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Sample records for activated receptor signaling

  1. Biased Signaling of Protease-activated Receptors

    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...

  2. Modulation of β-catenin signaling by glucagon receptor activation.

    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.

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

    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...

  4. Monocyte Signal Transduction Receptors in Active and Latent Tuberculosis

    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.

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

    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

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

    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.

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

    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.

  8. Activation of the chicken gonadotropin-inhibitory hormone receptor reduces gonadotropin releasing hormone receptor signaling.

    Shimizu, Mamiko; Bédécarrats, Grégoy Y

    2010-06-01

    Gonadotropin-inhibitory hormone (GnIH) is a hypothalamic peptide from the RFamide peptide family that has been identified in multiple avian species. Although GnIH has clearly been shown to reduce LH release from the anterior pituitary gland, its mechanism of action remains to be determined. The overall objectives of this study were (1) to characterize the GnIH receptor (GnIH-R) signaling pathway, (2) to evaluate potential interactions with gonadotropin releasing hormone type III receptor (GnRH-R-III) signaling, and (3) to determine the molecular mechanisms by which GnIH and GnRH regulate pituitary gonadotrope function during a reproductive cycle in the chicken. Using real-time PCR, we showed that in the chicken pituitary gland, GnIH-R mRNA levels fluctuate in an opposite manner to GnRH-R-III, with higher and lower levels observed during inactive and active reproductive stages, respectively. We demonstrated that the chicken GnIH-R signals by inhibiting adenylyl cyclase cAMP production, most likely by coupling to G(alphai). We also showed that this inhibition is sufficient to significantly reduce GnRH-induced cAMP responsive element (CRE) activation in a dose-dependent manner, and that the ratio of GnRH/GnIH receptors is a significant factor. We propose that in avian species, sexual maturation is characterized by a change in GnIH/GnRH receptor ratio, resulting in a switch in pituitary sensitivity from inhibitory (involving GnIH) to stimulatory (involving GnRH). In turn, decreasing GnIH-R signaling, combined with increasing GnRH-R-III signaling, results in significant increases in CRE activation, possibly initiating gonadotropin synthesis. PMID:20350548

  9. The angiotensin II type 1 receptor antagonist Losartan binds and activates bradykinin B2 receptor signaling

    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...

  10. Identification of key residues involved in the activation and signaling properties of dopamine D3 receptor.

    Kota, Kokila; Kuzhikandathil, Eldo V; Afrasiabi, Milad; Lacy, Brett; Kontoyianni, Maria; Crider, A Michael; Song, Daniel

    2015-09-01

    The dopamine D3 receptor exhibits agonist-dependent tolerance and slow response termination (SRT) signaling properties that distinguish it from the closely-related D2 receptors. While amino acid residues important for D3 receptor ligand binding have been identified, the residues involved in activation of D3 receptor signaling and induction of signaling properties have not been determined. In this paper, we used cis and trans isomers of a novel D3 receptor agonist, 8-OH-PBZI, and site-directed mutagenesis to identify key residues involved in D3 receptor signaling function. Our results show that trans-8-OH-PBZI, but not cis-8-OH-PBZI, elicit the D3 receptor tolerance and SRT properties. We show that while both agonists require a subset of residues in the orthosteric binding site of D3 receptors for activation of the receptor, the ability of the two isomers to differentially induce tolerance and SRT is mediated by interactions with specific residues in the sixth transmembrane helix and third extracellular loop of the D3 receptor. We also show that unlike cis-8-OH-PBZI, which is a partial agonist at the dopamine D2S receptor and full agonist at dopamine D2L receptor, trans-8-OH-PBZI is a full agonist at both D2S and D2L receptors. The different effect of the two isomers on D3 receptor signaling properties and D2S receptor activation correlated with differential effects of the isomers on agonist-induced mouse locomotor activity. The two isomers of 8-OH-PBZI represent novel pharmacological tools for in silico D3 and D2 receptor homology modeling and for determining the role of D3 receptor tolerance and SRT properties in signaling and behavior. PMID:26116441

  11. Signaling Mechanism of Cannabinoid Receptor-2 Activation-Induced β-Endorphin Release.

    Gao, Fang; Zhang, Ling-Hong; Su, Tang-Feng; Li, Lin; Zhou, Rui; Peng, Miao; Wu, Cai-Hua; Yuan, Xiao-Cui; Sun, Ning; Meng, Xian-Fang; Tian, Bo; Shi, Jing; Pan, Hui-Lin; Li, Man

    2016-08-01

    Activation of cannabinoid receptor-2 (CB2) results in β-endorphin release from keratinocytes, which then acts on primary afferent neurons to inhibit nociception. However, the underlying mechanism is still unknown. The CB2 receptor is generally thought to couple to Gi/o to inhibit cAMP production, which cannot explain the peripheral stimulatory effects of CB2 receptor activation. In this study, we found that in a keratinocyte cell line, the Gβγ subunits from Gi/o, but not Gαs, were involved in CB2 receptor activation-induced β-endorphin release. Inhibition of MAPK kinase, but not PLC, abolished CB2 receptor activation-induced β-endorphin release. Also, CB2 receptor activation significantly increased intracellular Ca(2+). Treatment with BAPTA-AM or thapsigargin blocked CB2 receptor activation-induced β-endorphin release. Using a rat model of inflammatory pain, we showed that the MAPK kinase inhibitor PD98059 abolished the peripheral effect of the CB2 receptor agonist on nociception. We thus present a novel mechanism of CB2 receptor activation-induced β-endorphin release through Gi/o-Gβγ-MAPK-Ca(2+) signaling pathway. Our data also suggest that stimulation of MAPK contributes to the peripheral analgesic effect of CB2 receptor agonists. PMID:26108183

  12. PROKR2 missense mutations associated with Kallmann syndrome impair receptor signalling activity.

    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

  13. Biased signalling from the glucocorticoid receptor: Renewed opportunity for tailoring glucocorticoid activity.

    Keenan, Christine R; Lew, Michael J; Stewart, Alastair G

    2016-07-15

    Recent landmark studies applying analytical pharmacology approaches to the glucocorticoid receptor (GR) have demonstrated that different ligands can cause differential activation of distinct GR-regulated genes. Drawing on concepts of signalling bias from the field of G protein-coupled receptor (GPCR) biology, we speculate that ligand-dependent differences in GR signalling can be considered analogous to GPCR biased signalling, and thus can be quantitatively analysed in a similar way. This type of approach opens up the possibility of using rational structure-based drug optimisation strategies to improve the therapeutic selectivity of glucocorticoid drugs to maximise their efficacy and minimise adverse effects. PMID:26898958

  14. Morbilliviruses Use Signaling Lymphocyte Activation Molecules (CD150) as Cellular Receptors

    Tatsuo, Hironobu; Ono, Nobuyuki; Yanagi, Yusuke

    2001-01-01

    Morbilliviruses comprise measles virus, canine distemper virus, rinderpest virus, and several other viruses that cause devastating human and animal diseases accompanied by severe immunosuppression and lymphopenia. Recently, we have shown that human signaling lymphocyte activation molecule (SLAM) is a cellular receptor for measles virus. In this study, we examined whether canine distemper and rinderpest viruses also use canine and bovine SLAMs, respectively, as cellular receptors. The Onderste...

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

    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....

  16. Ligand Perception, Activation, and Early Signaling of Plant Steroid Receptor Brassinosteroid Insensitive 1

    Jianjun Jiang; Chi Zhang; Xuelu Wang

    2013-01-01

    Leucine-rich repeat receptor-like kinases (LRR-RLKs) belong to a large group of cell surface proteins involved in many aspects of plant development and environmental responses in both monocots and dicots. Brassinosteroid insensitive 1 (BRI1), a member of the LRR X subfamily, was first identified through several forward genetic screenings for mutants insensitive to brassinosteroids (BRs), which are a class of plant-specific steroid hormones. Since its identification, BRI1 and its homologs had been proved as receptors perceiving BRs and initiating BR signaling. The co-receptor BRI1-associated kinase 1 and its homologs, and other BRI1 interacting proteins such as its inhibitor BRI1 kinase inhibitor 1 (BKI1) were identified by genetic and biochemical approaches. The detailed mechanisms of BR perception by BRI1 and the activation of BRI1 receptor complex have also been elucidated. Moreover, several mechanisms for termination of the activated BRI1 signaling were also discovered. In this review, we will focus on the recent advances on the mechanism of BRI1 phosphorylation and activation, the regulation of its receptor complex, the structure basis of BRI1 ectodomain and BR recognition, its direct substrates, and the termination of the activated BRI1 receptor complex.

  17. Receptor activity modifying proteins (RAMPs) interact with the VPAC1 receptor: evidence for differential RAMP modulation of multiple signalling pathways

    Full text: Receptor activity modifying proteins (RAMP) constitute a family of three accessory proteins that affect the expression and/or phenotype of the calcitonin receptor (CTR) or CTR-like receptor (CRLR). In this study we screened a range of class II G protein-coupled receptors (PTH1, PTH2, GHRH, VPAC1, VPAC2 receptors) for possible RAMP interactions by measurement of receptor-induced translocation of c-myc tagged RAMP1 or HA tagged RAMP3. Of these, only the VPAC1 receptor caused significant translocation of c-myc-RAMP1 or HA-RAMP3 to the cell surface. Co-transfection of VPAC1 and RAMPs did not alter 125I-VIP binding and specificity. VPAC1 receptor function was subsequently analyzed through parallel determinations of cAMP accumulation and phosphoinositide (PI) hydrolysis in the presence and absence of each of the three RAMPs. In contrast to CTR-RAMP interaction, where there was an increase in cAMP Pharmacologisand a decrease in PI hydrolysis, VPAC1-RAMP interaction was characterized by a specific increase in agonist-mediated PI hydrolysis when co-transfected with RAMP2. This change was due to an enhancement of Emax with no change in EC50 value for VIP. No significant change in cAMP accumulation was observed. This is the first demonstration of an interaction of RAMPs with a G protein-coupled receptor outside the CTR family and may suggest a more generalized role for RAMPs in modulating G protein-coupled receptor signaling. Copyright (2001) Australasian Society of Clinical and Experimental Pharmacologists and Toxicologists

  18. Active regulation of receptor ratios controls integration of quorum-sensing signals in Vibrio harveyi

    Teng, Shu-Wen; Schaffer, Jessica N; Tu, Kimberly C; Mehta, Pankaj; Lu, Wenyun; Ong, N P; Bassler, Bonnie L; Wingreen, Ned S

    2011-01-01

    Quorum sensing is a chemical signaling mechanism used by bacteria to communicate and orchestrate group behaviors. Multiple feedback loops exist in the quorum-sensing circuit of the model bacterium Vibrio harveyi. Using fluorescence microscopy of individual cells, we assayed the activity of the quorum-sensing circuit, with a focus on defining the functions of the feedback loops. We quantitatively investigated the signaling input–output relation both in cells with all feedback loops present as well as in mutants with specific feedback loops disrupted. We found that one of the feedback loops regulates receptor ratios to control the integration of multiple signals. Together, the feedback loops affect the input–output dynamic range of signal transmission and the noise in the output. We conclude that V. harveyi employs multiple feedback loops to simultaneously control quorum-sensing signal integration and to ensure signal transmission fidelity. PMID:21613980

  19. H(1)-Receptor activation triggers the endogenous nitric oxide signalling system in the rat submandibular gland.

    Borda, Enri; Stranieri, Graciela; Sterin-Borda, Leonor

    2002-01-01

    BACKGROUND: Histamine is released from mast cells by immunologic and non-immunologic stimuli during salivary gland inflammation, regulating salivary secretion. The receptor-secretory mechanism has not been studied in detail. AIMS: The studies reported were directed toward elucidating signal transduction/second messenger pathways within the rat submandibular gland associated with 2-thiazolylethylamine (ThEA)-induced H(1)-receptor responses. MATERIALS AND METHODS: To assess the H(1) receptor subtype expression in the rat submandibular gland, a radioligand binding assay was performed. The study also included inositolphosphates and cyclic GMP accumulation, protein kinase C and nitric oxide synthase activities, and amylase release. RESULTS: The histamine H(1) receptor subtype is expressed on the rat submandibular gland with high-affinity binding sites. The ThEA effect was associated with activation of phosphoinositide-specific phospholipase C, translocation of protein kinase C, stimulation of nitric oxide synthase activity and increased production of cyclic GMP. ThEA stimulation of nitric oxide synthase and cyclic GMP was blunted by agents able to interfere with calcium movilization, while a protein kinase C inhibitor was able to stimulate ThEA action. On the other hand, ThEA stimulation evoked amylase release via the H1 receptor but was not followed by the L-arginine/nitric oxide pathway activation. CONCLUSIONS: These results suggest that, apart from the effect of ThEA on amylase release, it also appears to be a vasoactive chemical mediator that triggers vasodilatation, modulating the course of inflammation. PMID:12581497

  20. Agmatine produces antidepressant-like effects by activating AMPA receptors and mTOR signaling.

    Neis, Vivian Binder; Moretti, Morgana; Bettio, Luis Eduardo B; Ribeiro, Camille M; Rosa, Priscila Batista; Gonçalves, Filipe Marques; Lopes, Mark William; Leal, Rodrigo Bainy; Rodrigues, Ana Lúcia S

