douglass, Erika J.; Uematsu, Satoshi; Akira, Shizuo; Woods, Amina S.; Gowda, D. Channe

    2016-01-01

    SUMMARY The proinflammatory cytokines produced by the innate immune system in response to pathogenic infection protect the host by controlling microbial growth. However, excessive proinflammatory responses could disrupt the host’s vital physiological functions, causing severe pathological conditions. In the case of Plasmodium falciparum, the protozoan parasite that causes fatal malaria in man, the glycosylphosphatidylinositol (GPI) anchors are thought to be the major factors that contribute to malaria pathogenesis through their ability to induce proinflammatory responses. In this study, we identified the receptors for P. falciparum GPI-induced cell signaling that leads to proinflammatory responses, and studied the GPI structure-activity relationship. The data show that GPI-signaling is mediated mainly through recognition by TLR2 and to a lesser extent by TLR4. The activity of sn-2 lyso GPIs is comparable to that of the intact GPIs, whereas the activity of Man3-GPIs is about 80% that of the intact GPIs. The GPIs with three (intact GPIs and Man3-GPIs) and two fatty acids (sn-2 lyso GPIs) appear to differ considerably in the requirement of the auxiliary receptor, TLR1 or TLR6, for recognition by TLR2. The former are preferentially recognized by TLR2/TLR1, whereas the latter are favored by TLR2/TLR6. However, the signaling pathways initiated by all three GPI types are similar, involving the MyD88-dependent activation of ERK, JNK and p38, and NF-κB signaling pathways. The signaling molecules of these pathways differentially contribute to the production of various cytokines and nitric oxide (Zhu, J., et al. (2004) J. Biol. Chem., accompanying manuscript). Our data also show that GPIs are degraded by the macrophage surface phospholipases, predominantly into inactive species, indicating that the host can regulate GPI activity, at least in part, by this mechanism. These results imply that macrophage surface phospholipases play important roles in the GPI-induced innate

  5. Retinoic acid receptor agonist Am80 inhibits CXCL2 production from microglial BV-2 cells via attenuation of NF-κB signaling.

    Science.gov (United States)

    Takaoka, Yuichiro; Takahashi, Moeka; Kurauchi, Yuki; Hisatsune, Akinori; Seki, Takahiro; Shudo, Koichi; Katsuki, Hiroshi

    2016-09-01

    Accumulating lines of evidence suggest that retinoic acid receptor agonists such as Am80 exerts anti-inflammatory actions in the central nervous system, although detailed mechanisms of the action remain largely unknown. Our previous findings suggest that Am80 provides therapeutic effect on intracerebral hemorrhage in mice via suppression of expression of chemokine (C-X-C motif) ligand 2 (CXCL2). Here we investigated the mechanisms of inhibitory action of Am80 on expression of CXCL2 and other pro-inflammatory factors in microglial BV-2 cells. Pretreatment with Am80 markedly suppressed lipopolysaccharide (LPS)-induced expression of CXCL2 mRNA and release of CXCL2 protein. Am80 had no effect on LPS-induced activation of p38 mitogen-activated protein kinase and extracellular signal-regulated kinase. On the other hand, Am80 prevented LPS-induced nuclear translocation of p65 subunit of NF-κB complex. In addition, total expression levels of p65 and IκBα proteins, as well as of mRNAs encoding p65 and IκBα, were lowered by Am80. Dependence of CXCL2 expression on NF-κB was confirmed by the effect of an NF-κB inhibitor caffeic acid phenethyl ester that abolished LPS-induced CXCL2 expression. Caffeic acid phenethyl ester also abolished LPS-induced expression of inducible nitric oxide synthase, interleukin-1β and tumor necrosis factor α, which may be relevant to the inhibitory effect of Am80 on expression of these pro-inflammatory factors. We additionally found that Am80 attenuated LPS-induced up-regulation of CD14, a co-receptor for Toll-like receptor 4 (TLR4). These results suggest that inhibitory effect on TLR4 signaling mediated by NF-κB pathway underlies the anti-inflammatory action of retinoic acid receptor agonists in microglia. PMID:27351827

  6. Receptors and signalling mechanisms in the procoagulant response of platelets.

    Science.gov (United States)

    Heemskerk, J W; Siljander, P R; Bevers, E M; Farndale, R W; Lindhout, T

    2000-09-01

    Platelets in an advanced stage of activation change from coagulation-inactive to coagulation-promoting cells. This procoagulant response is characterised by exposure of aminophospholipids, such as phosphatidylserine, to the platelet surface and by formation of microvesicles. Under specific conditions, when both signalling and adhesive platelet receptors are occupied, collagen and also thrombin are able to trigger this response. Thus, platelets express high coagulation-promoting activity only after interacting with multiple receptors. PMID:11083453

  7. The phosphatidylserine receptor TIM-4 does not mediate direct signaling.

    Science.gov (United States)

    Park, Daeho; Hochreiter-Hufford, Amelia; Ravichandran, Kodi S

    2009-02-24

    Engulfment of apoptotic cells is an active process coordinated by receptors on phagocytes and ligands on apoptotic cells [1]. Phosphatidylserine (PtdSer) is a key ligand on apoptotic cells, and recently three PtdSer recognition receptors have been identified, namely, TIM-4, BAI1, and Stabilin-2 [1-6]. Whereas BAI1 is dependent on the ELMO1/Dock180/Rac signaling module, and Stablilin-2 appears to use the intracellular adaptor GULP [2, 3, 7], little is known about how TIM-4 transduces signals downstream of PtdSer recognition [8]. To test the role of known engulfment signaling pathways in TIM-4-mediated engulfment, we used a combination of dominant-negative mutants, knockdown of specific signaling proteins, and knockout cell lines. TIM-4 appears to be largely independent of the two known engulfment signaling pathways [7, 9-17], yet the TIM-4-mediated uptake is inhibited by cytoskeleton disrupting drugs. Remarkably, a version of TIM-4 lacking its cytoplasmic tail promoted corpse uptake via PtdSer recognition. Moreover, replacement of the transmembrane region of TIM-4 with a glycophosphatidylinositol anchor still promoted engulfment comparable to wild-type TIM-4. Thus, the transmembrane region and cytoplasmic tail of TIM-4 are dispensable for apoptotic cell engulfment, and we propose that TIM-4 is a PtdSer tethering receptor without any direct signaling of its own. PMID:19217291

  8. BRI1-Associated Receptor Kinase 1 Regulates Guard Cell ABA Signaling Mediated by Open Stomata 1 in Arabidopsis.

    Science.gov (United States)

    Shang, Yun; Dai, Changbo; Lee, Myeong Min; Kwak, June M; Nam, Kyoung Hee

    2016-03-01

    Stomatal movements are critical in regulating gas exchange for photosynthesis and water balance between plant tissues and the atmosphere. The plant hormone abscisic acid (ABA) plays key roles in regulating stomatal closure under various abiotic stresses. In this study, we revealed a novel role of BAK1 in guard cell ABA signaling. We found that the brassinosteroid (BR) signaling mutant bak1 lost more water than wild-type plants and showed ABA insensitivity in stomatal closure. ABA-induced OST1 expression and reactive oxygen species (ROS) production were also impaired in bak1. Unlike direct treatment with H2O2, overexpression of OST1 did not completely rescue the insensitivity of bak1 to ABA. We demonstrated that BAK1 forms a complex with OST1 near the plasma membrane and that the BAK1/OST1 complex is increased in response to ABA in planta. Brassinolide, the most active BR, exerted a negative effect on ABA-induced formation of the BAK1/OST1 complex and OST1 expression. Moreover, we found that BAK1 and ABI1 oppositely regulate OST1 phosphorylation in vitro, and that ABI1 interacts with BAK1 and inhibits the interaction of BAK1 and OST1. Taken together, our results suggest that BAK1 regulates ABA-induced stomatal closure in guard cells. PMID:26724418

  9. Insulin-like growth factor 1 receptor and p38 mitogen-activated protein kinase signals inversely regulate signal transducer and activator of transcription 3 activity to control human dental pulp stem cell quiescence, propagation, and differentiation.

    Science.gov (United States)

    Vandomme, Jerome; Touil, Yasmine; Ostyn, Pauline; Olejnik, Cecile; Flamenco, Pilar; El Machhour, Raja; Segard, Pascaline; Masselot, Bernadette; Bailliez, Yves; Formstecher, Pierre; Polakowska, Renata

    2014-04-15

    Dental pulp stem cells (DPSCs) remain quiescent until activated in response to severe dental pulp damage. Once activated, they exit quiescence and enter regenerative odontogenesis, producing reparative dentin. The factors and signaling molecules that control the quiescence/activation and commitment to differentiation of human DPSCs are not known. In this study, we determined that the inhibition of insulin-like growth factor 1 receptor (IGF-1R) and p38 mitogen-activated protein kinase (p38 MAPK) signaling commonly activates DPSCs and promotes their exit from the G0 phase of the cell cycle as well as from the pyronin Y(low) stem cell compartment. The inhibition of these two pathways, however, inversely determines DPSC fate. In contrast to p38 MAPK inhibitors, IGF-1R inhibitors enhance dental pulp cell sphere-forming capacity and reduce the cells' colony-forming capacity without inducing cell death. The inverse cellular changes initiated by IGF-1R and p38 MAPK inhibitors were accompanied by inverse changes in the levels of active signal transducer and activator of transcription 3 (STAT3) factor, inactive glycogen synthase kinase 3, and matrix extracellular phosphoglycoprotein, a marker of early odontoblast differentiation. Our data suggest that there is cross talk between the IGF-1R and p38 MAPK signaling pathways in DPSCs and that the signals provided by these pathways converge at STAT3 and inversely regulate its activity to maintain quiescence or to promote self-renewal and differentiation of the cells. We propose a working model that explains the possible interactions between IGF-1R and p38 MAPK at the molecular level and describes the cellular consequences of these interactions. This model may inspire further fundamental study and stimulate research on the clinical applications of DPSC in cellular therapy and tissue regeneration. PMID:24266654

  10. Insulin-Like Growth Factor 1 Receptor and p38 Mitogen-Activated Protein Kinase Signals Inversely Regulate Signal Transducer and Activator of Transcription 3 Activity to Control Human Dental Pulp Stem Cell Quiescence, Propagation, and Differentiation

    Science.gov (United States)

    Vandomme, Jerome; Touil, Yasmine; Ostyn, Pauline; Olejnik, Cecile; Flamenco, Pilar; El Machhour, Raja; Segard, Pascaline; Masselot, Bernadette; Bailliez, Yves; Formstecher, Pierre

    2014-01-01

    Dental pulp stem cells (DPSCs) remain quiescent until activated in response to severe dental pulp damage. Once activated, they exit quiescence and enter regenerative odontogenesis, producing reparative dentin. The factors and signaling molecules that control the quiescence/activation and commitment to differentiation of human DPSCs are not known. In this study, we determined that the inhibition of insulin-like growth factor 1 receptor (IGF-1R) and p38 mitogen-activated protein kinase (p38 MAPK) signaling commonly activates DPSCs and promotes their exit from the G0 phase of the cell cycle as well as from the pyronin Ylow stem cell compartment. The inhibition of these two pathways, however, inversely determines DPSC fate. In contrast to p38 MAPK inhibitors, IGF-1R inhibitors enhance dental pulp cell sphere-forming capacity and reduce the cells' colony-forming capacity without inducing cell death. The inverse cellular changes initiated by IGF-1R and p38 MAPK inhibitors were accompanied by inverse changes in the levels of active signal transducer and activator of transcription 3 (STAT3) factor, inactive glycogen synthase kinase 3, and matrix extracellular phosphoglycoprotein, a marker of early odontoblast differentiation. Our data suggest that there is cross talk between the IGF-1R and p38 MAPK signaling pathways in DPSCs and that the signals provided by these pathways converge at STAT3 and inversely regulate its activity to maintain quiescence or to promote self-renewal and differentiation of the cells. We propose a working model that explains the possible interactions between IGF-1R and p38 MAPK at the molecular level and describes the cellular consequences of these interactions. This model may inspire further fundamental study and stimulate research on the clinical applications of DPSC in cellular therapy and tissue regeneration. PMID:24266654

  11. Reassessing ecdysteroidogenic cells from the cell membrane receptors' perspective.

    Science.gov (United States)

    Alexandratos, Alexandros; Moulos, Panagiotis; Nellas, Ioannis; Mavridis, Konstantinos; Dedos, Skarlatos G

    2016-01-01

    Ecdysteroids secreted by the prothoracic gland (PG) cells of insects control the developmental timing of their immature life stages. These cells have been historically considered as carrying out a single function in insects, namely the biochemical conversion of cholesterol to ecdysteroids and their secretion. A growing body of evidence shows that PG cells receive multiple cues during insect development so we tested the hypothesis that they carry out more than just one function in insects. We characterised the molecular nature and developmental profiles of cell membrane receptors in PG cells of Bombyx mori during the final larval stage and determined what receptors decode nutritional, developmental and physiological signals. Through iterative approaches we identified a complex repertoire of cell membrane receptors that are expressed in intricate patterns and activate previously unidentified signal transduction cascades in PG cells. The expression patterns of some of these receptors explain precisely the mechanisms that are known to control ecdysteroidogenesis. However, the presence of receptors for the notch, hedgehog and wingless signalling pathways and the expression of innate immunity-related receptors such as phagocytosis receptors, receptors for microbial ligands and Toll-like receptors call for a re-evaluation of the role these cells play in insects. PMID:26847502

  12. Receptor downregulation and desensitization enhance the information processing ability of signaling receptors

    Energy Technology Data Exchange (ETDEWEB)

    Shankaran, Harish; Wiley, H. S.; Resat, Haluk

    2007-11-09

    The activation of cell surface receptors in addition to initiating signaling events also triggers regulatory processes that restrict the duration of signaling. Acute attenuation of signaling can be accomplished either via ligand-induced internalization of receptors (receptor downregulation) or via ligand-induced receptor desensitization. These phenomena have traditionally been viewed in the context of “adaptation” wherein the receptor system enters a refractory state in the presence of sustained ligand stimuli and thereby prevents the cell from “over-responding” to the ligand. Here we use the epidermal growth factor receptor (EGFR) and G-protein coupled receptors (GPCR) as model systems to respectively examine the effects of downregulation and desensitization on the ability of signaling receptors to decode time-varying ligand stimuli. We show that downregulation and desensitization mechanisms can lead to tight and efficient input-output coupling thereby ensuring synchronous processing of ligand inputs. Frequency response analysis indicates that upstream elements of the EGFR and GPCR networks behave like low-pass filters. Receptor downregulation and desensitization increase the filter bandwidth thereby enabling the receptor systems to decode inputs in a wider frequency range. Further, system-theoretic analysis reveals that the receptor systems are analogous to classical mechanical over-damped oscillators. This analogy enables us to describe downregulation and desensitization as phenomena that make the systems more resilient in responding to ligand perturbations thereby improving the stability of the system resting state. We hypothesize that, in addition to serving as mechanisms for adaptation, receptor downregulation and desensitization play a critical role in temporal information processing.

  13. Luteolin decreases IGF-II production and downregulates insulin-like growth factor-I receptor signaling in HT-29 human colon cancer cells

    Directory of Open Access Journals (Sweden)

    Lim Do

    2012-01-01

    Full Text Available Abstract Background Luteolin is a 3',4',5,7-tetrahydroxyflavone found in various fruits and vegetables. We have shown previously that luteolin reduces HT-29 cell growth by inducing apoptosis and cell cycle arrest. The objective of this study was to examine whether luteolin downregulates the insulin-like growth factor-I receptor (IGF-IR signaling pathway in HT-29 cells. Methods In order to assess the effects of luteolin and/or IGF-I on the IGF-IR signaling pathway, cells were cultured with or without 60 μmol/L luteolin and/or 10 nmol/L IGF-I. Cell proliferation, DNA synthesis, and IGF-IR mRNA levels were evaluated by a cell viability assay, [3H]thymidine incorporation assays, and real-time polymerase chain reaction, respectively. Western blot analyses, immunoprecipitation, and in vitro kinase assays were conducted to evaluate the secretion of IGF-II, the protein expression and activation of IGF-IR, and the association of the p85 subunit of phophatidylinositol-3 kinase (PI3K with IGF-IR, the phosphorylation of Akt and extracellular signal-regulated kinase (ERK1/2, and cell division cycle 25c (CDC25c, and PI3K activity. Results Luteolin (0 - 60 μmol/L dose-dependently reduced the IGF-II secretion of HT-29 cells. IGF-I stimulated HT-29 cell growth but did not abrogate luteolin-induced growth inhibition. Luteolin reduced the levels of the IGF-IR precursor protein and IGF-IR transcripts. Luteolin reduced the IGF-I-induced tyrosine phosphorylation of IGF-IR and the association of p85 with IGF-IR. Additionally, luteolin inhibited the activity of PI3K activity as well as the phosphorylation of Akt, ERK1/2, and CDC25c in the presence and absence of IGF-I stimulation. Conclusions The present results demonstrate that luteolin downregulates the activation of the PI3K/Akt and ERK1/2 pathways via a reduction in IGF-IR signaling in HT-29 cells; this may be one of the mechanisms responsible for the observed luteolin-induced apoptosis and cell cycle arrest.

  14. Absence of Toll-Like Receptor 4 Signaling Results in Delayed Yersinia enterocolitica YopP-Induced Cell Death of Dendritic Cells

    Czech Academy of Sciences Publication Activity Database

    Gröbner, S.; Schulz, S.; Adkins, Irena; Gunst, D. S. J.; Waibel, M.; Wesselborg, S.; Borgmann, S.; Autenrieth, I. B.

    2007-01-01

    Roč. 75, č. 1 (2007), s. 512-517. ISSN 0019-9567 Institutional research plan: CEZ:AV0Z50200510 Keywords : yersinia enterocolitica * toll-like receptor 4 * dendritic cells Subject RIV: EC - Immunology Impact factor: 3.996, year: 2007

  15. The Selective Estrogen Receptor Modulator Raloxifene Regulates Arginine-Vasopressin Gene Expression in Human Female Neuroblastoma Cells Through G Protein-Coupled Estrogen Receptor and ERK Signaling.

    Science.gov (United States)

    Grassi, Daniela; Ghorbanpoor, Samar; Acaz-Fonseca, Estefania; Ruiz-Palmero, Isabel; Garcia-Segura, Luis M

    2015-10-01

    The selective estrogen receptor modulator raloxifene reduces blood pressure in hypertensive postmenopausal women. In the present study we have explored whether raloxifene regulates gene expression of arginine vasopressin (AVP), which is involved in the pathogenesis of hypertension. The effect of raloxifene was assessed in human female SH-SY5Y neuroblastoma cells, which have been recently identified as a suitable cellular model to study the estrogenic regulation of AVP. Raloxifene, within a concentration ranging from 10(-10) M to 10(-6) M, decreased the mRNA levels of AVP in SH-SY5Y cells with maximal effect at 10(-7) M. This effect of raloxifene was imitated by an agonist (±)-1-[(3aR*,4S*,9bS*)-4-(6-bromo-1,3-benzodioxol-5-yl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinolin-8-yl]-ethanone of G protein-coupled estrogen receptor-1 (GPER) and blocked by an antagonist (3aS*,4R*,9bR*)-4-(6-bromo-1,3-benzodioxol-5-yl)-3a,4,5,9b-3H-cyclopenta[c]quinoline of GPER and by GPER silencing. Raloxifene induced a time-dependent increase in the level of phosphorylated ERK1 and ERK2, by a mechanism blocked by the GPER antagonist. The treatment of SH-SY5Y cells with either a MAPK/ERK kinase 1/2-specific inhibitor (1,4-diamino-2, 3-dicyano-1,4-bis(2-aminophenylthio)butadine) or a protein kinase C inhibitor (sotrastaurin) blocked the effects of raloxifene on the phosphorylation of ERK1/2 and the regulation of AVP mRNA levels. These results reveal a mechanism mediating the regulation of AVP expression by raloxifene, involving the activation of GPER, which in turn activates protein kinase C, MAPK/ERK kinase, and ERK. The regulation of AVP by raloxifene and GPER may have implications for the treatment of blood hypertension(.). PMID:26200092

  16. Synthetic high-density lipoprotein-like nanoparticles potently inhibit cell signaling and production of inflammatory mediators induced by lipopolysaccharide binding Toll-like receptor 4.

    Science.gov (United States)

    Foit, Linda; Thaxton, C Shad

    2016-09-01

    Toll-like receptor 4 (TLR4) plays a critical role in the innate immune system. Stimulation of TLR4 occurs upon binding lipopolysaccharide (LPS), a component of Gram-negative bacterial cell walls. Due to the potency of the induced inflammatory response, there is a growing interest in agents that can most proximally modulate this LPS/TLR4 interaction to prevent downstream cell signaling events and the production of inflammatory mediators. Building on the natural ability of human high-density lipoprotein (HDL) to bind LPS, we synthesized a suite of HDL-like nanoparticles (HDL-like NP). We identified one HDL-like NP that was particularly effective at decreasing TLR4 signaling caused by addition of purified LPS or Gram-negative bacteria to model human cell lines or primary human peripheral blood cells. The HDL-like NP functioned to inhibit TLR4-dependent inflammatory response to LPS derived from multiple bacterial species. Mechanistically, data show that the NP mainly functions by scavenging and neutralizing the LPS toxin. Taken together, HDL-like NPs constitute a powerful endotoxin scavenger with the potential to significantly reduce LPS-mediated inflammation. PMID:27244690

  17. Expression of GABAergic receptors in mouse taste receptor cells.

    Directory of Open Access Journals (Sweden)

    Margaret R Starostik

    Full Text Available BACKGROUND: Multiple excitatory neurotransmitters have been identified in the mammalian taste transduction, with few studies focused on inhibitory neurotransmitters. Since the synthetic enzyme glutamate decarboxylase (GAD for gamma-aminobutyric acid (GABA is expressed in a subset of mouse taste cells, we hypothesized that other components of the GABA signaling pathway are likely expressed in this system. GABA signaling is initiated by the activation of either ionotropic receptors (GABA(A and GABA(C or metabotropic receptors (GABA(B while it is terminated by the re-uptake of GABA through transporters (GATs. METHODOLOGY/PRINCIPAL FINDINGS: Using reverse transcriptase-PCR (RT-PCR analysis, we investigated the expression of different GABA signaling molecules in the mouse taste system. Taste receptor cells (TRCs in the circumvallate papillae express multiple subunits of the GABA(A and GABA(B receptors as well as multiple GATs. Immunocytochemical analyses examined the distribution of the GABA machinery in the circumvallate papillae. Both GABA(A-and GABA(B- immunoreactivity were detected in the peripheral taste receptor cells. We also used transgenic mice that express green fluorescent protein (GFP in either the Type II taste cells, which can respond to bitter, sweet or umami taste stimuli, or in the Type III GAD67 expressing taste cells. Thus, we were able to identify that GABAergic receptors are expressed in some Type II and Type III taste cells. Mouse GAT4 labeling was concentrated in the cells surrounding the taste buds with a few positively labeled TRCs at the margins of the taste buds. CONCLUSIONS/SIGNIFICANCE: The presence of GABAergic receptors localized on Type II and Type III taste cells suggests that GABA is likely modulating evoked taste responses in the mouse taste bud.

  18. The miR9863 family regulates distinct Mla alleles in barley to attenuate NLR receptor-triggered disease resistance and cell-death signaling.

    Directory of Open Access Journals (Sweden)

    Jie Liu

    2014-12-01

    Full Text Available Barley (Hordeum vulgare L. Mla alleles encode coiled-coil (CC, nucleotide binding, leucine-rich repeat (NB-LRR receptors that trigger isolate-specific immune responses against the powdery mildew fungus, Blumeria graminis f. sp. hordei (Bgh. How Mla or NB-LRR genes in grass species are regulated at post-transcriptional level is not clear. The microRNA family, miR9863, comprises four members that differentially regulate distinct Mla alleles in barley. We show that miR9863 members guide the cleavage of Mla1 transcripts in barley, and block or reduce the accumulation of MLA1 protein in the heterologous Nicotiana benthamiana expression system. Regulation specificity is determined by variation in a unique single-nucleotide-polymorphism (SNP in mature miR9863 family members and two SNPs in the Mla miR9863-binding site that separates these alleles into three groups. Further, we demonstrate that 22-nt miR9863s trigger the biogenesis of 21-nt phased siRNAs (phasiRNAs and together these sRNAs form a feed-forward regulation network for repressing the expression of group I Mla alleles. Overexpression of miR9863 members specifically attenuates MLA1, but not MLA10-triggered disease resistance and cell-death signaling. We propose a key role of the miR9863 family in dampening immune response signaling triggered by a group of MLA immune receptors in barley.

  19. Inhibition of insulin-like growth factor-1 receptor signaling enhances growth-inhibitory and proapoptotic effects of gefitinib (Iressa) in human breast cancer cells

    International Nuclear Information System (INIS)

    Gefitinib (Iressa, ZD 1839, AstraZeneca) blocks the tyrosine kinase activity of the epidermal growth factor receptor (EGFR) and inhibits proliferation of several human cancer cell types including breast cancer. Phase II clinical trials with gefitinib monotherapy showed an objective response of 9 to 19% in non-small-cell lung cancer patients and less than 10% for breast cancer, and phase III results have indicated no benefit of gefitinib in combination with chemotherapy over chemotherapy alone. In order to improve the antineoplastic activity of gefitinib, we investigated the effects of blocking the signalling of the insulin-like growth factor 1 receptor (IGF-1R), a tyrosine kinase with a crucial role in malignancy that is coexpressed with EGFR in most human primary breast carcinomas. AG1024 (an inhibitor of IGF-1R) was used with gefitinib for treatment of MDA468, MDA231, SK-BR-3, and MCF-7 breast cancer lines, which express similar levels of IGF-1R but varying levels of EGFR. Proliferation assays, apoptosis induction studies, and Western blot analyses were conducted with cells treated with AG1024 and gefitinib as single agents and in combination. Gefitinib and AG1024 reduced proliferation in all lines when used as single agents, and when used in combination revealed an additive-to-synergistic effect on cell growth inhibition. Flow cytometry measurements of cells stained with annexin V-propidium iodide and cells stained for caspase-3 activation indicated that adding an IGF-1R-targeting strategy to gefitinib results in higher levels of apoptosis than are achieved with gefitinib alone. Gefitinib either reduced or completely inhibited p42/p44 Erk kinase phosphorylation, depending on the cell line, while Akt phosphorylation was reduced by a combination of the two agents. Overexpression of IGF-1R in SK-BR-3 cells was sufficient to cause a marked enhancement in gefitinib resistance. These results indicate that IGF-1R signaling reduces the antiproliferative effects of

  20. Modulation of the tyrosine kinase receptor Ret/glial cell-derived neurotrophic factor (GDNF) signaling: a new player in reproduction induced anterior pituitary plasticity?

    Science.gov (United States)

    Guillou, Anne; Romanò, Nicola; Bonnefont, Xavier; Le Tissier, Paul; Mollard, Patrice; Martin, Agnès O

    2011-02-01

    During gestation, parturition, and lactation, the endocrine axis of the dam must continually adapt to ensure the continual and healthy development of offspring. The anterior pituitary gland, which serves as the endocrine interface between the brain and periphery, undergoes adaptations that contribute to regulation of the reproductive axis. Growth factors and their receptors are potential candidates for intrapituitary and paracrine factors to participate in the functional and anatomical plasticity of the gland. We examined the involvement of the growth factor glial cell-derived neurotrophic factor (GDNF) and its receptor tyrosine kinase rearranged during transfection (Ret) in the physiological functional and anatomical plasticity of the anterior pituitary gland. We found that variations in both expression and subcellular localization of Ret during gestation and lactation are temporally correlated with changes in pituitary gland function. We showed that Ret/GDNF signaling could endorse two different functional roles depending on the physiological status. At the end of lactation and after weaning, Ret was colocalized with markers of apoptosis. We found that Ret could therefore act as a physiological dependence receptor capable of inducing apoptosis in the absence of GDNF. In addition, we identified the follicullostellate cell as a probable source for intrapituitary GDNF and proposed GDNF as a potential physiological modulator of endocrine cell function. During all stages studied, we showed that acute application of GDNF to pituitary slices was able to modulate both positively and negatively intracellular calcium activity. Altogether our results implicate Ret/GDNF as a potent pleiotropic factor able to influence pituitary physiology during a period of high plasticity. PMID:21239429

  1. Wnt signaling and stem cell control

    Institute of Scientific and Technical Information of China (English)

    Roel Nusse

    2008-01-01

    Wnt signaling has been implicated in the control over various types of stem cells and may act as a niche factor to maintain stem cells in a self-renewing state.As currently understood,Wnt proteins bind to receptors of the Frizzled and LRP families on the cell surface.Through several cytoplasmic relay components,the signal is transduced to B-catenin,which then enters the nucleus and forms a complex with TCF to activate transcription of Wnt target genes.Wnts can also signal through tyrosine kinase receptors,in particular the ROR and RYK receptors,leading to alternative modes of Wnt signaling.During the growth of tissues,these ligands and receptors are dynamically expressed,often transcriptionally controlled by Wnt signals themselves,to ensure the right balance between proliferation and differentiation.Isolated Wnt proteins are active on a variety of stem cells,including neural,mammary and embryonic stem cells.In general,Wnt proteins act to maintain the undifferentiated state of stem cells,while other growth factors instruct the cells to proliferate.These other factors include FGF and EGF,signaling through tyrosine kinase pathways.

  2. CD40-mediated tumor necrosis factor receptor-associated factor 3 signaling upregulates IL-4-induced germline Cepsilon transcription in a human B cell line.

    Science.gov (United States)

    Basaki, Yuji; Ikizawa, Koichi; Kajiwara, Keiichi; Yanagihara, Yukiyoshi

    2002-09-15

    Induction of germline C epsilon transcription in B cells by IL-4, which is a critical initiating step for IgE class switching, is enhanced by CD40 engagement. Although signaling by CD40 is initiated by the binding of tumor necrosis factor receptor-associated factor (TRAF) family members to its cytoplasmic domain, whether those TRAF family proteins mediate enhancement of germline Cepsilon transcription is not evident. We report here that CD40-induced TRAF3-dependent activation of mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase 1 (MEK1) is involved in the upregulation of IL-4-driven germline C epsilon transcription in a human Burkitt's lymphoma B cell line, DG75. Among the six known TRAF proteins, TRAF2, 3, 5, and 6 associated with CD40 in an unstimulated state, and the levels of these four proteins were unaffected by anti-CD40 stimulation. Antisense oligodeoxynucleotide (ODN) for TRAF3 inhibited CD40-induced activation of MEK1-ERK pathway by decreasing expression of TRAF3 protein, but antisense ODNs for TRAF2, 5, and 6 were ineffective. Furthermore, CD40-mediated enhancement of IL-4-driven germline C epsilon transcription was inhibited by antisense ODN for TRAF3 and by a MEK1 inhibitor, PD98059. These results suggest that in DG75 cells, TRAF3-induced MEK1 activation may be involved in CD40-mediated upregulation of IL-4-driven germline C epsilon transcription. PMID:12220533

  3. Therapeutic intervention of proanthocyanidins on the migration capacity of melanoma cells is mediated through PGE2 receptors and β-catenin signaling molecules.

    Science.gov (United States)

    Vaid, Mudit; Singh, Tripti; Prasad, Ram; Kappes, John C; Katiyar, Santosh K

    2015-01-01

    Melanoma is a highly aggressive form of skin cancer and a leading cause of death from skin diseases mainly due to its propensity to metastasis. Due to metastatic tendency, melanoma is often associated with activation of Wnt/β-catenin signaling mechanism. Blocking β-catenin activation may be a good strategy to block melanoma-associated mortality. We have shown earlier that grape seed proanthocyanidins (GSPs) inhibit melanoma cell migration via targeting cyclooxygenase-2 (COX-2) overexpression. Here we explored further whether inhibition of inflammatory mediators-mediated activation of β-catenin by GSPs is associated with the inhibition of melanoma cell migration. Our study revealed that PGE2 receptors (EP2 and EP4) agonists promote melanoma cell migration while PGE2 receptor antagonist suppressed the migration capacity of melanoma cells. GSPs treatment inhibit butaprost (EP2 agonist) or Cay10580 (EP4 agonist) induced migration of melanoma cells. Western blot analysis revealed that GSPs reduced cellular accumulation of β-catenin, and decreased the expressions of matrix metalloproteinase (MMP)-2, MMP-9 and MITF, downstream targets of β-catenin in melanoma cells. GSPs also reduced the protein expressions of PI3K and p-Akt in the same set of experiment. To verify that β-catenin is a specific molecular target of GSPs, we compared the effect of GSPs on cell migration of β-catenin-activated (Mel1241) and β-catenin-inactivated (Mel1011) melanoma cells. GSPs inhibit cell migration of Mel1241 cells but not of Mel1011 cells. Additionally, in vivo bioluminescence imaging data indicate that dietary administration of GSPs (0.5%, w/w) in supplementation with AIN76A control diet inhibited the migration/extravasation of intravenously injected melanoma cells in lungs of immune-compromised nude mice, and that this effect of GSPs was associated with an inhibitory effect on the activation of β-catenin and its downstream targets, such as MMPs, in lungs as a target organ. PMID

  4. Diverse FGF receptor signaling controls astrocyte specification and proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Kyungjun [School of Life Sciences, Gwangju Institute of Science and Technology, Oryong-dong, Buk-gu, Gwangju 500-712 (Korea, Republic of); Song, Mi-Ryoung, E-mail: msong@gist.ac.kr [School of Life Sciences, Gwangju Institute of Science and Technology, Oryong-dong, Buk-gu, Gwangju 500-712 (Korea, Republic of); Bioimaging Research Center and Cell Dynamics Research Center, Gwangju Institute of Science and Technology, Oryong-dong, Buk-gu, Gwangju 500-712 (Korea, Republic of)

    2010-05-07

    During CNS development, pluripotency neuronal progenitor cells give rise in succession to neurons and glia. Fibroblast growth factor-2 (FGF-2), a major signal that maintains neural progenitors in the undifferentiated state, is also thought to influence the transition from neurogenesis to gliogenesis. Here we present evidence that FGF receptors and underlying signaling pathways transmit the FGF-2 signals that regulate astrocyte specification aside from its mitogenic activity. Application of FGF-2 to cortical progenitors suppressed neurogenesis whereas treatment with an FGFR antagonist in vitro promoted neurogenesis. Introduction of chimeric FGFRs with mutated tyrosine residues into cortical progenitors and drug treatments to specifically block individual downstream signaling pathways revealed that the overall activity of FGFR rather than individual autophosphorylation sites is important for delivering signals for glial specification. In contrast, a signal for cell proliferation by FGFR was mainly delivered by MAPK pathway. Together our findings indicate that FGFR activity promotes astrocyte specification in the developing CNS.

  5. Diverse FGF receptor signaling controls astrocyte specification and proliferation

    International Nuclear Information System (INIS)

    During CNS development, pluripotency neuronal progenitor cells give rise in succession to neurons and glia. Fibroblast growth factor-2 (FGF-2), a major signal that maintains neural progenitors in the undifferentiated state, is also thought to influence the transition from neurogenesis to gliogenesis. Here we present evidence that FGF receptors and underlying signaling pathways transmit the FGF-2 signals that regulate astrocyte specification aside from its mitogenic activity. Application of FGF-2 to cortical progenitors suppressed neurogenesis whereas treatment with an FGFR antagonist in vitro promoted neurogenesis. Introduction of chimeric FGFRs with mutated tyrosine residues into cortical progenitors and drug treatments to specifically block individual downstream signaling pathways revealed that the overall activity of FGFR rather than individual autophosphorylation sites is important for delivering signals for glial specification. In contrast, a signal for cell proliferation by FGFR was mainly delivered by MAPK pathway. Together our findings indicate that FGFR activity promotes astrocyte specification in the developing CNS.

  6. σ1 Receptor Modulation of G-Protein-Coupled Receptor Signaling: Potentiation of Opioid Transduction Independent from Receptor Binding

    Science.gov (United States)

    Kim, Felix J.; Kovalyshyn, Ivanka; Burgman, Maxim; Neilan, Claire; Chien, Chih-Cheng

    2010-01-01

    σ Ligands modulate opioid actions in vivo, with agonists diminishing morphine analgesia and antagonists enhancing the response. Using human BE(2)-C neuroblastoma cells that natively express opioid receptors and human embryonic kidney (HEK) cells transfected with a cloned μ opioid receptor, we now demonstrate a similar modulation of opioid function, as assessed by guanosine 5′-O-(3-[35S]thio)triphosphate ([35S]GTPγS) binding, by σ1 receptors. σ Ligands do not compete opioid receptor binding. Administered alone, neither σ agonists nor antagonists significantly stimulated [35S]GTPγS binding. Yet σ receptor selective antagonists, but not agonists, shifted the EC50 of opioid-induced stimulation of [35S]GTPγS binding by 3- to 10-fold to the left. This enhanced potency was seen without a change in the efficacy of the opioid, as assessed by the maximal stimulation of [35S]GTPγS binding. σ1 Receptors physically associate with μ opioid receptors, as shown by coimmunoprecipitation studies in transfected HEK cells, implying a direct interaction between the proteins. Thus, σ receptors modulate opioid transduction without influencing opioid receptor binding. RNA interference knockdown of σ1 in BE(2)-C cells also potentiated μ opioid-induced stimulation of [35S]GTPγS binding. These modulatory actions are not limited to μ and δ opioid receptors. In mouse brain membrane preparations, σ1-selective antagonists also potentiated both opioid receptor and muscarinic acetylcholine receptor-mediated stimulation of [35S]GTPγS binding, suggesting a broader role for σ receptors in modulating G-protein-coupled receptor signaling. PMID:20089882

  7. Sigma 1 receptor modulation of G-protein-coupled receptor signaling: potentiation of opioid transduction independent from receptor binding.

    Science.gov (United States)

    Kim, Felix J; Kovalyshyn, Ivanka; Burgman, Maxim; Neilan, Claire; Chien, Chih-Cheng; Pasternak, Gavril W

    2010-04-01

    sigma Ligands modulate opioid actions in vivo, with agonists diminishing morphine analgesia and antagonists enhancing the response. Using human BE(2)-C neuroblastoma cells that natively express opioid receptors and human embryonic kidney (HEK) cells transfected with a cloned mu opioid receptor, we now demonstrate a similar modulation of opioid function, as assessed by guanosine 5'-O-(3-[(35)S]thio)triphosphate ([(35)S]GTP gamma S) binding, by sigma(1) receptors. sigma Ligands do not compete opioid receptor binding. Administered alone, neither sigma agonists nor antagonists significantly stimulated [(35)S]GTP gamma S binding. Yet sigma receptor selective antagonists, but not agonists, shifted the EC(50) of opioid-induced stimulation of [(35)S]GTP gamma S binding by 3- to 10-fold to the left. This enhanced potency was seen without a change in the efficacy of the opioid, as assessed by the maximal stimulation of [(35)S]GTP gamma S binding. sigma(1) Receptors physically associate with mu opioid receptors, as shown by coimmunoprecipitation studies in transfected HEK cells, implying a direct interaction between the proteins. Thus, sigma receptors modulate opioid transduction without influencing opioid receptor binding. RNA interference knockdown of sigma(1) in BE(2)-C cells also potentiated mu opioid-induced stimulation of [(35)S]GTP gamma S binding. These modulatory actions are not limited to mu and delta opioid receptors. In mouse brain membrane preparations, sigma(1)-selective antagonists also potentiated both opioid receptor and muscarinic acetylcholine receptor-mediated stimulation of [(35)S]GTP gamma S binding, suggesting a broader role for sigma receptors in modulating G-protein-coupled receptor signaling. PMID:20089882

  8. Erythropoietin signaling promotes transplanted progenitor cell survival

    OpenAIRE

    Jia, Yi; Warin, Renaud; Yu, Xiaobing; Epstein, Reed; Noguchi, Constance Tom

    2009-01-01

    We examine the potential for erythropoietin signaling to promote donor cell survival in a model of myoblast transplantation. Expression of a truncated erythropoietin receptor in hematopoietic stem cells has been shown to promote selective engraftment in mice. We previously demonstrated expression of endogenous erythropoietin receptor on murine myoblasts, and erythropoietin treatment can stimulate myoblast proliferation and delay differentiation. Here, we report that enhanced erythropoietin re...

  9. PAX5 promotes pre-B cell proliferation by regulating the expression of pre-B cell receptor and its downstream signaling.

    Science.gov (United States)

    Xue, Kai; Song, Jiazhe; Yang, Yan; Li, Zhi; Wu, Chunhua; Jin, Jinhua; Li, Wenzhe

    2016-05-01

    PAX5 is indispensable for the commitment of early lymphoid progenitors to the B cell lineage as well as for the development of B cells. Although previous studies have indicated that the Pax5-conditional-knockout mouse exhibited dedifferentiation of mature B cell and the development of aggressive lymphomas, the changes of Pax5 gene expressions in pre-B cells have not been analyzed. To understand the functional importance of Pax5 gene in the proliferation and survival of pre-B cells, we established a Pax5-knockdown model using 70Z/3 pre-B cell line. Pax5 knockdown 70Z/3 cells (70Z/3-KD cells) showed down-regulations of pre-BCR compounds such as CD19, BLNK, Id2 and λ5. The signaling via pre-BCRs was significantly diminished in the 70Z/3-KD cells, and this alteration was normalized by restored Pax5 gene expression. Loss of PAX5 reduced the growth rates in the 70Z/3-KD cells, compared to the mock cells. Meanwhile, the proliferation of pre-B cells was reduced by the knockdown of Pax5 gene. Moreover, further examinations showed that PAX5 was also activated in B cell acute lymphoblastic leukemia (B-ALL) as a cell proliferation enhancer. These findings suggested that pax5 is critically important for the proliferation and survival of pre-B cells. PMID:27016671

  10. Unifying mechanism for metals in toxicity, carcinogenicity and therapeutic action: integrated approach involving electron transfer, oxidative stress, antioxidants, cell signaling and receptors.

    Science.gov (United States)

    Kovacic, Peter; Somanathan, Ratnasamy

    2010-04-01

    This comprehensive review of biometals involves the unifying theme of electron transfer, reactive oxygen species, and oxidative stress (OS) applied to toxicity, carcinogenicity, and therapeutic action. The beneficial effect of antioxidants supports the participation of OS. The metals involved are mainly in the heavier category. An important aspect is the favorable reduction potential exhibited by the bioactive materials that permits redox cycling in vivo with the generation of oxy radicals. The basic mechanistic theme is applicable to other electron transfer (ET) functionalities. Appreciable evidence indicates the participation of cell signaling in various ways. Also, a simplifying framework is provided based on radical species and electrochemistry (ET and molecular electrostatic potential). This review also discusses receptor participation with focus on binding to proteins. Resultant physiological effects are summarized. The overview provides an integrated approach to metal bioactivation. PMID:20144140

  11. Penicillin binding proteins as danger signals: meningococcal penicillin binding protein 2 activates dendritic cells through Toll-like receptor 4.

    Directory of Open Access Journals (Sweden)

    Marcelo Hill

    Full Text Available Neisseria meningitidis is a human pathogen responsible for life-threatening inflammatory diseases. Meningococcal penicillin-binding proteins (PBPs and particularly PBP2 are involved in bacterial resistance to β-lactams. Here we describe a novel function for PBP2 that activates human and mouse dendritic cells (DC in a time and dose-dependent manner. PBP2 induces MHC II (LOGEC50 = 4.7 µg/ml ± 0.1, CD80 (LOGEC50 = 4.88 µg/ml ± 0.15 and CD86 (LOGEC50 = 5.36 µg/ml ± 0.1. This effect was abolished when DCs were co-treated with anti-PBP2 antibodies. PBP2-treated DCs displayed enhanced immunogenic properties in vitro and in vivo. Furthermore, proteins co-purified with PBP2 showed no effect on DC maturation. We show through different in vivo and in vitro approaches that this effect is not due to endotoxin contamination. At the mechanistic level, PBP2 induces nuclear localization of p65 NF-kB of 70.7 ± 5.1% cells versus 12 ± 2.6% in untreated DCs and needs TLR4 expression to mature DCs. Immunoprecipitation and blocking experiments showed thatPBP2 binds TLR4. In conclusion, we describe a novel function of meningococcal PBP2 as a pathogen associated molecular pattern (PAMP at the host-pathogen interface that could be recognized by the immune system as a danger signal, promoting the development of immune responses.

  12. IgE receptor signaling in food allergy pathogenesis.

    Science.gov (United States)

    Oettgen, Hans C; Burton, Oliver T

    2015-10-01

    The pathogenesis of food allergy remains poorly understood. Recent advances in the use of murine models have led to discoveries that mast cells and IgE receptor signaling not only drive immediate hypersensitivity reactions but also exert an immunoregulatory function, promoting the development of allergic sensitivity to foods. We review the evidence that IgE, IgE receptors, key signaling kinases and mast cells impair oral tolerance to ingested foods, preventing the induction of regulatory T cells (Treg) and promoting the acquisition of pro-allergic T helper (Th) 2 responses. We discuss innovative strategies that that could be implemented to counteract these immunoregulatory effects of IgE-mediated mast cell activation, and potentially reverse established sensitization, curing food allergy. PMID:26296054

  13. The role of CXC-chemokine receptor CXCR2 and suppressor of cytokine signaling-3 (SOCS-3) in renal cell carcinoma

    International Nuclear Information System (INIS)

    Chemokine receptor signaling pathways are implicated in the pathobiology of renal cell carcinoma (RCC). However, the clinical relevance of CXCR2 receptor, mediating the effects of all angiogenic chemokines, remains unclear. SOCS (suppressor of cytokine signaling)-3 is a negative regulator of cytokine-driven responses, contributing to interferon-α resistance commonly used to treat advanced RCC with limited information regarding its expression in RCC. In this study, CXCR2 and SOCS-3 were immunohistochemically investigated in 118 RCC cases in relation to interleukin (IL)-6 and (IL)-8, their downstream transducer phosphorylated (p-)STAT-3, and VEGF expression, being further correlated with microvascular characteristics, clinicopathological features and survival. In 30 cases relationships with hypoxia-inducible factors, i.e. HIF-1a, p53 and NF-κΒ (p65/RelA) were also examined. Validation of immunohistochemistry and further investigation of downstream transducers, p-JAK2 and p-c-Jun were evaluated by Western immunoblotting in 5 cases. Both CXCR2 and IL-8 were expressed by the neoplastic cells their levels being interrelated. CXCR2 strongly correlated with the levels of HIF-1a, p53 and p65/RelA in the neoplastic cells. Although SOCS-3 was simultaneously expressed with p-STAT-3, its levels tended to show an inverse relationship with p-JAK-2 and p-c-Jun in Western blots and were positively correlated with HIF-1a, p53 and p65/p65/RelA expression. Neither CXCR2 nor SOCS-3 correlated with the extent of microvascular network. IL-8 and CXCR2 expression was associated with high grade, advanced stage and the presence/number of metastases but only CXCR2 adversely affected survival in univariate analysis. Elevated SOCS-3 expression was associated with progression, the presence/number of metastasis and shortened survival in both univariate and multivariate analysis. Our findings implicate SOCS-3 overexpression in RCC metastasis and biologic aggressiveness advocating its

  14. The erbB3- and IGF-1 receptor-initiated signaling pathways exhibit distinct effects on lapatinib sensitivity against trastuzumab-resistant breast cancer cells.

    Science.gov (United States)

    Lyu, Hui; Yang, Xiao He; Edgerton, Susan M; Thor, Ann D; Wu, Xiaoying; He, Zhimin; Liu, Bolin

    2016-01-19

    Both erbB3 and IGF-1 receptor (IGF-1R) have been shown to play an important role in trastuzumab resistance. However, it remains unclear whether erbB3- and IGF-1R-initiated signaling pathways possess distinct effects on the sensitivity of lapatinib, a dual tyrosine kinase inhibitor against both EGFR and erbB2, in trastuzumab-resistant breast cancer. Here, we show that the trastuzumab-resistant SKBR3-pool2 and BT474-HR20 breast cancer sublines, as compared the parental SKBR3 and BT474 cells, respectively, exhibit refractoriness to lapatinib. Knockdown of erbB3 inhibited Akt in SKBR3-pool2 and BT474-HR20 cells, significantly increased lapatinib efficacy, and dramatically re-sensitized the cells to lapatinib-induced apoptosis. In contrast, specific knockdown of IGF-1R did not alter the cells' responsiveness to lapatinib. While the levels of phosphorylated Src (P-Src) were reduced upon IGF-1R downregulation, the P-Akt levels remained unchanged. Furthermore, a specific inhibitor of Akt, but not Src, significantly enhanced lapatinib-mediated anti-proliferative/anti-survival effects on SKBR3-pool2 and BT474-HR20 cells. These data indicate that erbB3 signaling is critical for both trastuzumab and lapatinib resistances mainly through the PI-3K/Akt pathway, whereas IGF-1R-initiated Src activation results in trastuzumab resistance without affecting lapatinib sensitivity. Our findings may facilitate the development of precision therapeutic regimens for erbB2-positive breast cancer patients who become resistant to erbB2-targeted therapy. PMID:26621843

  15. Signal transduction by the platelet-derived growth factor receptor

    International Nuclear Information System (INIS)

    The mitogenic effects of platelet-derived growth factor (PDGF) are mediated by the PDGF receptor. The mouse PDGF receptor was recently purified on the basis of its ability to become tyrosine phosphorylated in response to the A-B human platelet form of PDGF, and the receptor amino acid sequence was determined from a full-length cDNA clone. Both the human and mouse receptor cDNA sequences have been expressed in Chinese hamster ovary fibroblast (CHO) cells that normally lack PDGF receptors. This paper summarizes recent results using this system to study signal transduction by the PDGF receptor. Some of the findings show that the KI domain of the PDGF receptor plays an important role in the stimulation of DNA synthesis by PDGF. Surprisingly, the kinase insert region is not essential for PDGF stimulation of PtdIns turnover, pH change, increase in cellular calcium, and receptor autophosphorylation. In addition, PDGF stimulates a conformational change in the receptor

  16. Human herpesvirus 6 infection impairs Toll-like receptor signaling

    OpenAIRE

    Ochi Toshiki; Suemori Koichiro; An Jun; Fujiwara Hiroshi; Tanimoto Kazushi; Murakami Yuichi; Hasegawa Hitoshi; Yasukawa Masaki

    2010-01-01

    Abstract Human herpesvirus 6 (HHV-6) has a tropism for immunocompetent cells, including T lymphocytes, monocytes/macrophages, and dendritic cells (DCs) suggesting that HHV-6 infection affects the immunosurveillance system. Toll-like receptor (TLR) system plays an important role in innate immunity against various pathogens. In the present study, we investigated the effect of HHV-6 infection on the expression and intracellular signaling of TLRs in DCs. Although expression levels of TLRs were no...

  17. The molecular mechanisms behind receptor signaling

    OpenAIRE

    Blain, Katherine Yoshie

    2010-01-01

    Cell membranes are a crucial component to the life of a cell. The membrane defines boundaries, provides structural elements, and contains proteins that serve as sensor receptors to transmit external cues across the phospholipid bilayer, either from the environment to the cell's interior or from the cytosol to a particular sub- cellular compartment. One type of proteins found spanning these lipid bilayers are known as the integral membrane proteins (IMPs). Since the dysfunction of this class o...

  18. Hypoxia perturbs aryl hydrocarbon receptor signaling and CYP1A1 expression induced by PCB 126 in human skin and liver-derived cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Vorrink, Sabine U. [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Severson, Paul L. [Department of Pharmacology and Toxicology, The University of Arizona, Tucson, AZ (United States); Kulak, Mikhail V. [Department of Surgery, The University of Iowa, Iowa City, IA (United States); Futscher, Bernard W. [Department of Pharmacology and Toxicology, The University of Arizona, Tucson, AZ (United States); Domann, Frederick E., E-mail: frederick-domann@uiowa.edu [Interdisciplinary Graduate Program in Human Toxicology, The University of Iowa, Iowa City, IA (United States); Department of Radiation Oncology, The University of Iowa, Iowa City, IA (United States); Department of Surgery, The University of Iowa, Iowa City, IA (United States)

    2014-02-01

    The aryl hydrocarbon receptor (AhR) is an important mediator of toxic responses after exposure to xenobiotics including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and dioxin-like polychlorinated biphenyls (PCBs). Activation of AhR responsive genes requires AhR dimerization with the aryl hydrocarbon receptor nuclear translocator (ARNT), a heterodimeric partner also shared by the hypoxia-inducible factor-1α (HIF-1α) protein. TCDD-stimulated AhR transcriptional activity can be influenced by hypoxia; however, it less well known whether hypoxia interferes with AhR transcriptional transactivation in the context of PCB-mediated AhR activation in human cells. Elucidation of this interaction is important in liver hepatocytes which extensively metabolize ingested PCBs and experience varying degrees of oxygen tension during normal physiologic function. This study was designed to assess the effect of hypoxia on AhR transcriptional responses after exposure to 3,3′,4,4′,5-pentachlorobiphenyl (PCB 126). Exposure to 1% O{sub 2} prior to PCB 126 treatment significantly inhibited CYP1A1 mRNA and protein expression in human HepG2 and HaCaT cells. CYP1A1 transcriptional activation was significantly decreased upon PCB 126 stimulation under conditions of hypoxia. Additionally, hypoxia pre-treatment reduced PCB 126 induced AhR binding to CYP1 target gene promoters. Importantly, ARNT overexpression rescued cells from the inhibitory effect of hypoxia on XRE-luciferase reporter activity. Therefore, the mechanism of interference of the signaling crosstalk between the AhR and hypoxia pathways appears to be at least in part dependent on ARNT availability. Our results show that AhR activation and CYP1A1 expression induced by PCB 126 were significantly inhibited by hypoxia and hypoxia might therefore play an important role in PCB metabolism and toxicity. - Highlights: • Significant crosstalk exists between AhR and HIF-1α signaling. • Hypoxia perturbs PCB 126 induced AhR function and

  19. Hypoxia perturbs aryl hydrocarbon receptor signaling and CYP1A1 expression induced by PCB 126 in human skin and liver-derived cell lines

    International Nuclear Information System (INIS)

    The aryl hydrocarbon receptor (AhR) is an important mediator of toxic responses after exposure to xenobiotics including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and dioxin-like polychlorinated biphenyls (PCBs). Activation of AhR responsive genes requires AhR dimerization with the aryl hydrocarbon receptor nuclear translocator (ARNT), a heterodimeric partner also shared by the hypoxia-inducible factor-1α (HIF-1α) protein. TCDD-stimulated AhR transcriptional activity can be influenced by hypoxia; however, it less well known whether hypoxia interferes with AhR transcriptional transactivation in the context of PCB-mediated AhR activation in human cells. Elucidation of this interaction is important in liver hepatocytes which extensively metabolize ingested PCBs and experience varying degrees of oxygen tension during normal physiologic function. This study was designed to assess the effect of hypoxia on AhR transcriptional responses after exposure to 3,3′,4,4′,5-pentachlorobiphenyl (PCB 126). Exposure to 1% O2 prior to PCB 126 treatment significantly inhibited CYP1A1 mRNA and protein expression in human HepG2 and HaCaT cells. CYP1A1 transcriptional activation was significantly decreased upon PCB 126 stimulation under conditions of hypoxia. Additionally, hypoxia pre-treatment reduced PCB 126 induced AhR binding to CYP1 target gene promoters. Importantly, ARNT overexpression rescued cells from the inhibitory effect of hypoxia on XRE-luciferase reporter activity. Therefore, the mechanism of interference of the signaling crosstalk between the AhR and hypoxia pathways appears to be at least in part dependent on ARNT availability. Our results show that AhR activation and CYP1A1 expression induced by PCB 126 were significantly inhibited by hypoxia and hypoxia might therefore play an important role in PCB metabolism and toxicity. - Highlights: • Significant crosstalk exists between AhR and HIF-1α signaling. • Hypoxia perturbs PCB 126 induced AhR function and target

  20. Role of inositol phospholipid signaling in natural killer cell biology

    OpenAIRE

    Gumbleton, Matthew; Kerr, William G.

    2013-01-01

    Natural killer (NK) cells are important for host defense against malignancy and infection. At a cellular level NK cells are activated when signals from activating receptors exceed signaling from inhibitory receptors. At a molecular level NK cells undergo an education process to both prevent autoimmunity and acquire lytic capacity. Mouse models have shown important roles for inositol phospholipid signaling in lymphocytes. NK cells from mice with deletion in different members of the inositol ph...

  1. Sweet Taste Receptor Expressed in Pancreatic β-Cells Activates the Calcium and Cyclic AMP Signaling Systems and Stimulates Insulin Secretion

    OpenAIRE

    Nakagawa, Yuko; Nagasawa, Masahiro; Yamada, Satoko; Hara, Akemi; Mogami, Hideo; Nikolaev, Viacheslav O.; Lohse, Martin J.; Shigemura, Noriatsu; Ninomiya, Yuzo; Kojima, Itaru

    2009-01-01

    Background Sweet taste receptor is expressed in the taste buds and enteroendocrine cells acting as a sugar sensor. We investigated the expression and function of the sweet taste receptor in MIN6 cells and mouse islets. Methodology/Principal Findings The expression of the sweet taste receptor was determined by RT–PCR and immunohistochemistry. Changes in cytoplasmic Ca2+ ([Ca2+]c) and cAMP ([cAMP]c) were monitored in MIN6 cells using fura-2 and Epac1-camps. Activation of protein kinase C was mo...

  2. Minocycline suppresses interleukine-6, its receptor system and signaling pathways and impairs migration, invasion and adhesion capacity of ovarian cancer cells: in vitro and in vivo studies.