    2016-06-01

    The activation of AMPA receptors and mTOR signaling has been reported as mechanisms underlying the antidepressant effects of fast-acting agents, specially the NMDA receptor antagonist ketamine. In the present study, oral administration of agmatine (0.1mg/kg), a neuromodulator that has been reported to modulate NMDA receptors, caused a significant reduction in the immobility time of mice submitted to the tail suspension test (TST), an effect prevented by the administration of DNQX (AMPA receptor antagonist, 2.5μg/site, i.c.v.), BDNF antibody (1μg/site, i.c.v.), K-252a (TrkB receptor antagonist, 1μg/site, i.c.v.), LY294002 (PI3K inhibitor, 10nmol/site, i.c.v.) or rapamycin (selective mTOR inhibitor, 0.2nmol/site, i.c.v.). Moreover, the administration of lithium chloride (non-selective GSK-3β inhibitor, 10mg/kg, p.o.) or AR-A014418 (selective GSK-3β inhibitor, 0.01μg/site, i.c.v.) in combination with a sub-effective dose of agmatine (0.0001mg/kg, p.o.) reduced the immobility time in the TST when compared with either drug alone. Furthermore, increased immunocontents of BDNF, PSD-95 and GluA1 were found in the prefrontal cortex of mice just 1h after agmatine administration. These results indicate that the antidepressant-like effect of agmatine in the TST may be dependent on the activation of AMPA and TrkB receptors, PI3K and mTOR signaling as well as inhibition of GSK-3β, and increase in synaptic proteins. The results contribute to elucidate the complex signaling pathways involved in the antidepressant effect of agmatine and reinforce the pivotal role of these molecular targets for antidepressant responses. PMID:27061850

  1. SGIP1 alters internalization and modulates signaling of activated cannabinoid receptor 1 in a biased manner.

    Hájková, Alena; Techlovská, Šárka; Dvořáková, Michaela; Chambers, Jayne Nicole; Kumpošt, Jiří; Hubálková, Pavla; Prezeau, Laurent; Blahos, Jaroslav

    2016-08-01

    Many diseases of the nervous system are accompanied by alterations in synaptic functions. Synaptic plasticity mediated by the endogenous cannabinoid system involves the activation of the cannabinoid receptor 1 (CB1R). The principles of CB1R signaling must be understood in detail for its therapeutic exploration. We detected the Src homology 3-domain growth factor receptor-bound 2-like (endophilin) interacting protein 1 (SGIP1) as a novel CB1R partner. SGIP1 is functionally linked to clathrin-mediated endocytosis and its overexpression in animals leads to an energy regulation imbalance resulting in obesity. We report that SGIP1 prevents the endocytosis of activated CB1R and that it alters signaling via the CB1R in a biased manner. CB1R mediated G-protein activation is selectively influenced by SGIP1, β-arrestin associated signaling is changed profoundly, most likely as a consequence of the prevention of the receptor's internalization elicited by SGIP1. PMID:26970018

  2. Essential biphasic role for JAK3 catalytic activity in IL-2 receptor signaling.

    Smith, Geoffrey A; Uchida, Kenji; Weiss, Arthur; Taunton, Jack

    2016-05-01

    To drive lymphocyte proliferation and differentiation, common γ-chain (γc) cytokine receptors require hours to days of sustained stimulation. JAK1 and JAK3 kinases are found together in all γc-receptor complexes, but how their respective catalytic activities contribute to signaling over time is not known. Here we dissect the temporal requirements for JAK3 kinase activity with a selective covalent inhibitor (JAK3i). By monitoring phosphorylation of the transcription factor STAT5 over 20 h in CD4(+) T cells stimulated with interleukin 2 (IL-2), we document a second wave of signaling that is much more sensitive to JAK3i than the first wave. Selective inhibition of this second wave is sufficient to block cyclin expression and entry to S phase. An inhibitor-resistant JAK3 mutant (C905S) rescued all effects of JAK3i in isolated T cells and in mice. Our chemical genetic toolkit elucidates a biphasic requirement for JAK3 kinase activity in IL-2-driven T cell proliferation and will find broad utility in studies of γc-receptor signaling. PMID:27018889

  3. A novel bile acid-activated vitamin D receptor signaling in human hepatocytes.

    Han, Shuxin; Li, Tiangang; Ellis, Ewa; Strom, Stephen; Chiang, John Y L

    2010-06-01

    Vitamin D receptor (VDR) is activated by natural ligands, 1alpha, 25-dihydroxy-vitamin D(3) [1alpha,25(OH)(2)-D(3)] and lithocholic acid (LCA). Our previous study shows that VDR is expressed in human hepatocytes, and VDR ligands inhibit bile acid synthesis and transcription of the gene encoding cholesterol 7alpha-hydroxylase (CYP7A1). Primary human hepatocytes were used to study LCA and 1alpha,25(OH)(2)-D(3) activation of VDR signaling. Confocal immunofluorescent microscopy imaging and immunoblot analysis showed that LCA and 1alpha, 25(OH)(2)-D(3) induced intracellular translocation of VDR from the cytosol to the nucleus and also plasma membrane where VDR colocalized with caveolin-1. VDR ligands induced tyrosine phosphorylation of c-Src and VDR and their interaction. Inhibition of c-Src abrogated VDR ligand-dependent inhibition of CYP7A1 mRNA expression. Kinase assays showed that VDR ligands specifically activated the c-Raf/MEK1/2/extracellular signal-regulated kinase (ERK) 1/2 pathway, which stimulates serine phosphorylation of VDR and hepatocyte nuclear factor-4alpha, and their interaction. Mammalian two-hybrid assays showed a VDR ligand-dependent interaction of nuclear receptor corepressor-1 and silencing mediator of retinoid and thyroid with VDR/retinoid X receptor-alpha (RXRalpha). Chromatin immunoprecipitation assays revealed that an ERK1/2 inhibitor reversed VDR ligand-induced recruitment of VDR, RXRalpha, and corepressors to human CYP7A1 promoter. In conclusion, VDR ligands activate membrane VDR signaling to activate the MEK1/2/ERK1/2 pathway, which stimulates nuclear VDR/RXRalpha recruitment of corepressors to inhibit CYP7A1 gene transcription in human hepatocytes. This membrane VDR-signaling pathway may be activated by bile acids to inhibit bile acid synthesis as a rapid response to protect hepatocytes from cholestatic liver injury. PMID:20371703

  4. A Novel Bile Acid-Activated Vitamin D Receptor Signaling in Human Hepatocytes

    Han, Shuxin; Li, Tiangang; Ellis, Ewa; Strom, Stephen; Chiang, John Y. L.

    2010-01-01

    Vitamin D receptor (VDR) is activated by natural ligands, 1α, 25-dihydroxy-vitamin D3 [1α,25(OH)2-D3] and lithocholic acid (LCA). Our previous study shows that VDR is expressed in human hepatocytes, and VDR ligands inhibit bile acid synthesis and transcription of the gene encoding cholesterol 7α-hydroxylase (CYP7A1). Primary human hepatocytes were used to study LCA and 1α,25(OH)2-D3 activation of VDR signaling. Confocal immunofluorescent microscopy imaging and immunoblot analysis showed that ...

  5. Intercellular calcium signaling occurs between human osteoblasts and osteoclasts and requires activation of osteoclast P2X7 receptors

    Jørgensen, Niklas R; Henriksen, Zanne; Sørensen, Ole;

    2002-01-01

    Signaling between osteoblasts and osteoclasts is important in bone homeostasis. We previously showed that human osteoblasts propagate intercellular calcium signals via two mechanisms: autocrine activation of P2Y receptors, and gap junctional communication. In the current work we identified...... mechanically induced intercellular calcium signaling between osteoblasts and osteoclasts and among osteoclasts. Intercellular calcium responses in osteoclasts required P2 receptor activation but not gap junctional communication. Pharmacological studies and reverse transcriptase-PCR amplification demonstrated...... that human osteoclasts expressed functional P2Y1 receptors, but, unexpectedly, desensitization of P2Y1 did not block calcium signaling to osteoclasts. We also found that osteoclasts expressed functional P2X7 receptors and showed that pharmacological inhibition of these receptors blocked calcium...

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

    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.

  7. Activation of aryl hydrocarbon receptor signaling by extracts of teak and other wood dusts.

    Wilson, Mark J; Sabbioni, Gabriele; Rando, Roy; Miller, Charles A

    2015-12-01

    Wood dusts, as a group, are categorized as known human carcinogens, but the risks of exposure to specific types of wood dusts and the carcinogenic chemicals they contain are not well studied. Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is linked to the carcinogenic action of specific classes of chemicals. Here we examined whether chemicals in various wood dusts had the potential to activate AhR signaling as a potential toxic mechanism of action. We found that methanol extracts of teak, walnut, mahogany, and poplar dusts contained a wide range of AhR ligand activity, whereas extracts of oak, pine, and other softwoods did not contain appreciable activity. Teak dust extract, being particularly potent, was subjected to chemical analysis. The 2-methylanthraquinone (2-MAQ) accounted for the AhR ligand activity and was present at an average concentration of 0.27 parts per hundred in teak dust. Pure 2-MAQ potently induced AhR signaling (EC50 115 nM), confirming that this was the active ligand. Aqueous extracts of teak dust made using yeast or mammalian cell culture medium also contained robust AhR activity, suggesting the 2-MAQ ligand is soluble at bioactive concentrations in physiologically relevant fluids. The high concentration and potency of 2-MAQ in teak wood suggest it may mediate toxic effects through activation of AhR signaling in exposed wood workers. PMID:24898320

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

    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

  9. Roles of activin receptor-like kinase 7 signaling and its target, peroxisome proliferator-activated receptor γ, in lean and obese adipocytes

    Yogosawa, Satomi; Izumi, Tetsuro

    2013-01-01

    We recently discovered a novel signaling pathway involving activin receptor-like kinase 7 (ALK7), one of the type I transforming growth factor-β receptors. ALK7 and activated Smads 2, 3, and 4 inhibit the master regulators of adipogenesis, CCAAT/enhancer-binding protein α (C/EBPα) and peroxisome proliferator-activated receptor γ (PPARγ) specifically in differentiated adipocytes, but surprisingly increase both the adipocyte size and lipid content by suppressing lipolysis. Here, we show that, a...

  10. Nuclear Factor-κB: Activation and Regulation during Toll-like Receptor Signaling

    Ruaidhrí J. Carmody; Youhai H. Chen

    2007-01-01

    Toll-like receptors (TLRs) recognize distinct microbial components to initiate the innate and adaptive immune responses. TLR activation culminates in the expression of appropriate pro-inflammatory and immunomodulatory factors to meet pathogenic challenges. The transcription factor NF-κB is the master regulator of all TLR-induced responses and its activation is the pivotal event in TLR-mediated activation of the innate immune response. Many of the key molecular events required for TLR-induced NF-κB activation have been elucidated. However, much remain to be learned about the ability of TLRs to generate pathogen-specific responses using a limited number of transcription factors. This review will focus on our current understanding of NF-κB activation by TLRs and potential mechanisms for achieving a signal-specific response through NF-κB.

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

    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...

  12. Extracellular Ca2+ is a danger signal activating the NLRP3 inflammasome through G protein-coupled calcium sensing receptors

    Rossol, Manuela; Pierer, Matthias; Raulien, Nora; Quandt, Dagmar; Meusch, Undine; Rothe, Kathrin; Schubert, Kristin; Schöneberg, Torsten; Schaefer, Michael; Krügel, Ute; Smajilovic, Sanela; Bräuner-Osborne, Hans; Baerwald, Christoph; Wagner, Ulf

    2012-01-01

    calcium activates the NLRP3 inflammasome via stimulation of G protein-coupled calcium sensing receptors. Activation is mediated by signalling through the calcium-sensing receptor and GPRC6A via the phosphatidyl inositol/Ca(2+) pathway. The resulting increase in the intracellular calcium concentration...... this effect was inhibited in GPRC6A(-/-) mice. Our results demonstrate that G-protein-coupled receptors can activate the inflammasome, and indicate that increased extracellular calcium has a role as a danger signal and amplifier of inflammation....

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

    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.

  14. Interferon lambda 4 signals via the IFNλ receptor to regulate antiviral activity against HCV and coronaviruses

    Hamming, Ole Jensen; Terczynska-Dyla, Ewa; Vieyres, Gabrielle;

    2013-01-01

    The IFNL4 gene is a recently discovered type III interferon, which in a significant fraction of the human population harbours a frameshift mutation abolishing the IFNλ4 ORF. The expression of IFNλ4 is correlated with both poor spontaneous clearance of hepatitis C virus (HCV) and poor response...... to treatment with type I interferon. Here, we show that the IFNL4 gene encodes an active type III interferon, named IFNλ4, which signals through the IFNλR1 and IL-10R2 receptor chains. Recombinant IFNλ4 is antiviral against both HCV and coronaviruses at levels comparable to IFNλ3. However, the secretion...

  15. Interleukin-2 activity can be fine tuned with engineered receptor signaling clamps.

    Mitra, Suman; Ring, Aaron M; Amarnath, Shoba; Spangler, Jamie B; Li, Peng; Ju, Wei; Fischer, Suzanne; Oh, Jangsuk; Spolski, Rosanne; Weiskopf, Kipp; Kohrt, Holbrook; Foley, Jason E; Rajagopalan, Sumati; Long, Eric O; Fowler, Daniel H; Waldmann, Thomas A; Garcia, K Christopher; Leonard, Warren J

    2015-05-19

    Interleukin-2 (IL-2) regulates lymphocyte function by signaling through heterodimerization of the IL-2Rβ and γc receptor subunits. IL-2 is of considerable therapeutic interest, but harnessing its actions in a controllable manner remains a challenge. Previously, we have engineered an IL-2 "superkine" with enhanced affinity for IL-2Rβ. Here, we describe next-generation IL-2 variants that function as "receptor signaling clamps." They retained high affinity for IL-2Rβ, inhibiting binding of endogenous IL-2, but their interaction with γc was weakened, attenuating IL-2Rβ-γc heterodimerization. These IL-2 analogs acted as partial agonists and differentially affected lymphocytes poised at distinct activation thresholds. Moreover, one variant, H9-RETR, antagonized IL-2 and IL-15 better than blocking antibodies against IL-2Rα or IL-2Rβ. Furthermore, this mutein prolonged survival in a model of graft-versus-host disease and blocked spontaneous proliferation of smoldering adult T cell leukemia (ATL) T cells. This receptor-clamping approach might be a general mechanism-based strategy for engineering cytokine partial agonists for therapeutic immunomodulation. PMID:25992859

  16. Melanocortin-4 receptor activation inhibits c-Jun N-terminal kinase activity and promotes insulin signaling

    Chai, Biaoxin; Li, Ji-Yao; Zhang, Weizhen; Wang, Hui; Mulholland, Michael W.