    Directory of Open Access Journals (Sweden)

    Parvin Ataie-Kachoie

    Full Text Available Interleukin (IL-6 has been shown to be a major contributing factor in growth and progression of ovarian cancer. The cytokine exerts pro-tumorigenic activity through activation of several signaling pathways in particular signal transducer and activator of transcription (STAT3 and extracellular signal-regulated kinase (ERK1/2. Hence, targeting IL-6 is becoming increasingly attractive as a treatment option in ovarian cancer. Here, we investigated the effects of minocycline on IL-6 and its signaling pathways in ovarian cancer. In vitro, minocycline was found to significantly suppress both constitutive and IL-1β or 4-hydroxyestradiol (4-OH-E2-stimulated IL-6 expression in human ovarian cancer cells; OVCAR-3, SKOV-3 and CAOV-3. Moreover, minocycline down-regulated two major components of IL-6 receptor system (IL-6Rα and gp130 and blocked the activation of STAT3 and ERK1/2 pathways leading to suppression of the downstream product MCL-1. In female nude mice bearing intraperitoneal OVCAR-3 tumors, acute administration (4 and 24 h of minocycline (30 mg/kg led to suppression of IL-6. Even single dose of minocycline was effective at significantly lowering plasma and tumor IL-6 levels. In line with this, tumoral expression of p-STAT3, p-ERK1/2 and MCL-1 were decreased in minocycline-treated mice. Evaluation of the functional implication of minocycline on metastatic activity revealed the capacity of minocycline to inhibit cellular migration, invasion and adhesion associated with down-regulation of matrix metalloproteinases (MMP-2 and 9. Thus, the data suggest a potential role for minocycline in suppressing IL-6 expression and activity. These effects may prove to be an important attribute to the upcoming clinical trials of minocycline in ovarian cancer.

  3. VISFATIN (NAMPT) Improves In Vitro IGF1-Induced Steroidogenesis and IGF1 Receptor Signaling Through SIRT1 in Bovine Granulosa Cells.

    Science.gov (United States)

    Reverchon, Maxime; Rame, Christelle; Bunel, Audrey; Chen, Wenyong; Froment, Pascal; Dupont, Joëlle

    2016-03-01

    VISFATIN is a novel adipokine, also known as a nicotinamide phosphorybosyltransferase (NAMPT), that is able to modulate different processes, including lipid and glucose metabolism, oxidative stress, inflammation, and insulin resistance. Recent data suggest that it also plays a role in reproductive function in rats, humans, and chickens. Here we identified VISFATIN in the bovine ovary and investigated the in vitro effects of this hormone on granulosa cell steroidogenesis and proliferation and oocyte maturation. By RT-PCR, immunoblotting, and immunohistochemistry, we found VISFATIN in various ovarian cells, including granulosa and theca cells, corpus luteum, and oocytes. In cultured bovine granulosa cells, we showed that IGF1 (10(-8) M) and VISFATIN (10 and 100 ng/ml) but not FSH (10(-8) M) increased mRNA expression levels of NAMPT after 48 h of stimulation. Moreover, we observed that human recombinant VISFATIN (hVisf, 10 ng/ml, 48 h) increased the release of progesterone and estradiol secretion, and this was associated with an increase in the protein level of STAR, the HSD3B activity, and the phosphorylation levels of IGF1R and MAPK ERK1/2 in the presence or absence of IGF1 (10(-8) M). All these effects were abolished when NAMPT was knocked down and when the sirtuin pharmacological inhibitors CHIC-35 (60 nM) and EX-527 (0.5 μM) were preincubated in bovine granulosa cells. Thus, in cultured bovine granulosa cells, VISFATIN improves basal and IGF1-induced steroidogenesis and IGF1 receptor signaling through SIRT1. PMID:26792944

  4. Nitric Oxide-Releasing Aspirin Suppresses NF-κB Signaling in Estrogen Receptor Negative Breast Cancer Cells in Vitro and in Vivo

    Directory of Open Access Journals (Sweden)

    Niharika Nath

    2015-07-01

    Full Text Available Estrogen receptor negative (ER(− breast cancer is aggressive, responds poorly to current treatments and has a poor prognosis. The NF-κB signaling pathway is implicated in ER(− tumorigenesis. Aspirin (ASA is chemopreventive against ER(+ but not for ER(− breast cancers. Nitric oxide-releasing aspirin (NO-ASA is a safer ASA where ASA is linked to an NO-releasing moiety through a spacer. In vitro, we investigated anti-proliferation effects of NO-ASA (para- and meta-isomers against ER(− breast cancer cells MDA-MB-231 and SK-BR-23, effects on NF-κB signaling, and reactive oxygen species by standard techniques. In vivo, effects of NO-ASA were evaluated in a mouse xenograft model using MDA-MB-231 cells. p-NO-ASA inhibited the growth of MDA-MB-231 and SK-BR-3 cells at 24 h, the respective IC50s were 13 ± 2 and 17 ± 2 μM; ASA had an IC50 of >3000 μM in both cell lines. The IC50s for m-NO-ASA in MDA-MB-231 and SK-BR-3 were 173 ± 15 and 185 ± 12 μM, respectively, therefore, implying p-NO-ASA as a stronger inhibitor of growth p-NO-ASA reduced cell growth by inhibiting proliferation, inducing apoptosis and causing G0/G1 cell cycle block. Activation of NF-κB was inhibited by both isomers as demonstrated by decreases in NF-κB-DNA binding and luciferase activity at 24 h, However, m-NO-ASA produced transient effects at 3 h such as increased NF-κB-DNA-binding, increased levels of nuclear p50, even though both isomers inhibited IκB degradation. Increase in nuclear p50 by m-NO-ASA was associated with translocation of p50 in to the nucleus as observed by immunoflouresence at 3 h. NO-ASA induced reactive oxygen species (ROS as evidenced by overall increases in both H2DCFDA (2′,7′-dichlorodihydrofluorescein and DHE (dihydroethidium-derived fluorescence. Inhibition of ROS by N-acetyl-cysteine reversed the m-NO-ASA-mediated translocation of p50 in to the nucleus. In xenografts, p-NO-ASA inhibited tumor growth by inhibiting proliferation (PCNA and

  5. Janus kinases in immune cell signaling

    OpenAIRE

    Ghoreschi, Kamran; Laurence, Arian; O’Shea, John J.

    2009-01-01

    The Janus family kinases (Jaks), Jak1, Jak2, Jak3, and Tyk2, form one subgroup of the non-receptor protein tyrosine kinases. They are involved in cell growth, survival, development, and differentiation of a variety of cells but are critically important for immune cells and hematopoietic cells. Data from experimental mice and clinical observations have unraveled multiple signaling events mediated by Jak in innate and adaptive immunity. Deficiency of Jak3 or Tyk2 results in defined clinical dis...

  6. Activation of P2X7 and P2Y11 purinergic receptors inhibits migration and normalizes tumor-derived endothelial cells via cAMP signaling.

    Science.gov (United States)

    Avanzato, D; Genova, T; Fiorio Pla, A; Bernardini, M; Bianco, S; Bussolati, B; Mancardi, D; Giraudo, E; Maione, F; Cassoni, P; Castellano, I; Munaron, L

    2016-01-01

    Purinergic signaling is involved in inflammation and cancer. Extracellular ATP accumulates in tumor interstitium, reaching hundreds micromolar concentrations, but its functional role on tumor vasculature and endothelium is unknown. Here we show that high ATP doses (>20 μM) strongly inhibit migration of endothelial cells from human breast carcinoma (BTEC), but not of normal human microvascular EC. Lower doses (1-10 mm result ineffective. The anti-migratory activity is associated with cytoskeleton remodeling and is significantly prevented by hypoxia. Pharmacological and molecular evidences suggest a major role for P2X7R and P2Y11R in ATP-mediated inhibition of TEC migration: selective activation of these purinergic receptors by BzATP mimics the anti-migratory effect of ATP, which is in turn impaired by their pharmacological or molecular silencing. Downstream pathway includes calcium-dependent Adenilyl Cyclase 10 (AC10) recruitment, cAMP release and EPAC-1 activation. Notably, high ATP enhances TEC-mediated attraction of human pericytes, leading to a decrease of endothelial permeability, a hallmark of vessel normalization. Finally, we provide the first evidence of in vivo P2X7R expression in blood vessels of murine and human breast carcinoma. In conclusion, we have identified a purinergic pathway selectively acting as an antiangiogenic and normalizing signal for human tumor-derived vascular endothelium. PMID:27586846

  7. Evidence for cross-talk between M2 and M3 muscarinic acetylcholine receptors in the regulation of second messenger and extracellular signal-regulated kinase signalling pathways in Chinese hamster ovary cells

    OpenAIRE

    Hornigold, David C; Mistry, Rajendra; Raymond, Pamela D; Blank, Jonathan L; John Challiss, R A

    2003-01-01

    We have examined possible mechanisms of cross-talk between the Gq/11-linked M3 muscarinic acetylcholine (mACh) receptor and the Gi/o-linked M2 mACh receptor by stable receptor coexpression in Chinese hamster ovary (CHO) cells. A number of second messenger (cyclic AMP, Ins(1,4,5)P3) and mitogen-activated protein kinase (ERK and JNK) responses stimulated by the mACh receptor agonist methacholine were examined in CHO-m2m3 cells and compared to those stimulated in CHO-m2 and CHO-m3 cell-lines, ex...

  8. Activation of the GLP-1 Receptor Signalling Pathway: A Relevant Strategy to Repair a Deficient Beta-Cell Mass

    Directory of Open Access Journals (Sweden)

    Bernard Portha

    2011-01-01

    Full Text Available Recent preclinical studies in rodent models of diabetes suggest that exogenous GLP-1R agonists and DPP-4 inhibitors have the ability to increase islet mass and preserve beta-cell function, by immediate reactivation of beta-cell glucose competence, as well as enhanced beta-cell proliferation and neogenesis and promotion of beta-cell survival. These effects have tremendous implication in the treatment of T2D because they directly address one of the basic defects in T2D, that is, beta-cell failure. In human diabetes, however, evidence that the GLP-1-based drugs alter the course of beta-cell function remains to be found. Several questions surrounding the risks and benefits of GLP-1-based therapy for the diabetic beta-cell mass are discussed in this review and require further investigation.

  9. Physician Education: The Erythropoietin Receptor and Signal Transduction.

    Science.gov (United States)

    Yoshimura; Arai

    1996-01-01

    receptor gene was cloned by D'Andrea and coworkers in 1989 from murine erythroleukemia cells [1]. It became clear that the EPO receptor belongs to the cytokine receptor family that comprises receptors for the various interleukins, GM-CSF, granulocyte colony-stimulating factor (G-CSF), growth hormone and prolactin. The special characteristic of this family of receptors is that they are switched on (i.e., the receptor is activated) and transduce signals to the interior of the cell by the formation of homo- or hetero-oligomers (dimers or trimers). Moreover, hetero-oligomers of these receptors share a common receptor subunit. As shown in Figure 2, the IL-3, IL-5 and GM-CSF receptors have a common &bgr; subunit, and their ligand specificity is determined by the &agr; subunit. In the same manner, the IL-6, LIF and oncostatin M (OSM) receptors all share gp130, which is the &bgr; subunit of the IL-6 receptor. The IL-2, IL-4 and IL-7 receptors all share the &ggr; subunit of the IL-2 receptor. All the above receptors are activated by the formation of hetero-oligomers, but the G-CSF receptor, EPO receptor, and growth hormone receptor are activated by the formation of homodimers of the same types of molecules [2]. We can see that groups of cytokines such as the interleukins that affect a relatively wide range of cells and have redundant biological activity create this redundancy through the common use of a single receptor subunit. On the other hand, EPO and G-CSF act with high specificity on a relatively limited range of cells, so it was probably unnecessary for their receptors to share one of the subunits. EPO RECEPTOR AND JAK2 KINASE: The signal for cellular proliferation and differentiation into erythroblasts is thought to originate at the EPO receptor. The cytoplasmic domain of the EPO receptor can be divided into two major regions. Roughly half of the cytoplasmic domain, the part lying nearest the plasma membrane, is required for generating the signals for proliferation and

  10. G-Protein-Coupled Lysophosphatidic Acid Receptors and Their Regulation of AKT Signaling

    OpenAIRE

    Anjum Riaz; Ying Huang; Staffan Johansson

    2016-01-01

    A hallmark of G-protein-coupled receptors (GPCRs) is their ability to recognize and respond to chemically diverse ligands. Lysophospholipids constitute a relatively recent addition to these ligands and carry out their biological functions by activating G-proteins coupled to a large family of cell-surface receptors. This review aims to highlight salient features of cell signaling by one class of these receptors, known as lysophosphatidic acid (LPA) receptors, in the context of phosphatidylinos...

  11. Nuclear receptor TLX inhibits TGF-β signaling in glioblastoma.

    Science.gov (United States)

    Johansson, Erik; Zhai, Qiwei; Zeng, Zhao-Jun; Yoshida, Takeshi; Funa, Keiko

    2016-05-01

    TLX (also called NR2E1) is an orphan nuclear receptor that maintains stemness of neuronal stem cells. TLX is highly expressed in the most malignant form of glioma, glioblastoma multiforme (GBM), and is important for the proliferation and maintenance of the stem/progenitor cells of the tumor. Transforming Growth Factor-β (TGF-β) is a cytokine regulating many different cellular processes such as differentiation, migration, adhesion, cell death and proliferation. TGF-β has an important function in cancer where it can work as either a tumor suppressor or oncogene, depending on the cancer type and stage of tumor development. Since glioblastoma often have dysfunctional TGF-β signaling we wanted to find out if there is any interaction between TLX and TGF-β in glioblastoma cells. We demonstrate that knockdown of TLX enhances the canonical TGF-β signaling response in glioblastoma cell lines. TLX physically interacts with and stabilizes Smurf1, which can ubiquitinate and target TGF-β receptor II for degradation, whereas knockdown of TLX leads to stabilization of TGF-β receptor II, increased nuclear translocation of Smad2/3 and enhanced expression of TGF-β target genes. The interaction between TLX and TGF-β may play an important role in the regulation of proliferation and tumor-initiating properties of glioblastoma cells. PMID:27048878

  12. Nonredundant roles of Src-family kinases and Syk in the initiation of B-cell antigen receptor signaling

    Czech Academy of Sciences Publication Activity Database

    Štěpánek, Ondřej; Dráber, Peter; Drobek, Aleš; Hořejší, Václav; Brdička, Tomáš

    2013-01-01

    Roč. 190, č. 4 (2013), s. 1807-1818. ISSN 0022-1767 R&D Projects: GA ČR(CZ) GBP302/12/G101; GA ČR GAP302/12/1712 Institutional support: RVO:68378050 Keywords : BCR signaling * Src family kinases * Syk Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.362, year: 2013

  13. Gefitinib Radiosensitizes Stem-Like Glioma Cells: Inhibition of Epidermal Growth Factor Receptor-Akt-DNA-PK Signaling, Accompanied by Inhibition of DNA Double-Strand Break Repair

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Khong Bee, E-mail: dmskkb@nccs.com.sg [Brain Tumour Research Laboratory, Division of Medical Sciences, National Cancer Centre Singapore (Singapore); Zhu Congju; Wong Yinling; Gao Qiuhan; Ty, Albert; Wong, Meng Cheong [Brain Tumour Research Laboratory, Division of Medical Sciences, National Cancer Centre Singapore (Singapore)

    2012-05-01

    Purpose: We compared radiosensitivity of brain tumor stem cells (BTSCs) with matched nonstem glioma cells, and determined whether gefitinib enhanced BTSC radiosensitivity by inhibiting epidermal growth factor receptor (EGFR)-Akt-DNA-dependent protein kinase (DNA-PK) signaling, followed by enhanced DNA double-stand breaks (DSBs) and inhibition of DSB repair. Methods and Materials: Radiosensitivity of stem-like gliomaspheres and nonstem glioma cells (obtained at patient neurosurgical resection) were evaluated by clonogenic assays, {gamma}-H{sub 2}AX immunostaining and cell cycle distribution. Survival of irradiated and nonirradiated NOD-SCID mice intracranially implanted with stem-like gliomaspheres were monitored. Glioma cells treated with gefitinib, irradiation, or both were assayed for clonogenic survival, {gamma}-H{sub 2}AX immunostaining, DNA-PKcs expression, and phosphorylation of EGFR and Akt. Results: Stem-like gliomaspheres displayed BTSC characteristics of self-renewal; differentiation into lineages of neurons, oligodendrocytes, and astrocytes; and initiation of glioma growth in NOD-SCID mice. Irradiation dose-dependently reduced clonogenic survival, induced G{sub 2}/M arrest and increased {gamma}-H{sub 2}AX immunostaining of nonstem glioma cells, but not stem-like gliomaspheres. There was no difference in survival of irradiated and nonirradiated mice implanted with stem-like gliomaspheres. The addition of gefitinib significantly inhibited clonogenic survival, increased {gamma}-H{sub 2}AX immunostaining, and reduced DNA-PKcs expression of irradiated stem-like gliomaspheres, without affecting irradiated-nonstem glioma cells. Gefitinib alone, and when combined with irradiation, inhibited phosphorylation of EGFR (Y1068 and Y1045) and Akt (S473) in stem-like gliomaspheres. In nonstem glioma cells, gefitinib alone inhibited EGFR Y1068 phosphorylation, with further inhibition by combined gefitinib and irradiation. Conclusions: Stem-like gliomaspheres are

  14. Gefitinib Radiosensitizes Stem-Like Glioma Cells: Inhibition of Epidermal Growth Factor Receptor-Akt-DNA-PK Signaling, Accompanied by Inhibition of DNA Double-Strand Break Repair

    International Nuclear Information System (INIS)

    Purpose: We compared radiosensitivity of brain tumor stem cells (BTSCs) with matched nonstem glioma cells, and determined whether gefitinib enhanced BTSC radiosensitivity by inhibiting epidermal growth factor receptor (EGFR)–Akt-DNA–dependent protein kinase (DNA-PK) signaling, followed by enhanced DNA double-stand breaks (DSBs) and inhibition of DSB repair. Methods and Materials: Radiosensitivity of stem-like gliomaspheres and nonstem glioma cells (obtained at patient neurosurgical resection) were evaluated by clonogenic assays, γ-H2AX immunostaining and cell cycle distribution. Survival of irradiated and nonirradiated NOD-SCID mice intracranially implanted with stem-like gliomaspheres were monitored. Glioma cells treated with gefitinib, irradiation, or both were assayed for clonogenic survival, γ-H2AX immunostaining, DNA-PKcs expression, and phosphorylation of EGFR and Akt. Results: Stem-like gliomaspheres displayed BTSC characteristics of self-renewal; differentiation into lineages of neurons, oligodendrocytes, and astrocytes; and initiation of glioma growth in NOD-SCID mice. Irradiation dose-dependently reduced clonogenic survival, induced G2/M arrest and increased γ-H2AX immunostaining of nonstem glioma cells, but not stem-like gliomaspheres. There was no difference in survival of irradiated and nonirradiated mice implanted with stem-like gliomaspheres. The addition of gefitinib significantly inhibited clonogenic survival, increased γ-H2AX immunostaining, and reduced DNA-PKcs expression of irradiated stem-like gliomaspheres, without affecting irradiated-nonstem glioma cells. Gefitinib alone, and when combined with irradiation, inhibited phosphorylation of EGFR (Y1068 and Y1045) and Akt (S473) in stem-like gliomaspheres. In nonstem glioma cells, gefitinib alone inhibited EGFR Y1068 phosphorylation, with further inhibition by combined gefitinib and irradiation. Conclusions: Stem-like gliomaspheres are resistant to irradiation-induced cytotoxicity, G2/M

  15. p21-activated kinase group II small compound inhibitor GNE-2861 perturbs estrogen receptor alpha signaling and restores tamoxifen-sensitivity in breast cancer cells

    OpenAIRE

    Zhuang, Ting; Zhu, Jian; Li, Zhilun; Lorent, Julie; Zhao, Chunyan; Dahlman-Wright, Karin; Strömblad, Staffan

    2015-01-01

    Estrogen receptor alpha (ERα) is highly expressed in most breast cancers. Consequently, ERα modulators, such as tamoxifen, are successful in breast cancer treatment, although tamoxifen resistance is commonly observed. While tamoxifen resistance may be caused by altered ERα signaling, the molecular mechanisms regulating ERα signaling and tamoxifen resistance are not entirely clear. Here, we found that PAK4 expression was consistently correlated to poor patient outcome in endocrine treated and ...

  16. Activation of type 5 metabotropic glutamate receptor promotes the proliferation of rat retinal progenitor cell via activation of the PI-3-K and MAPK signaling pathways.

    Science.gov (United States)

    Zhang, Z; Hu, F; Liu, Y; Ma, B; Chen, X; Zhu, K; Shi, Y; Wei, T; Xing, Y; Gao, Y; Lu, H; Liu, Y; Kang, Q

    2016-05-13

    The metabotropic glutamate receptor 5 (mGluR5) regulates neurogenesis in the brain, but the effect of mGluR5 on retinal progenitor cells (RPCs) remains unknown. In this study, we found that mGluR5 promoted the proliferation of rat RPCs with activation of the phosphatidylinositol-3-kinase (PI-3-K) and mitogen-activated protein kinase (MAPK) signaling pathways in vitro. The mGluR5 agonist (S)-3,5-dihydroxyphenylglycine hydrate (DHPG) increased the cellular viability in a concentration- and time-dependent manner, whereas the mGluR5 antagonist 6-methyl-2-(phenylethynyl)pyridine hydrochloride (MTEP) had the opposite effect, as shown by 3-((2-methyl-1,3-thiazol-4-yl)ethynyl)pyridine hydrochloride (MTT) assay. Treatment with DHPG (100μM) also promoted the proliferation of RPCs, as indicated by 5-Bromo-2-deoxyUridine (BrdU) staining and flow cytometry, and likewise, MTEP (100μM) and mGluR5 knockdown abolished the action of mGluR5 activity. Western blot demonstrated that the activation of mGluR5 enhanced the expression of Cyclin D1 and the phosphorylation level of PKC however, MTEP or mGluR5 knockdown also abrogated the effect of DHPG on RPCs. Furthermore, we found that activation of the extracellular signal-regulated protein kinase (ERK) and protein kinase B (AKT) signaling pathways was involved in the proliferation of RPC. After DHPG treatment, the levels of both p-ERK1/2 and p-AKT increased in a time-dependent manner. Then we used MTEP, mGluR5 knockdown, the ERK1/2 inhibitor U0126 and the AKT inhibitor LY294002 to pretreat the cells, and all of them clearly eliminated the influence of DHPG. These results demonstrated that mGluR5 regulates neurogenesis in RPCs through the MAPK and PI-3-K signaling pathways, and these findings may motivate a pharmacological study investigating a potential mechanism for the treatment of retinal diseases such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD). PMID:26902516

  17. Intestinal epithelial vitamin D receptor signaling inhibits experimental colitis

    OpenAIRE

    Liu, Weicheng; Chen, Yunzi; Golan, Maya Aharoni; Annunziata, Maria L.; Du, Jie; Dougherty, Urszula; Kong, Juan; Musch, Mark; Huang, Yong; Pekow, Joel; Zheng, Changqing; Bissonnette, Marc; Hanauer, Stephen B.; Li, Yan Chun

    2013-01-01

    The inhibitory effects of vitamin D on colitis have been previously documented. Global vitamin D receptor (VDR) deletion exaggerates colitis, but the relative anticolitic contribution of epithelial and nonepithelial VDR signaling is unknown. Here, we showed that colonic epithelial VDR expression was substantially reduced in patients with Crohn’s disease or ulcerative colitis. Moreover, targeted expression of human VDR (hVDR) in intestinal epithelial cells (IECs) protected mice from developing...

  18. MHC class II molecules deliver costimulatory signals in human T cells through a functional linkage with IL-2-receptors

    DEFF Research Database (Denmark)

    Odum, Niels; Kanner, S B; Ledbetter, J A;

    1993-01-01

    a regulatory function in T cell activation. Here, we show that cross-linking HLA-DR and -DP but not -DQ molecules by immobilized mAb enhanced proliferative T cell responses to IL-2. In contrast, class II stimulation had no effect on IL-4-induced proliferation. The costimulatory effect was most......Ab induced tyrosine phosphorylation of specific substrates including PLC-gamma 1. Combined stimulation of IL-2R and class II molecules had an additive effect on tyrosine phosphorylation. Pretreatment of T cells with a protein tyrosine kinase inhibitor, herbimycin A, inhibited IL-2 and class II...

  19. The role of MAPK in CD4{sup +} T cells toll-like receptor 9-mediated signaling following HHV-6 infection

    Energy Technology Data Exchange (ETDEWEB)

    Chi, Jing [Department of Microbiology and Immunology, Nanjing Medical University, Nanjing 210029, Jiangsu Province (China); Wang, Fang [Department of Laboratory Medicine, the First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing 210029, Jiangsu Province (China); Li, Lingyun [Department of Developmental Genetics, Nanjing Medical University, Nanjing 210029, Jiangsu Province (China); Feng, Dongju [Department of Microbiology and Immunology, Nanjing Medical University, Nanjing 210029, Jiangsu Province (China); Qin, Jian [College of Foreign Languages, Hehai University, Nanjing 210029, Jiangsu Province (China); Xie, Fangyi; Zhou, Feng; Chen, Yun; Wang, Jinfeng [Department of Microbiology and Immunology, Nanjing Medical University, Nanjing 210029, Jiangsu Province (China); Yao, Kun, E-mail: yaokun@njmu.edu.cn [Department of Microbiology and Immunology, Nanjing Medical University, Nanjing 210029, Jiangsu Province (China)

    2012-01-05

    Human herpesvirus-6 (HHV-6) is an important immunosuppressive and immunomodulatory virus that primarily infects immune cells (mainly CD4{sup +} T cells) and strongly suppresses the proliferation of infected cells. Toll-like receptors are pattern-recognition receptors essential for the development of an appropriate innate immune defense against infection. To understand the role of CD4{sup +} T cells in the innate response to HHV-6 infection and the involvement of TLRs, we used an in vitro infection model and observed that the infection of CD4{sup +} T cells resulted in the activation of JNK/SAPK via up-regulation of toll-like receptor 9 (TLR9). Associated with JNK activation, annexin V-PI staining indicated that HHV-6A was a strong inducer of apoptosis. Apoptotic response associated cytokines, IL-6 and TNF-{alpha} also induced by HHV-6A infection.

  20. Physical and functional association of the cbl protooncogen product with an src-family protein tyrosine kinase, p53/56lyn, in the B cell antigen receptor-mediated signaling.

    Science.gov (United States)

    Tezuka, T; Umemori, H; Fusaki, N; Yagi, T; Takata, M; Kurosaki, T; Yamamoto, T

    1996-02-01

    To identify novel signal transducers involved in signaling mediated by the Src-family protein tyrosine kinases (PTKs), we used a yeast two-hybrid system with a probe corresponding to the regulatory region of p56lyn, a member of Src-family PTKs. One of the isolated clones contained the COOH-terminal 470 amino acid residues of p120c-cbl, the product of the cellular homologue of the v-cbl retroviral oncogene. p120c-cbl is a cytoplasmic protein with nuclear protein-like motifs. Here we show in vivo association of p120c-cbl with p53/56lyn. After stimulation of the B cell antigen receptor (BCR), p120c-cbl was rapidly tyrosine phosphorylated. Studies with lyn- or syk-negative chicken B cells demonstrated that p53/56lyn, but not p72syk, was crucial for tyrosine phosphorylation of p120c-cbl upon stimulation of the BCR. We also show the importance of p59fyn in tyrosine phosphorylation of p120c-cbl in the T-cell receptor-mediated signaling using fyn-overexpressing T cell hybridomas and splenic T cells from fyn-deficient mice. These results suggest that p120c-cbl is an important substrate of Src-family PTKs in the intracellular signaling mediated by the antigen receptors PMID:8627181

  1. INHIBITION OF PROTEIN TYROSINE PHOSPHATASE ACTIVITY MEDIATES EPIDERMAL GROWTH FACTOR RECEPTOR SIGNALING IN HUMAN AIRWAY EPITHELIAL CELLS

    Science.gov (United States)

    Epidemiological studies have implicated zinc in the toxicity of ambient particulate matter (PM) inhalation. We previously showed that exposure to metal-laden PM inhibits protein tyrosine phosphatase (PTP) activity in human primary bronchial epithelial cells (HAEC) and leads t...

  2. Inflammation enhances myeloid-derived suppressor cell cross-talk by signaling through Toll-like receptor 4

    OpenAIRE

    Bunt, Stephanie K.; Clements, Virginia K.; Hanson, Erica M.; Sinha, Pratima; Ostrand-Rosenberg, Suzanne

    2009-01-01

    Myeloid-derived suppressor cells (MDSC) are potent inhibitors of anti-tumor immunity that facilitate tumor progression by blocking the activation of CD4+ and CD8+ T cells and by promoting a type 2 immune response through their production of IL-10 and down-regulation of macrophage production of IL-12. MDSC accumulate in many cancer patients and are a significant impediment to active cancer immunotherapies. Chronic inflammation has been shown recently to enhance the accumulation of MDSC and to ...

  3. Nitric oxide from inflammatory origin impairs neural stem cell proliferation by inhibiting epidermal growth factor receptor signaling

    OpenAIRE

    Bruno Pereira Carreira; Maria Inês Morte; Ana Isabel Santos; Ana Sofia Lourenço; António Francisco Ambrósio; Carvalho, Caetana M.; Araújo, Inês M.

    2014-01-01

    Neuroinflammation is characterized by activation of microglial cells, followed by production of nitric oxide (NO), which may have different outcomes on neurogenesis, favoring or inhibiting this process. In the present study, we investigated how the inflammatory mediator NO can affect proliferation of neural stem cells (NSCs), and explored possible mechanisms underlying this effect. We investigated which mechanisms are involved in the regulation of NSC proliferation following treatment with an...

  4. Isosilybin A Induces Apoptosis in Human Prostate Cancer Cells via Targeting Akt, NF-κB and Androgen Receptor Signaling

    OpenAIRE

    DEEP, GAGAN; Gangar, Subhash Chander; Oberlies, Nicholas H.; Kroll, David J.; Agarwal, Rajesh

    2010-01-01

    Prostate cancer (PCA) is second most malignancy in American men. Advanced stage PCA cells possess unlimited replication potential as well as resistance to apoptosis. Therefore, targeting survival mechanisms and activating apoptotic machinery in PCA cells using non-toxic phytochemicals is suggested as an attractive strategy against this deadly malignancy. In the present study, we assessed the effect of one such botanical agent, namely isosilybin A, on apoptotic machinery and key members of cel...

  5. Cell transformation mediated by the Epstein-Barr virus G protein-coupled receptor BILF1 is dependent on constitutive signaling

    DEFF Research Database (Denmark)

    Lyngaa, Rikke Birgitte; Nørregaard, K.; Kristensen, Martin; Kubale, V.; Rosenkilde, M.M.; Kledal, Thomas N

    2010-01-01

    Epstein-Barr virus (EBV) open reading frame BILF1 encodes a seven trans-membrane (TM) G protein-coupled receptor that signals with high constitutive activity through G alpha(i) (Beisser et al., 2005; Paulsen et al., 2005). In this paper, the transforming potential of BILF1 is investigated in vitro...... an intermediately active triple-mutated receptor possessed an intermediate transforming potential. Furthermore, BILF1 expression induced vascular endothelial growth factor secretion in a constitutively active manner. In nude mice, BILF1 promoted tumor formation in 90% of cases, ORF74 (from Kaposi......'s sarcoma-associated herpes virus) in 100% of cases, whereas the signaling-deficient receptor resulted in tumor establishment in 40% of cases. These data suggest that BILF1, when expressed during EBV infection, could indeed be involved in the pathogenesis of EBV-associated diseases and malignancies...

  6. Escherichia coli Nissle 1917 Distinctively Modulates T-Cell Cycling and Expansion via Toll-Like Receptor 2 Signaling

    OpenAIRE

    Sturm, Andreas; Rilling, Klaus; Baumgart, Daniel C.; Gargas, Konstantinos; Abou-Ghazalé, Tay; Raupach, Bärbel; Eckert, Jana; Schumann, Ralf R.; Enders, Corinne; Sonnenborn, Ulrich; Wiedenmann, Bertram; Dignass, Axel U

    2005-01-01

    Although the probiotic Escherichia coli strain Nissle 1917 has been proven to be efficacious for the treatment of inflammatory bowel diseases, the underlying mechanisms of action still remain elusive. The aim of the present study was to analyze the effects of E. coli Nissle 1917 on cell cycling and apoptosis of peripheral blood and lamina propria T cells (PBT and LPT, respectively). Anti-CD3-stimulated PBT and LPT were treated with E. coli Nissle 1917-conditioned medium (E. coli Nissle 1917-C...

  7. Leukocyte transmigration into tissue-engineered constructs is influenced by endothelial cells through Toll-like receptor signaling

    OpenAIRE

    Bartaula-Brevik, Sushma; Pedersen, Torbjorn Østvik; Blois, Anna L; Papadakou, Panagiota; Finne-Wistrand, Anna; Xue, Ying; Bolstad, Anne Isine; Mustafa, Kamal Babikeir Eln

    2014-01-01

    Introduction Inflammation plays a crucial role in tissue regeneration, wound healing, and the success of tissue-engineered constructs. The aim of this study was to investigate the influence of human umbilical vein endothelial cells (ECs) on leukocyte transmigration when co-cultured with primary human bone marrow-derived multipotent stromal cells (MSCs). Methods MSCs with and without ECs were cultured in poly (L-lactide-co-1, 5-dioxepan-2-one) (poly (LLA-co-DXO)) scaffolds for 1 week in vit...

  8. Radically altered T cell receptor signaling in glycopeptide-specific T cell hybridoma induced by antigen with minimal differences in the glycan group

    DEFF Research Database (Denmark)

    Jensen, T; Nielsen, M; Gad, Monika;

    2001-01-01

    A T cell hybridoma raised against the synthetic glycopeptide T(72)(Tn) was used to study whether the initial TCR signaling events are markedly different when the hybridoma is stimulated with glycopeptides closely related to the cognate glycopeptide antigen. T(72)(Tn) has an alpha-D-GalNAc group O......-linked to the central threonine in the decapeptide VITAFTEGLK, and the hybridoma is known to be highly specific for this carbohydrate group. T(72)(Tn)-pulsed APC induced tyrosine phosphorylation of the TCR-zeta 21- and 23-kDa proteins and the downstream p42/44 MAP kinase and strong IL-2 secretion. APC...

  9. Characterization of prolactin-releasing peptide: Binding, signaling and hormone secretion in rodent pituitary cell lines endogenously expressing its receptor

    Czech Academy of Sciences Publication Activity Database

    Maixnerová, Jana; Špolcová, Andrea; Pýchová, Miroslava; Blechová, Miroslava; Elbert, Tomáš; Řezáčová, M.; Železná, Blanka; Maletínská, Lenka

    2011-01-01

    Roč. 32, č. 4 (2011), s. 811-817. ISSN 0196-9781 R&D Projects: GA ČR GAP303/10/1368 Institutional research plan: CEZ:AV0Z40550506 Keywords : PrRP * pituitary cell lines * food intake Subject RIV: CC - Organic Chemistry Impact factor: 2.434, year: 2011

  10. Hinokitiol, a metal chelator derived from natural plants, suppresses cell growth and disrupts androgen receptor signaling in prostate carcinoma cell lines

    International Nuclear Information System (INIS)

    Hinokitiol (β-thujaplicin), a troplone-related compound found in the heartwood of cupressaceous plants, strongly inhibits the proliferation of a broad range of tumor cell lines. This is the first report to demonstrate that hinokitiol, a metal chelator derived from natural plants, suppresses cell growth and disrupts AR signaling in prostate carcinoma cell lines. Our present studies indicate that hinokitiol suppresses androgen/AR-mediated cell growth and androgen-stimulated DNA synthesis by [3H]thymidine incorporation in a dose- and time-dependent manner. Hinokitiol simultaneously suppresses the intracellular and secreted PSA levels, a marker for the progression of prostate cancer. Hinokitiol significantly represses the AR mRNA and protein expression in a dose- and time-dependent manner. Additionally, the ligand-binding assay shows that hinokitiol blocks binding of the synthetic androgen [3H]R1881 to AR in LNCaP cells. These findings collectively suggest that hinokitiol is potentially effective against prostate cancer in vitro, and thus it might become a novel chemopreventive or chemotherapeutic agent for prostate cancer

  11. Characterization of Thyrotropin Receptor Antibody-Induced Signaling Cascades

    OpenAIRE

    Morshed, Syed A.; Latif, Rauf; Davies, Terry F.

    2008-01-01

    The TSH receptor (TSHR) is constitutively active and is further enhanced by TSH ligand binding or by stimulating TSHR antibodies (TSHR-Abs) as seen in Graves’ disease. TSH is known to activate the thyroid epithelial cell via both Gαs-cAMP/protein kinase A/ERK and Gαq-Akt/protein kinase C coupled signaling networks. The recent development of monoclonal antibodies to the TSHR has enabled us to investigate the hypothesis that different TSHR-Abs may have unique signaling imprints that differ from...

  12. JAK-STAT and G-protein-coupled receptor signaling pathways are frequently altered in epitheliotropic intestinal T-cell lymphoma.

    Science.gov (United States)

    Nairismägi, M-L; Tan, J; Lim, J Q; Nagarajan, S; Ng, C C Y; Rajasegaran, V; Huang, D; Lim, W K; Laurensia, Y; Wijaya, G C; Li, Z M; Cutcutache, I; Pang, W L; Thangaraju, S; Ha, J; Khoo, L P; Chin, S T; Dey, S; Poore, G; Tan, L H C; Koh, H K M; Sabai, K; Rao, H-L; Chuah, K L; Ho, Y-H; Ng, S-B; Chuang, S-S; Zhang, F; Liu, Y-H; Pongpruttipan, T; Ko, Y H; Cheah, P-L; Karim, N; Chng, W-J; Tang, T; Tao, M; Tay, K; Farid, M; Quek, R; Rozen, S G; Tan, P; Teh, B T; Lim, S T; Tan, S-Y; Ong, C K

    2016-06-01

    Epitheliotropic intestinal T-cell lymphoma (EITL, also known as type II enteropathy-associated T-cell lymphoma) is an aggressive intestinal disease with poor prognosis and its molecular alterations have not been comprehensively characterized. We aimed to identify actionable easy-to-screen alterations that would allow better diagnostics and/or treatment of this deadly disease. By performing whole-exome sequencing of four EITL tumor-normal pairs, followed by amplicon deep sequencing of 42 tumor samples, frequent alterations of the JAK-STAT and G-protein-coupled receptor (GPCR) signaling pathways were discovered in a large portion of samples. Specifically, STAT5B was mutated in a remarkable 63% of cases, JAK3 in 35% and GNAI2 in 24%, with the majority occurring at known activating hotspots in key functional domains. Moreover, STAT5B locus carried copy-neutral loss of heterozygosity resulting in the duplication of the mutant copy, suggesting the importance of mutant STAT5B dosage for the development of EITL. Dysregulation of the JAK-STAT and GPCR pathways was also supported by gene expression profiling and further verified in patient tumor samples. In vitro overexpression of GNAI2 mutants led to the upregulation of pERK1/2, a member of MEK-ERK pathway. Notably, inhibitors of both JAK-STAT and MEK-ERK pathways effectively reduced viability of patient-derived primary EITL cells, indicating potential therapeutic strategies for this neoplasm with no effective treatment currently available. PMID:26854024

  13. Thyroid Hormone Receptor-β (TRβ) Mediates Runt-Related Transcription Factor 2 (Runx2) Expression in Thyroid Cancer Cells: A Novel Signaling Pathway in Thyroid Cancer.

    Science.gov (United States)

    Carr, Frances E; Tai, Phillip W L; Barnum, Michael S; Gillis, Noelle E; Evans, Katherine G; Taber, Thomas H; White, Jeffrey H; Tomczak, Jennifer A; Jaworski, Diane M; Zaidi, Sayyed K; Lian, Jane B; Stein, Janet L; Stein, Gary S

    2016-08-01

    Dysregulation of the thyroid hormone receptor (TR)β is common in human cancers. Restoration of functional TRβ delays tumor progression in models of thyroid and breast cancers implicating TRβ as a tumor suppressor. Conversely, aberrant expression of the runt-related transcription factor 2 (Runx2) is established in the progression and metastasis of thyroid, breast, and other cancers. Silencing of Runx2 diminishes tumor invasive characteristics. With TRβ as a tumor suppressor and Runx2 as a tumor promoter, a compelling question is whether there is a functional relationship between these regulatory factors in thyroid tumorigenesis. Here, we demonstrated that these proteins are reciprocally expressed in normal and malignant thyroid cells; TRβ is high in normal cells, and Runx2 is high in malignant cells. T3 induced a time- and concentration-dependent decrease in Runx2 expression. Silencing of TRβ by small interfering RNA knockdown resulted in a corresponding increase in Runx2 and Runx2-regulated genes, indicating that TRβ levels directly impact Runx2 expression and associated epithelial to mesenchymal transition molecules. TRβ specifically bound to 3 putative thyroid hormone-response element motifs within the Runx2-P1 promoter ((-)105/(+)133) as detected by EMSA and chromatin immunoprecipitation. TRβ suppressed Runx2 transcriptional activities, thus confirming TRβ regulation of Runx2 at functional thyroid hormone-response elements. Significantly, these findings indicate that a ratio of the tumor-suppressor TRβ and tumor-promoting Runx2 may reflect tumor aggression and serve as biomarkers in biopsy tissues. The discovery of this TRβ-Runx2 signaling supports the emerging role of TRβ as a tumor suppressor and reveals a novel pathway for intervention. PMID:27253998

  14. Sweet taste receptor expressed in pancreatic beta-cells activates the calcium and cyclic AMP signaling systems and stimulates insulin secretion.

    Directory of Open Access Journals (Sweden)

    Yuko Nakagawa

    Full Text Available BACKGROUND: Sweet taste receptor is expressed in the taste buds and enteroendocrine cells acting as a sugar sensor. We investigated the expression and function of the sweet taste receptor in MIN6 cells and mouse islets. METHODOLOGY/PRINCIPAL FINDINGS: The expression of the sweet taste receptor was determined by RT-PCR and immunohistochemistry. Changes in cytoplasmic Ca(2+ ([Ca(2+](c and cAMP ([cAMP](c were monitored in MIN6 cells using fura-2 and Epac1-camps. Activation of protein kinase C was monitored by measuring translocation of MARCKS-GFP. Insulin was measured by radioimmunoassay. mRNA for T1R2, T1R3, and gustducin was expressed in MIN6 cells. In these cells, artificial sweeteners such as sucralose, succharin, and acesulfame-K increased insulin secretion and augmented secretion induced by glucose. Sucralose increased biphasic increase in [Ca(2+](c. The second sustained phase was blocked by removal of extracellular calcium and addition of nifedipine. An inhibitor of inositol(1, 4, 5-trisphophate receptor, 2-aminoethoxydiphenyl borate, blocked both phases of [Ca(2+](c response. The effect of sucralose on [Ca(2+](c was inhibited by gurmarin, an inhibitor of the sweet taste receptor, but not affected by a G(q inhibitor. Sucralose also induced sustained elevation of [cAMP](c, which was only partially inhibited by removal of extracellular calcium and nifedipine. Finally, mouse islets expressed T1R2 and T1R3, and artificial sweeteners stimulated insulin secretion. CONCLUSIONS: Sweet taste receptor is expressed in beta-cells, and activation of this receptor induces insulin secretion by Ca(2+ and cAMP-dependent mechanisms.

  15. Steroid Hormone Receptor Signals as Prognosticators for Urothelial Tumor

    Directory of Open Access Journals (Sweden)

    Hiroki Ide

    2015-01-01

    Full Text Available There is a substantial amount of preclinical or clinical evidence suggesting that steroid hormone receptor-mediated signals play a critical role in urothelial tumorigenesis and tumor progression. These receptors include androgen receptor, estrogen receptors, glucocorticoid receptor, progesterone receptor, vitamin D receptor, retinoid receptors, peroxisome proliferator-activated receptors, and others including orphan receptors. In particular, studies using urothelial cancer tissue specimens have demonstrated that elevated or reduced expression of these receptors as well as alterations of their upstream or downstream pathways correlates with patient outcomes. This review summarizes and discusses available data suggesting that steroid hormone receptors and related signals serve as biomarkers for urothelial carcinoma and are able to predict tumor recurrence or progression.

  16. Cytokine signalling in embryonic stem cells

    DEFF Research Database (Denmark)

    Kristensen, David Møbjerg; Kalisz, Mark; Nielsen, Jens Høiriis

    2006-01-01

    Cytokines play a central role in maintaining self-renewal in mouse embryonic stem (ES) cells through a member of the interleukin-6 type cytokine family termed leukemia inhibitory factor (LIF). LIF activates the JAK-STAT3 pathway through the class I cytokine receptor gp130, which forms a trimeric...... pathways seem to converge on c-myc as a common target to promote self-renewal. Whereas LIF does not seem to stimulate self-renewal in human embryonic stem cells it cannot be excluded that other cytokines are involved. The pleiotropic actions of the increasing number of cytokines and receptors signalling...... via JAKs, STATs and SOCS exhibit considerable redundancy, compensation and plasticity in stem cells in accordance with the view that stem cells are governed by quantitative variations in strength and duration of signalling events known from other cell types rather than qualitatively different stem...

  17. Cell death sensitization of leukemia cells by opioid receptor activation

    Science.gov (United States)

    Friesen, Claudia; Roscher, Mareike; Hormann, Inis; Fichtner, Iduna; Alt, Andreas; Hilger, Ralf A.; Debatin, Klaus-Michael; Miltner, Erich

    2013-01-01

    Cyclic AMP (cAMP) regulates a number of cellular processes and modulates cell death induction. cAMP levels are altered upon stimulation of specific G-protein-coupled receptors inhibiting or activating adenylyl cyclases. Opioid receptor stimulation can activate inhibitory Gi-proteins which in turn block adenylyl cyclase activity reducing cAMP. Opioids such as D,L-methadone induce cell death in leukemia cells. However, the mechanism how opioids trigger apoptosis and activate caspases in leukemia cells is not understood. In this study, we demonstrate that downregulation of cAMP induced by opioid receptor activation using the opioid D,L-methadone kills and sensitizes leukemia cells for doxorubicin treatment. Enhancing cAMP levels by blocking opioid-receptor signaling strongly reduced D,L-methadone-induced apoptosis, caspase activation and doxorubicin-sensitivity. Induction of cell death in leukemia cells by activation of opioid receptors using the opioid D,L-methadone depends on critical levels of opioid receptor expression on the cell surface. Doxorubicin increased opioid receptor expression in leukemia cells. In addition, the opioid D,L-methadone increased doxorubicin uptake and decreased doxorubicin efflux in leukemia cells, suggesting that the opioid D,L-methadone as well as doxorubicin mutually increase their cytotoxic potential. Furthermore, we found that opioid receptor activation using D,L-methadone alone or in addition to doxorubicin inhibits tumor growth significantly in vivo. These results demonstrate that opioid receptor activation via triggering the downregulation of cAMP induces apoptosis, activates caspases and sensitizes leukemia cells for doxorubicin treatment. Hence, opioid receptor activation seems to be a promising strategy to improve anticancer therapies. PMID:23633472

  18. Novel method for the study of receptor Ca2+ signalling exemplified by the NK1 receptor

    DEFF Research Database (Denmark)

    Heding, A; Elling, C E; Schwartz, T W

    We have used a novel technology (NovoStar from BMG Labtechnologies) for the study of the Ca2+ signalling of the human tackykinin NK1 (hNK-I receptor). The NovoStar is a microplate reader based on fluorescence and luminescence. The instrument implements a robotic pipettor arm and two microplate...... carriers, typically one for samples and one for cells. The robotic pipettor arm can transfer sample (agonist or antagonist) from the sample plate or other liquid containers to the cell plate, facilitating the study of Ca2+ signalling to such a degree that the instrument can be used for Medium Throughput...

  19. Prostaglandin EP2 receptor signaling protects human trabecular meshwork cells from apoptosis induced by ER stress through down-regulation of p53.

    Science.gov (United States)

    Kalouche, Georges; Boucher, Céline; Coste, Annick; Debussche, Laurent; Orsini, Cécile; Baudouin, Christophe; Debeir, Thomas; Vigé, Xavier; Rostène, William

    2016-09-01

    E-prostanoid receptor subtype 2 (EP2) agonists are currently under clinical development as hypotensive agents for the treatment of ocular hypertension. However, the effects of EP2 receptor agonists on trabecular meshwork (TM) alterations leading to primary open-angle glaucoma (POAG) are still unknown. Here, we evaluated whether EP2 receptor activation exhibits protective functions on TM cell death induced by endoplasmic reticulum (ER) stress. We show that the EP2 receptor agonist butaprost protects TM cell death mediated by the ER stress inducer tunicamycin through a cyclic AMP (cAMP)-dependent mechanism, but independent of the classical cAMP sensors, protein kinase A and exchange proteins activated by cAMP. The ER stress-induced intrinsic apoptosis inhibited by the EP2 receptor agonist was correlated with a decreased accumulation of the cellular stress sensor p53. In addition, p53 down-regulation was associated with inhibition of its transcriptional activity, which led to decreased expression of the pro-apoptotic p53-upregulated modulator of apoptosis (PUMA). The stabilization of p53 by nutlin-3a abolished butaprost-mediated cell death protection. In conclusion, we showed that EP2 receptor activation protects against ER stress-dependent mitochondrial apoptosis through down-regulation of p53. The specific inhibition of this pathway could reduce TM alterations observed in POAG patients. PMID:27321910

  20. The Role of Cgrp-Receptor Component Protein (Rcp in Cgrp-Mediated Signal Transduction

    Directory of Open Access Journals (Sweden)

    M. A. Prado

    2001-01-01

    Full Text Available The calcitonin gene-related peptide (CGRP-receptor component protein (RCP is a 17-kDa intracellular peripheral membrane protein required for signal transduction at CGRP receptors. To determine the role of RCP in CGRP-mediated signal transduction, RCP was depleted from NIH3T3 cells using antisense strategy. Loss of RCP protein correlated with loss of cAMP production by CGRP in the antisense cells. In contrast, loss of RCP had no effect on CGRP-mediated binding; therefore RCP is not acting as a chaperone for the CGRP receptor. Instead, RCP is a novel signal transduction molecule that couples the CGRP receptor to the cellular signal transduction machinery. RCP thus represents a prototype for a new class of signal transduction proteins that are required for regulation of G protein-coupled receptors.

  1. Deletion of angiotensin II type 2 receptor accelerates adipogenesis in murine mesenchymal stem cells via Wnt10b/beta-catenin signaling.

    Science.gov (United States)

    Matsushita, Kenichi; Wu, Yaojiong; Pratt, Richard E; Dzau, Victor J

    2016-08-01

    Recent evidence suggests that the renin-angiotensin system (RAS) has a vital role in adipocyte biology and the pathophysiology of metabolic syndrome. Obesity is the main culprit of metabolic syndrome; and mesenchymal stem cells (MSCs) have been forwarded as a major source of adipocyte generation. Previously, we reported that MSCs have a local RAS and that pharmacological blockade of angiotensin II type 2 receptor (AT2R) promotes adipogenesis in human MSCs. However, the definitive roles of AT2R and how AT2R functions in adipogenesis remains unknown. To this end, we employed AT2R-null murine MSCs to characterize how AT2R affects the differentiation of MSCs into adipocytes. Murine MSCs were isolated from AT2R-null mice and wild-type littermates, grown to confluency, and then differentiated into adipocytes. Adipogenesis was quantitated by assessing the lipid droplet accumulation. Using the lipophilic fluorescent dye, the AT2R-null cells showed significantly increased total fluorescence (261.6±49.6% vs littermate) on day 7. Oil red O staining followed by extraction of the absorbed dye and measurement of the absorbance on day 14 also exhibited significantly increased lipid droplet accumulation in the AT2R-null cells (202.7±14.1% vs littermate). We also examined the expression of adipogenic marker genes by quantitative RT-PCR. The AT2R-null group exhibited significantly increased expression of PPAR-gamma, fatty acid synthase, and adiponectin (vs littermate). We further examined the role of Wnt10b/beta-catenin signaling, which reportedly has an important inhibitory role in adipogenesis. The AT2R-null group exhibited significantly decreased Wnt10b expression accompanied by decreased beta-catenin (vs littermate). Our results thus revealed that the AT2R inhibits adipogenic differentiation in murine MSCs. Moreover, this inhibitory effect is associated with Wnt10b/beta-catenin signaling. These results provide important insights into the pathophysiology of obesity and obesity

  2. Expression of Functional Sphingosine-1 Phosphate Receptor-1 Is Reduced by B Cell Receptor Signaling and Increased by Inhibition of PI3 Kinase δ but Not SYK or BTK in Chronic Lymphocytic Leukemia Cells

    OpenAIRE

    Till, Kathleen J.; Andrew R Pettitt; Slupsky, Joseph R.

    2015-01-01

    BCR signaling pathway inhibitors such as ibrutinib, idelalisib, and fostamatinib (respective inhibitors of Bruton’s tyrosine kinase, PI3Kδ, and spleen tyrosine kinase) represent a significant therapeutic advance in B cell malignancies, including chronic lymphocytic leukemia (CLL). These drugs are distinctive in increasing blood lymphocytes while simultaneously shrinking enlarged lymph nodes, suggesting anatomical redistribution of CLL cells from lymph nodes into the blood. However, the mechan...

  3. Gene expression changes induced by estrogen and selective estrogen receptor modulators in primary-cultured human endometrial cells: signals that distinguish the human carcinogen tamoxifen

    International Nuclear Information System (INIS)

    Tamoxifen has long been the endocrine treatment of choice for women with breast cancer and is now employed for prophylactic use in women at high risk from breast cancer. Other selective estrogen receptor modulators (SERMs), such as raloxifene, mimic some of tamoxifen's beneficial effects and, like tamoxifen, exhibit a complex mixture of organ-specific estrogen agonist and antagonistic properties. However, accompanying the positive effects of tamoxifen has been the emergence of evidence for an increased risk of endometrial cancer associated with its use. A more complete understanding of the mechanism(s) of SERM carcinogenicity and endometrial effects is therefore required. We have sought to compare and characterise the transcript profile of tamoxifen, raloxifene and the agonist estradiol in human endometrial cells. Using primary cultures of human endometria, to best emulate the in vivo responses in a manageable in vitro system, we have shown 230 significant changes in gene expression for epithelial cultures and 83 in stromal cultures, either specific to 17β-estradiol, tamoxifen or raloxifene, or changed across more than one of the treatments. Considering the transcriptome as a whole, the endometrial responses to raloxifene or tamoxifen were more similar than either drug was to 17β-estradiol. Treatment of endometrial cultures with tamoxifen resulted in the largest number of gene changes relative to control cultures and a high proportion of genes associated with regulation of gene transcription, cell-cycle control and signal transduction. Tamoxifen-specific changes that might point towards mechanisms for its proliferative response in the endometrium included changes in retinoblastoma and c-myc binding proteins, the APCL, dihydrofolate reductase (DHFR) and E2F1 genes and other transcription factors. Tamoxifen was also found to give rise to the highest number of gene expression changes common to those that characterise malignant endometria. It is anticipated that this

  4. A Stochastic Model of the Germinal Center Integrating Local Antigen Competition, Individualistic T-B Interactions, and B Cell Receptor Signaling.

    Science.gov (United States)

    Wang, Peng; Shih, Chang-Ming; Qi, Hai; Lan, Yue-Heng

    2016-08-15

    The germinal center (GC) reaction underlies productive humoral immunity by orchestrating competition-based affinity maturation to produce plasma cells and memory B cells. T cells are limiting in this process. How B cells integrate signals from T cells and BCRs to make fate decisions while subjected to a cyclic selection process is not clear. In this article, we present a spatiotemporally resolved stochastic model that describes cell behaviors as rate-limited stochastic reactions. We hypothesize a signal integrator protein integrates follicular helper T (Tfh)- and Ag-derived signals to drive different B cell fates in a probabilistic manner and a dedicated module of Tfh interaction promoting factors control the efficiency of contact-dependent Tfh help delivery to B cells. Without assuming deterministic affinity-based decisions or temporal event sequence, this model recapitulates GC characteristics, highlights the importance of efficient T cell help delivery during individual contacts with B cells and intercellular positive feedback for affinity maturation, reveals the possibility that antagonism between BCR signaling and T cell help accelerates affinity maturation, and suggests that the dichotomy between affinity and magnitude of GC reaction can be avoided by tuning the efficiency of contact-dependent help delivery during reiterative T-B interactions. PMID:27421481

  5. Signal transduction and chemotaxis in mast cells.

    Science.gov (United States)

    Draber, Petr; Halova, Ivana; Polakovicova, Iva; Kawakami, Toshiaki

    2016-05-01

    Mast cells play crucial roles in both innate and adaptive arms of the immune system. Along with basophils, mast cells are essential effector cells for allergic inflammation that causes asthma, allergic rhinitis, food allergy and atopic dermatitis. Mast cells are usually increased in inflammatory sites of allergy and, upon activation, release various chemical, lipid, peptide and protein mediators of allergic reactions. Since antigen/immunoglobulin E (IgE)-mediated activation of these cells is a central event to trigger allergic reactions, innumerable studies have been conducted on how these cells are activated through cross-linking of the high-affinity IgE receptor (FcεRI). Development of mature mast cells from their progenitor cells is under the influence of several growth factors, of which the stem cell factor (SCF) seems to be the most important. Therefore, how SCF induces mast cell development and activation via its receptor, KIT, has been studied extensively, including a cross-talk between KIT and FcεRI signaling pathways. Although our understanding of the signaling mechanisms of the FcεRI and KIT pathways is far from complete, pharmaceutical applications of the knowledge about these pathways are underway. This review will focus on recent progresses in FcεRI and KIT signaling and chemotaxis. PMID:25941081

  6. Palmitoylation of Estrogen Receptors Is Essential for Neuronal Membrane Signaling

    OpenAIRE

    Meitzen, John; Luoma, Jessie I.; Boulware, Marissa I.; Hedges, Valerie L.; Peterson, Brittni M.; Tuomela, Krista; Britson, Kyla A.; Mermelstein, Paul G.

    2013-01-01

    In addition to activating nuclear estrogen receptor signaling, 17β-estradiol can also regulate neuronal function via surface membrane receptors. In various brain regions, these actions are mediated by the direct association of estrogen receptors (ERs) activating metabotropic glutamate receptors (mGluRs). These ER/mGluR signaling partners are organized into discrete functional microdomains via caveolin proteins. A central question that remains concerns the underlying mechanism by which these s...