    2009-01-01

    The melanocortin system is crucial to regulation of energy homeostasis. The melanocortin receptor type 4 (MC4R) modulates insulin signaling via effects on c-Jun N-terminal kinase (JNK). The melanocortin agonist NDP-MSH dose-dependently inhibited JNK activity in HEK293 cells stably expressing the human MC4R; effects were reversed by melanocortin receptor antagonist. NDP-MSH time- and dose-dependently inhibited IRS-1ser307 phosphorylation, effects also reversed by a specific melanocortin recept...

  17. Dioxin mediates downregulation of the reduced folate carrier transport activity via the arylhydrocarbon receptor signalling pathway

    Dioxins such as 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD) are common environmental contaminants known to regulate several genes via activation of the transcription factor aryl hydrocarbon receptor (AhR) associated with the development of numerous adverse biological effects. However, comparatively little is known about the molecular mechanisms by which dioxins display their toxic effects in vertebrates. The 5' untranslated region of the hepatocellular Reduced folate carrier (Rfc1; Slc19a1) exhibits AhR binding sites termed dioxin responsive elements (DRE) that have as yet only been found in the promoter region of prototypical TCDD target genes. Rfc1 mediated transport of reduced folates and antifolate drugs such as methotrexate (MTX) plays an essential role in physiological folate homeostasis and MTX cancer chemotherapy. In order to determine whether this carrier represents a target gene of dioxins we have investigated the influence of TCDD on functional Rfc1 activity in rat liver. Pre-treatment of rats with TCDD significantly diminished hepatocellular Rfc1 uptake activity in a time- and dose-dependent manner. In further mechanistic studies we demonstrated that this reduction was due to TCDD-dependent activation of the AhR signalling pathway. We additionally showed that binding of the activated receptor to DRE motifs in the Rfc1 promoter resulted in downregulation of Rfc1 gene expression and reduced carrier protein levels. As downregulation of pivotal Rfc1 activity results in functional folate deficiency associated with an elevated risk of cardiovascular diseases or carcinogenesis, our results indicate that deregulation of this essential transport pathway represents a novel regulatory mechanism how dioxins display their toxic effects through the Ah receptor.

  18. The Relaxin Receptor (RXFP1) Utilizes Hydrophobic Moieties on a Signaling Surface of Its N-terminal Low Density Lipoprotein Class A Module to Mediate Receptor Activation*

    Kong, Roy C. K.; Petrie, Emma J.; Mohanty, Biswaranjan; Ling, Jason; Lee, Jeremy C. Y.; Gooley, Paul R.; Bathgate, Ross A. D.

    2013-01-01

    The peptide hormone relaxin is showing potential as a treatment for acute heart failure. Although it is known that relaxin mediates its actions through the G protein-coupled receptor relaxin family peptide receptor 1 (RXFP1), little is known about the molecular mechanisms by which relaxin binding results in receptor activation. Previous studies have highlighted that the unique N-terminal low density lipoprotein class A (LDLa) module of RXFP1 is essential for receptor activation, and it has been hypothesized that this module is the true “ligand” of the receptor that directs the conformational changes necessary for G protein coupling. In this study, we confirmed that an RXFP1 receptor lacking the LDLa module binds ligand normally but cannot signal through any characterized G protein-coupled receptor signaling pathway. Furthermore, we comprehensively examined the contributions of amino acids in the LDLa module to RXFP1 activity using both gain-of-function and loss-of-function mutational analysis together with NMR structural analysis of recombinant LDLa modules. Gain-of-function studies with an inactive RXFP1 chimera containing the LDLa module of the human LDL receptor (LB2) demonstrated two key N-terminal regions of the module that were able to rescue receptor signaling. Loss-of-function mutations of residues in these regions demonstrated that Leu-7, Tyr-9, and Lys-17 all contributed to the ability of the LDLa module to drive receptor activation, and judicious amino acid substitutions suggested this involves hydrophobic interactions. Our results demonstrate that these key residues contribute to interactions driving the active receptor conformation, providing further evidence of a unique mode of G protein-coupled receptor activation. PMID:23926099

  19. The relaxin receptor (RXFP1) utilizes hydrophobic moieties on a signaling surface of its N-terminal low density lipoprotein class A module to mediate receptor activation.

    Kong, Roy C K; Petrie, Emma J; Mohanty, Biswaranjan; Ling, Jason; Lee, Jeremy C Y; Gooley, Paul R; Bathgate, Ross A D

    2013-09-27

    The peptide hormone relaxin is showing potential as a treatment for acute heart failure. Although it is known that relaxin mediates its actions through the G protein-coupled receptor relaxin family peptide receptor 1 (RXFP1), little is known about the molecular mechanisms by which relaxin binding results in receptor activation. Previous studies have highlighted that the unique N-terminal low density lipoprotein class A (LDLa) module of RXFP1 is essential for receptor activation, and it has been hypothesized that this module is the true "ligand" of the receptor that directs the conformational changes necessary for G protein coupling. In this study, we confirmed that an RXFP1 receptor lacking the LDLa module binds ligand normally but cannot signal through any characterized G protein-coupled receptor signaling pathway. Furthermore, we comprehensively examined the contributions of amino acids in the LDLa module to RXFP1 activity using both gain-of-function and loss-of-function mutational analysis together with NMR structural analysis of recombinant LDLa modules. Gain-of-function studies with an inactive RXFP1 chimera containing the LDLa module of the human LDL receptor (LB2) demonstrated two key N-terminal regions of the module that were able to rescue receptor signaling. Loss-of-function mutations of residues in these regions demonstrated that Leu-7, Tyr-9, and Lys-17 all contributed to the ability of the LDLa module to drive receptor activation, and judicious amino acid substitutions suggested this involves hydrophobic interactions. Our results demonstrate that these key residues contribute to interactions driving the active receptor conformation, providing further evidence of a unique mode of G protein-coupled receptor activation. PMID:23926099

  20. Potentiation of acid-sensing ion channel activity by peripheral group I metabotropic glutamate receptor signaling.

    Gan, Xiong; Wu, Jing; Ren, Cuixia; Qiu, Chun-Yu; Li, Yan-Kun; Hu, Wang-Ping

    2016-05-01

    Glutamate activates peripheral group I metabotropic glutamate receptors (mGluRs) and contributes to inflammatory pain. However, it is still not clear the mechanisms are involved in group I mGluR-mediated peripheral sensitization. Herein, we report that group I mGluRs signaling sensitizes acid-sensing ion channels (ASICs) in dorsal root ganglion (DRG) neurons and contributes to acidosis-evoked pain. DHPG, a selective group I mGluR agonist, can potentiate the functional activity of ASICs, which mediated the proton-induced events. DHPG concentration-dependently increased proton-gated currents in DRG neurons. It shifted the proton concentration-response curve upwards, with a 47.3±7.0% increase of the maximal current response to proton. Group I mGluRs, especially mGluR5, mediated the potentiation of DHPG via an intracellular cascade. DHPG potentiation of proton-gated currents disappeared after inhibition of intracellular Gq/11 proteins, PLCβ, PKC or PICK1 signaling. Moreover, DHPG enhanced proton-evoked membrane excitability of rat DRG neurons and increased the amplitude of the depolarization and the number of spikes induced by acid stimuli. Finally, peripherally administration of DHPG dose-dependently exacerbated nociceptive responses to intraplantar injection of acetic acid in rats. Potentiation of ASIC activity by group I mGluR signaling in rat DRG neurons revealed a novel peripheral mechanism underlying group I mGluRs involvement in hyperalgesia. PMID:26946972

  1. Melanocortin-1 receptor-mediated signalling pathways activated by NDP-MSH and HBD3 ligands

    Beaumont, Kimberley A.; Smit, Darren J.; Liu, Yan Yan; Chai, Eric; Patel, Mira P.; Millhauser, Glenn L.; Smith, Jennifer J.; Alewood, Paul F.; Sturm, Richard A.

    2012-01-01

    Binding of melanocortin peptide agonists to the melanocortin-1 receptor of melanocytes results in eumelanin production, whereas binding of the agouti signalling protein inverse agonist results in pheomelanin synthesis. Recently, a novel melanocortin-1 receptor ligand was reported. A β-defensin gene mutation was found to beresponsible for black coat colour in domestic dogs. Notably, the human equivalent, β-defensin 3, was found to bind with high affinity to the melanocortin-1 receptor; however...

  2. Polysaccharide of Dendrobium huoshanense activates macrophages via toll-like receptor 4-mediated signaling pathways.

    Xie, Song-Zi; Hao, Ran; Zha, Xue-Qiang; Pan, Li-Hua; Liu, Jian; Luo, Jian-Ping

    2016-08-01

    The present work aimed at investigating the pattern recognition receptor (PRR) and immunostimulatory mechanism of a purified Dendrobium huoshanense polysaccharide (DHP). We found that DHP could bind to the surface of macrophages and stimulate macrophages to secrete NO, TNF-α and IL-1β. To unravel the mechanism for the binding of DHP to macrophages, flow cytometry, confocal laser-scanning microscopy, affinity electrophoresis, SDS-PAGE and western blotting were employed to verify the type of PRR responsible for the recognition of DHP by RAW264.7 macrophages and peritoneal macrophages of C3H/HeN and C3H/HeJ macrophages. Results showed that toll-like receptor 4 (TLR4) was an essential receptor for macrophages to directly bind DHP. Further, the phosphorylation of ERK, JNK, Akt and p38 were observed to be time-dependently promoted by DHP, as well as the nuclear translocation of NF-κB p65. These results suggest that DHP activates macrophages via its direct binding to TLR4 to trigger TLR4 signaling pathways. PMID:27112877

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

    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

  4. Attenuation of signaling pathways stimulated by pathologically activated FGF-receptor 2 mutants prevents craniosynostosis.

    Eswarakumar, V P; Ozcan, F; Lew, E D; Bae, J H; Tomé, F; Booth, C J; Adams, D J; Lax, I; Schlessinger, J

    2006-12-01

    Craniosynostosis, the fusion of one or more of the sutures of the skull vault before the brain completes its growth, is a common (1 in 2,500 births) craniofacial abnormality, approximately 20% of which occurrences are caused by gain-of-function mutations in FGF receptors (FGFRs). We describe a genetic and pharmacological approach for the treatment of a murine model system of Crouzon-like craniosynostosis induced by a dominant mutation in Fgfr2c. Using genetically modified mice, we demonstrate that premature fusion of sutures mediated by Crouzon-like activated Fgfr2c mutant is prevented by attenuation of signaling pathways by selective uncoupling between the docking protein Frs2alpha and activated Fgfr2c, resulting in normal skull development. We also demonstrate that attenuation of Fgfr signaling in a calvaria organ culture with an Fgfr inhibitor prevents premature fusion of sutures without adversely affecting calvaria development. These experiments show that attenuation of FGFR signaling by pharmacological intervention could be applied for the treatment of craniosynostosis or other severe bone disorders caused by mutations in FGFRs that currently have no treatment. PMID:17132737

  5. Aberrant Activation of the RANK Signaling Receptor Induces Murine Salivary Gland Tumors.

    Maria M Szwarc

    Full Text Available Unlike cancers of related exocrine tissues such as the mammary and prostate gland, diagnosis and treatment of aggressive salivary gland malignancies have not markedly advanced in decades. Effective clinical management of malignant salivary gland cancers is undercut by our limited knowledge concerning the key molecular signals that underpin the etiopathogenesis of this rare and heterogeneous head and neck cancer. Without knowledge of the critical signals that drive salivary gland tumorigenesis, tumor vulnerabilities cannot be exploited that allow for targeted molecular therapies. This knowledge insufficiency is further exacerbated by a paucity of preclinical mouse models (as compared to other cancer fields with which to both study salivary gland pathobiology and test novel intervention strategies. Using a mouse transgenic approach, we demonstrate that deregulation of the Receptor Activator of NFkB Ligand (RANKL/RANK signaling axis results in rapid tumor development in all three major salivary glands. In line with its established role in other exocrine gland cancers (i.e., breast cancer, the RANKL/RANK signaling axis elicits an aggressive salivary gland tumor phenotype both at the histologic and molecular level. Despite the ability of this cytokine signaling axis to drive advanced stage disease within a short latency period, early blockade of RANKL/RANK signaling markedly attenuates the development of malignant salivary gland neoplasms. Together, our findings have uncovered a tumorigenic role for RANKL/RANK in the salivary gland and suggest that targeting this pathway may represent a novel therapeutic intervention approach in the prevention and/or treatment of this understudied head and neck cancer.

  6. Binding of receptor-recognized forms of alpha2-macroglobulin to the alpha2-macroglobulin signaling receptor activates phosphatidylinositol 3-kinase.