  7. Ric-8A, a Gα protein guanine nucleotide exchange factor potentiates taste receptor signaling

    Directory of Open Access Journals (Sweden)

    Claire J Fenech

    2009-10-01

    Full Text Available Taste receptors for sweet, bitter and umami tastants are G-protein coupled receptors (GPCRs. While much effort has been devoted to understanding G-protein-receptor interactions and identifying the components of the signalling cascade downstream of these receptors, at the level of the G-protein the modulation of receptor signal transduction remains relatively unexplored. In this regard a taste-specific regulator of G-protein signaling (RGS, RGS21, has recently been identified. To study whether guanine nucleotide exchange factors (GEFs are involved in the transduction of the signal downstream of the taste GPCRs we investigated the expression of Ric-8A and Ric-8B in mouse taste cells and their interaction with G-protein subunits found in taste buds. Mammalian Ric-8 proteins were initially identified as potent GEFs for a range of Gα subunits and Ric-8B has recently been shown to amplify olfactory signal transduction. We find that both Ric-8A and Ric-8B are expressed in a large portion of taste bud cells and that most of these cells contain IP3R-3 a marker for sweet, umami and bitter taste receptor cells. Ric-8A interacts with Gα-gustducin and Gαi2 through which it amplifies the signal transduction of hTas2R16, a receptor for bitter compounds. Overall, these findings are consistent with a role for Ric-8 in mammalian taste signal transduction.

  8. Recent advances in the Okamoto model: the CD38-cyclic ADP-ribose signal system and the regenerating gene protein (Reg)-Reg receptor system in beta-cells.

    Science.gov (United States)

    Okamoto, Hiroshi; Takasawa, Shin

    2002-12-01

    Twenty years ago, we first proposed our hypothesis on beta-cell damage and its prevention (the Okamoto model), according to which poly(ADP-ribose) synthetase/polymerase (PARP) activation is critically involved in the consumption of NAD(+), leading to energy depletion and cell death by necrosis. Recently, the model was reconfirmed by results using PARP knockout mice and has been recognized as providing the basis for necrotic death of various cells and tissues. Based on the model, we proposed two signal systems in beta-cells: one is the CD38-cyclic ADP-ribose (cADPR) signal system for insulin secretion, and the other is the regenerating gene protein (Reg)-Reg receptor system for beta-cell regeneration. The physiological and pathological significance of the two signal systems in a variety of cells and tissues as well as in pancreatic beta-cells has recently been recognized. Here, we describe the Okamoto model and its descendents, the CD38-cADPR signal system and the Reg-Reg receptor system, focusing on recent advances and how their significance came to light. Because PARP is involved in Reg gene transcription to induce beta-cell regeneration, and the PARP activation reduces the cellular NAD(+) to decrease the formation of cADPR (a second messenger for insulin secretion) and further to cause necrotic beta-cell death, PARP and its inhibitors have key roles in the induction of beta-cell regeneration, the maintenance of insulin secretion, and the prevention of beta-cell death. PMID:12475791

  9. Opioid receptors: Structural and mechanistic insights into pharmacology and signaling.

    Science.gov (United States)

    Shang, Yi; Filizola, Marta

    2015-09-15

    Opioid receptors are important drug targets for pain management, addiction, and mood disorders. Although substantial research on these important subtypes of G protein-coupled receptors has been conducted over the past two decades to discover ligands with higher specificity and diminished side effects, currently used opioid therapeutics remain suboptimal. Luckily, recent advances in structural biology of opioid receptors provide unprecedented insights into opioid receptor pharmacology and signaling. We review here a few recent studies that have used the crystal structures of opioid receptors as a basis for revealing mechanistic details of signal transduction mediated by these receptors, and for the purpose of drug discovery. PMID:25981301

  10. Inhibition of insulin-like growth factor-1 receptor signaling enhances growth-inhibitory and proapoptotic effects of gefitinib (Iressa) in human breast cancer cells

    OpenAIRE

    Camirand, Anne; Zakikhani, Mahvash; Young, Fiona; Pollak, Michael

    2005-01-01

    Introduction Gefitinib (Iressa, ZD 1839, AstraZeneca) blocks the tyrosine kinase activity of the epidermal growth factor receptor (EGFR) and inhibits proliferation of several human cancer cell types including breast cancer. Phase II clinical trials with gefitinib monotherapy showed an objective response of 9 to 19% in non-small-cell lung cancer patients and less than 10% for breast cancer, and phase III results have indicated no benefit of gefitinib in combination with chemotherapy over chemo...

  11. 受体的适应性变化及细胞信号转导规律与运动的联系%Association of receptor adaptative change and cell signal transduction with exercise

    Institute of Scientific and Technical Information of China (English)

    李宏伟; 狄朝晖

    2011-01-01

    背景:受体调控和转换信号并启动细胞内信号转导,使靶细胞产生生物学效应.受体调节是机体对运动适应的重要调节方式.目的:总结运动对受体的影响、受体的适应性变化及运动受体细胞信号转导规律.方法:由第一作者用计算机检索中国期刊全文数据库(CNKI:2000/2010)和Medline数据库(2000/2010),检索词分别为"受体,运动,适应"和"receptor regulation,exercise,adaption".共检索到137篇文章,按纳入和排除标准对文献进行筛选,共纳入37篇文章.从受体概念、受体的分类和功能,受体的功能与调节,运动对受体的影响等方面进行总结,对受体在运动中的适应性变化和调节机制等方面进行探讨.结果与结论:运动训练引起瘦素受体和胰岛素受体上调,使瘦素和胰岛素分泌发生适应性改变.适宜运动使雄激素受体结合容量和受体数量提高,长期大运动量训练和力竭使雄激素受体结合容量和受体数量下降,意味着保护细胞免受过量或长期刺激而导致生理功能紊乱.急性应激使机体各组织中糖皮质激素受体含量减少,对于防止物质代谢和能量代谢紊乱,维持机体内环境稳定有重要意义.%BACKGROUND: The receptor regulates and switches signal and starts intracellular signal transduction, which causes target cellsto produce biological effects. Receptor regulation is an important regulation mode for organism to motor adaptation.OBJECTIVE: To summarize the effects of motor on receptor, the adaptati ve change of receptor, and signal transduction rule ofmotor receptor cells.METHODS: A computer-based retrieval was performed by the first author to search manuscripts in the CNKI (2000/2010) andMedline database (2000/2010) with the key word “receptor regulation, exercise, adaption”. A total of 137 manuscripts wereincluded. According to inclusion and exclusion criteria, totally 37 manuscripts were included in the final analysis. The

  12. Distribution of natural killer cell receptors in HIV infected individuals

    Institute of Scientific and Technical Information of China (English)

    JIANG Yong-jun; SHANG Hong; ZHANG Zi-ning; DIAO Ying-ying; GENG Wen-qing; DAI Di; LIU Jing; WANG Ya-nan; ZHANG Min; HAN Xiao-xu

    2007-01-01

    @@ Natural killer (NK) cells are bone marrow derived,large granular lymphocytes, comprising approximately 10% to 20% of the mononuclear cell fraction in normal peripheral blood. They form a part of the first line defense mechanism against tumoural and viral spreading.1-4 Unlike T and B cells, NK cells do not require gene rearrangement for assembly of their receptor genes; rather, NK cells discriminate potential target cells based on the levels of self major histocompatibility complex (MHC) class Ⅰ expression on such cells.5,6 There are two kinds of NK cell receptors.2,7,8 Inhibitory receptors recognize MHC class Ⅰ molecules and deliver a downregulatory signal that inactivates the lyric machinery of NK cells. Stimulatory receptors expressed by NK cells deliver an activation signal.

  13. Activation of histamine H4 receptors decreases epithelial-to-mesenchymal transition progress by inhibiting transforming growth factor-β1 signalling pathway in non-small cell lung cancer.

    Science.gov (United States)

    Cai, Wen-Ke; Hu, Jing; Li, Teng; Meng, Jing-Ru; Ma, Xue; Yin, Sun-Jun; Zhao, Can-Hu; He, Gong-Hao; Xu, Gui-Li

    2014-04-01

    Previous investigations found that epithelial-to-mesenchymal transition (EMT) was an important character of non-small cell lung cancer (NSCLC) and it was also suggested that histamine H4 receptors may have a role in preventing EMT progress in certain kind of tumours. However, the effect of H4 receptor activation on EMT progress of NSCLC and its potential mechanisms remain unclear. Therefore, we performed both in vitro and in vivo experiments to explore the effects of specific H4 receptor agonist 4-methylhistamine and antagonist JNJ7777120 on EMT progress. We showed the expression of H4 receptors in NSCLC and found that 4-methylhistamine increased the expression of the epithelial marker E-cadherin and decreased the expression of Vimentin, the mesenchymal marker, in both NSCLC cell lines and xenograft NSCLC tumours. Pretreatment with JNJ7777120 or H4 receptor gene silencing decreased while overexpression of H4 receptors facilitated this effect of 4-methylhistamine. Furthermore, we showed that down-regulation of cyclic adenosine monophosphate (cAMP) was the secondary signalling after H4 receptor activation, which in turn resulted in inactivation of transforming growth factor-β1 (TGF-β1) pathway and down-regulation of several important EMT inducing factors such as ZEB1, Snail and Slug. In conclusion, these findings revealed the anti-EMT effect of histamine H4 receptor activation in NSCLC, which provide novel insight into the development mechanism of NSCLC; and H4 receptors may be a new therapeutic target for NSCLC treatment. PMID:24447834

  14. HBpF-proBDNF: A New Tool for the Analysis of Pro-Brain Derived Neurotrophic Factor Receptor Signaling and Cell Biology.

    Science.gov (United States)

    Gaub, Perrine; de Léon, Andrès; Gibon, Julien; Soubannier, Vincent; Dorval, Geneviève; Séguéla, Philippe; Barker, Philip A

    2016-01-01

    Neurotrophins activate intracellular signaling pathways necessary for neuronal survival, growth and apoptosis. The most abundant neurotrophin in the adult brain, brain-derived neurotrophic factor (BDNF), is first synthesized as a proBDNF precursor and recent studies have demonstrated that proBDNF can be secreted and that it functions as a ligand for a receptor complex containing p75NTR and sortilin. Activation of proBDNF receptors mediates growth cone collapse, reduces synaptic activity, and facilitates developmental apoptosis of motoneurons but the precise signaling cascades have been difficult to discern. To address this, we have engineered, expressed and purified HBpF-proBDNF, an expression construct containing a 6X-HIS tag, a biotin acceptor peptide (BAP) sequence, a PreScission™ Protease cleavage site and a FLAG-tag attached to the N-terminal part of murine proBDNF. Intact HBpF-proBDNF has activities indistinguishable from its wild-type counterpart and can be used to purify proBDNF signaling complexes or to monitor proBDNF endocytosis and retrograde transport. HBpF-proBDNF will be useful for characterizing proBDNF signaling complexes and for deciphering the role of proBDNF in neuronal development, synapse function and neurodegenerative disease. PMID:26950209

  15. Biased signaling through G-protein-coupled PROKR2 receptors harboring missense mutations.

    Science.gov (United States)

    Sbai, Oualid; Monnier, Carine; Dodé, Catherine; Pin, Jean-Philippe; Hardelin, Jean-Pierre; Rondard, Philippe

    2014-08-01

    Various missense mutations in the gene coding for prokineticin receptor 2 (PROKR2), a G-protein-coupled receptor, have been identified in patients with Kallmann syndrome. However, the functional consequences of these mutations on the different signaling pathways of this receptor have not been studied. We first showed that the wild-type PROKR2 can activate different G-protein subtypes (Gq, Gs, and Gi/o) and recruit β-arrestins in transfected HEK-293 cells. We then examined, for each of these signaling pathways, the effects of 9 mutations that did not significantly impair cell surface targeting or ligand binding of the receptor. Four mutant receptors showing defective Gq signaling (R85C, R85H, R164Q, and V331M) could still recruit β-arrestins on ligand activation, which may cause biased signaling in vivo. Conversely, the R80C receptor could activate the 3 types of G proteins but could not recruit β-arrestins. Finally, the R268C receptor could recruit β-arrestins and activate the Gq and Gs signaling pathways but could not activate the Gi/o signaling pathway. Our results validate the concept that mutations in the genes encoding membrane receptors can bias downstream signaling in various ways, possibly leading to pathogenic and, perhaps in some cases, protective (e.g., R268C) effects. PMID:24830383

  16. Receptor Expression in Rat Skeletal Muscle Cell Cultures

    Science.gov (United States)

    Young, Ronald B.

    1996-01-01

    One on the most persistent problems with long-term space flight is atrophy of skeletal muscles. Skeletal muscle is unique as a tissue in the body in that its ability to undergo atrophy or hypertrophy is controlled exclusively by cues from the extracellular environment. The mechanism of communication between muscle cells and their environment is through a group of membrane-bound and soluble receptors, each of which carries out unique, but often interrelated, functions. The primary receptors include acetyl choline receptors, beta-adrenergic receptors, glucocorticoid receptors, insulin receptors, growth hormone (i.e., somatotropin) receptors, insulin-like growth factor receptors, and steroid receptors. This project has been initiated to develop an integrated approach toward muscle atrophy and hypertrophy that takes into account information on the populations of the entire group of receptors (and their respective hormone concentrations), and it is hypothesized that this information can form the basis for a predictive computer model for muscle atrophy and hypertrophy. The conceptual basis for this project is illustrated in the figure below. The individual receptors are shown as membrane-bound, with the exception of the glucocorticoid receptor which is a soluble intracellular receptor. Each of these receptors has an extracellular signalling component (e.g., innervation, glucocorticoids, epinephrine, etc.), and following the interaction of the extracellular component with the receptor itself, an intracellular signal is generated. Each of these intracellular signals is unique in its own way; however, they are often interrelated.

  17. Cancer Cell Invasion in Three-dimensional Collagen Is Regulated Differentially by Gα13 Protein and Discoidin Domain Receptor 1-Par3 Protein Signaling.

    Science.gov (United States)

    Chow, Christina R; Ebine, Kazumi; Knab, Lawrence M; Bentrem, David J; Kumar, Krishan; Munshi, Hidayatullah G

    2016-01-22

    Cancer cells can invade in three-dimensional collagen as single cells or as a cohesive group of cells that require coordination of cell-cell junctions and the actin cytoskeleton. To examine the role of Gα13, a G12 family heterotrimeric G protein, in regulating cellular invasion in three-dimensional collagen, we established a novel method to track cell invasion by membrane type 1 matrix metalloproteinase-expressing cancer cells. We show that knockdown of Gα13 decreased membrane type 1 matrix metalloproteinase-driven proteolytic invasion in three-dimensional collagen and enhanced E-cadherin-mediated cell-cell adhesion. E-cadherin knockdown reversed Gα13 siRNA-induced cell-cell adhesion but failed to reverse the effect of Gα13 siRNA on proteolytic invasion. Instead, concurrent knockdown of E-cadherin and Gα13 led to an increased number of single cells rather than groups of cells. Significantly, knockdown of discoidin domain receptor 1 (DDR1), a collagen-binding protein that also co-localizes to cell-cell junctions, reversed the effects of Gα13 knockdown on cell-cell adhesion and proteolytic invasion in three-dimensional collagen. Knockdown of the polarity protein Par3, which can function downstream of DDR1, also reversed the effects of Gα13 knockdown on cell-cell adhesion and proteolytic invasion in three-dimensional collagen. Overall, we show that Gα13 and DDR1-Par3 differentially regulate cell-cell junctions and the actin cytoskeleton to mediate invasion in three-dimensional collagen. PMID:26589794

  18. Type I interferon production during herpes simplex virus infection is controlled by cell-type-specific viral recognition through Toll-like receptor 9, the mitochondrial antiviral signaling protein pathway, and novel recognition systems

    DEFF Research Database (Denmark)

    Rasmussen, Simon Brandtoft; Sørensen, Louise Nørgaard; Malmgaard, Lene; Ank, Nina; Baines, JD; Chen, ZJ; Paludan, Søren Riis

    2007-01-01

    Recognition of viruses by germ line-encoded pattern recognition receptors of the innate immune system is essential for rapid production of type I interferon (IFN) and early antiviral defense. We investigated the mechanisms of viral recognition governing production of type I IFN during herpes...... fibroblasts, where the virus was able to replicate, HSV-induced IFN-alpha/beta production was dependent on both viral entry and replication, and ablated in cells unable to signal through the mitochondrial antiviral signaling protein pathway. Thus, during an HSV infection in vivo, multiple mechanisms of...

  19. Toll-like receptor signaling in primary immune deficiencies.

    Science.gov (United States)

    Maglione, Paul J; Simchoni, Noa; Cunningham-Rundles, Charlotte

    2015-11-01

    Toll-like receptors (TLRs) recognize common microbial or host-derived macromolecules and have important roles in early activation of the immune system. Patients with primary immune deficiencies (PIDs) affecting TLR signaling can elucidate the importance of these proteins to the human immune system. Defects in interleukin-1 receptor-associated kinase-4 and myeloid differentiation factor 88 (MyD88) lead to susceptibility to infections with bacteria, while mutations in nuclear factor-κB essential modulator (NEMO) and other downstream mediators generally induce broader susceptibility to bacteria, viruses, and fungi. In contrast, TLR3 signaling defects are specific for susceptibility to herpes simplex virus type 1 encephalitis. Other PIDs induce functional alterations of TLR signaling pathways, such as common variable immunodeficiency in which plasmacytoid dendritic cell defects enhance defective responses of B cells to shared TLR agonists. Dampening of TLR responses is seen for TLRs 2 and 4 in chronic granulomatous disease (CGD) and X-linked agammaglobulinemia (XLA). Enhanced TLR responses, meanwhile, are seen for TLRs 5 and 9 in CGD, TLRs 4, 7/8, and 9 in XLA, TLRs 2 and 4 in hyper IgE syndrome, and for most TLRs in adenosine deaminase deficiency. PMID:25930993

  20. Up-regulation of SOX9 in sertoli cells from testiculopathic patients accounts for increasing anti-mullerian hormone expression via impaired androgen receptor signaling.

    Directory of Open Access Journals (Sweden)

    Kuo-Chung Lan

    Full Text Available BACKGROUND: Testosterone provokes Sertoli cell maturation and represses AMH production. In adult patients with Sertoli-cells-only syndrome (SCOS and androgen insensitivity syndrome (AIS, high level of AMH expression is detected in Sertoli cells due to defect of androgen/AR signaling. OBJECTIVE: We postulated that up-regulation of SOX9 due to impairment of androgen/AR signaling in Sertoli cells might explain why high level of anti-Mullerian hormone (AMH expression occur in these testiculopathic patients. METHODS: Biological research of testicular specimens from men with azoospermia or mouse. The serum hormone levels were studied in 23 men with obstructive azoospermia, 33 men with SCOS azoospermia and 21 volunteers with normal seminograms during a period of 4 years. Immunohistochemical staining and reverse-transcription PCR were used to examine the relationships among AR, SOX9 and AMH expression in adult human and mouse testes. The ability of AR to repress the expression of SOX9 and AMH was evaluated in vitro in TM4 Sertoli cells and C3H10T1/2 cells. RESULTS: SCOS specimens showed up-regulation of SOX9 and AMH proteins but down-regulation of AR proteins in Sertoli cells. The mRNA levels of AR were significantly lower and the SOX9, AMH mRNA levels higher in all SCOS patients compared to controls (P< 0.05. The testosterone levels in the SCOS patients were within the normal range, but most were below the median of the controls. Furthermore, our in vitro cell line experiments demonstrated that androgen/AR signaling suppressed the gene and protein levels of AMH via repression of SOX9. CONCLUSIONS: Our data show that the functional androgen/AR signaling to repress SOX9 and AMH expression is essential for Sertoli cell maturation. Impairment of androgen/AR signaling promotes SOX9-mediated AMH production, accounts for impairments of Sertoli cells in SCOS azoospermic patients.

  1. Effect of hyperprolactinemia on PRL-receptor expression and activation of Stat and Mapk cell signaling in the prostate of long-term sexually-active rats.

    Science.gov (United States)

    Pascual-Mathey, Luz I; Rojas-Duran, Fausto; Aranda-Abreu, Gonzalo E; Manzo, Jorge; Herrera-Covarrubias, Deissy; Muñoz-Zavaleta, David A; Garcia, Luis I; Hernandez, Ma Elena

    2016-04-01

    The abnormal elevation of serum PRL, referred to as hyperprolactinemia (HyperPRL), produces alterations in several reproductive parameters of male rats such as penile erection or decreased tendency to reach ejaculation. Additionally, this situation produces a significant modification of prostate histology, as observed in the epithelial structure and alveolar area, which could reach a level of hyperplasia in the long-term. In this tissue, HyperPRL produces an increase in expression of PRL receptors and activation of the Stat3 signaling pathway that is correlated with the evolution of prostate pathologies. However, the impact of HyperPRL in long-term sexually active male rats is unknown. In this work, using constantly copulating Wistar male rats with induced HyperPRL, we analyzed the level of serum PRL, the effect on prostate PRL receptors, and activation of pStat3, pStat5 and Mapk signaling pathways. Two procedures to induce HyperPRL were employed, comprising daily IP administration or adenohypophysis transplant, and although neither affected the execution of sexual behavior, the serum PRL profile following successive ejaculations was affected. Messenger RNA expression of the short and long isoforms of the PRL receptor at the ventral prostate was affected in different ways depending on the procedure to induce HyperPRL. The ventral prostate did not show any modification in terms of activation of the pStat5 signaling pathway in subjects with daily administration of PRL, although this was significantly increased in ADH transplanted subjects in the second and fourth consecutive ejaculation. A similar profile was found for the pStat3 pathway which additionally showed a significant increase in the third and fourth ejaculation of daily-injected subjects. The Mapk signaling pathway did not show any modifications in subjects with daily administration of PRL, but showed a significant increase in the second and third ejaculations of subjects with ADH transplants. Thus

  2. N-Acetylglucosamine Functions in Cell Signaling

    Directory of Open Access Journals (Sweden)

    James B. Konopka

    2012-01-01

    Full Text Available The amino sugar N-acetylglucosamine (GlcNAc is well known for the important structural roles that it plays at the cell surface. It is a key component of bacterial cell wall peptidoglycan, fungal cell wall chitin, and the extracellular matrix of animal cells. Interestingly, recent studies have also identified new roles for GlcNAc in cell signaling. For example, GlcNAc stimulates the human fungal pathogen Candida albicans to undergo changes in morphogenesis and expression of virulence genes. Pathogenic E. coli responds to GlcNAc by altering the expression of fimbriae and CURLI fibers that promote biofilm formation and GlcNAc stimulates soil bacteria to undergo changes in morphogenesis and production of antibiotics. Studies with animal cells have revealed that GlcNAc influences cell signaling through the posttranslational modification of proteins by glycosylation. O-linked attachment of GlcNAc to Ser and Thr residues regulates a variety of intracellular proteins, including transcription factors such as NFκB, c-myc, and p53. In addition, the specificity of Notch family receptors for different ligands is altered by GlcNAc attachment to fucose residues in the extracellular domain. GlcNAc also impacts signal transduction by altering the degree of branching of N-linked glycans, which influences cell surface signaling proteins. These emerging roles of GlcNAc as an activator and mediator of cellular signaling in fungi, animals, and bacteria will be the focus of this paper.

  3. Testin, a novel binding partner of the calcium-sensing receptor, enhances receptor-mediated Rho-kinase signalling

    International Nuclear Information System (INIS)

    Highlights: → A yeast two-hybrid screen revealed testin bound to the calcium-sensing receptor. → The second zinc finger of LIM domain 1 of testin is critical for interaction. → Testin bound to a region of the receptor tail important for cell signalling. → Testin and receptor interaction was confirmed in mammalian (HEK293) cells. → Overexpression of testin enhanced receptor-mediated Rho signalling in HEK293 cells. -- Abstract: The calcium-sensing receptor (CaR) plays an integral role in calcium homeostasis and the regulation of other cellular functions including cell proliferation and cytoskeletal organisation. The multifunctional nature of the CaR is manifested through ligand-dependent stimulation of different signalling pathways that are also regulated by partner binding proteins. Following a yeast two-hybrid library screen using the intracellular tail of the CaR as bait, we identified several novel binding partners including the focal adhesion protein, testin. Testin has not previously been shown to interact with cell surface receptors. The sites of interaction between the CaR and testin were mapped to the membrane proximal region of the receptor tail and the second zinc-finger of LIM domain 1 of testin, the integrity of which was found to be critical for the CaR-testin interaction. The CaR-testin association was confirmed in HEK293 cells by coimmunoprecipitation and confocal microscopy studies. Ectopic expression of testin in HEK293 cells stably expressing the CaR enhanced CaR-stimulated Rho activity but had no effect on CaR-stimulated ERK signalling. These results suggest an interplay between the CaR and testin in the regulation of CaR-mediated Rho signalling with possible effects on the cytoskeleton.

  4. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Orfali, Nina [Cork Cancer Research Center, University College Cork, Cork (Ireland); Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States); McKenna, Sharon L. [Cork Cancer Research Center, University College Cork, Cork (Ireland); Cahill, Mary R. [Department of Hematology, Cork University Hospital, Cork (Ireland); Gudas, Lorraine J., E-mail: ljgudas@med.cornell.edu [Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States); Mongan, Nigel P., E-mail: nigel.mongan@nottingham.ac.uk [Faculty of Medicine and Health Science, School of Veterinary Medicine and Science, University of Nottingham, LE12 5RD (United Kingdom); Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA. (United States)

    2014-05-15

    Retinoids are a family of signaling molecules derived from vitamin A with well established roles in cellular differentiation. Physiologically active retinoids mediate transcriptional effects on cells through interactions with retinoic acid (RARs) and retinoid-X (RXR) receptors. Chromosomal translocations involving the RARα gene, which lead to impaired retinoid signaling, are implicated in acute promyelocytic leukemia (APL). All-trans-retinoic acid (ATRA), alone and in combination with arsenic trioxide (ATO), restores differentiation in APL cells and promotes degradation of the abnormal oncogenic fusion protein through several proteolytic mechanisms. RARα fusion-protein elimination is emerging as critical to obtaining sustained remission and long-term cure in APL. Autophagy is a degradative cellular pathway involved in protein turnover. Both ATRA and ATO also induce autophagy in APL cells. Enhancing autophagy may therefore be of therapeutic benefit in resistant APL and could broaden the application of differentiation therapy to other cancers. Here we discuss retinoid signaling in hematopoiesis, leukemogenesis, and APL treatment. We highlight autophagy as a potential important regulator in anti-leukemic strategies. - Highlights: • Normal and aberrant retinoid signaling in hematopoiesis and leukemia is reviewed. • We suggest a novel role for RARα in the development of X-RARα gene fusions in APL. • ATRA therapy in APL activates transcription and promotes onco-protein degradation. • Autophagy may be involved in both onco-protein degradation and differentiation. • Pharmacologic autophagy induction may potentiate ATRA's therapeutic effects.

  5. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia

    International Nuclear Information System (INIS)

    Retinoids are a family of signaling molecules derived from vitamin A with well established roles in cellular differentiation. Physiologically active retinoids mediate transcriptional effects on cells through interactions with retinoic acid (RARs) and retinoid-X (RXR) receptors. Chromosomal translocations involving the RARα gene, which lead to impaired retinoid signaling, are implicated in acute promyelocytic leukemia (APL). All-trans-retinoic acid (ATRA), alone and in combination with arsenic trioxide (ATO), restores differentiation in APL cells and promotes degradation of the abnormal oncogenic fusion protein through several proteolytic mechanisms. RARα fusion-protein elimination is emerging as critical to obtaining sustained remission and long-term cure in APL. Autophagy is a degradative cellular pathway involved in protein turnover. Both ATRA and ATO also induce autophagy in APL cells. Enhancing autophagy may therefore be of therapeutic benefit in resistant APL and could broaden the application of differentiation therapy to other cancers. Here we discuss retinoid signaling in hematopoiesis, leukemogenesis, and APL treatment. We highlight autophagy as a potential important regulator in anti-leukemic strategies. - Highlights: • Normal and aberrant retinoid signaling in hematopoiesis and leukemia is reviewed. • We suggest a novel role for RARα in the development of X-RARα gene fusions in APL. • ATRA therapy in APL activates transcription and promotes onco-protein degradation. • Autophagy may be involved in both onco-protein degradation and differentiation. • Pharmacologic autophagy induction may potentiate ATRA's therapeutic effects

  6. Monocyte Signal Transduction Receptors in Active and Latent Tuberculosis

    Directory of Open Access Journals (Sweden)

    Magdalena Druszczynska

    2013-01-01

    Full Text Available The mechanisms that promote either resistance or susceptibility to TB disease remain insufficiently understood. Our aim was to compare the expression of cell signaling transduction receptors, CD14, TLR2, CD206, and β2 integrin LFA-1 on monocytes from patients with active TB or nonmycobacterial lung disease and healthy individuals with M.tb latency and uninfected controls to explain the background of the differences between clinical and subclinical forms of M.tb infection. A simultaneous increase in the expression of the membrane bound mCD14 receptor and LFA-1 integrin in patients with active TB may be considered a prodrome of breaking immune control by M.tb bacilli in subjects with the latent TB and absence of clinical symptoms.

  7. Signal interaction of Hedgehog/GLI and epidermal growth factor receptor signaling in cancer development

    International Nuclear Information System (INIS)

    The subject of this PhD thesis is based on the cooperation of Hedgehog (HH)/GLI with epidermal growth factor receptor (EGFR) signaling synergistically promoting oncogenic transformation and cancer growth. In previous studies we have demonstrated that the HH/GLI and EGFR signaling pathways interact synergistically resulting not only in selective induction of HH/GLI-EGFR target genes, but also in the onset of oncogenic transformation and tumor formation (Kasper, Schnidar et al. 2006; Schnidar, Eberl et al. 2009). However, the molecular key mediators acting downstream of HH/GLI and EGFR signal cooperation were largely unknown and the in vivo evidence for the therapeutic relevance of HH/GLI and EGFR signal cooperation in HH-associated cancers was lacking. During my PhD thesis I could demonstrate that the integration of EGFR and HH/GLI signaling involves activation of RAS/MEK/ERK and JUN/AP1 signaling in response to EGFR activation. Furthermore I succeeded in identifying genes, including stem cell- (SOX2, SOX9), tumor growth- (JUN, TGFA, FGF19) and metastasis-associated genes (SPP1/osteopontin, CXCR4) that showed synergistic transcriptional activation by HH/GLI-EGFR signal integration. Importantly, I could demonstrate that these genes arrange themselves within a stable interdependent signaling network, which is required for in vivo growth of basal cell carcinoma (BCC) and tumor-initiating pancreatic cancer cells. These data validate EGFR signaling as additional drug target in HH/GLI driven cancers and provide new therapeutic strategies based on combined targeting of cooperative HH/GLI-EGFR signaling and selected downstream target genes (Eberl, Klingler et al. 2012). (author)

  8. Real-time trafficking and signaling of the glucagon-like peptide-1 receptor

    DEFF Research Database (Denmark)

    Roed, Sarah Noerklit; Wismann, Pernille; Underwood, Christina Rye;

    2014-01-01

    . A fundamental mechanism controlling the signaling capacity of GPCRs is the post-endocytic trafficking of receptors between recycling and degradative fates. Here, we combined microscopy with novel real-time assays to monitor both receptor trafficking and signaling in living cells. We find that the...... human GLP-1R internalizes rapidly and with similar kinetics in response to equipotent concentrations of GLP-1 and the stable GLP-1 analogues exendin-4 and liraglutide. Receptor internalization was confirmed in mouse pancreatic islets. GLP-1R is shown to be a recycling receptor with faster recycling...

  9. Nicotine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone induce cyclooxygenase-2 activity in human gastric cancer cells: Involvement of nicotinic acetylcholine receptor (nAChR) and β-adrenergic receptor signaling pathways

    International Nuclear Information System (INIS)

    Induction of cyclooxygenase-2 (COX-2) associates with cigarette smoke exposure in many malignancies. Nicotine and its derivative, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), are the two important components in cigarette smoke that contributes to cancer development. However, the molecular mechanism(s) by which nicotine or NNK promotes gastric carcinogenesis remains largely unknown. We found that nicotine and NNK significantly enhanced cell proliferation in AGS cells that expressed both alpha7 nicotinic acetylcholine receptor (α7 nAChR) and β-adrenergic receptors. Treatment of cells with α-bungarotoxin (α-BTX, α7nAChR antagonist) or propranolol (β-adrenergic receptor antagonist) blocked NNK-induced COX-2/PGE2 and cell proliferation, while nicotine-mediated cell growth and COX-2/PGE2 induction can only be suppressed by propranolol, but not α-BTX. Moreover, in contrast to the dependence of growth promoting effect of nicotine on Erk activation, inhibitor of p38 mitogen-activated protein kinase (MAPK) repressed NNK-induced COX-2 upregulation and resulted in suppression of cell growth. In addition, nicotine and NNK mediated COX-2 induction via different receptors to modulate several G1/S transition regulatory proteins and promote gastric cancer cell growth. Selective COX-2 inhibitor (SC-236) caused G1 arrest and abrogated nicotine/NNK-induced cell proliferation. Aberrant expression of cyclin D1 and other G1 regulatory proteins are reversed by blockade of COX-2. These results pointed to the importance of adrenergic and nicotinic receptors in gastric tumor growth through MAPK/COX-2 activation, which may perhaps provide a chemoprevention strategy for cigarette smoke-related gastric carcinogenesis

  10. Nuclear Receptor Signaling Atlas: Opening Access to the Biology of Nuclear Receptor Signaling Pathways.

    Directory of Open Access Journals (Sweden)

    Lauren B Becnel

    Full Text Available Signaling pathways involving nuclear receptors (NRs, their ligands and coregulators, regulate tissue-specific transcriptomes in diverse processes, including development, metabolism, reproduction, the immune response and neuronal function, as well as in their associated pathologies. The Nuclear Receptor Signaling Atlas (NURSA is a Consortium focused around a Hub website (www.nursa.org that annotates and integrates diverse 'omics datasets originating from the published literature and NURSA-funded Data Source Projects (NDSPs. These datasets are then exposed to the scientific community on an Open Access basis through user-friendly data browsing and search interfaces. Here, we describe the redesign of the Hub, version 3.0, to deploy "Web 2.0" technologies and add richer, more diverse content. The Molecule Pages, which aggregate information relevant to NR signaling pathways from myriad external databases, have been enhanced to include resources for basic scientists, such as post-translational modification sites and targeting miRNAs, and for clinicians, such as clinical trials. A portal to NURSA's Open Access, PubMed-indexed journal Nuclear Receptor Signaling has been added to facilitate manuscript submissions. Datasets and information on reagents generated by NDSPs are available, as is information concerning periodic new NDSP funding solicitations. Finally, the new website integrates the Transcriptomine analysis tool, which allows for mining of millions of richly annotated public transcriptomic data points in the field, providing an environment for dataset re-use and citation, bench data validation and hypothesis generation. We anticipate that this new release of the NURSA database will have tangible, long term benefits for both basic and clinical research in this field.

  11. Nuclear Receptor Signaling Atlas: Opening Access to the Biology of Nuclear Receptor Signaling Pathways

    Science.gov (United States)

    Becnel, Lauren B.; Darlington, Yolanda F.; Ochsner, Scott A.; Easton-Marks, Jeremy R.; Watkins, Christopher M.; McOwiti, Apollo; Kankanamge, Wasula H.; Wise, Michael W.; DeHart, Michael; Margolis, Ronald N.; McKenna, Neil J.

    2015-01-01

    Signaling pathways involving nuclear receptors (NRs), their ligands and coregulators, regulate tissue-specific transcriptomes in diverse processes, including development, metabolism, reproduction, the immune response and neuronal function, as well as in their associated pathologies. The Nuclear Receptor Signaling Atlas (NURSA) is a Consortium focused around a Hub website (www.nursa.org) that annotates and integrates diverse ‘omics datasets originating from the published literature and NURSA-funded Data Source Projects (NDSPs). These datasets are then exposed to the scientific community on an Open Access basis through user-friendly data browsing and search interfaces. Here, we describe the redesign of the Hub, version 3.0, to deploy “Web 2.0” technologies and add richer, more diverse content. The Molecule Pages, which aggregate information relevant to NR signaling pathways from myriad external databases, have been enhanced to include resources for basic scientists, such as post-translational modification sites and targeting miRNAs, and for clinicians, such as clinical trials. A portal to NURSA’s Open Access, PubMed-indexed journal Nuclear Receptor Signaling has been added to facilitate manuscript submissions. Datasets and information on reagents generated by NDSPs are available, as is information concerning periodic new NDSP funding solicitations. Finally, the new website integrates the Transcriptomine analysis tool, which allows for mining of millions of richly annotated public transcriptomic data points in the field, providing an environment for dataset re-use and citation, bench data validation and hypothesis generation. We anticipate that this new release of the NURSA database will have tangible, long term benefits for both basic and clinical research in this field. PMID:26325041

  12. Self-enforcing Feedback Activation between BCL6 and Pre-B Cell Receptor Signaling Defines a Distinct Subtype of Acute Lymphoblastic Leukemia

    Science.gov (United States)

    Geng, Huimin; Hurtz, Christian; Lenz, Kyle B.; Chen, Zhengshan; Baumjohann, Dirk; Thompson, Sarah; Goloviznina, Natalya; Chen, Wei-Yi; Huan, Jianya; LaTocha, Dorian; Ballabio, Erica; Xiao, Gang; Lee, Jae-Woong; Deucher, Anne; Qi, Zhongxia; Park, Eugene; Huang, Chuanxin; Nahar, Rahul; Kweon, Soo-Mi; Shojaee, Seyedmehdi; Chan, Lai N.; Yu, Jingwei; Kornblau, Steven M.; Bijl, Janetta J.; Ye, B. Hilda; Ansel, Mark; Paietta, Elisabeth; Melnick, Ari; Hunger, Stephen P.; Kurre, Peter; Tyner, Jeffrey W.; Loh, Mignon L.; Roeder, Robert G.; Druker, Brian J.; Burger, Jan. A.; Milne, Thomas A.; Chang, Bill H.; Müschen, Markus

    2015-01-01

    SUMMARY Studying 830 pre-B ALL cases from four clinical trials, we found that human ALL can be divided into two fundamentally distinct subtypes based on pre-BCR function. While absent in the majority of ALL cases, tonic pre-BCR signaling was found in 112 cases (13.5%). In these cases, tonic pre-BCR signaling induced activation of BCL6, which in turn increased pre-BCR signaling output at the transcriptional level. Interestingly, inhibition of pre-BCR-related tyrosine kinases reduced constitutive BCL6 expression and selectively killed patient-derived pre-BCR+ ALL cells. These findings identify a genetically and phenotypically distinct subset of human ALL that critically depends on tonic pre-BCR signaling. In vivo treatment studies suggested that pre-BCR tyrosine kinase inhibitors are useful for the treatment of patients with pre-BCR+ ALL. PMID:25759025

  13. The Myc-miR-17-92 axis amplifies B-cell receptor signaling via inhibition of ITIM proteins: a novel lymphomagenic feed-forward loop

    OpenAIRE

    Psathas, James N.; Doonan, Patrick J.; Raman, Pichai; Freedman, Bruce D.; Minn, Andy J.; Thomas-Tikhonenko, Andrei

    2013-01-01

    Myc amplifies BCR signaling and increases its own levels via upregulation of miR-17∼92 and subsequent targeting of ITIM proteins.Inhibition of miR-17∼92 in DLBCL cell lines dampens the BCR response; DLBCL tumors of the BCR subtype have elevated levels of miR-17∼92.

  14. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia.

    LENUS (Irish Health Repository)

    Orfali, Nina

    2014-05-15

    Retinoids are a family of signaling molecules derived from vitamin A with well established roles in cellular differentiation. Physiologically active retinoids mediate transcriptional effects on cells through interactions with retinoic acid (RARs) and retinoid-X (RXR) receptors. Chromosomal translocations involving the RARα gene, which lead to impaired retinoid signaling, are implicated in acute promyelocytic leukemia (APL). All-trans-retinoic acid (ATRA), alone and in combination with arsenic trioxide (ATO), restores differentiation in APL cells and promotes degradation of the abnormal oncogenic fusion protein through several proteolytic mechanisms. RARα fusion-protein elimination is emerging as critical to obtaining sustained remission and long-term cure in APL. Autophagy is a degradative cellular pathway involved in protein turnover. Both ATRA and ATO also induce autophagy in APL cells. Enhancing autophagy may therefore be of therapeutic benefit in resistant APL and could broaden the application of differentiation therapy to other cancers. Here we discuss retinoid signaling in hematopoiesis, leukemogenesis, and APL treatment. We highlight autophagy as a potential important regulator in anti-leukemic strategies.

  15. Intracellular Signals of T Cell Costimulation

    Institute of Scientific and Technical Information of China (English)

    Jianxun Song; Fengyang Tylan Lei; Xiaofang Xiong; Rizwanul Haque

    2008-01-01

    Ligation of T cell receptor (TCR) alone is insufficient to induce full activation of T lymphocytes. Additional ligand-receptor interactions (costimulation) on antigen presenting cells (APCs) and T cells are required. T cell costimulation has been shown to be essential for eliciting efficient T cell responses, involving all phases during T cell development. However, the mechanisms by which costimulation affects the function of T cells still need to be elucidated. In recent years, advances have been made in studies of costimulation as potential therapies in cancer, infectious disease as well as autoimmune disease. In this review, we discussed intracellular costimulation signals that regulate T cell proliferation, cell cycle progression, cytokine production, survival, and memory development. In general, the pathway of phosphoinositide-3 kinase (PBK)/protein kinase B (PKB, also known as Akt)/nuclear factor κB (NF-κB) might be central to many costimulatory effects. Through these pathways, costimulation controls T-cell expansion and proliferation by maintenance of survivin and aurora B expression, and sustains long-term T-cell survival and memory development by regulating the expression of bci-2 family members. Cellular & Molecular Immunology.2008;5(4):239-247.

  16. CD54/intercellular adhesion molecule 1 and major histocompatibility complex II signaling induces B cells to express interleukin 2 receptors and complements help provided through CD40 ligation

    DEFF Research Database (Denmark)

    Poudrier, J; Owens, T

    1994-01-01

    We have examined signaling roles for CD54 intercellular adhesion molecule 1 and major histocompatibility complex (MHC) II as contact ligands during T help for B cell activation. We used a T helper 1 (Th1)-dependent helper system that was previously shown to be contact as well as interleukin 2 (IL-2......) dependent to demonstrate the relative roles of CD54, MHC II, and CD40 signaling in the events leading to the induction of B cell proliferation and responsiveness to IL-2. Paraformaldehyde-fixed activated Th1-induced expression of IL-2R alpha, IL-2R beta, and B7, and upregulated MHC II and CD54 on B cells...

  17. Benzophenone-1 stimulated the growth of BG-1 ovarian cancer cells by cell cycle regulation via an estrogen receptor alpha-mediated signaling pathway in cellular and xenograft mouse models

    International Nuclear Information System (INIS)

    Highlights: ► BP-1 induced cell growth was reversed by an ER antagonist in BG-1 cells. ► BP-1 up-regulated the mRNA expression of cyclin D1. ► Up-regulation of cyclin D1 by BP-1 was reversed by an ER antagonist. ► BP-1 is a potential endocrine disruptor that exerts estrogenic effects. - Abstract: 2,4-Dihydroxybenzophenone (benzophenone-1; BP-1) is an UV stabilizer primarily used to prevent polymer degradation and deterioration in quality due to UV irradiation. Recently, BP-1 has been reported to bioaccumulate in human bodies by absorption through the skin and has the potential to induce health problems including endocrine disruption. In the present study, we examined the xenoestrogenic effect of BP-1 on BG-1 human ovarian cancer cells expressing estrogen receptors (ERs) and relevant xenografted animal models in comparison with 17-β estradiol (E2). In in vitro cell viability assay, BP-1 (10−8–10−5 M) significantly increased BG-1 cell growth the way E2 did. The mechanism underlying the BG-1 cell proliferation was proved to be related with the up-regulation of cyclin D1, a cell cycle progressor, by E2 or BP-1. Both BP-1 and E2 induced cell growth and up-regulation of cyclin D1 were reversed by co-treatment with ICI 182,780, an ER antagonist, suggesting that BP-1 may mediate the cancer cell proliferation via an ER-dependent pathway like E2. On the other hand, the expression of p21, a regulator of cell cycle progression at G1 phase, was not altered by BP-1 though it was down-regulated by E2. In xenograft mouse models transplanted with BG-1 cells, BP-1 or E2 treatment significantly increased the tumor mass formation compared to a vehicle (corn oil) within 8 weeks. In histopathological analysis, the tumor sections of E2 or BP-1 group displayed extensive cell formations with high density and disordered arrangement, which were supported by the increased number of BrdUrd positive nuclei and the over-expression of cyclin D1 protein. Taken together, these

  18. Single-walled carbon nanotube exposure induces membrane rearrangement and suppression of receptor-mediated signalling pathways in model mast cells

    Science.gov (United States)

    Umemoto, Eric Y.; Speck, Mark; Shimoda, Lori M.N.; Kahue, Kara; Sung, Carl; Stokes, Alexander J.; Turner, Helen

    2014-01-01

    Carbon nanotubes (CNT) are environmental challenges to the respiratory and gastrointestinal mucosa, and to the dermal immune system. Mast cells (MC) are pro-inflammatory immunocytes that reside at these interfaces with the environment. Mast cells are sources of pro-inflammatory mediators (histamine, serotonin, matrix-active proteases, eicosanoids, prostanoids, cytokines and chemokines), which are released in a calcium-dependent manner following immunological challenge or physico-chemical stimulation. Since C-60 fullerenes, which share geometry with CNT, are suppressive of mast cell-driven inflammatory responses, we explored the effects of unmodified SWCNT aggregates on mast cell signaling pathways, phenotype and pro-inflammatory function. We noted SWCNT suppression of antigen-induced signalling pathways and pro-inflammatory degranulation responses. Mast cells recognize unmodified SWCNT by remodeling the plasma membrane, disaggregating the cortical actin cytoskeleton and relocalizing clathrin. Clathrin was also identified as a component of an affinity-purified ‘interactome’ isolated from MC using an SWCNT affinity matrix for mast cell lysates. Together these data are consistent with the ability of SWCNT to suppress mast cell pro-inflammatory function via a novel recognition mechanism. PMID:24910985

  19. Cordycepin Suppresses Thymic Stromal Lymphopoietin Expression via Blocking Caspase-1 and Receptor-Interacting Protein 2 Signaling Pathways in Mast Cells.

    Science.gov (United States)

    Yoou, Myoung-schook; Jin, Mu Hyun; Lee, So Young; Lee, Sang Hwa; Kim, Byunghyun; Roh, Seok Seon; Choi, In Hwa; Lee, Myeong Soo; Kim, Hyung-Min; Jeong, Hyun-Ja

    2016-01-01

    Cordycepin (3'-deoxyadenosine) is one of the active components isolated from Cordyceps militaris, and has been shown to have anti-inflammatory, anti-oxidant, anti-aging, and anti-cancer effects. Mast cell-derived thymic stromal lymphopoietin (TSLP) plays an important role in the pathogenesis of allergic inflammatory reactions. Here, we investigated the regulatory effect and mechanisms of cordycepin on the expression of TSLP in the human mast cell line, HMC-1 cells, and in the human keratinocyte cell line, HaCaT cells. Cordycepin significantly decreased the production and mRNA expression of TSLP through the inhibition of caspase-1 and nuclear factor-κB activation. Cordycepin also significantly reduced the phosphorylation of receptor-interacting protein 2 and inhibitory kappa B (IκB) kinase β. Cordycepin significantly decreased the production and mRNA expression of interleukin (IL)-8, IL-1β, IL-6, and tumor necrosis factor-α in activated HMC-1 cells. Moreover, cordycepin significantly decreased the levels of TSLP in activated HaCaT cells. Our studies suggest that cordycepin can be applied to the treatment of allergic inflammatory diseases exacerbated by TSLP. PMID:26725432

  20. Glutamate Delta-1 Receptor Regulates Metabotropic Glutamate Receptor 5 Signaling in the Hippocampus.

    Science.gov (United States)

    Suryavanshi, Pratyush S; Gupta, Subhash C; Yadav, Roopali; Kesherwani, Varun; Liu, Jinxu; Dravid, Shashank M

    2016-08-01

    The delta family of ionotropic glutamate receptors consists of glutamate delta-1 (GluD1) and glutamate delta-2 receptors. We have previously shown that GluD1 knockout mice exhibit features of developmental delay, including impaired spine pruning and switch in the N-methyl-D-aspartate receptor subunit, which are relevant to autism and other neurodevelopmental disorders. Here, we identified a novel role of GluD1 in regulating metabotropic glutamate receptor 5 (mGlu5) signaling in the hippocampus. Immunohistochemical analysis demonstrated colocalization of mGlu5 with GluD1 punctas in the hippocampus. Additionally, GluD1 protein coimmunoprecipitated with mGlu5 in the hippocampal membrane fraction, as well as when overexpressed in human embryonic kidney 293 cells, demonstrating that GluD1 and mGlu5 may cooperate in a signaling complex. The interaction of mGlu5 with scaffold protein effector Homer, which regulates mechanistic target of rapamycin (mTOR) signaling, was abnormal both under basal conditions and in response to mGlu1/5 agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) in GluD1 knockout mice. The basal levels of phosphorylated mTOR and protein kinase B, the signaling proteins downstream of mGlu5 activation, were higher in GluD1 knockout mice, and no further increase was induced by DHPG. We also observed higher basal protein translation and an absence of DHPG-induced increase in GluD1 knockout mice. In accordance with a role of mGlu5-mediated mTOR signaling in synaptic plasticity, DHPG-induced internalization of surface α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor subunits was impaired in the GluD1 knockout mice. These results demonstrate that GluD1 interacts with mGlu5, and loss of GluD1 impairs normal mGlu5 signaling potentially by dysregulating coupling to its effector. These studies identify a novel role of the enigmatic GluD1 subunit in hippocampal function. PMID:27231330

  1. Molecular mechanisms of alternative estrogen receptor signaling

    OpenAIRE

    Björnström, Linda

    2003-01-01

    Estrogen is a key regulator of growth, differentiation and function in a broad range of target tissues, including the male and female reproductive tracts, mammary gland, bone, brain and the cardiovascular system. The biological effects of estrogen are mediated through estrogen receptor a (ERalpha) and estrogen receptor beta (ERbeta), which belong to a large superfamily of nuclear receptors that act as ligand-activated transcription factors. The classical mechanism of ER acti...

  2. MAPK Cascades in Guard Cell Signal Transduction.

    Science.gov (United States)

    Lee, Yuree; Kim, Yun Ju; Kim, Myung-Hee; Kwak, June M

    2016-01-01

    Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions. PMID:26904052

  3. Novel roles of nuclear angiotensin receptors and signaling mechanisms.

    Science.gov (United States)

    Gwathmey, TanYa M; Alzayadneh, Ebaa M; Pendergrass, Karl D; Chappell, Mark C

    2012-03-01

    The renin-angiotensin system (RAS) constitutes an important hormonal system in the physiological regulation of blood pressure. The dysregulation of the RAS is considered a major influence in the development and progression of cardiovascular disease and other pathologies. Indeed, experimental and clinical evidence indicates that blockade of this system with angiotensin-converting enzyme (ACE) inhibitors or angiotensin type 1 receptor (AT1R) antagonists is an effective therapy to attenuate hypertension and diabetic renal injury, and to improve heart failure. Originally defined as a circulating system, multiple tissues express a complete RAS, and compelling evidence now favors an intracellular system involved in cell signaling and function. Within the kidney, intracellular expression of the three predominant ANG receptor subtypes is evident in the nuclear compartment. The ANG type 1 receptor (AT1R) is coupled to the generation of reactive oxygen species (ROS) through the activation of phosphoinositol-3 kinase (PI3K) and PKC. In contrast, both ANG type 2 (AT2R) and ANG-(1-7) (AT7R) receptors stimulate nitric oxide (NO) formation, which may involve nuclear endothelial NO synthase (eNOS). Moreover, blockade of either ACE2-the enzyme that converts ANG II to ANG-(1-7)-or the AT7 receptor exacerbates the ANG II-ROS response on renal nuclei. Finally, in a model of fetal programmed hypertension, the nuclear ROS response to ANG II is enhanced, while both AT2 and AT7 stimulation of NO is attenuated, suggesting that an imbalance in the intracellular RAS may contribute to the development of programming events. We conclude that a functional intracellular or nuclear RAS may have important implications in the therapeutic approaches to cardiovascular disease. PMID:22170620

  4. Primary Cilia, Signaling Networks and Cell Migration

    DEFF Research Database (Denmark)

    Veland, Iben Rønn

    controls directional cell migration as a physiological response. The ciliary pocket is a membrane invagination with elevated activity of clathrin-dependent endocytosis (CDE). In paper I, we show that the primary cilium regulates TGF-β signaling and the ciliary pocket is a compartment for CDE......-dependent regulation of signal transduction. Upon ligand-binding and activation in the cilium, TGFβ receptors accumulate and are internalized at the ciliary base together with Smad2/3 transcription factors that are phosphorylated here and translocated to the nucleus for target gene expression. These processes depend...... migration. A number of central Wnt components localize to the fibroblast primary cilium, including the Wnt5a-receptor, Fzd3, and Dvl proteins. Inversin-deficient MEFs have an elevated expression of canonical Wnt-associated genes and proteins, in addition to dysregulation of components in non-canonical Wnt...

  5. The Na+/H+ Exchanger Regulatory Factor Stabilizes Epidermal Growth Factor Receptors at the Cell Surface

    OpenAIRE

    Lazar, Cheri S.; Cresson, Catherine M.; Lauffenburger, Douglas A.; Gill, Gordon N.

    2004-01-01

    Ligand binding to cell surface receptors initiates both signal transduction and endocytosis. Although signaling may continue within the endocytic compartment, down-regulation is the major mechanism that controls the concentration of cell surface receptors, their ability to receive environmental signals, and the ultimate strength of biological signaling. Internalization, recycling, and trafficking of receptor tyrosine kinases (RTKs) within the endosome compartment are each regulated to control...

  6. G Protein-coupled Receptors and Resistance to Inhibitors of Cholinesterase-8A (Ric-8A) Both Regulate the Regulator of G Protein Signaling 14 (RGS14)·Gαi1 Complex in Live Cells*

    OpenAIRE

    Vellano, Christopher P.; Maher, Ellen M.; Hepler, John R.; Blumer, Joe B.

    2011-01-01

    Background: Regulator of G protein signaling 14 (RGS14) is a G protein regulatory (GPR) protein that participates in unconventional G protein signaling independent of G protein-coupled receptors (GPCRs).

  7. Persistent cAMP Signaling by Internalized LH Receptors in Ovarian Follicles.

    Science.gov (United States)

    Lyga, Sandra; Volpe, Silvia; Werthmann, Ruth C; Götz, Konrad; Sungkaworn, Titiwat; Lohse, Martin J; Calebiro, Davide

    2016-04-01

    A crucial event in female reproduction occurs at midcycle, when a LH peak induces the final maturation of ovarian follicles. LH signals via a G protein-coupled receptor selectively expressed in the outermost follicular cell layers. However, how LH signals are relayed inside these cells and finally to the oocyte is incompletely understood. Here, we monitored LH signaling in intact ovarian follicles of transgenic mice expressing a fluorescent cAMP sensor. We found that LH stimulation induces 2 phases of cAMP signaling in all cell layers surrounding the oocyte. Interfering with LH receptor internalization abolished the second, persistent cAMP phase and partially inhibited oocyte meiosis resumption. These data suggest that persistent cAMP signals from internalized LH receptors contribute to transmitting LH effects inside follicle cells and ultimately to the oocyte. Thus, this study indicates that the recently proposed paradigm of cAMP signaling by internalized G protein-coupled receptors is implicated in receptor function and is physiologically relevant. PMID:26828746

  8. Pharmacology of bile acid receptors: Evolution of bile acids from simple detergents to complex signaling molecules.

    Science.gov (United States)

    Copple, Bryan L; Li, Tiangang

    2016-02-01

    For many years, bile acids were thought to only function as detergents which solubilize fats and facilitate the uptake of fat-soluble vitamins in the intestine. Many early observations; however, demonstrated that bile acids regulate more complex processes, such as bile acids synthesis and immune cell function through activation of signal transduction pathways. These studies were the first to suggest that receptors may exist for bile acids. Ultimately, seminal studies by many investigators led to the discovery of several bile acid-activated receptors including the farnesoid X receptor, the vitamin D receptor, the pregnane X receptor, TGR5, α5 β1 integrin, and sphingosine-1-phosphate receptor 2. Several of these receptors are expressed outside of the gastrointestinal system, indicating that bile acids may have diverse functions throughout the body. Characterization of the functions of these receptors over the last two decades has identified many important roles for these receptors in regulation of bile acid synthesis, transport, and detoxification; regulation of glucose utilization; regulation of fatty acid synthesis and oxidation; regulation of immune cell function; regulation of energy expenditure; and regulation of neural processes such as gastric motility. Through these many functions, bile acids regulate many aspects of digestion ranging from uptake of essential vitamins to proper utilization of nutrients. Accordingly, within a short time period, bile acids moved beyond simple detergents and into the realm of complex signaling molecules. Because of the important processes that bile acids regulate through activation of receptors, drugs that target these receptors are under development for the treatment of several diseases, including cholestatic liver disease and metabolic syndrome. In this review, we will describe the various bile acid receptors, the signal transduction pathways activated by these receptors, and briefly discuss the physiological processes that

  9. Dopamine receptor repertoire of human granulosa cells

    Directory of Open Access Journals (Sweden)

    Kunz Lars

    2007-10-01

    Full Text Available Abstract Background High levels of dopamine (DA were described in human ovary and recently evidence for DA receptors in granulosa and luteal cells has been provided, as well. However, neither the full repertoire of ovarian receptors for DA, nor their specific role, is established. Human granulosa cells (GCs derived from women undergoing in vitro fertilization (IVF are an adequate model for endocrine cells of the follicle and the corpus luteum and were therefore employed in an attempt to decipher their DA receptor repertoire and functionality. Methods Cells were obtained from patients undergoing IVF and examined using cDNA-array, RT-PCR, Western blotting and immunocytochemistry. In addition, calcium measurements (with FLUO-4 were employed. Expression of two DA receptors was also examined by in-situ hybridization in rat ovary. Effects of DA on cell viability and cell volume were studied by using an ATP assay and an electronic cell counter system. Results We found members of the two DA receptor families (D1- and D2 -like associated with different signaling pathways in human GCs, namely D1 (as expected and D5 (both are Gs coupled and linked to cAMP increase and D2, D4 (Gi/Gq coupled and linked to IP3/DAG. D3 was not found. The presence of the trophic hormone hCG (10 IU/ml in the culture medium for several days did not alter mRNA (semiquantitative RT-PCR or protein levels (immunocytochemistry/Western blotting of D1,2,4,5 DA receptors. Expression of prototype receptors for the two families, D1 and D2, was furthermore shown in rat granulosa and luteal cells by in situ hybridization. Among the DA receptors found in human GCs, D2 expression was marked both at mRNA and protein levels and it was therefore further studied. Results of additional RT-PCR and Western blots showed two splice variants (D2L, D2S. Irrespective of these variants, D2 proved to be functional, as DA raised intracellular calcium levels. This calcium mobilizing effect of DA was observed

  10. PROKR2 missense mutations associated with Kallmann syndrome impair receptor signalling activity.

    Science.gov (United States)

    Monnier, Carine; Dodé, Catherine; Fabre, Ludovic; Teixeira, Luis; Labesse, Gilles; Pin, Jean-Philippe; Hardelin, Jean-Pierre; Rondard, Philippe

    2009-01-01

    Kallmann syndrome (KS) combines hypogonadism due to gonadotropin-releasing hormone deficiency, and anosmia or hyposmia, related to defective olfactory bulb morphogenesis. In a large series of KS patients, ten different missense mutations (p.R85C, p.R85H, p.R164Q, p.L173R, p.W178S, p.Q210R, p.R268C, p.P290S, p.M323I, p.V331M) have been identified in the gene encoding the G protein-coupled receptor prokineticin receptor-2 (PROKR2), most often in the heterozygous state. Many of these mutations were, however, also found in clinically unaffected individuals, thus raising the question of their actual implication in the KS phenotype. We reproduced each of the ten mutations in a recombinant murine Prokr2, and tested their effects on the signalling activity in transfected HEK-293 cells, by measuring intracellular calcium release upon ligand-activation of the receptor. We found that all mutated receptors except one (M323I) had decreased signalling activities. These could be explained by different defective mechanisms. Three mutations (L173R, W178S, P290S) impaired cell surface-targeting of the receptor. One mutation (Q210R) abolished ligand-binding. Finally, five mutations (R85C, R85H, R164Q, R268C, V331M) presumably impaired G protein-coupling of the receptor. In addition, when wild-type and mutant receptors were coexpressed in HEK-293 cells, none of the mutant receptors that were retained within the cells did affect cell surface-targeting of the wild-type receptor, and none of the mutant receptors properly addressed at the plasma membrane did affect wild-type receptor signalling activity. This argues against a dominant negative effect of the mutations in vivo. PMID:18826963

  11. The angiotensin II type 1 receptor antagonist Losartan binds and activates bradykinin B2 receptor signaling

    DEFF Research Database (Denmark)

    Bonde, Marie Mi; Olsen, Kristine Boisen; Erikstrup, Niels;

    2011-01-01

    The angiotensin II type 1 receptor (AT1R) blocker (ARB) Losartan has cardioprotective effects during ischemia-reperfusion injury and inhibits reperfusion arrhythmias -effects that go beyond the benefits of lowering blood pressure. The renin-angiotensin and kallikrein-kinin systems are intricately...... connected and some of the cardioprotective effects of Losartan are abolished by blocking the bradykinin B2 receptor (B2R) signaling. In this study, we investigated the ability of six clinically available ARBs to specifically bind and activate the B2R. First, we investigated their ability to activate...... phosphoinositide (PI) hydrolysis in COS-7 cells transiently expressing the B2R. We found that only Losartan activated the B2R, working as a partial agonist compared to the endogenous ligand bradykinin. This effect was blocked by the B2R antagonist HOE 140. A competitive binding analysis revealed that Losartan does...