    Misra, U K; Pizzo, S V

    1998-05-29

    Ligation of the alpha2-macroglobulin (alpha2M) signaling receptor by receptor-recognized forms of alpha2M (alpha2M*) initiates mitogenesis secondary to increased intracellular Ca2+. We report here that ligation of the alpha2M signaling receptor also causes a 1. 5-2.5-fold increase in wortmannin-sensitive phosphatidylinositol 3-kinase (PI3K) activity as measured by the quantitation of phosphatidylinositol 3,4,5-trisphosphate (PIP3). PIP3 formation was alpha2M* concentration-dependent with a maximal response at approximately 50 pM ligand concentration. The peak formation of PIP3 occurred at 10 min of incubation. The alpha2M receptor binding fragment mutant K1370R which binds to the alpha2M signaling receptor activating the signaling cascade, increased PIP3 formation by 2-fold. The mutant K1374A, which binds very poorly to the alpha2M signaling receptor, did not cause any increase in PIP3 formation. alpha2M*-induced DNA synthesis was inhibited by wortmannin. 1, 2Bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acetoxymethylester a chelator of intracellular Ca2+, drastically reduced alpha2M*-induced increases in PIP3 formation. We conclude that PI3K is involved in alpha2M*-induced mitogenesis in macrophages and intracellular Ca2+ plays a role in PI3K activation. PMID:9593670

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

    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...

  8. Dopamine D2 receptor-mediated Akt/PKB signalling: initiation by the D2S receptor and role in quinpirole-induced behavioural activation

    Jin‑Chung Chen; Nan‑Yu Ruan; Tzu‑Yung Lin; Han‑Ting Chen

    2012-01-01

    The short and long isoforms of the dopamine D2 receptor (D2S and D2L respectively) are highly expressed in the striatum. Functional D2 receptors activate an intracellular signalling pathway that includes a cAMP-independent route involving Akt/GSK3 (glycogen synthase kinase 3). To investigate the Akt/GSK3 response to the seldom-studied D2S receptor, we established a rat D2S receptor-expressing cell line [HEK (human embryonic kidney)-293/rD2S]. We found that in HEK-293/rD2S cells, the D2/D3 ago...

  9. A novel signaling pathway of tissue kallikrein in promoting keratinocyte migration: Activation of proteinase-activated receptor 1 and epidermal growth factor receptor

    Biological functions of tissue kallikrein (TK, KLK1) are mainly mediated by kinin generation and subsequent kinin B2 receptor activation. In this study, we investigated the potential role of TK and its signaling pathways in cultured human keratinocyte migration and in a rat skin wound healing model. Herein, we show that TK promoted cell migration and proliferation in a concentration- and time-dependent manner. Inactive TK or kinin had no significant effect on cell migration. Interestingly, cell migration induced by active TK was not blocked by icatibant or L-NAME, indicating an event independent of kinin B2 receptor and nitric oxide formation. TK's stimulatory effect on cell migration was inhibited by small interfering RNA for proteinase-activated receptor 1 (PAR1), and by PAR1 inhibitor. TK-induced migration was associated with increased phosphorylation of epidermal growth factor receptor (EGFR) and extracellular signal-regulated kinase (ERK), which was blocked by inhibition of protein kinase C (PKC), Src, EGFR and ERK. TK-induced cell migration and EGFR phosphorylation were blocked by metalloproteinase (MMP) inhibitor, heparin, and antibodies against EGFR external domain, heparin-binding EGF-like growth factor (HB-EGF) and amphiregulin (AR). Local application of TK promoted skin wound healing in rats, whereas icatibant and EGFR inhibitor blocked TK's effect. Skin wound healing was further delayed by aprotinin and neutralizing TK antibody. This study demonstrates a novel role of TK in skin wound healing and uncovers new signaling pathways mediated by TK in promoting keratinocyte migration through activation of the PAR1-PKC-Src-MMP pathway and HB-EGF/AR shedding-dependent EGFR transactivation.

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

    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.

  11. Olfactory receptor signaling.

    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

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

    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

  13. Valerian inhibits rat hepatocarcinogenesis by activating GABA(A receptor-mediated signaling.

    Anna Kakehashi

    Full Text Available Valerian is widely used as a traditional medicine to improve the quality of sleep due to interaction of several active components with the γ-aminobutyric acid (GABA A receptor (GABA(AR system. Recently, activation of GABA signaling in stem cells has been reported to suppress cell cycle progression in vivo. Furthermore, possible inhibitory effects of GABA(AR agonists on hepatocarcinogenesis have been reported. The present study was performed to investigate modulating effects of Valerian on hepatocarcinogenesis using a medium-term rat liver bioassay. Male F344 rats were treated with one of the most powerful Valerian species (Valeriana sitchensis at doses of 0, 50, 500 and 5000 ppm in their drinking water after initiation of hepatocarcinogenesis with diethylnitrosamine (DEN. Formation of glutathione S-transferase placental form positive (GST-P(+ foci was significantly inhibited by Valerian at all applied doses compared with DEN initiation control rats. Generation of 8-hydroxy-2'-deoxyguanosine in the rat liver was significantly suppressed by all doses of Valerian, likely due to suppression of Nrf2, CYP7A1 and induction of catalase expression. Cell proliferation was significantly inhibited, while apoptosis was induced in areas of GST-P(+ foci of Valerian groups associated with suppression of c-myc, Mafb, cyclin D1 and induction of p21(Waf1/Cip1, p53 and Bax mRNA expression. Interestingly, expression of the GABA(AR alpha 1 subunit was observed in GST-P(+ foci of DEN control rats, with significant elevation associated with Valerian treatment. These results indicate that Valerian exhibits inhibitory effects on rat hepatocarcinogenesis by inhibiting oxidative DNA damage, suppressing cell proliferation and inducing apoptosis in GST-P(+ foci by activating GABA(AR-mediated signaling.

  14. Valerian inhibits rat hepatocarcinogenesis by activating GABA(A) receptor-mediated signaling.

    Kakehashi, Anna; Kato, Ayumi; Ishii, Naomi; Wei, Min; Morimura, Keiichirou; Fukushima, Shoji; Wanibuchi, Hideki

    2014-01-01

    Valerian is widely used as a traditional medicine to improve the quality of sleep due to interaction of several active components with the γ-aminobutyric acid (GABA) A receptor (GABA(A)R) system. Recently, activation of GABA signaling in stem cells has been reported to suppress cell cycle progression in vivo. Furthermore, possible inhibitory effects of GABA(A)R agonists on hepatocarcinogenesis have been reported. The present study was performed to investigate modulating effects of Valerian on hepatocarcinogenesis using a medium-term rat liver bioassay. Male F344 rats were treated with one of the most powerful Valerian species (Valeriana sitchensis) at doses of 0, 50, 500 and 5000 ppm in their drinking water after initiation of hepatocarcinogenesis with diethylnitrosamine (DEN). Formation of glutathione S-transferase placental form positive (GST-P(+)) foci was significantly inhibited by Valerian at all applied doses compared with DEN initiation control rats. Generation of 8-hydroxy-2'-deoxyguanosine in the rat liver was significantly suppressed by all doses of Valerian, likely due to suppression of Nrf2, CYP7A1 and induction of catalase expression. Cell proliferation was significantly inhibited, while apoptosis was induced in areas of GST-P(+) foci of Valerian groups associated with suppression of c-myc, Mafb, cyclin D1 and induction of p21(Waf1/Cip1), p53 and Bax mRNA expression. Interestingly, expression of the GABA(A)R alpha 1 subunit was observed in GST-P(+) foci of DEN control rats, with significant elevation associated with Valerian treatment. These results indicate that Valerian exhibits inhibitory effects on rat hepatocarcinogenesis by inhibiting oxidative DNA damage, suppressing cell proliferation and inducing apoptosis in GST-P(+) foci by activating GABA(A)R-mediated signaling. PMID:25419570

  15. Fluorescence/bioluminescence resonance energy transfer techniques to study G-protein-coupled receptor activation and signaling.

    Lohse, Martin J; Nuber, Susanne; Hoffmann, Carsten

    2012-04-01

    Fluorescence and bioluminescence resonance energy transfer (FRET and BRET) techniques allow the sensitive monitoring of distances between two labels at the nanometer scale. Depending on the placement of the labels, this permits the analysis of conformational changes within a single protein (for example of a receptor) or the monitoring of protein-protein interactions (for example, between receptors and G-protein subunits). Over the past decade, numerous such techniques have been developed to monitor the activation and signaling of G-protein-coupled receptors (GPCRs) in both the purified, reconstituted state and in intact cells. These techniques span the entire spectrum from ligand binding to the receptors down to intracellular second messengers. They allow the determination and the visualization of signaling processes with high temporal and spatial resolution. With these techniques, it has been demonstrated that GPCR signals may show spatial and temporal patterning. In particular, evidence has been provided for spatial compartmentalization of GPCRs and their signals in intact cells and for distinct physiological consequences of such spatial patterning. We review here the FRET and BRET technologies that have been developed for G-protein-coupled receptors and their signaling proteins (G-proteins, effectors) and the concepts that result from such experiments. PMID:22407612

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

    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.

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

    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

  18. Daple is a novel non-receptor GEF required for trimeric G protein activation in Wnt signaling.

    Aznar, Nicolas; Midde, Krishna K; Dunkel, Ying; Lopez-Sanchez, Inmaculada; Pavlova, Yelena; Marivin, Arthur; Barbazán, Jorge; Murray, Fiona; Nitsche, Ulrich; Janssen, Klaus-Peter; Willert, Karl; Goel, Ajay; Abal, Miguel; Garcia-Marcos, Mikel; Ghosh, Pradipta

    2015-01-01

    Wnt signaling is essential for tissue homeostasis and its dysregulation causes cancer. Wnt ligands trigger signaling by activating Frizzled receptors (FZDRs), which belong to the G-protein coupled receptor superfamily. However, the mechanisms of G protein activation in Wnt signaling remain controversial. In this study, we demonstrate that FZDRs activate G proteins and trigger non-canonical Wnt signaling via the Dishevelled-binding protein, Daple. Daple contains a Gα-binding and activating (GBA) motif, which activates Gαi proteins and an adjacent domain that directly binds FZDRs, thereby linking Wnt stimulation to G protein activation. This triggers non-canonical Wnt responses, that is, suppresses the β-catenin/TCF/LEF pathway and tumorigenesis, but enhances PI3K-Akt and Rac1 signals and tumor cell invasiveness. In colorectal cancers, Daple is suppressed during adenoma-to-carcinoma transformation and expressed later in metastasized tumor cells. Thus, Daple activates Gαi and enhances non-canonical Wnt signaling by FZDRs, and its dysregulation can impact both tumor initiation and progression to metastasis. PMID:26126266

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

    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

  20. S6K1 Negatively Regulates TAK1 Activity in the Toll-Like Receptor Signaling Pathway

    Kim, So Yong; Baik, Kyung-Hwa; Baek, Kwan-Hyuck; Chah, Kyong-Hwa; Kim, Kyung Ah; Moon, Gyuyoung; Jung, Eunyu; Kim, Seong-Tae; Shim, Jae-Hyuck; Greenblatt, Matthew B.; Chun, Eunyoung; Lee, Ki-Young

    2014-01-01

    Transforming growth factor β (TGF-β)-activated kinase 1 (TAK1) is a key regulator in the signals transduced by proinflammatory cytokines and Toll-like receptors (TLRs). The regulatory mechanism of TAK1 in response to various tissue types and stimuli remains incompletely understood. Here, we show that ribosomal S6 kinase 1 (S6K1) negatively regulates TLR-mediated signals by inhibiting TAK1 activity. S6K1 overexpression causes a marked reduction in NF-κB and AP-1 activity induced by stimulation...

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

    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.

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

    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

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

    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...

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

    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...

  5. Blood coagulation-(in)dependent protease activated receptor signaling in pancreatic cancer

    K. Shi

    2015-01-01

    There is no doubt that pancreatic cancer is one of the most lethal diseases in general and current treatment options are limited. The studies described in this thesis pinpoint Protease activated receptors (PARs) as key players in pancreatic cancer progression. Inhibition of PAR-1 may be clinical rel

  6. The phytosulfokine (PSK) receptor is capable of guanylate cyclase activity and enabling cyclic GMP-dependent signaling in plants

    Kwezi, Lusisizwe

    2011-04-19

    Phytosulfokines (PSKs) are sulfated pentapeptides that stimulate plant growth and differentiation mediated by the PSK receptor (PSKR1), which is a leucine-rich repeat receptor-like kinase. We identified a putative guanylate cyclase (GC) catalytic center in PSKR1 that is embedded within the kinase domain and hypothesized that the GC works in conjunction with the kinase in downstream PSK signaling. We expressed the recombinant complete kinase (cytoplasmic) domain of AtPSKR1 and show that it has serine/threonine kinase activity using the Ser/Thr peptide 1 as a substrate with an approximate Km of 7.5 μM and Vmax of 1800 nmol min-1 mg-1 of protein. This same recombinant protein also has GC activity in vitro that is dependent on the presence of either Mg2+ or Mn2+. Overexpression of the full-length AtPSKR1 receptor in Arabidopsis leaf protoplasts raised the endogenous basal cGMP levels over 20-fold, indicating that the receptor has GC activity in vivo. In addition, PSK-α itself, but not the non-sulfated backbone, induces rapid increases in cGMP levels in protoplasts. Together these results indicate that the PSKR1 contains dual GC and kinase catalytic activities that operate in vivo and that this receptor constitutes a novel class of enzymes with overlapping catalytic domains. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.

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

    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.

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

    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.

  9. α1A-adrenergic receptor induces activation of extracellular signal-regulated kinase 1/2 through endocytic pathway.

    Fei Liu

    Full Text Available G protein-coupled receptors (GPCRs activate mitogen-activated protein kinases through a number of distinct pathways in cells. Increasing evidence has suggested that endosomal signaling has an important role in receptor signal transduction. Here we investigated the involvement of endocytosis in α(1A-adrenergic receptor (α(1A-AR-induced activation of extracellular signal-regulated kinase 1/2 (ERK1/2. Agonist-mediated endocytic traffic of α(1A-AR was assessed by real-time imaging of living, stably transfected human embryonic kidney 293A cells (HEK-293A. α(1A-AR was internalized dynamically in cells with agonist stimulation, and actin filaments regulated the initial trafficking of α(1A-AR. α(1A-AR-induced activation of ERK1/2 but not p38 MAPK was sensitive to disruption of endocytosis, as demonstrated by 4°C chilling, dynamin mutation and treatment with cytochalasin D (actin depolymerizing agent. Activation of protein kinase C (PKC and C-Raf by α(1A-AR was not affected by 4°C chilling or cytochalasin D treatment. U73122 (a phospholipase C [PLC] inhibitor and Ro 31-8220 (a PKC inhibitor inhibited α(1B-AR- but not α(1A-AR-induced ERK1/2 activation. These data suggest that the endocytic pathway is involved in α(1A-AR-induced ERK1/2 activation, which is independent of G(q/PLC/PKC signaling.

  10. Transferrin receptor facilitates TGF-β and BMP signaling activation to control craniofacial morphogenesis.

    Lei, R; Zhang, K; Liu, K; Shao, X; Ding, Z; Wang, F; Hong, Y; Zhu, M; Li, H; Li, H

    2016-01-01

    The Pierre Robin Sequence (PRS), consisting of cleft palate, glossoptosis and micrognathia, is a common human birth defect. However, how this abnormality occurs remains largely unknown. Here we report that neural crest cell (NCC)-specific knockout of transferrin receptor (Tfrc), a well known transferrin transporter protein, caused micrognathia, cleft palate, severe respiratory distress and inability to suckle in mice, which highly resemble human PRS. Histological and anatomical analysis revealed that the cleft palate is due to the failure of palatal shelves elevation that resulted from a retarded extension of Meckel's cartilage. Interestingly, Tfrc deletion dramatically suppressed both transforming growth factor-β (TGF-β) and bone morphogenetic protein (BMP) signaling in cranial NCCs-derived mandibular tissues, suggesting that Tfrc may act as a facilitator of these two signaling pathways during craniofacial morphogenesis. Together, our study uncovers an unknown function of Tfrc in craniofacial development and provides novel insight into the etiology of PRS. PMID:27362800

  11. 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.

    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

  12. Xingshentongqiao Decoction Mediates Proliferation, Apoptosis, Orexin-A Receptor and Orexin-B Receptor Messenger Ribonucleic Acid Expression and Represses Mitogen-activated Protein Kinase Signaling

    Yuanli Dong

    2015-01-01

    Full Text Available Background: Hypocretin (HCRT signaling plays an important role in the pathogenesis of narcolepsy and can be significantly influenced by Chinese herbal therapy. Our previous study showed that xingshentongqiao decoction (XSTQ is clinically effective for the treatment of narcolepsy. To determine whether XSTQ improves narcolepsy by modulating HCRT signaling, we investigated its effects on SH-SY5Y cell proliferation, apoptosis, and HCRT receptor 1/2 (orexin receptor 1 [OX1R] and orexin receptor 2 [OX2R] expression. The signaling pathways involved in these processes were also assessed. Methods: The effects of XSTQ on proliferation and apoptosis in SH-SY5Y cells were assessed using cell counting kit-8 and annexin V-fluorescein isothiocyanate assays. OX1R and OX2R expression was assessed by quantitative real-time polymerase chain reaction analysis. Western blotting for mitogen-activated protein kinase (MAPK pathway activation was performed to further assess the signaling mechanism of XSTQ. Results: XSTQ reduced the proliferation and induced apoptosis of SH-SY5Y cells. This effect was accompanied by the upregulation of OX1R and OX2R expression and the reduced phosphorylation of extracellular signal-regulated kinase (Erk 1/2, p38 MAPK and c-Jun N-terminal kinase (JNK. Conclusions: XSTQ inhibits proliferation and induces apoptosis in SH-SY5Y cells. XSTQ also promotes OX1R and OX2R expression. These effects are associated with the repression of the Erk1/2, p38 MAPK, and JNK signaling pathways. These results define a molecular mechanism for XSTQ in regulating HCRT and MAPK activation, which may explain its ability to treat narcolepsy.

  13. The Receptor for Activated C Kinase in Plant Signaling: Tale of a Promiscuous Little Molecule

    Islas-Flores, Tania; Rahman, Ahasanur; Ullah, Hemayet; Villanueva, Marco A.

    2015-01-01

    Two decades after the first report of the plant homolog of the Receptor for Activated C Kinase 1 (RACK1) in cultured tobacco BY2 cells, a significant advancement has been made in the elucidation of its cellular and molecular role. The protein is now implicated in many biological functions including protein translation, multiple hormonal responses, developmental processes, pathogen infection resistance, environmental stress responses, and miRNA production. Such multiple functional roles are co...

  14. Gq/5-HT2c receptor signals activate a local GABAergic inhibitory feedback circuit to modulate serotonergic firing and anxiety in mice

    Spoida, Katharina; Masseck, Olivia A.; Deneris, Evan S.; Herlitze, Stefan

    2014-01-01

    Serotonin is an important modulator of anxious states. Serotonergic neurotransmission in the dorsal raphe nuclei is known to be regulated by different presynaptic and postsynaptic feedback mechanisms involving G protein-coupled receptor signals, but the influence of such feedback mechanisms on anxiety-related behavior has not been investigated until now. We found that optogenetic activation of the Gq-coupled receptor signals in 5-HT2c receptor locations in GABAergic neurons in dorsal raphe nu...

  15. Selective estrogen receptor modulator BC-1 activates antioxidant signaling pathway in vitro via formation of reactive metabolites

    Bo-lan YU; Zi-xin MAI; Xu-xiang LIU; Zhao-feng HUANG

    2013-01-01

    Aim:Benzothiophene compounds are selective estrogen receptor modulators (SERMs),which are recently found to activate antioxidant signaling.In this study the molecular mechanisms of antioxidant signaling activation by benzothiophene compound BC-1 were investigated.Methods:HepG2 cells were stably transfected with antioxidant response element (ARE)-Iuciferase reporter (HepG2-ARE cells).The expression of nuclear factor erythroid 2-related factor 2 (Nrf2) in HepG2-ARE cells was suppressed using siRNA.The metabolites of BC-1 in rat liver microsome incubation were analyzed using LC-UV and LC-MS.Results:Addition of BC-1 (5 μmol/L) in HepG2-ARE cells resulted in a 17-fold increase of ARE-luciferase activity.Pretreatment with the estrogen receptor agonist E2 (5 μmol/L) or antagonist ICl 182,780 (5 μmol/L) did not affect BC-1-induced ARE-luciferase activity.However,transfection of the cells with anti-Nrf2 siRNA suppressed this effect by 79%.Addition of BC-1 in rat microsome incubation resulted in formation of di-quinone methides and o-quinones,followed by formation of GSH conjugates.BC-1 analogues with hydrogen (BC-2) or fluorine (BC-3) at the 4' position did not form the di-quinone methides.Both BC-2 and BC-3 showed comparable estrogenic activity with BC-1,but did not induce ARE-luciferase activity in HepG2-ARE cells.Conclusion:Benzothiophene compound BC-1 activates ARE signaling via reactive metabolite formation that is independent of estrogen receptors.

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

    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...

  17. Prostaglandin Receptor Signaling in Disease

    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.

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

    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

  19. Nuclear Receptor Signaling Atlas (NURSA)

    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...

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

    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

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

    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...

  2. GABA-A receptor-mediated signaling alters the structure of spontaneous activity in the developing retina

    Wang, Chih-Tien; Blankenship, Aaron G.; Anishchenko, Anastasia; Elstrott, Justin; Fikhman, Michael; Nakanishi, Shigetada; Feller, Marla B

    2007-01-01

    Ambient GABA modulates firing patterns in adult neural circuits by tonically activating extrasynaptic GABA-A receptors. Here, we demonstrate that during a developmental period when activation of GABA-A receptors causes membrane depolarization, tonic activation of GABA-A receptors blocks all spontaneous activity recorded in retinal ganglion cells (RGCs) and starburst amacrine cells (SACs). Bath application of the GABA-A receptor agonist muscimol blocked spontaneous correlated increases in intr...

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

    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. Dopamine D2 receptor activates extracellular signal-regulated kinase through the specific region in the third cytoplasmic loop.

    Takeuchi, Yusuke; Fukunaga, Kohji

    2004-06-01

    To investigate whether the third cytoplasmic loop and the C-terminal cytoplasmic tail of dopamine D(2) receptor (D2R) are involved in extracellular signal-regulated kinase (ERK) activation and subsequent regulation of transcription factors, we established NG108-15 cells stably expressing D2LR and D2SR deleted 40 amino acid residues in the third cytoplasmic loop (NGD2LR-3rd-dele and NGD2SR-3rd-dele) or the C-terminal cytoplasmic tail (NGD2LR-C-dele and NGD2SR-C-dele) and evaluated these receptors' functions using luciferase reporter gene assay. Immunocytochemical studies showed similar intracellular distributions of D2LR-3rd-dele and D2SR-3rd-dele to D2LR and D2SR, respectively. Quinpirole-induced inhibition of forskolin-induced cyclic AMP responsive element (CRE) activation was not affected by the deletion of 40 amino acid residues. However, nuclear factor-kappa B (NF-kappaB) activation by D2R-3rd-dele was largely attenuated compared to that by D2R. Similarly, ERK or serum-responsive element (SRE) activation by quinpirole treatment was totally abolished in NGD2R-3rd-dele cells. Moreover, D2R-C-dele was diffusely distributed or clustered in the cell bodies and lost the receptor functions. Taken together, the 40 amino acid residues in the third cytoplasmic loop are essential for the ERK activation but not for inhibition of adenylyl cyclase through Gi/o proteins. In addition, the C-terminal cytoplasmic tail is essential for membrane association of D2Rs to elicit the receptor functions. PMID:15189353

  5. Agonist-induced activation of histamine H3 receptor signals to extracellular signal-regulated kinases 1 and 2 through PKC-, PLD-, and EGFR-dependent mechanisms.

    Lai, Xiangru; Ye, Lingyan; Liao, Yuan; Jin, Lili; Ma, Qiang; Lu, Bing; Sun, Yi; Shi, Ying; Zhou, Naiming

    2016-04-01

    The histamine H3 receptor (H3R), abundantly expressed in the central and the peripheral nervous system, has been recognized as a promising target for the treatment of various important CNS diseases including narcolepsy, Alzheimer's disease, and attention deficit hyperactivity disorder. The H3R acts via Gi/o -proteins to inhibit adenylate cyclase activity and modulate MAPK activity. However, the underlying molecular mechanisms for H3R mediation of the activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) remain to be elucidated. In this study, using HEK293 cells stably expressing human H3R and mouse primary cortical neurons endogenously expressing mouse H3R, we found that the H3R-mediated activation of ERK1/2 was significantly blocked by both the pertussis toxin and the MEK1/2 inhibitor U0126. Upon stimulation by H3R agonist histamine or imetit, H3R was shown to rapidly induce ERK1/2 phosphorylation via PLC/PKC-, PLDs-, and epidermal growth factor receptor (EGFR) transactivation-dependent pathways. Furthermore, it was also indicated that while the βγ-subunits play a key role in H3R-activated ERK1/2 phosphorylation, β-arrestins were not required for ERK1/2 activation. In addition, when the cultured mouse cortical neurons were exposed to oxygen and glucose deprivation conditions (OGD), imetit exhibited neuroprotective properties through the H3R. Treatment of cells with the inhibitor UO126 abolished these protective effects. This suggests a possible neuroprotective role of the H3R-mediated ERK1/2 pathway under hypoxia conditions. These observations may provide new insights into the pharmacological effects and the physiological functions modulated by the H3R-mediated activation of ERK1/2. Histamine H3 receptors are abundantly expressed in the brain and play important roles in various CNS physiological functions. However, the underlying mechanisms for H3R-induced activation of extracellular signal-regulated kinase (ERK)1/2 remain largely unknown. Here

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

    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

  7. Neu1 desialylation of sialyl alpha-2,3-linked beta-galactosyl residues of TOLL-like receptor 4 is essential for receptor activation and cellular signaling.

    Amith, Schammim Ray; Jayanth, Preethi; Franchuk, Susan; Finlay, Trisha; Seyrantepe, Volkan; Beyaert, Rudi; Pshezhetsky, Alexey V; Szewczuk, Myron R

    2010-02-01

    The ectodomain of TOLL-like receptors (TLR) is highly glycosylated with several N-linked gylcosylation sites located in the inner concave surface. The precise role of these sugar N-glycans in TLR receptor activation is unknown. Recently, we have shown that Neu1 sialidase and not Neu2, -3 and -4 forms a complex with TLR-2, -3 and -4 receptors on the cell-surface membrane of naïve and activated macrophage cells (Glycoconj J DOI 10.1007/s10719-009-9239-8). Activation of Neu1 is induced by TLR ligands binding to their respective receptors. Here, we show that endotoxin lipopolysaccharide (LPS)-induced MyD88/TLR4 complex formation and subsequent NFkappaB activation is dependent on the removal of alpha-2,3-sialyl residue linked to beta-galactoside of TLR4 by the Neu1 activity associated with LPS-stimulated live primary macrophage cells, macrophage and dendritic cell lines but not with primary Neu1-deficient macrophage cells. Exogenous alpha-2,3 sialyl specific neuraminidase (Streptoccocus pneumoniae) and wild-type T. cruzi trans-sialidase (TS) but not the catalytically inactive mutant TSAsp98-Glu mediate TLR4 dimerization to facilitate MyD88/TLR4 complex formation and NFkappaB activation similar to those responses seen with LPS. These same TLR ligand-induced NFkappaB responses are not observed in TLR deficient HEK293 cells, but are re-established in HEK293 cells stably transfected with TLR4/MD2, and are significantly inhibited by alpha-2,3-sialyl specific Maackia amurensis (MAL-2) lectin, alpha-2,3-sialyl specific galectin-1 and neuraminidase inhibitor Tamiflu but not by alpha-2,6-sialyl specific Sambucus nigra lectin (SNA). Taken together, the findings suggest that Neu1 desialylation of alpha-2,3-sialyl residues of TLR receptors enables in removing a steric hinderance to receptor association for TLR activation and cellular signaling. PMID:19796680

  8. Optogenetic Activation of Adenosine A2A Receptor Signaling in the Dorsomedial Striatopallidal Neurons Suppresses Goal-Directed Behavior.