  12. Thyroid hormone receptor β1 suppresses proliferation and migration by inhibiting PI3K/Akt signaling in human colorectal cancer cells.

    Science.gov (United States)

    Zhu, Lei; Tian, Guangang; Yang, Qin; De, Gejing; Zhang, Zhigang; Wang, Yahui; Nie, Huizhen; Zhang, Yanli; Yang, Xiaomei; Li, Jun

    2016-09-01

    Thyroid hormone receptor β1 (TRβ1) is a ligand‑dependent transcription factor that belongs to the superfamily of nuclear receptors. TRβ1 has been found to act as a tumor suppressor in many solid tumors including breast cancer and hepatocellular carcinoma, but its role in the progression of human colorectal cancer (CRC) remains unclear. In this study, microarray data analysis revealed that TRβ1 mRNA was downregulated in CRC tumors compared with that in the normal counterparts in both The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) datasets. Using a CRC tissue microarray (TMA), we confirmed that the expression of TRβ1 was decreased in human CRC tumor tissues in contrast to normal colorectal mucosal tissues. Notably, the TRβ1 expression was strongly correlated with tumor size (p=0.045). Furthermore, we found that CRC cell proliferation and migration were significantly inhibited by TRβ1 overexpression in vitro. Mechanistic studies indicated that activated phosphorylated Akt was clearly suppressed by TRβ1 in the CRC tissues and cells. In conclusion, this study provides evidence that TRβ1 plays a critical role in the progression of CRC via the PI3K/Akt pathway, and the TRβ1 gene may represent a novel target for CRC therapeutics. PMID:27431682

  13. Continuous requirement for the T cell receptor for regulatory T cell function

    OpenAIRE

    Levine, Andrew G; Arvey, Aaron; Jin, Wei; Rudensky, Alexander Y.

    2014-01-01

    Foxp3+ regulatory T cells (Treg cells) maintain immunological tolerance and their deficiency results in fatal multi-organ autoimmunity. Although heightened T cell receptor (TCR) signaling is critical for the differentiation of Treg cells, the role of TCR signaling in Treg cell function remains largely unknown. Here we demonstrate inducible ablation of the TCR results in Treg cell dysfunction which cannot be attributed to impaired Foxp3 expression, decreased expression of Treg cell signature g...

  14. Sensing of cell death by myeloid C-type lectin receptors

    Science.gov (United States)

    Sancho, David; Reis e Sousa, Caetano

    2015-01-01

    Molecules associated with dead or dying cells can be detected by receptors on macrophages and dendritic cells. Signals from these receptors impact myeloid cell function and play a role in determining whether death is silent or proinflammatory, tolerogenic or immunogenic. Prominent among myeloid receptors detecting dead cells are C-type lectin receptors (CLRs). Signals from these receptors variably induce endocytosis of cell corpses, corpse degradation, retrieval of dead cell-associated antigens and/or modulation of immune responses. The sensing of tissue damage by myeloid CLRs complements detection of pathogens in immunity and represents an ancient response aimed at restoring tissue homeostasis. PMID:23332826

  15. Effects of the group I metabotropic glutamate receptor agonist, DHPG, and injection stress on striatal cell signaling in food-restricted and ad libitum fed rats

    Directory of Open Access Journals (Sweden)

    Carr Kenneth D

    2004-12-01

    Full Text Available Abstract Background Chronic food restriction augments the rewarding effect of centrally administered psychostimulant drugs and this effect may involve a previously documented upregulation of D-1 dopamine receptor-mediated MAP kinase signaling in nucleus accumbens (NAc and caudate-putamen (CPu. Psychostimulants are known to induce striatal glutamate release, and group I metabotropic glutamate receptors (mGluR have been implicated in the cellular and behavioral responses to amphetamine. The purpose of the present study was to evaluate whether chronic food restriction increases striatal MAP kinase signaling in response to the group I mGluR agonist, DHPG. Results Western immunoblotting was used to demonstrate that intracerebroventricular (i.c.v. injection of DHPG (500 nmol produces greater activation of ERK1/2 and CREB in CPu and NAc of food-restricted as compared to ad libitum fed rats. Fos-immunostaining induced by DHPG was also stronger in CPu and NAc core of food-restricted relative to ad libitum fed rats. However, i.c.v. injection of saline-vehicle produced greater activation of ERK1/2 and CREB in CPu and NAc of food-restricted relative to ad libitum fed rats, and this difference was not seen when subjects received no i.c.v. injection prior to sacrifice. In addition, although DHPG activated Akt, there was no difference in Akt activation between feeding groups. To probe whether the augmented ERK1/2 and CREB activation in vehicle-injected food-restricted rats are mediated by one or more GluR types, effects of an NMDA antagonist (MK-801, 100 nmol, AMPA antagonist (DNQX, 10 nmol, and group I mGluR antagonist (AIDA, 100 nmol were compared to saline-vehicle. Antagonist injections did not diminish activation of ERK1/2 or CREB. Conclusions These results indicate that a group I mGluR agonist induces phosphorylation of Akt, ERK1/2 and CREB in both CPu and NAc. However, group I mGluR-mediated signaling may not be upregulated in food-restricted rats

  16. A subset of human pancreatic beta cells express functional CD14 receptors: a signaling pathway fot beta cell-related glycolipids, sulfatide and beta-galactosylceramide

    Czech Academy of Sciences Publication Activity Database

    Osterbye, T.; Funda, David; Fundová, Petra; Mansson, J.-E.; Tlaskalová-Hogenová, Helena; Buschard, K.

    2010-01-01

    Roč. 26, č. 8 (2010), s. 656-667. E-ISSN 1520-7560 R&D Projects: GA ČR GA303/06/1329; GA ČR GA310/07/0414; GA ČR GA310/09/1640 Institutional research plan: CEZ:AV0Z50200510 Keywords : human beta-cell * cd14 * innate immunity Subject RIV: EE - Microbiology, Virology

  17. Effects of thermal fluctuation and the receptor-receptor interaction in bacterial chemotactic signalling and adaptation

    OpenAIRE

    Shi, Yu

    2001-01-01

    Bacterial chemotaxis is controlled by the conformational changes of the receptors, in response to the change of the ambient chemical concentration. In a statistical mechanical approach, the signalling due to the conformational changes is a thermodynamic average quantity, dependent on the temperature and the total energy of the system, including both ligand-receptor interaction and receptor-receptor interaction. This physical theory suggests to biology a new understanding of cooperation in lig...

  18. Progesterone in pregnancy; receptor-ligand interaction and signaling pathways.

    Science.gov (United States)

    Szekeres-Bartho, Julia; Halasz, Melinda; Palkovics, Tamas

    2009-12-01

    Progesterone is indispensable in creating a suitable endometrial environment for implantation, and also for the maintenance of pregnancy. Successful pregnancy depends on an appropriate maternal immune response to the fetus. Along with its endocrine effects, progesterone also acts as an "immunosteroid", by contributing to the establishment of a pregnancy protective immune milieu. Progesterone plays a role in uterine homing of NK cells and upregulates HLA-G gene expression, the ligand for NK inhibitory and activating receptors. At high concentrations, progesterone is a potent inducer of Th2-type cytokines as well as of LIF and M-CSF production by T cells. A protein called progesterone-induced blocking factor (PIBF), by inducing a Th2-dominant cytokine production mediates the immunological effects of progesterone. PIBF binds to a novel type of the IL-4 receptor and signals via the Jak/STAT pathway, to induce a number of genes, that not only affect the immune response, but might also play a role in trophoblast invasiveness. PMID:19880194

  19. T cell traffic signals

    OpenAIRE

    Van Epps, Heather L.

    2005-01-01

    In 1990, Charles Mackay and colleagues combined classical physiology with modern molecular biology to provide the first concrete evidence that naive and memory T cells follow distinct migratory routes out of the bloodstream— a discovery that helped invigorate the field of lymphocyte homing.

  20. Interactions of the ubiquitous octamer-binding transcription factor-1 with both the signal transducer and activator of transcription 5 and the glucocorticoid receptor mediate prolactin and glucocorticoid-induced β-casein gene expression in mammary epithelial cells.

    Science.gov (United States)

    Qian, Xi; Zhao, Feng-Qi

    2013-03-01

    Regulation of milk protein gene expression by lactogenic hormones (prolactin and glucocorticoids) provides an attractive model for studying the mechanisms by which protein and steroid hormones synergistically regulate gene expression. β-Casein is one of the major milk proteins and its expression in mammary epithelial cells is stimulated by lactogenic hormones. The signal transducer and activator of transcription 5 and glucocorticoid receptor are essential downstream mediators of prolactin and glucocorticoid signaling, respectively. Previous studies have shown that mutating the octamer-binding site of the β-casein gene proximal promoter dramatically reduces the hormonal induction of the promoter activity. However, little is known about the underlying molecular mechanisms. In this report, we show that lactogenic hormones rapidly induce the binding of octamer-binding transcription factor-1 to the β-casein promoter and this induction is not mediated by either increasing the expression of octamer-binding transcription factor-1 or inducing its translocation to the nucleus. Rather, lactogenic hormones induce physical interactions between the octamer-binding transcription factor-1, signal transducer and activator of transcription 5, and glucocorticoid receptor to form a ternary complex, and these interactions enhance or stabilize the binding of these transcription factors to the promoter. Abolishing these interactions significantly reduces the hormonal induction of β-casein gene transcription. Thus, our study indicates that octamer-binding transcription factor-1 may serve as a master regulator that facilitates the DNA binding of both signal transducer and activator of transcription 5 and glucocorticoid receptor in hormone-induced β-casein expression, and defines a novel mechanism of regulation of tissue-specific gene expression by the ubiquitous octamer-binding transcription factor-1. PMID:23313770

  1. Berberine attenuates high glucose-induced fibrosis by activating the G protein-coupled bile acid receptor TGR5 and repressing the S1P2/MAPK signaling pathway in glomerular mesangial cells.

    Science.gov (United States)

    Yang, Zhiying; Li, Jie; Xiong, Fengxiao; Huang, Junying; Chen, Cheng; Liu, Peiqing; Huang, Heqing

    2016-08-15

    Berberine (BBR) exerts powerful renoprotective effects on diabetic nephropathy (DN), but the underlying mechanisms remain unclear. We previously demonstrated that activation of the G protein-coupled bile acid receptor TGR5 ameliorates diabetic nephropathy by inhibiting the activation of the sphingosine 1-phosphate (S1P)/sphingosine 1-phosphate receptor 2 (S1P2) signaling pathway. In this study, we explored the role of TGR5 in the BBR-induced downregulation of sphingosine 1-phosphate receptor 2 (S1P2)/mitogen-activated protein kinase (MAPK)-mediated fibrosis in glomerular mesangial cells (GMCs). Results showed that, BBR suppressed the expression of FN, ICAM-1, and TGF-β1 in high-glucose cultures of GMCs, and the phosphorylation level of c-Jun/c-Fos was downregulated. The high glucose lowered TGR5 expression in a time-dependent manner; this effect was reversed by BBR in a dose-dependent manner. The TGR5 agonist INT-777 decreased the high glucose-induced FN, ICAM-1, and TGF-β1 protein contents. In addition, TGR5 siRNA blocked S1P2 degradation by BBR. And MAPK signaling, which plays important regulatory roles in the pathological progression of DN, was activated by TGR5 siRNA. Apart from this, MAPK signaling as well as FN, ICAM-1, and TGF-β1 suppressed by BBR under high glucose conditions were limited by TGR5 depletion. Thus, BBR decreases FN, ICAM-1, and TGF-β1 levels under high glucose conditions in GMCs possibly by activating TGR5 and inhibiting S1P2/MAPK signaling. PMID:27292312

  2. Retinoic acid signalling in thymocytes regulates T cell development

    DEFF Research Database (Denmark)

    Wendland, Kerstin; Sitnik, Katarzyna Maria; Kotarsky, Knut;

    The Vitamin A derivative retinoic acid (RA) has emerged as an important regulator of peripheral T cell responses. However, whether there is endogenous retinoic acid receptor (RAR) signaling in developing thymocytes and the potential impact of such signals in thymocyte development remains unclear...

  3. DMPD: Signal transduction by the lipopolysaccharide receptor, Toll-like receptor-4. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15379975 Signal transduction by the lipopolysaccharide receptor, Toll-like receptor...-4. Palsson-McDermott EM, O'Neill LA. Immunology. 2004 Oct;113(2):153-62. (.png) (.svg) (.html) (.csml) Show Signal... transduction by the lipopolysaccharide receptor, Toll-like receptor-4. PubmedID 15379975 Title Signal

  4. P2X receptor-mediated ATP purinergic signaling in health and disease

    Directory of Open Access Journals (Sweden)

    Jiang LH

    2012-09-01

    Full Text Available Lin-Hua JiangSchool of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds, United KingdomAbstract: Purinergic P2X receptors are plasma membrane proteins present in a wide range of mammalian cells where they act as a cellular sensor, enabling cells to detect and respond to extracellular adenosine triphosphate (ATP, an important signaling molecule. P2X receptors function as ligand-gated Ca2+-permeable cationic channels that open upon ATP binding to elevate intracellular Ca2+ concentrations and cause membrane depolarization. In response to sustained activation, P2X receptors induce formation of a pore permeable to large molecules. P2X receptors also interact with distinct functional proteins and membrane lipids to form specialized signaling complexes. Studies have provided compelling evidence to show that such P2X receptor-mediated ATP-signaling mechanisms determine and regulate a growing number and diversity of important physiological processes, including neurotransmission, muscle contraction, and cytokine release. There is accumulating evidence to support strong causative relationships of altered receptor expression and function with chronic pain, inflammatory diseases, cancers, and other pathologies or diseases. Numerous high throughput screening drug discovery programs and preclinical studies have thus far demonstrated the proof of concepts that the P2X receptors are druggable targets and selective receptor antagonism is a promising therapeutics approach. This review will discuss the recent progress in understanding the mammalian P2X receptors with respect to the ATP-signaling mechanisms, physiological and pathophysiological roles, and development and preclinical studies of receptor antagonists.Keywords: extracellular ATP, ion channel, large pore, signaling complex, chronic pain, inflammatory diseases

  5. The role of GPCR dimerisation/oligomerisation in receptor signalling.

    Science.gov (United States)

    Milligan, G; Canals, M; Pediani, J D; Ellis, J; Lopez-Gimenez, J F

    2006-01-01

    (Fotiadis et al. 2004). Only for relatively few other GPCRs are details of the molecular basis of dimerisation available but within this limited data set, recent studies on the dopamine D2 receptor suggest a means by which information on the binding of an agonist can be transmitted between the two elements of the dimer via the dimer interface (Guo et al. 2005). Although the availability of cDNAs encoding molecularly defined GPCRs has allowed high-throughput screening for ligands that modulate GPCR function, this is performed almost exclusively in heterologous cell lines transfected to express only the specific GPCR of interest. Given that the human genome contains some 400-450 genes encoding non-chemosensory GPCRs, it is clear that any individual cell of the body may express a considerable number of GPCRs. Interactions between these, either via hetero-dimerisation, via heterologous desensitisation or via the integration of downstream signals can potentially alter the pharmacology, sensitivity and function of receptor agonists and hence produce varied responses. In this article, we will use specific examples to consider the role of homo-dimerisation/oligomerisation in GPCR function and whether either direct hetero-dimerisation or heterologous desensitisation between pairs of co-expressed GPCRs affects the function of the receptor pairs. PMID:17703581

  6. Ectodysplasin A (EDA) - EDA receptor signalling and its pharmacological modulation.

    Science.gov (United States)

    Kowalczyk-Quintas, Christine; Schneider, Pascal

    2014-04-01

    The TNF family ligand ectodysplasin A (EDA) regulates the induction, morphogenesis and/or maintenance of skin-derived structures such as teeth, hair, sweat glands and several other glands. Deficiencies in the EDA - EDA receptor (EDAR) signalling pathway cause hypohidrotic ectodermal dysplasia (HED). This syndrome is characterized by the absence or malformation of several skin-derived appendages resulting in hypotrychosis, hypodontia, heat-intolerance, dry skin and dry eyes, susceptibility to airways infections and crusting of various secretions. The EDA-EDAR system is an important effector of canonical Wnt signalling in developing skin appendages. It functions by stimulating NF-κB-mediated transcription of effectors or inhibitors of the Wnt, Sonic hedgehog (SHH), fibroblast growth factor (FGF) and transforming growth factor beta (TGFβ) pathways that regulate interactions within or between epithelial and mesenchymal cells and tissues. In animal models of Eda-deficiency, soluble EDAR agonists can precisely correct clinically relevant symptoms with low side effects even at high agonist doses, indicating that efficient negative feedback signals occur in treated tissues. Hijacking of the placental antibody transport system can help deliver active molecules to developing foetuses in a timely manner. EDAR agonists may serve to treat certain forms of ectodermal dysplasia. PMID:24508088

  7. Cell signalling and phospholipid metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Boss, W.F.

    1990-01-01

    These studies explored whether phosphoinositide (PI) has a role in plants analogous to its role in animal cells. Although no parallel activity of PI in signal transduction was found in plant cells, activity of inositol phospholipid kinase was found to be modulated by light and by cell wall degrading enzymes. These studies indicate a major role for inositol phospholipids in plant growth and development as membrane effectors but not as a source of second messengers.

  8. Fibroblast Growth Factor Receptor Signaling in Oligodendrocytes Regulates Myelin Sheath Thickness

    OpenAIRE

    Furusho, M.; Dupree, J. L.; Nave, K-A; Bansal, R.

    2012-01-01

    Formation of the central nervous system (CNS) white matter is developmentally tightly regulated, but the molecules and mechanisms of myelination control in the postnatal CNS are poorly understood. Here, we show that myelin growth is controlled by Fibroblast Growth Factor (FGF) signaling, originally identified as a proliferative signal for oligodendrocyte precursor cells (OPC) in vitro. We created two lines of mice lacking both FGF-receptor 1 (Fgfr1) and Fgfr2 in oligodendrocyte lineage cells ...

  9. Role of Phospholipase C-L2, a Novel Phospholipase C-Like Protein That Lacks Lipase Activity, in B-Cell Receptor Signaling

    OpenAIRE

    Takenaka, Kei; Fukami, Kiyoko; Otsuki, Makiko; Nakamura, Yoshikazu; Kataoka, Yuki; Wada, Mika; Tsuji, Kohichiro; Nishikawa, Shin-ichi; Yoshida, Nobuaki; Takenawa, Tadaomi

    2003-01-01

    Phospholipase C (PLC) plays important roles in phosphoinositide turnover by regulating the calcium-protein kinase C signaling pathway. PLC-L2 is a novel PLC-like protein which lacks PLC activity, although it is very homologous with PLC δ. PLC-L2 is expressed in hematopoietic cells, but its physiological roles and intracellular functions in the immune system have not yet been clarified. To elucidate the physiological function of PLC-L2, we generated mice which had a genetic PLC-L2 deficiency. ...

  10. Modulation of β-catenin signaling by glucagon receptor activation.

    Directory of Open Access Journals (Sweden)

    Jiyuan Ke

    Full Text Available The glucagon receptor (GCGR is a member of the class B G protein-coupled receptor family. Activation of GCGR by glucagon leads to increased glucose production by the liver. Thus, glucagon is a key component of glucose homeostasis by counteracting the effect of insulin. In this report, we found that in addition to activation of the classic cAMP/protein kinase A (PKA pathway, activation of GCGR also induced β-catenin stabilization and activated β-catenin-mediated transcription. Activation of β-catenin signaling was PKA-dependent, consistent with previous reports on the parathyroid hormone receptor type 1 (PTH1R and glucagon-like peptide 1 (GLP-1R receptors. Since low-density-lipoprotein receptor-related protein 5 (Lrp5 is an essential co-receptor required for Wnt protein mediated β-catenin signaling, we examined the role of Lrp5 in glucagon-induced β-catenin signaling. Cotransfection with Lrp5 enhanced the glucagon-induced β-catenin stabilization and TCF promoter-mediated transcription. Inhibiting Lrp5/6 function using Dickkopf-1(DKK1 or by expression of the Lrp5 extracellular domain blocked glucagon-induced β-catenin signaling. Furthermore, we showed that Lrp5 physically interacted with GCGR by immunoprecipitation and bioluminescence resonance energy transfer assays. Together, these results reveal an unexpected crosstalk between glucagon and β-catenin signaling, and may help to explain the metabolic phenotypes of Lrp5/6 mutations.

  11. Human herpesvirus 6 infection impairs Toll-like receptor signaling

    Directory of Open Access Journals (Sweden)

    Ochi Toshiki

    2010-05-01

    Full Text Available Abstract Human herpesvirus 6 (HHV-6 has a tropism for immunocompetent cells, including T lymphocytes, monocytes/macrophages, and dendritic cells (DCs suggesting that HHV-6 infection affects the immunosurveillance system. Toll-like receptor (TLR system plays an important role in innate immunity against various pathogens. In the present study, we investigated the effect of HHV-6 infection on the expression and intracellular signaling of TLRs in DCs. Although expression levels of TLRs were not decreased or slightly elevated following HHV-6 infection, the amounts of cytokines produced following stimulation with ligands for TLRs appeared to be dramatically decreased in HHV-6-infected DCs as compared to mock-infected DCs. Similarly, phosphorylation levels of TAK-1, IκB kinase, and IκB-α following stimulation of HHV-6-infected DCs with lipopolysaccharide, which is the ligand for TLR4, appeared to be decreased. These data show that HHV-6 impairs intracellular signaling through TLRs indicating the novel mechanism of HHV-6-mediated immunomodulation.

  12. Regulator of G protein signaling 2 (RGS2 and RGS4 form distinct G protein-dependent complexes with protease activated-receptor 1 (PAR1 in live cells.

    Directory of Open Access Journals (Sweden)

    Sungho Ghil

    Full Text Available Protease-activated receptor 1 (PAR1 is a G-protein coupled receptor (GPCR that is activated by natural proteases to regulate many physiological actions. We previously reported that PAR1 couples to Gi, Gq and G12 to activate linked signaling pathways. Regulators of G protein signaling (RGS proteins serve as GTPase activating proteins to inhibit GPCR/G protein signaling. Some RGS proteins interact directly with certain GPCRs to modulate their signals, though cellular mechanisms dictating selective RGS/GPCR coupling are poorly understood. Here, using bioluminescence resonance energy transfer (BRET, we tested whether RGS2 and RGS4 bind to PAR1 in live COS-7 cells to regulate PAR1/Gα-mediated signaling. We report that PAR1 selectively interacts with either RGS2 or RGS4 in a G protein-dependent manner. Very little BRET activity is observed between PAR1-Venus (PAR1-Ven and either RGS2-Luciferase (RGS2-Luc or RGS4-Luc in the absence of Gα. However, in the presence of specific Gα subunits, BRET activity was markedly enhanced between PAR1-RGS2 by Gαq/11, and PAR1-RGS4 by Gαo, but not by other Gα subunits. Gαq/11-YFP/RGS2-Luc BRET activity is promoted by PAR1 and is markedly enhanced by agonist (TFLLR stimulation. However, PAR1-Ven/RGS-Luc BRET activity was blocked by a PAR1 mutant (R205A that eliminates PAR1-Gq/11 coupling. The purified intracellular third loop of PAR1 binds directly to purified His-RGS2 or His-RGS4. In cells, RGS2 and RGS4 inhibited PAR1/Gα-mediated calcium and MAPK/ERK signaling, respectively, but not RhoA signaling. Our findings indicate that RGS2 and RGS4 interact directly with PAR1 in Gα-dependent manner to modulate PAR1/Gα-mediated signaling, and highlight a cellular mechanism for selective GPCR/G protein/RGS coupling.

  13. Neuromedin B receptors regulate EGF receptor tyrosine phosphorylation in lung cancer cells

    OpenAIRE

    Moody, Terry W.; Berna, Marc J.; Mantey, Samuel; Sancho, Veronica; Ridnour, Lisa; Wink, David A.; Chan, Daniel; Giaccone, Giuseppe; Jensen, Robert T.

    2010-01-01

    Neuromedin B (NMB), a member of the bombesin family of peptides, is an autocrine growth factor for many lung cancer cells. The present study investigated the ability of NMB to cause transactivation of the epidermal growth factor (EGF) receptor in lung cancer cells. By Western blot, addition of NMB or related peptides to NCI-H1299 human non-small cell lung cancer (NSCLC) cells, caused phosphorylation of Tyr1068 of the EGF receptor. The signal was amplified using NCI-H1299 cells stably transect...

  14. 4-tert-Octylphenol stimulates the expression of cathepsins in human breast cancer cells and xenografted breast tumors of a mouse model via an estrogen receptor-mediated signaling pathway

    International Nuclear Information System (INIS)

    Highlights: ► Cathepsins B and D were markedly enhanced by octylphenol (OP) in MCF-7 cells. ► OP may accelerate breast cancer cell growth and cathepsins via ER-mediated signaling. ► Breast cancer cells exposed with OP to mouse model were more aggressive. ► OP can promote metastasis through the amplification of cathepsins B and D via ER-mediated signaling pathway. -- Abstract: Endocrine disrupting chemicals (EDCs) are defined as environmental compounds that modulate steroid hormone receptor-dependent responses an abnormal manner, resulting in adverse health problems for humans such as cancer growth and metastasis. Cathepsins are proteases that have been implicated in cancer progression. However, there have been few studies about the association between cathepsins and estrogenic chemicals during the cancer progression. In this study, we examined the effect(s) of 4-tert-octylphenol (OP), a potent EDC, on the expression of cathepsins B and D in human MCF-7 breast cancer cells and a xenograft mouse model. Treatment with OP significantly induced the proliferation MCF-7 cells in an MTT assay. In addition, the expression of cathepsins B and D was markedly enhanced in MCF-7 cells at both the transcriptional and the translational levels following treatment with E2 or OP up to 48 h. These results demonstrated the ability of OP to disrupt normal transcriptional regulation of cathepsins B and D in human breast cancer cells. However, the effects of OP on cell growth or overexpression of cathepsins by inhibiting ER-mediated signaling were abolished by an ER antagonist and siRNA specific for ERα. In conclusion, our findings suggest that OP at 10−6 M, like E2, may accelerate breast cancer cell proliferation and the expression of cathepsins through an ER-mediated signaling pathway. In addition, the breast cancer cells exposed with OP to a xenograft mouse model were more aggressive according to our histological analysis and showed markedly increased expression of cathepsin

  15. Toll-like receptor-2 mediates mycobacteria-induced proinflammatory signaling in macrophages

    OpenAIRE

    Underhill, David M; Ozinsky, Adrian; Smith, Kelly D.; Aderem, Alan

    1999-01-01

    The recognition of mycobacterial cell wall components causes macrophages to secrete tumor necrosis factor α (TNF-α) and other cytokines that are essential for the development of a protective inflammatory response. We show that toll-like receptors are required for the induction of TNF-α in macrophages by Mycobacterium tuberculosis. Expression of a dominant negative form of MyD88 (a signaling component required for toll-like receptor signaling) in a mouse macrophage cell line blocks TNF-α produ...