    Li, Yan; He, Yan; Chen, Mozi; Pu, Zhilan; Chen, Li; Li, Ping; Li, Bo; Li, Haiyan; Huang, Zhi-Li; Li, Zhihui; Chen, Jiang-Fan

    2016-03-01

    The striatum has an essential role in neural control of instrumental behaviors by reinforcement learning. Adenosine A(2A) receptors (A(2A)Rs) are highly enriched in the striatopallidal neurons and are implicated in instrumental behavior control. However, the temporal importance of the A(2A)R signaling in relation to the reward and specific contributions of the striatopallidal A(2A)Rs in the dorsolateral striatum (DLS) and the dorsomedial striatum (DMS) to the control of instrumental learning are not defined. Here, we addressed temporal relationship and sufficiency of transient activation of optoA(2A)R signaling precisely at the time of the reward to the control of instrumental learning, using our newly developed rhodopsin-A2AR chimeras (optoA(2A)R). We demonstrated that transient light activation of optoA(2A)R signaling in the striatopallidal neurons in 'time-locked' manner with the reward delivery (but not random optoA(2A)R activation) was sufficient to change the animal's sensitivity to outcome devaluation without affecting the acquisition or extinction phases of instrumental learning. We further demonstrated that optogenetic activation of striatopallidal A(2A)R signaling in the DMS suppressed goal-directed behaviors, as focally genetic knockdown of striatopallidal A(2A)Rs in the DMS enhanced goal-directed behavior by the devaluation test. By contrast, optogenetic activation or focal AAV-Cre-mediated knockdown of striatopallidal A(2A)R in the DLS had relatively limited effects on instrumental learning. Thus, the striatopallidal A(2A)R signaling in the DMS exerts inhibitory and predominant control of goal-directed behavior by acting precisely at the time of reward, and may represent a therapeutic target to reverse abnormal habit formation that is associated with compulsive obsessive disorder and drug addiction. PMID:26216520

  9. Erythropoietin receptor signaling is membrane raft dependent.

    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.

  10. Inhibition of P2Y6 receptor-mediated phospholipase C activation and Ca(2+) signalling by prostaglandin E2 in J774 murine macrophages.

    Ito, Masaaki; Matsuoka, Isao

    2015-02-15

    Extracellular nucleotides act as inflammatory mediators through activation of multiple purinoceptors. Under inflammatory conditions, the purinergic signalling is affected by various inflammatory mediators. We previously showed that prostaglandin (PG) E2 suppressed the elevation of intracellular Ca(2+) concentration ([Ca(2+)]i) stimulated by P2X4, P2Y2, and P2Y6 receptors in J774 murine macrophages. In this study, we examined the mechanism of PGE2 inhibitory effects on P2Y6 receptor-mediated function in J774 cells. The P2Y6 receptor agonist UDP induced a sustained elevation of [Ca(2+)]i by stimulating the phospholipase C (PLC) signalling pathway. PGE2 inhibited [Ca(2+)]i elevation and phosphatidylinositol (PI) hydrolysis in a concentration-dependent manner. J774 cells highly expressed the E-type prostanoid 2 (EP2) receptor subtype, a Gs-coupled receptor. PGE2 and a selective EP2 receptor agonist caused cyclic AMP (cAMP) accumulation in J774 cells. The inhibitory effects of PGE2 on P2Y6 receptor-mediated responses were mimicked by the selective EP2 receptor agonist. Although EP2 receptor is linked to adenylyl cyclase activation, PGE2-induced inhibition of Ca(2+) response and PI hydrolysis could not be mimicked by a lipophilic cAMP derivative, dibutyryl cAMP, or an adenylyl cyclase activator, forskolin. The inhibition of UDP-induced PLC activation by PGE2 was not affected by down-regulation of protein kinase C by phorbol-12-myristate-13-acetate treatment. PGE2 inhibited PLC activation induced by aluminium fluoride, but not by the Ca(2+)-ionophore, ionomycin. Finally, the inhibition of UDP-induced PLC activation by PGE2 was impaired by Gs knockdown using siRNA. These results suggest that EP2 receptor activation in macrophages negatively controls the Gq/11-PLC signalling through a Gs-mediated, but cAMP-independent signalling mechanism. PMID:25614334

  11. Sphingosine 1-phosphate (S1P)/S1P receptor 1 signaling regulates receptor activator of NF-{kappa}B ligand (RANKL) expression in rheumatoid arthritis

    Takeshita, Harunori [Division of Rheumatology, Department of Internal Medicine, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan); Kitano, Masayasu, E-mail: mkitano6@hyo-med.ac.jp [Division of Rheumatology, Department of Internal Medicine, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan); Iwasaki, Tsuyoshi [Department of Pharmacy, Hyogo University of Health Sciences, 1-3-6 Minatojima Kobe, Hyogo 650-8530 (Japan); Kitano, Sachie; Tsunemi, Sachi; Sato, Chieri; Sekiguchi, Masahiro; Azuma, Naoto [Division of Rheumatology, Department of Internal Medicine, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan); Miyazawa, Keiji [Discovery Research III, Research and Development, Kissei Pharmaceutical Company, 4365-1 Hodakakashiwara, Azumino, Nagano 399-8304 (Japan); Hla, Timothy [Center for Vascular Biology, Department of Pathology and Laboratory Medicine, Weill Medical College of Cornell University, 1300 York Avenue, Box 69, NY 10065 (United States); Sano, Hajime [Division of Rheumatology, Department of Internal Medicine, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan)

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer MH7A cells and CD4{sup +} T cells expressed S1P1 and RANKL. Black-Right-Pointing-Pointer S1P increased RANKL expression in MH7A cells and CD4{sup +} T cells. Black-Right-Pointing-Pointer The effect of S1P in MH7A cells was inhibited by specific Gi/Go inhibitors. -- Abstract: Sphingosine 1-phosphate (S1P)/S1P receptor 1 (S1P1) signaling plays an important role in synovial cell proliferation and inflammatory gene expression by rheumatoid arthritis (RA) synoviocytes. The purpose of this study is to clarify the role of S1P/S1P1 signaling in the expression of receptor activator of NF-{kappa}B ligand (RANKL) in RA synoviocytes and CD4{sup +} T cells. We demonstrated MH7A cells, a human RA synovial cell line, and CD4{sup +} T cells expressed S1P1 and RANKL. Surprisingly, S1P increased RANKL expression in MH7A cells and CD4{sup +} T cells in a dose-dependent manner. Moreover, S1P enhanced RANKL expression induced by stimulation with TNF-{alpha} in MH7A cells and CD4{sup +} T cells. These effects of S1P in MH7A cells were inhibited by pretreatment with PTX, a specific Gi/Go inhibitor. These findings suggest that S1P/S1P1 signaling may play an important role in RANKL expression by MH7A cells and CD4{sup +} T cells. S1P/S1P1 signaling of RA synoviocytes is closely connected with synovial hyperplasia, inflammation, and RANKL-induced osteoclastogenesis in RA. Thus, regulation of S1P/S1P1 signaling may become a novel therapeutic target for RA.

  12. Muscarinic Receptor Signaling in Colon Cancer

    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.

  13. Muscarinic Receptor Signaling in Colon Cancer

    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.

  14. Muscarinic Receptor Signaling in Colon Cancer

    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

  15. Calycosin promotes angiogenesis involving estrogen receptor and mitogen-activated protein kinase (MAPK signaling pathway in zebrafish and HUVEC.

    Jing Yan Tang

    Full Text Available BACKGROUND: Angiogenesis plays an important role in a wide range of physiological processes, and many diseases are associated with the dysregulation of angiogenesis. Radix Astragali is a Chinese medicinal herb commonly used for treating cardiovascular disorders and has been shown to possess angiogenic effect in previous studies but its active constituent and underlying mechanism remain unclear. The present study investigates the angiogenic effects of calycosin, a major isoflavonoid isolated from Radix Astragali, in vitro and in vivo. METHODOLOGY: Tg(fli1:EGFP and Tg(fli1:nEGFP transgenic zebrafish embryos were treated with different concentrations of calycosin (10, 30, 100 microM from 72 hpf to 96 hpf prior morphological observation and angiogenesis phenotypes assessment. Zebrafish embryos were exposed to calycosin (10, 100 microM from 72 hpf to 78 hpf before gene-expression analysis. The effects of VEGFR tyrosine kinase inhibitor on calycosin-induced angiogenesis were studied using 72 hpf Tg(fli1:EGFP and Tg(fli1:nEGFP zebrafish embryos. The pro-angiogenic effects of calycosin were compared with raloxifene and tamoxifen in 72 hpf Tg(fli1:EGFP zebrafish embryos. The binding affinities of calycosin to estrogen receptors (ERs were evaluated by cell-free and cell-based estrogen receptor binding assays. Human umbilical vein endothelial cell cultures (HUVEC were pretreated with different concentrations of calycosin (3, 10, 30, 100 microM for 48 h then tested for cell viability and tube formation. The role of MAPK signaling in calycosin-induced angiogenesis was evaluated using western blotting. CONCLUSION: Calycosin was shown to induce angiogenesis in human umbilical vein endothelial cell cultures (HUVEC in vitro and zebrafish embryos in vivo via the up-regulation of vascular endothelial growth factor (VEGF, VEGFR1 and VEGFR2 mRNA expression. It was demonstrated that calycosin acted similar to other selective estrogen receptor modulators (SERMs, such

  16. Selective Activator of the Glucocorticoid Receptor Compound A Dissociates Therapeutic and Atrophogenic Effects of Glucocorticoid Receptor Signaling in Skin

    Klopot, Anna; Baida, Gleb; Bhalla, Pankaj; Haegeman, Guy; Budunova, Irina

    2015-01-01

    Background: Glucocorticoids are effective anti-inflammatory drugs widely used in dermatology and for the treatment of blood cancer patients. Unfortunately, chronic treatment with glucocorticoids results in serious metabolic and atrophogenic adverse effects including skin atrophy. Glucocorticoids act via the glucocorticoid receptor (GR), a transcription factor that causes either gene transactivation (TA) or transrepression (TR). Compound A (CpdA), a novel non-steroidal GR ligand, does not prom...

  17. The Receptor for Activated C Kinase in Plant Signaling: Tale of a Promiscuous Little Molecule

    Tania Islas Flores

    2015-12-01

    Full Text Available Two decades after the first report of the plant homologue of the Receptor for Activated C Kinase 1 (RACK1 in cultured tobacco BY2 cells, a significant advancement has been made in the elucidation of its celular and molecular role. The protein is now implicated in many biological functions including protein translation, multiple hormonal responses, developmental processes, pathogen infection resistance, environmental stress responses, and miRNA production. Such multiple functional roles are consistent with the scaffolding nature of the plant RACK1 protein. A significant advance was achieved when the β-propeller structure of the Arabidopsis RACK1A isoform was elucidated, thus revealing that its conserved seven WD repeats also assembled into this typical topology. From its crystal structure, it became apparent that it shares the structural platform for the interaction with ligands identified in other systems such as mammals. Although RACK1 proteins maintain conserved Protein Kinase C binding sites, the lack of a bona fide PKC adds complexity and enigma to the nature of the ligand partners with which RACK1 interacts in plants. Nevertheless, ligands recently identified using the split-ubiquitin based and conventional yeast two-hybrid assays, have revealed that plant RACK1 is involved in several processes that include defense response, drought and salt stress, ribosomal function, cell wall biogenesis, and photosynthesis. The information acquired indicates that, in spite of the high degree of conservation of its structure, the functions of the plant RACK1 homologue appear to be distinct and diverse from those in yeast, mammals, insects, etc. In this review, we take a critical look at the novel information regarding the many functions in which plant RACK1 has been reported to participate, with a special emphasis on the information on its currently identified and missing ligand partners.

  18. The Receptor for Activated C Kinase in Plant Signaling: Tale of a Promiscuous Little Molecule.

    Islas-Flores, Tania; Rahman, Ahasanur; Ullah, Hemayet; Villanueva, Marco A

    2015-01-01

    Two decades after the first report of the plant homolog of the Receptor for Activated C Kinase 1 (RACK1) in cultured tobacco BY2 cells, a significant advancement has been made in the elucidation of its cellular and molecular role. The protein is now implicated in many biological functions including protein translation, multiple hormonal responses, developmental processes, pathogen infection resistance, environmental stress responses, and miRNA production. Such multiple functional roles are consistent with the scaffolding nature of the plant RACK1 protein. A significant advance was achieved when the β-propeller structure of the Arabidopsis RACK1A isoform was elucidated, thus revealing that its conserved seven WD repeats also assembled into this typical topology. From its crystal structure, it became apparent that it shares the structural platform for the interaction with ligands identified in other systems such as mammals. Although RACK1 proteins maintain conserved Protein Kinase C binding sites, the lack of a bona fide PKC adds complexity and enigma to the nature of the ligand partners with which RACK1 interacts in plants. Nevertheless, ligands recently identified using the split-ubiquitin based and conventional yeast two-hybrid assays, have revealed that plant RACK1 is involved in several processes that include defense response, drought and salt stress, ribosomal function, cell wall biogenesis, and photosynthesis. The information acquired indicates that, in spite of the high degree of conservation of its structure, the functions of the plant RACK1 homolog appear to be distinct and diverse from those in yeast, mammals, insects, etc. In this review, we take a critical look at the novel information regarding the many functions in which plant RACK1 has been reported to participate, with a special emphasis on the information on its currently identified and missing ligand partners. PMID:26697044

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

    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

  20. Differential extracellular signal-regulated kinases 1 and 2 activation by the angiotensin type 1 receptor supports distinct phenotypes of cardiac myocytes

    Aplin, Mark; Christensen, Gitte Lund; Schneider, Mikael;

    2007-01-01

    The angiotensin II (AngII) type 1 receptor (AT(1)R) is a seven-transmembrane receptor well established to activate extracellular signal-regulated kinases 1 and 2 (ERK1/2) by discrete G protein-dependent and beta-arrestin2-dependent pathways. The biological importance of this, however, remains obs...... obscure. Application of the modified analogue [Sar(1), Ile(4), Ile(8)]-AngII ([SII] AngII) allowed us to dissect the two pathways of ERK1/2 activation in native cardiac myocytes. Although cytosol-retained, the beta-arrestin2-bound pool of ERK1/2 represents an active signalling component...

  1. Spinal 5-HT3 receptor activation induces behavioral hypersensitivity via a neuronal-glial-neuronal signaling cascade

    Gu, Ming; Miyoshi, Kan; Dubner, Ronald; Wei GUO; Zou, Shiping; Ren, Ke; Noguchi, Koichi; Wei, Feng

    2011-01-01

    Recent studies indicate that the descending serotonin (5-HT) system from the rostral ventromedial medulla (RVM) in brainstem and the 5-HT3 receptor subtype in the spinal dorsal horn are involved in enhanced descending pain facilitation after tissue and nerve injury. However, the mechanisms underlying the activation of the 5-HT3 receptor and its contribution to facilitation of pain remain unclear. In the present study, activation of spinal 5-HT3 receptor by intrathecal injection of a selective...

  2. Muscarinic receptor signaling and colon cancer progression

    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.

  3. Palmitoylation of Estrogen Receptors Is Essential for Neuronal Membrane Signaling

    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...

  4. Phosphoproteomic analysis of adhesion receptor signalling

    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...

  5. Autoantibody-mediated angiotensin receptor activation contributes to preeclampsia through TNF-alpha signaling

    Irani, Roxanna A.; Zhang, Yujin; Zhou, Cissy Chenyi; Blackwell, Sean C.; Hicks, M. John; Ramin, Susan M.; Kellems, Rodney E.; Xia, Yang

    2010-01-01

    Preeclampsia is a prevalent life-threatening hypertensive disorder of pregnancy whose pathophysiology remains largely undefined. Recently, a circulating maternal autoantibody, the angiotensin II type I receptor agonistic autoantibody (AT1-AA), has emerged as a contributor to disease features. Increased circulating maternal tumor necrosis factor alpha (TNF-α) is also associated with the disease, however it is unknown if this factor directly contributes to preeclamptic symptoms. Here we report ...

  6. Peroxisome proliferator-activated receptor γ is expressed in hippocampal neurons and its activation prevents β-amyloid neurodegeneration: role of Wnt signaling

    The molecular pathogenesis of Alzheimer's disease (AD) involves the participation of the amyloid-β-peptide (Aβ), which plays a critical role in the neurodegeneration that triggers the disease. Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors, which are members of the nuclear receptor family. We report here that (1) PPARγ is present in rat hippocampal neurons in culture. (2) Activation of PPARγ by troglitazone and rosiglitazone protects rat hippocampal neurons against Aβ-induced neurodegeneration, as shown by the 3-[4,5 -2yl]-2,5-diphenyltetrazolium bromide (MTT) reduction assay, immunofluorescence using an anti-heavy neurofilament antibody, and quantitative electron microscopy. (3) Hippocampal neurons treated with several PPARγ agonists, including troglitazone, rosiglitazone, and ciglitazone, prevent the excitotoxic Aβ-induced rise in bulk-free Ca2+. (4) PPARγ activation results in the modulation of Wnt signaling components, including the inhibition of glycogen synthase kinase-3β (GSK-3β) and an increase of the cytoplasmic and nuclear β-catenin levels. We conclude that the activation of PPARγ prevents Aβ-induced neurodegeneration by a mechanism that may involve a cross talk between neuronal PPARγ and the Wnt signaling pathway. More important, the fact that the activation of PPARγ attenuated Aβ-dependent neurodegeneration opens the possibility to fight AD from a new therapeutic perspective

  7. Promyelocytic leukemia zinc finger protein activates GATA4 transcription and mediates cardiac hypertrophic signaling from angiotensin II receptor 2.

    Ning Wang

    Full Text Available BACKGROUND: Pressure overload and prolonged angiotensin II (Ang II infusion elicit cardiac hypertrophy in Ang II receptor 1 (AT(1 null mouse, whereas Ang II receptor 2 (AT(2 gene deletion abolishes the hypertrophic response. The roles and signals of the cardiac AT(2 receptor still remain unsettled. Promyelocytic leukemia zinc finger protein (PLZF was shown to bind to the AT(2 receptor and transmit the hypertrophic signal. Using PLZF knockout mice we directed our studies on the function of PLZF concerning the cardiac specific transcription factor GATA4, and GATA4 targets. METHODOLOGY AND PRINCIPAL FINDINGS: PLZF knockout and age-matched wild-type (WT mice were treated with Ang II, infused at a rate of 4.2 ng·kg(-1·min(-1 for 3 weeks. Ang II elevated systolic blood pressure to comparable levels in PLZF knockout and WT mice (140 mmHg. WT mice developed prominent cardiac hypertrophy and fibrosis after Ang II infusion. In contrast, there was no obvious cardiac hypertrophy or fibrosis in PLZF knockout mice. An AT(2 receptor blocker given to Ang II-infused wild type mice prevented hypertrophy, verifying the role of AT(2 receptor for cardiac hypertrophy. Chromatin immunoprecipitation and electrophoretic mobility shift assay showed that PLZF bound to the GATA4 gene regulatory region. A Luciferase assay verified that PLZF up-regulated GATA4 gene expression and the absence of PLZF expression in vivo produced a corresponding repression of GATA4 protein. CONCLUSIONS: PLZF is an important AT(2 receptor binding protein in mediating Ang II induced cardiac hypertrophy through an AT(2 receptor-dependent signal pathway. The angiotensin II-AT(2-PLZF-GATA4 signal may further augment Ang II induced pathological effects on cardiomyocytes.

  8. 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.

    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

  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

    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

  10. Steroid Hormone Receptor Signals as Prognosticators for Urothelial Tumor

    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.

  11. Effect of hyperprolactinemia on PRL-receptor expression and activation of Stat and Mapk cell signaling in the prostate of long-term sexually-active rats.

    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

  12. Plasmin-induced migration requires signaling through protease-activated receptor 1 and integrin alpha(9)beta(1).

    Majumdar, Mousumi; Tarui, Takehiko; Shi, Biao; Akakura, Nobuaki; Ruf, Wolfram; Takada, Yoshikazu

    2004-09-01

    Plasmin is a major extracellular protease that elicits intracellular signals to mediate platelet aggregation, chemotaxis of peripheral blood monocytes, and release of arachidonate and leukotriene from several cell types in a G protein-dependent manner. Angiostatin, a fragment of plasmin(ogen), is a ligand and an antagonist for integrin alpha(9)beta(1). Here we report that plasmin specifically interacts with alpha(9)beta(1) and that plasmin induces of cells expressing migration recombinant alpha(9)beta(1) (alpha(9)-Chinese hamster ovary (CHO) cells). Migration was dependent on an interaction of the kringle domains of plasmin with alpha(9)beta(1) as well as the catalytic activity of plasmin. Angiostatin, representing the kringle domains of plasmin, alone did not induce the migration of alpha(9)-CHO cells, but simultaneous activation of the G protein-coupled protease-activated receptor (PAR)-1 with an agonist peptide induced the migration on angiostatin, whereas PAR-2 or PAR-4 agonist peptides were without effect. Furthermore, a small chemical inhibitor of PAR-1 (RWJ 58259) and a palmitoylated PAR-1-blocking peptide inhibited plasmin-induced migration of alpha(9)-CHO cells. These results suggest that plasmin induces migration by kringle-mediated binding to alpha(9)beta(1) and simultaneous proteolytic activation of PAR-1. PMID:15247268

  13. The oxytocin receptor antagonist, Atosiban, activates pro-inflammatory pathways in human amnion via G(αi) signalling.

    Kim, Sung Hye; MacIntyre, David A; Hanyaloglu, Aylin C; Blanks, Andrew M; Thornton, Steven; Bennett, Phillip R; Terzidou, Vasso

    2016-01-15

    Oxytocin (OT) plays an important role in the onset of human labour by stimulating uterine contractions and promoting prostaglandin/inflammatory cytokine synthesis in amnion via oxytocin receptor (OTR) coupling. The OTR-antagonist, Atosiban, is widely used as a tocolytic for the management of acute preterm labour. We found that in primary human amniocytes, Atosiban (10 μM) signals via PTX-sensitive Gαi to activate transcription factor NF-κB p65, ERK1/2, and p38 which subsequently drives upregulation of the prostaglandin synthesis enzymes, COX-2 and phospho-cPLA2 and excretion of prostaglandins (PGE2) (n = 6; p Atosiban treatment increased expression and excretion of the inflammatory cytokines, IL-6 and CCL5. We also showed that OT-simulated activation of NF-κB, ERK1/2, and p38 and subsequent prostaglandin and inflammatory cytokine synthesis is via Gαi-2 and Gαi-3 but not Gαq, and is not inhibited by Atosiban. Activation or exacerbation of inflammation is not a desirable effect of tocolytics. Therefore therapeutic modulation of the OT/OTR system for clinical management of term/preterm labour should consider the effects of differential G-protein coupling of the OTR and the role of OT or selective OTR agonists/antagonists in activating proinflammatory pathways. PMID:26586210

  14. Protease-activated receptors (PARs) in cancer: Novel biased signaling and targets for therapy.

    Bar-Shavit, R; Maoz, M; Kancharla, A; Jaber, M; Agranovich, D; Grisaru-Granovsky, S; Uziely, B

    2016-01-01

    Despite the fact that G protein-coupled receptors (GPCRs) mediate numerous physiological processes and represent targets for therapeutics for a vast array of diseases, their role in tumor biology is under appreciated. Protease-activated receptors (PARs) form a family which belongs to GPCR class A. PAR1&2 emerge with a central role in epithelial malignancies. Although the part of PAR1&2 in cancer is on the rise, their underlying signaling events are poorly understood. We review hereby past, present, and future cancer-associated PAR biology. Mainly, their role in physiological (placenta-cytotophobalst) and patho-physiological invasion processes. The identification and characterization of signal pleckstrin homology (PH)-domain-binding motifs established critical sites for breast cancer growth in PAR1&2. Among the proteins found to harbor important PH-domains and are involved in PAR biology are Akt/PKB as also Etk/Bmx and Vav3. A point mutation in PAR2, H349A, but not R352A, abrogated PH-protein association and is sufficient to markedly reduce PAR2-instigated breast tumor growth in vivo as also placental extravillous trophoblast (EVT) invasion in vitro is markedly reduced. Similarly, the PAR1 mutant hPar1-7A, which is unable to bind PH-domain, inhibits mammary tumors and EVT invasion, endowing these motifs with physiological significance and underscoring the importance of these previously unknown PAR1 and PAR2 PH-domain-binding motifs in both pathological and physiological invasion processes. PMID:26928551

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

    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 ...

  16. Immune receptor signaling: from ubiquitination to NF-κB activation

    Shao-Cong Sun

    2012-01-01

    The immune system functions as a dynamic and sophisticated network that ensures efficient response to foreign antigens and tolerance to self-tissues.1 The function of the immune system relies on signal transduction that connects immune cells to the extracellular environment and mediates communication among the different types of immune cells.

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

    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. Regulation of endothelial protein C receptor shedding by cytokines is mediated through differential activation of MAP kinase signaling pathways

    The endothelial protein C receptor (EPCR) plays a pivotal role in coagulation, inflammation, cell proliferation, and cancer, but its activity is markedly changed by ectodomain cleavage and release as the soluble protein (sEPCR). In this study we examined the mechanisms involved in the regulation of EPCR shedding in human umbilical endothelial cells (HUVEC). Interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α), but not interferon-γ and interleukin-6, suppressed EPCR mRNA transcription and cell-associated EPCR expression in HUVEC. The release of sEPCR induced by IL-1β and TNF-α correlated with activation of p38 MAPK and c-Jun N-terminal kinase (JNK). EPCR shedding was also induced by phorbol 12-myristate 13-acetate, ionomycin, anisomycin, thiol oxidants or alkylators, thrombin, and disruptors of lipid rafts. Both basal and induced shedding of EPCR was blocked by the metalloproteinase inhibitors, TAPI-0 and GM6001, and by the reduced non-protein thiols, glutathione, dihydrolipoic acid, dithiothreitol, and N-acetyl-L-cysteine. Because other antioxidants and scavengers of reactive oxygen species failed to block the cleavage of EPCR, a direct suppression of metalloproteinase activity seems responsible for the observed effects of reduced thiols. In summary, the shedding of EPCR in HUVEC is effectively regulated by IL-1β and TNF-α, and downstream by MAP kinase signaling pathways and metalloproteinases.