  16. Tumor Necrosis Factor Receptor-associated Protein 1 (TRAP1) Mutation and TRAP1 Inhibitor Gamitrinib-triphenylphosphonium (G-TPP) Induce a Forkhead Box O (FOXO)-dependent Cell Protective Signal from Mitochondria.

    Science.gov (United States)

    Kim, Hyunjin; Yang, Jinsung; Kim, Min Ju; Choi, Sekyu; Chung, Ju-Ryung; Kim, Jong-Min; Yoo, Young Hyun; Chung, Jongkyeong; Koh, Hyongjong

    2016-01-22

    TRAP1 (tumor necrosis factor receptor-associated protein 1), a mitochondrial Hsp90 family chaperone, has been identified as a critical regulator of cell survival and bioenergetics in tumor cells. To discover novel signaling networks regulated by TRAP1, we generated Drosophila TRAP1 mutants. The mutants successfully developed into adults and produced fertile progeny, showing that TRAP1 is dispensable in development and reproduction. Surprisingly, mutation or knockdown of TRAP1 markedly enhanced Drosophila survival under oxidative stress. Moreover, TRAP1 mutation ameliorated mitochondrial dysfunction and dopaminergic (DA) neuron loss induced by deletion of a familial Parkinson disease gene PINK1 (Pten-induced kinase 1) in Drosophila. Gamitrinib-triphenylphosphonium, a mitochondria-targeted Hsp90 inhibitor that increases cell death in HeLa and MCF7 cells, consistently inhibited cell death induced by oxidative stress and mitochondrial dysfunction induced by PINK1 mutation in mouse embryonic fibroblast cells and DA cell models such as SH-SY5Y and SN4741 cells. Additionally, gamitrinib-triphenylphosphonium also suppressed the defective locomotive activity and DA neuron loss in Drosophila PINK1 null mutants. In further genetic analyses, we showed enhanced expression of Thor, a downstream target gene of transcription factor FOXO, in TRAP1 mutants. Furthermore, deletion of FOXO almost nullified the protective roles of TRAP1 mutation against oxidative stress and PINK1 mutation. These results strongly suggest that inhibition of the mitochondrial chaperone TRAP1 generates a retrograde cell protective signal from mitochondria to the nucleus in a FOXO-dependent manner. PMID:26631731

  17. Application of computational approaches to study signalling networks of nuclear and Tyrosine kinase receptors

    Directory of Open Access Journals (Sweden)

    Rebaï Ahmed

    2010-10-01

    Full Text Available Abstract Background Nuclear receptors (NRs and Receptor tyrosine kinases (RTKs are essential proteins in many cellular processes and sequence variations in their genes have been reported to be involved in many diseases including cancer. Although crosstalk between RTK and NR signalling and their contribution to the development of endocrine regulated cancers have been areas of intense investigation, the direct coupling of their signalling pathways remains elusive. In our understanding of the role and function of nuclear receptors on the cell membrane the interactions between nuclear receptors and tyrosine kinase receptors deserve further attention. Results We constructed a human signalling network containing nuclear receptors and tyrosine kinase receptors that identified a network topology involving eleven highly connected hubs. We further developed an integrated knowledge database, denominated NR-RTK database dedicated to human RTKs and NRs and their vertebrate orthologs and their interactions. These interactions were inferred using computational tools and those supported by literature evidence are indicated. NR-RTK database contains links to other relevant resources and includes data on receptor ligands. It aims to provide a comprehensive interaction map that identifies complex dynamics and potential crosstalk involved. Availability: NR-RTK database is accessible at http://www.bioinfo-cbs.org/NR-RTK/ Conclusions We infer that the NR-RTK interaction network is scale-free topology. We also uncovered the key receptors mediating the signal transduction between these two types of receptors. Furthermore, NR-RTK database is expected to be useful for researchers working on various aspects of the molecular basis of signal transduction by RTKs and NRs. Reviewers This article was reviewed by Professor Paul Harrison (nominated by Dr. Mark Gerstein, Dr. Arcady Mushegian and Dr. Anthony Almudevar.

  18. Functional significance of erythropoietin receptor on tumor cells

    Institute of Scientific and Technical Information of China (English)

    Kodetthoor B Udupa

    2006-01-01

    Erythropoietin (Epo) is the regulator of red blood cell formation. Its receptor (EpoR) is now found in many cells and tissues of the body. EpoR is also shown to occur in tumor cells and Epo enhances the proliferation of these cells through cell signaling. EpoR antagonist can reduce the growth of the tumor in vivo. In view of our current knowledge of Epo, its recombinant forms and receptor,use of Epo in cancer patients to enhance the recovery of hematocrit after chemotherapy treatment has to be carefully evaluated.

  19. Advances of Targeted Therapy Based on Estrogen Receptor Signaling Pathway 
in Lung Cancer

    OpenAIRE

    Xu, Liqiang; Liao, Yongde; Hexiao TANG; Zhang, Chao; Liu, Zhaoguo

    2011-01-01

    Increasing evidence indicates that estrogen promotes tumor growth in both estrogen target organs and non-target organs. Estrogen regulates cell proliferation and differentiation via two different receptors, estrogen receptors α and β (ERα and ERβ). In recent decades, with the clarification of the ERα-mediated signaling pathways in breast cancer, targeted therapy through these pathways have successfully been used in clinical application. Tamoxifen, the classic representative, is a selective es...

  20. Role of serotonergic signaling in GABAA receptor phosphorylation and functional expression

    OpenAIRE

    Vithlani, M.

    2009-01-01

    γ-aminobutyric acid type-A (GABAA) receptors are heteropentameric ligand-gated chloride channels that mediate the majority of fast synaptic inhibition in the brain. Emerging evidence indicates that their functional expression is subject to dynamic modulation by phosphorylation. However, the cell signaling molecules responsible for regulating GABAA receptor phosphorylation and thus the efficacy of neuronal inhibition remain to be identified. The β subunits are of particular interest in this co...

  1. Insulin glulisine: insulin receptor signaling characteristics in vivo.

    Science.gov (United States)

    Hennige, Anita M; Lehmann, Rainer; Weigert, Cora; Moeschel, Klaus; Schäuble, Myriam; Metzinger, Elisabeth; Lammers, Reiner; Häring, Hans-Ulrich

    2005-02-01

    In recent years, recombinant DNA technology has been used to design insulin molecules that overcome the limitations of regular insulin in mealtime supplementation. However, safety issues have been raised with these alternatives, as the alteration of the three-dimensional structure may alter the interaction with the insulin and/or IGF-I receptors and therefore lead to the activation of alternate metabolic as well as mitogenic signaling pathways. It is therefore essential to carefully study acute and long-term effects in a preclinical state, as insulin therapy is meant to be a lifelong treatment. In this study, we determined in vivo the insulin receptor signaling characteristics activated by insulin glulisine (Lys(B3), Glu(B29)) at the level of insulin receptor phosphorylation, insulin receptor substrate phosphorylation, and downstream signaling elements such as phosphatidylinositol (PI) 3-kinase, AKT, and mitogen-activated protein kinase. C57BL/6 mice were injected with insulin glulisine or regular insulin and Western blot analysis was performed for liver and muscle tissue. The extent and time course of insulin receptor phosphorylation and activation of downstream signaling elements after insulin glulisine treatment was similar to that of human regular insulin in vivo. Moreover, insulin signaling in hypothalamic tissue determined by PI 3-kinase activity was comparable. Therefore, insulin glulisine may be a useful tool for diabetes treatment. PMID:15677493

  2. Documentation of angiotensin II receptors in glomerular epithelial cells

    Science.gov (United States)

    Sharma, M.; Sharma, R.; Greene, A. S.; McCarthy, E. T.; Savin, V. J.; Cowley, A. W. (Principal Investigator)

    1998-01-01

    Angiotensin II decreases glomerular filtration rate, renal plasma flow, and glomerular capillary hydraulic conductivity. Although angiotensin II receptors have been demonstrated in mesangial cells and proximal tubule cells, the presence of angiotensin II receptors in glomerular epithelial cells has not previously been shown. Previously, we have reported that angiotensin II caused an accumulation of cAMP and a reorganization of the actin cytoskeleton in cultured glomerular epithelial cells. Current studies were conducted to verify the presence of angiotensin II receptors by immunological and non-peptide receptor ligand binding techniques and to ascertain the activation of intracellular signal transduction in glomerular epithelial cells in response to angiotensin II. Confluent monolayer cultures of glomerular epithelial cells were incubated with angiotensin II, with or without losartan and/or PD-123,319 in the medium. Membrane vesicle preparations were obtained by homogenization of washed cells followed by centrifugation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of membrane proteins followed by multiscreen immunoblotting was used to determine the presence of angiotensin II receptor type 1 (AT1) or type 2 (AT2). Angiotensin II-mediated signal transduction in glomerular epithelial cells was studied by measuring the levels of cAMP, using radioimmunoassay. Results obtained in these experiments showed the presence of both AT1 and AT2 receptor types in glomerular epithelial cells. Angiotensin II was found to cause an accumulation of cAMP in glomerular epithelial cells, which could be prevented only by simultaneous use of losartan and PD-123,319, antagonists for AT1 and AT2, respectively. The presence of both AT1 and AT2 receptors and an increase in cAMP indicate that glomerular epithelial cells respond to angiotensin II in a manner distinct from that of mesangial cells or proximal tubular epithelial cells. Our results suggest that glomerular epithelial

  3. Nanoconjugation prolongs endosomal signaling of the epidermal growth factor receptor and enhances apoptosis

    Science.gov (United States)

    Wu, L.; Xu, F.; Reinhard, B. M.

    2016-07-01

    mediated apoptosis at effective concentrations that do not induce apoptosis in the case of free EGF. Overall, these findings indicate nanoconjugation as a rational strategy for modifying signaling that acts by modulating the temporo-spatial distribution of the activated EGF-EGFR ligand-receptor complex. Electronic supplementary information (ESI) available: DLS data of NP-EGF in growth medium; MTT cell viability assay; validation of MW-NP uptake; positive controls for pharmacological inhibitors; EEA1 background for NP-EGF incubated with cell lysate; phosphorylation for EGF-Alexa647; live cell dynamic colocalization movie of MDA-MB-468 cells expressing Rab5a-GFP (green) 4.5 h after exposure to 8 pM NP-EGF (red). See DOI: 10.1039/c6nr02974d

  4. SOCS proteins in regulation of receptor tyrosine kinase signaling

    DEFF Research Database (Denmark)

    Kazi, Julhash U.; Kabir, Nuzhat N.; Flores Morales, Amilcar;

    2014-01-01

    signaling mediated by RTKs must be tightly regulated by interacting proteins including protein-tyrosine phosphatases and ubiquitin ligases. The suppressors of cytokine signaling (SOCS) family proteins are well-known negative regulators of cytokine receptors signaling consisting of eight structurally similar...... proteins, SOCS1-7, and cytokine-inducible SH2-containing protein (CIS). A key feature of this family of proteins is the presence of an SH2 domain and a SOCS box. Recent studies suggest that SOCS proteins also play a role in RTK signaling. Activation of RTK results in transcriptional activation of SOCS......-encoding genes. These proteins associate with RTKs through their SH2 domains and subsequently recruit the E3 ubiquitin machinery through the SOCS box, and thereby limit receptor stability by inducing ubiquitination. In a similar fashion, SOCS proteins negatively regulate mitogenic signaling by RTKs. It is also...

  5. Evolution of retinoic acid receptors and retinoic acid signaling.

    Science.gov (United States)

    Gutierrez-Mazariegos, Juliana; Schubert, Michael; Laudet, Vincent

    2014-01-01

    Retinoic acid (RA) is a vitamin A-derived morphogen controlling important developmental processes in vertebrates, and more generally in chordates, including axial patterning and tissue formation and differentiation. In the embryo, endogenous RA levels are controlled by RA synthesizing and degrading enzymes and the RA signal is transduced by two retinoid receptors: the retinoic acid receptor (RAR) and the retinoid X receptor (RXR). Both RAR and RXR are members of the nuclear receptor superfamily of ligand-activated transcription factors and mainly act as heterodimers to activate the transcription of target genes in the presence of their ligand, all-trans RA. This signaling pathway was long thought to be a chordate innovation, however, recent findings of gene homologs involved in RA signaling in the genomes of a wide variety of non-chordate animals, including ambulacrarians (sea urchins and acorn worms) and lophotrochozoans (annelids and mollusks), challenged this traditional view and suggested that the RA signaling pathway might have a more ancient evolutionary origin than previously thought. In this chapter, we discuss the evolutionary history of the RA signaling pathway, and more particularly of the RARs, which might have experienced independent gene losses and duplications in different animal lineages. In sum, the available data reveal novel insights into the origin of the RA signaling pathway as well as into the evolutionary history of the RARs. PMID:24962881

  6. Role of CD4 molecule in the induction of interleukin 2 and interleukin 2 receptor in class II major histocompatibility complex-restricted antigen-specific T helper clones. T cell receptor/CD3 complex transmits CD4-dependent and CD4-independent signals.

    OpenAIRE

    Oyaizu, N; Chirmule, N; Pahwa, S.

    1992-01-01

    The CD4 molecule plays an essential role in antigen-induced activation of T helper (Th) cells, but its contribution to signal transduction events resulting in physiologic T cell function is ill defined. By utilizing anti-CD4 monoclonal antibodies (MAbs) that recognize distinct epitopes of CD4, we have investigated the role of CD4 molecule in antigen-induced interleukin 2 (IL-2) and IL-2 receptor (IL-2R) alpha chain expression in class II major histocompatibility complex-restricted antigen-spe...

  7. NMU signaling promotes endometrial cancer cell progression by modulating adhesion signaling.

    Science.gov (United States)

    Lin, Ting-Yu; Wu, Fang-Ju; Chang, Chia-Lin; Li, Zhongyou; Luo, Ching-Wei

    2016-03-01

    Neuromedin U (NMU) was originally named based on its strong uterine contractile activity, but little is known regarding its signaling/functions in utero. We identified that NMU and one of its receptors, NMUR2, are not only present in normal uterine endometrium but also co-expressed in endometrial cancer tissues, where the NMU level is correlated with the malignant grades and survival of patients. Cell-based assays further confirmed that NMU signaling can promote cell motility and proliferation of endometrial cancer cells derived from grade II tumors. Activation of NMU pathway in these endometrial cancer cells is required in order to sustain expression of various adhesion molecules, such as CD44 and integrin alpha1, as well as production of their corresponding extracellular matrix ligands, hyaluronan and collagen IV; it also increased the activity of SRC and its downstream proteins RHOA and RAC1. Thus, it is concluded that NMU pathway positively controls the adhesion signaling-SRC-Rho GTPase axis in the tested endometrial cancer cells and that changes in cell motility and proliferation can occur when there is manipulation of NMU signaling in these cells either in vitro or in vivo. Intriguingly, this novel mechanism also explains how NMU signaling promotes the EGFR-driven and TGFβ receptor-driven mesenchymal transitions. Through the above axis, NMU signaling not only can promote malignancy of the tested endometrial cancer cells directly, but also helps these cells to become more sensitive to niche growth factors in their microenvironment. PMID:26849234

  8. Vitamin D Receptor Signaling Inhibits Atherosclerosis in Mice

    OpenAIRE

    Szeto, Frances L.; Reardon, Catherine A.; Yoon, Dosuk; Wang, Youli; Wong, Kari E.; Chen, Yunzi; Kong, Juan; Shu Q. Liu; Thadhani, Ravi; Getz, Godfrey S; Li, Yan Chun

    2012-01-01

    Although vitamin D has been implicated in cardiovascular protection, few studies have addressed the role of vitamin D receptor (VDR) in atherosclerosis. Here we investigate the effect of inactivation of the VDR signaling on atherogenesis and the antiatherosclerotic mechanism of vitamin D. Low density lipoprotein receptor (LDLR)−/−/VDR−/− mice exhibited site-specific accelerated atherogenesis, accompanied by increases in adhesion molecules and proinflammatory cytokines in the aorta and cholest...

  9. Chromatin remodeling by curcumin alters endogenous aryl hydrocarbon receptor signaling.

    Science.gov (United States)

    Mohammadi-Bardbori, Afshin; Akbarizadeh, Amin Reza; Delju, Fatemeh; Rannug, Agneta

    2016-05-25

    The aim of this study was to gain more information about the mechanisms that regulate expression of the aryl hydrocarbon receptor (AHR) target gene CYP1A1. Human hepatoma cells (HepG2 and Huh7) and human immortalized keratinocytes (HaCaT) were treated with different concentrations of the dietary polyphenolic compound curcumin (CUR) alone or in combination with the natural AHR agonist 6-formylindolo[3,2-b]carbazole (FICZ). In an earlier study, we described that CUR can activate the AHR indirectly by inhibiting metabolic clearance of FICZ. Here, we measured cell viability, activation of AHR signaling, oxidative stress and histone modifying activities in response to CUR at concentrations ranging from 0.1 to 50 μM. We observed apparent non-linear responses on cell viability and activation of AHR signaling. The CYP1A1 expression and the CYP1A1 enzyme activity in the presence of CUR reflected the histone acetylation efficiency observed in nuclear extracts. At the lowest concentration, CUR significantly decreased histone deacetylase activity and increased the FICZ-induced CYP1A1 activity. In contrast, at the highest concentration, CUR increased the formation of reactive oxygen species, significantly inhibited histone acetylation, and temporally decreased FICZ-induced CYP1A1 activity. The results suggest that CUR can both increase and decrease the accessibility of DNA and thereby influence transcriptional responses to the ligand-activated AHR. This suggestion was supported by the fact that chromatin remodeling treatments with trichostatin A, p300, or 5-aza-dC increased CYP1A1 transcription. We conclude that the AHR-dependent transcriptional efficiency is modified by factors that influence the cellular redox status and the chromatin structure. PMID:27041069

  10. Estrogen and pure antiestrogen fulvestrant (ICI 182 780) augment cell–matrigel adhesion of MCF-7 breast cancer cells through a novel G protein coupled estrogen receptor (GPR30)-to-calpain signaling axis

    International Nuclear Information System (INIS)

    Fulvestrant (ICI 182 780, ICI) has been used in treating patients with hormone-sensitive breast cancer, yet initial or acquired resistance to endocrine therapies frequently arises and, in particular, cancer recurs as metastasis. We demonstrate here that both 17-beta-estradiol (E2) and ICI enhance cell adhesion to matrigel in MCF-7 breast cancer cells, with increased autolysis of calpain 1 (large subunit) and proteolysis of focal adhesion kinase (FAK), indicating calpain activation. Additionally, either E2 or ICI induced down-regulation of estrogen receptor α without affecting G protein coupled estrogen receptor 30 (GPR30) expression. Interestingly, GPR30 agonist G1 triggered calpain 1 autolysis but not calpain 2, whereas ER agonist diethylstilbestrol caused no apparent calpain autolysis. Furthermore, the actions of E2 and ICI on calpain and cell adhesion were tremendously suppressed by G15, or knockdown of GPR30. E2 and ICI also induced phosphorylation of extracellular regulated protein kinases 1 and 2 (ERK1/2), and suppression of ERK1/2 phosphorylation by U0126 profoundly impeded calpain activation triggered by estrogenic and antiestrogenic stimulations indicating implication of ERK1/2 in the GPR30-mediated action. Lastly, the E2- or ICI-induced cell adhesion was dramatically impaired by calpain-specific inhibitors, ALLN or calpeptin, suggesting requirement of calpain in the GPR30-associated action. These data show that enhanced cell adhesion by E2 and ICI occurs via a novel GPR30-ERK1/2-calpain pathway. Our results indicate that targeting the GPR30 signaling may be a potential strategy to reduce metastasis and improve the efficacy of antiestrogens in treatment of advanced breast cancer. - Highlights: • Estrogen and ICI augment adhesion to matrigel with calpain activation in MCF-7 cells. • GPR30 mediates cell–matrigel adhesion and calpain activation via ERK1/2. • Calpain is required in the cell–matrigel adhesion induced by E2 and ICI

  11. Estrogen and pure antiestrogen fulvestrant (ICI 182 780) augment cell–matrigel adhesion of MCF-7 breast cancer cells through a novel G protein coupled estrogen receptor (GPR30)-to-calpain signaling axis

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Yan; Li, Zheng; He, Yan; Shang, Dandan; Pan, Jigang; Wang, Hongmei; Chen, Huamei; Zhu, Zhuxia [Department of Physiology/Cancer Research Group, Guiyang Medical University School of Basic Medicine, 9 Beijing Road, Guiyang 550004, Guizhou (China); Wan, Lei [Department of Pharmacology, Guiyang Medical University School of Basic Medicine, 9 Beijing Road, Guiyang 550004, Guizhou (China); Wang, Xudong, E-mail: xdwang@gmc.edu.cn [Department of Physiology/Cancer Research Group, Guiyang Medical University School of Basic Medicine, 9 Beijing Road, Guiyang 550004, Guizhou (China)

    2014-03-01

    Fulvestrant (ICI 182 780, ICI) has been used in treating patients with hormone-sensitive breast cancer, yet initial or acquired resistance to endocrine therapies frequently arises and, in particular, cancer recurs as metastasis. We demonstrate here that both 17-beta-estradiol (E2) and ICI enhance cell adhesion to matrigel in MCF-7 breast cancer cells, with increased autolysis of calpain 1 (large subunit) and proteolysis of focal adhesion kinase (FAK), indicating calpain activation. Additionally, either E2 or ICI induced down-regulation of estrogen receptor α without affecting G protein coupled estrogen receptor 30 (GPR30) expression. Interestingly, GPR30 agonist G1 triggered calpain 1 autolysis but not calpain 2, whereas ER agonist diethylstilbestrol caused no apparent calpain autolysis. Furthermore, the actions of E2 and ICI on calpain and cell adhesion were tremendously suppressed by G15, or knockdown of GPR30. E2 and ICI also induced phosphorylation of extracellular regulated protein kinases 1 and 2 (ERK1/2), and suppression of ERK1/2 phosphorylation by U0126 profoundly impeded calpain activation triggered by estrogenic and antiestrogenic stimulations indicating implication of ERK1/2 in the GPR30-mediated action. Lastly, the E2- or ICI-induced cell adhesion was dramatically impaired by calpain-specific inhibitors, ALLN or calpeptin, suggesting requirement of calpain in the GPR30-associated action. These data show that enhanced cell adhesion by E2 and ICI occurs via a novel GPR30-ERK1/2-calpain pathway. Our results indicate that targeting the GPR30 signaling may be a potential strategy to reduce metastasis and improve the efficacy of antiestrogens in treatment of advanced breast cancer. - Highlights: • Estrogen and ICI augment adhesion to matrigel with calpain activation in MCF-7 cells. • GPR30 mediates cell–matrigel adhesion and calpain activation via ERK1/2. • Calpain is required in the cell–matrigel adhesion induced by E2 and ICI.

  12. Cannabinoid receptor-interacting protein Crip1a modulates CB1 receptor signaling in mouse hippocampus.

    Science.gov (United States)

    Guggenhuber, Stephan; Alpar, Alan; Chen, Rongqing; Schmitz, Nina; Wickert, Melanie; Mattheus, Tobias; Harasta, Anne E; Purrio, Martin; Kaiser, Nadine; Elphick, Maurice R; Monory, Krisztina; Kilb, Werner; Luhmann, Heiko J; Harkany, Tibor; Lutz, Beat; Klugmann, Matthias

    2016-05-01

    The cannabinoid type 1 receptor (Cnr1, CB1R) mediates a plethora of physiological functions in the central nervous system as a presynaptic modulator of neurotransmitter release. The recently identified cannabinoid receptor-interacting protein 1a (Cnrip1a, CRIP1a) binds to the C-terminal domain of CB1R, a region known to be important for receptor desensitization and internalization. Evidence that CRIP1a and CB1R interact in vivo has been reported, but the neuroanatomical distribution of CRIP1a is unknown. Moreover, while alterations of hippocampal CRIP1a levels following limbic seizures indicate a role in controlling excessive neuronal activity, the physiological function of CRIP1a in vivo has not been investigated. In this study, we analyzed the spatial distribution of CRIP1a in the hippocampus and examined CRIP1a as a potential modulator of CB1R signaling. We found that Cnrip1a mRNA is co-expressed with Cnr1 mRNA in pyramidal neurons and interneurons of the hippocampal formation. CRIP1a protein profiles were largely segregated from CB1R profiles in mossy cell terminals but not in hippocampal CA1 region. CB1R activation induced relocalization to close proximity with CRIP1a. Adeno-associated virus-mediated overexpression of CRIP1a specifically in the hippocampus revealed that CRIP1a modulates CB1R activity by enhancing cannabinoid-induced G protein activation. CRIP1a overexpression extended the depression of excitatory currents by cannabinoids in pyramidal neurons of the hippocampus and diminished the severity of chemically induced acute epileptiform seizures. Collectively, our data indicate that CRIP1a enhances hippocampal CB1R signaling in vivo. PMID:25772509

  13. Functional erythropoietin receptors on human tumor cells

    International Nuclear Information System (INIS)

    Erythropoietin (EPO) is the principal regulator of red blood cell survival, growth and maturation and has achieved great clinical utility for the correction of anemia associated with renal failure, cancer and chemotherapy, and stem cell transplantation. EPO increasingly is being recognized as a pleiotrophic growth factor, having actions on nonhematopoietic cells as well. Both EPO and erythropoietin receptor (EPO-R) expression have been associated with cells of the endothelium, retina, central nervous system, gastrointestinal tract and female reproductive system. The role of EPO in these nonhematopoietic sites is not thoroughly understood and in some instances may be site-specific. Promotion of angiogenesis and blood vessel integrity, increased cell proliferation, prevention of apoptosis, and protection against ischemic damage in the presence of hypoxia have all been described as possible functions of EPO in one or more of these cell types. On the other hand, EPO-R also have been identified on a variety of tumor cells (while in some cases not on the adjacent normal tissue), and several reports have suggested a role for EPO in the direct stimulation of cancer cell growth in vivo and in vitro. Among those tumor cells on which we and others have identified functional EPO-R are breast and ovarian cancer cells. Additionally, the work presented here describes the first evidence that transformed prostate epithelial cells, prostate cancer cell lines, and both normal and cancerous prostate tissue express EPO-R. All of the EPO-R bearing prostate cell lines tested underwent a significant dose-dependent proliferative response to EPO, and EPO triggered intracellular signaling in the cells as evidenced by protein phosphorylation. The results implicate EPO in the biology of both normal and malignant prostate cells and suggest the need for careful evaluation of the use of recombinant EPO as a therapeutic agent in prostate cancer

  14. Patency of Litomosoides sigmodontis infection depends on Toll-like receptor 4 whereas Toll-like receptor 2 signalling influences filarial-specific CD4(+) T-cell responses.

    Science.gov (United States)

    Rodrigo, Maria B; Schulz, Sandy; Krupp, Vanessa; Ritter, Manuel; Wiszniewsky, Katharina; Arndts, Kathrin; Tamadaho, Ruth S E; Endl, Elmar; Hoerauf, Achim; Layland, Laura E

    2016-04-01

    BALB/c mice develop a patent state [release of microfilariae (Mf), the transmission life-stage, into the periphery] when exposed to the rodent filariae Litomosoides sigmodontis. Interestingly, only a portion of the infected mice become patent, which reflects the situation in human individuals infected with Wuchereria bancrofti. Since those individuals had differing filarial-specific profiles, this study compared differences in immune responses between Mf(+) and Mf(-) infected BALB/c mice. We demonstrate that cultures of total spleen or mediastinal lymph node cells from Mf(+) mice produce significantly more interleukin-5 (IL-5) to filarial antigens but equal levels of IL-10 when compared with Mf(-) mice. However, isolated CD4(+) T cells from Mf(+) mice produced significantly higher amounts of all measured cytokines, including IL-10, when compared with CD4(+) T-cell responses from Mf(-) mice. Since adaptive immune responses are influenced by triggering the innate immune system we further studied the immune profiles and parasitology in infected Toll-like receptor-2-deficient (TLR2(-/-)) and TLR4(-/-) BALB/c mice. Ninety-three per cent of L. sigmodontis-exposed TLR4(-/-) BALB/c mice became patent (Mf(+)) although worm numbers remained comparable to those in Mf(+) wild-type controls. Lack of TLR2 had no influence on patency outcome or worm burden but infected Mf(+) mice had significantly lower numbers of Foxp3(+) regulatory T cells and dampened peripheral immune responses. Interestingly