  19. Regulation of endothelial protein C receptor shedding by cytokines is mediated through differential activation of MAP kinase signaling pathways

    Menschikowski, Mario, E-mail: Mario.Menschikowski@uniklinikum-dresden.de [Institute of Clinical Chemistry and Laboratory Medicine, Technical University of Dresden, Medical Faculty ' Carl Gustav Carus' , Fetscherstrasse 74, D-01307 Dresden (Germany); Hagelgans, Albert; Eisenhofer, Graeme; Siegert, Gabriele [Institute of Clinical Chemistry and Laboratory Medicine, Technical University of Dresden, Medical Faculty ' Carl Gustav Carus' , Fetscherstrasse 74, D-01307 Dresden (Germany)

    2009-09-10

    The endothelial protein C receptor (EPCR) plays a pivotal role in coagulation, inflammation, cell proliferation, and cancer, but its activity is markedly changed by ectodomain cleavage and release as the soluble protein (sEPCR). In this study we examined the mechanisms involved in the regulation of EPCR shedding in human umbilical endothelial cells (HUVEC). Interleukin-1{beta} (IL-1{beta}) and tumor necrosis factor-{alpha} (TNF-{alpha}), but not interferon-{gamma} and interleukin-6, suppressed EPCR mRNA transcription and cell-associated EPCR expression in HUVEC. The release of sEPCR induced by IL-1{beta} and TNF-{alpha} correlated with activation of p38 MAPK and c-Jun N-terminal kinase (JNK). EPCR shedding was also induced by phorbol 12-myristate 13-acetate, ionomycin, anisomycin, thiol oxidants or alkylators, thrombin, and disruptors of lipid rafts. Both basal and induced shedding of EPCR was blocked by the metalloproteinase inhibitors, TAPI-0 and GM6001, and by the reduced non-protein thiols, glutathione, dihydrolipoic acid, dithiothreitol, and N-acetyl-L-cysteine. Because other antioxidants and scavengers of reactive oxygen species failed to block the cleavage of EPCR, a direct suppression of metalloproteinase activity seems responsible for the observed effects of reduced thiols. In summary, the shedding of EPCR in HUVEC is effectively regulated by IL-1{beta} and TNF-{alpha}, and downstream by MAP kinase signaling pathways and metalloproteinases.

  20. The haematopoietic GTPase RhoH modulates IL3 signalling through regulation of STAT activity and IL3 receptor expression

    Gündogdu Mehtap S

    2010-08-01

    Full Text Available Abstract Background RhoH is a constitutively active member of the family of Rho GTPases. Its expression is restricted to the haematopoietic lineage, where it serves as a positive regulator for T cell selection and mast cell function and as a negative regulator for growth-related functions in other lineages. Here, we examined the activation of signal transducer and activator of transcription (STAT proteins in response to stimulation with interleukin 3 (IL3. Results Using the murine IL3-dependent cell line BaF3 we investigated the influence of RhoH protein expression levels on IL3-mediated cellular responses. RhoH overexpressing cells showed lower sensitivity to IL3 and decreased STAT5 activation. SiRNA-mediated repression of RhoH gene expression led to an increase in proliferation and STAT5 activity which correlated with an increased number of IL3 receptor α chain molecules, also known as CD123, expressed at the cell surface. Interestingly, these findings could be reproduced using human THP-1 cells as a model system for acute myeloid leukaemia, where low RhoH levels are known to be an unfavourable prognostic marker. Overexpression of RhoH on the other hand caused an induction of STAT1 activity and western blot analysis revealed that activated STAT1 is phosphorylated on Tyr701. STAT1 is known to induce apoptosis or cell cycle arrest and we detected an upregulation of cyclin-dependent kinase inhibitors (CDKI p21Cip1 and p27Kip1 in RhoH overexpressing BaF3 cells. Conclusions We propose that RhoH functions as a negative regulator for IL3-induced signals through modulation of the JAK-STAT pathway. High levels of RhoH allow the IL3-dependent activation of STAT1 causing decreased proliferation through upregulation of p21Cip1 and p27Kip1. Low RhoH levels on the other hand led to an upregulation of IL3-dependent cell growth, STAT5 activity and an increase of CD123 surface expression, linking RhoH to a CD123/STAT5 phenotype that has been described in AML

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

    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

  2. Activation of the signaling cascade in response to T lymphocyte receptor stimulation and prostanoids in a case of cutaneous lupus

    Ana Maria Abreu-Velez

    2011-01-01

    Full Text Available Context: Discoid lupus erythematosus is a chronic inflammatory skin disorder presenting with scarring lesions that occur predominately on sun exposed areas of the face and scalp. Case Report: A 22-year-old male was evaluated after presenting with reddish-purple, atrophic and erythematous plaques on the scalp, with loss of hair within the plaques. Biopsies for hematoxylin and eosin examination, direct immunofluorescence and immunohistochemistry analysis were performed. The hematoxylin and eosin staining demonstrated classic features of cutaneous lupus. Direct immunofluorescence revealed strong deposits of immunoglobulins IgG and IgM, fibrinogen and Complement/C3, present in 1 a shaggy pattern at the epidermal basement membrane zone, and 2 focal pericytoplasmic and perinuclear staining within epidermal keratynocytes. Immunohistochemisty staining revealed strongly positive staining with antibodies to cyclooxygenase-2, Zeta-chain-associated protein kinase 70, and HLA-DPDQDR in areas where the inflammatory infiltrate was predominant, as well as around dermal blood vessels and within the dermal extracellular matrix. Conclusions : Noting the autoimmune nature of lupus and its strong inflammatory component, we present a patient with active discoid lupus erythematosus and strong expression of Zeta-chain-associated protein kinase 70, cyclo-oxygenase-2, and HLA-DPDQDR in the inflammatory areas. We suggest that these molecules may play a significant role in the immune response of discoid cutaneous lupus, possibly including 1 the biosynthesis of the prostanoids and 2 activation of the signaling cascade in response to T-lymphocyte receptor stimulation.

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

    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

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

    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. Membrane Trafficking of Death Receptors: Implications on Signalling

    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.

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

    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.

  7. Complexity of Receptor Tyrosine Kinase Signal Processing

    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

  8. Monoclonal anti-β1-adrenergic receptor antibodies activate G protein signaling in the absence of β-arrestin recruitment

    Hutchings, Catherine J; Cseke, Gabriella; Osborne, Greg; Woolard, Jeanette; Zhukov, Andrei; Koglin, Markus; Jazayeri, Ali; Pandya-Pathak, Jahnavi; Langmead, Christopher J.; Hill, Stephen J.; Weir, Malcolm; Marshall, Fiona H.

    2013-01-01

    Thermostabilized G protein-coupled receptors used as antigens for in vivo immunization have resulted in the generation of functional agonistic anti-β1-adrenergic (β1AR) receptor monoclonal antibodies (mAbs). The focus of this study was to examine the pharmacology of these antibodies to evaluate their mechanistic activity at β1AR. Immunization with the β1AR stabilized receptor yielded five stable hybridoma clones, four of which expressed functional IgG, as determined in cell-based assays used ...

  9. AT(1) receptor Gαq protein-independent signalling transcriptionally activates only a few genes directly, but robustly potentiates gene regulation from the β2-adrenergic receptor

    Christensen, Gitte Lund; Knudsen, Steen; Schneider, Mikael; Aplin, Mark; Gammeltoft, Steen; Sheikh, Søren P; Hansen, Jakob L

    2011-01-01

    potentiated β2-adrenergic receptor-stimulated gene expression. These novel findings indicate that the Gαq protein-independent signalling mainly modifies the transcriptional response governed by other signalling pathways, while direct induction of gene expression by the AT(1)R is dependent on classical Gαq......-independent signalling from the AT(1)R interact with transcriptional regulators and promote phosphorylation of nuclear proteins. However, the relative contribution of Gαq protein-independent signalling in AT(1)R mediated transcriptional regulation remains elusive. We here present a comprehensive comparative analysis of...... Gαq protein-dependent and -independent regulation of AT(1)R mediated gene expression. We found angiotensin II to regulate 212 genes, whereas Gαq-independent signalling obtained with the biased agonist, SII angiotensin II only regulated few genes. Interestingly, SII angiotensin II, like Ang II vastly...

  10. Ca 2+ signaling by plant Arabidopsis thaliana Pep peptides depends on AtPepR1, a receptor with guanylyl cyclase activity, and cGMP-activated Ca 2+ channels

    Qia, Zhi

    2010-11-18

    A family of peptide signaling molecules (AtPeps) and their plasma membrane receptor AtPepR1 are known to act in pathogendefense signaling cascades in plants. Little is currently known about the molecular mechanisms that link these signaling peptides and their receptor, a leucine-rich repeat receptor-like kinase, to downstream pathogen-defense responses. We identify some cellular activities of these molecules that provide the context for a model for their action in signaling cascades. AtPeps activate plasma membrane inwardly conducting Ca 2+ permeable channels in mesophyll cells, resulting in cytosolic Ca 2+ elevation. This activity is dependent on their receptor as well as a cyclic nucleotide-gated channel (CNGC2). We also show that the leucine-rich repeat receptor- like kinase receptor AtPepR1 has guanylyl cyclase activity, generating cGMP from GTP, and that cGMP can activate CNGC2- dependent cytosolic Ca 2+ elevation. AtPep-dependent expression of pathogen-defense genes (PDF1.2, MPK3, and WRKY33) is mediated by the Ca 2+ signaling pathway associated with AtPep peptides and their receptor. The work presented here indicates that extracellular AtPeps, which can act as danger-associated molecular patterns, signal by interaction with their receptor, AtPepR1, a plasma membrane protein that can generate cGMP. Downstream from AtPep and AtPepR1 in a signaling cascade, the cGMP-activated channel CNGC2 is involved in AtPep- and AtPepR1-dependent inward Ca 2+ conductance and resulting cytosolic Ca 2+ elevation. The signaling cascade initiated by AtPeps leads to expression of pathogen- defense genes in a Ca 2+-dependent manner.

  11. 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

    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

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

    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...

  13. The human angiotensin AT(1) receptor supports G protein-independent extracellular signal-regulated kinase 1/2 activation and cellular proliferation

    Hansen, Jakob Lerche; Aplin, Mark; Hansen, Jonas Tind; Christensen, Gitte Lund; Bonde, Marie Mi; Schneider, Mikael; Haunsø, Stig; Schiffer, Hans H; Burstein, Ethan S; Weiner, David M; Sheikh, Søren P

    AT(1) receptor signalling is illustrated by the common use of angiotensin AT(1) receptor-inverse agonists in clinical practice. It is well established that rodent orthologues of the angiotensin AT(1) receptor can selectively signal through G protein-dependent and -independent mechanisms in......(1) receptor actions. However, it is currently unknown whether the human angiotensin AT(1) receptor can signal through G protein-independent mechanisms - and if so, what the physiological impact of such signalling is. We have performed a detailed pharmacological analysis of the human angiotensin AT(1......) receptor using a battery of angiotensin analogues and registered drugs targeting this receptor. We show that the human angiotensin AT(1) receptor signals directly through G protein-independent pathways and supports NIH3T3 cellular proliferation. The realization of G protein-independent signalling by the...

  14. Essential Oil of Pinus koraiensis Exerts Antiobesic and Hypolipidemic Activity via Inhibition of Peroxisome Proliferator-Activated Receptors Gamma Signaling

    Hyun-Suk Ko; Hyo-Jeong Lee; Hyo-Jung Lee; Eun Jung Sohn; Miyong Yun; Min-Ho Lee; Sung-Hoon Kim

    2013-01-01

    Our group previously reported that essential oil of Pinus koraiensis (EOPK) exerts antihyperlipidemic effects via upregulation of low-density lipoprotein receptor and inhibition of acyl-coenzyme A. In the present study, we investigated the antiobesity and hypolipidemic mechanism of EOPK using in vitro 3T3-L1 cells and in vivo HFD-fed rats. EOPK markedly suppressed fat accumulation and intracellular triglyceride associated with downregulation of adipogenic transcription factor expression, incl...

  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

    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

    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. The Contribution of Proteinase-Activated Receptors to Intracellular Signaling, Transcellular Transport and Autophagy in Alzheimer´s Disease

    Matěj, R.; Rohan, Z.; Holada, K.; Olejár, Tomáš

    2015-01-01

    Roč. 12, č. 1 (2015), s. 2-12. ISSN 1567-2050 Institutional support: RVO:67985823 Keywords : Alzheimer ´s Disease * autophagy * proteinase-activated receptors Subject RIV: EA - Cell Biology Impact factor: 3.889, year: 2014

  18. The Vitamin D Receptor Is Required for Activation of cWnt and Hedgehog Signaling in Keratinocytes

    Lisse, Thomas S.; Saini, Vaibhav; Zhao, Hengguang; Luderer, Hilary F; Gori, Francesca; Demay, Marie B.

    2014-01-01

    Alopecia (hair loss) in vitamin D receptor (VDR)-null mice is due to absence of ligand-independent actions of the VDR that are required for initiation of postmorphogenic hair cycles. Investigations were undertaken to determine whether the VDR is required for the induction of signaling pathways that play an important role in this process. The induction of cWnt and hedgehog target genes that characterizes early anagen was found to be dramatically attenuated in VDR−/− mice, relative to wild-type...

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

    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.

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

    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...

  1. Modulation of constitutive activity and signaling bias of the ghrelin receptor by conformational constraint in the second extracellular loop

    Mokrosinski, Jacek; Frimurer, Thomas M; Sivertsen, Bjoern; Schwartz, Thue W; Holst, Birgitte

    2012-01-01

    mutational analysis of a number of potential partners in the extracellular loops and outer parts of the transmembrane segments. Systematic probing of position 204(C+6) with amino acid residues of different physicochemical properties indicated that a positively charged Lys surprisingly provided similar...... of extracellular loop 2 and that mutations or ligand binding which constrains this segment and thereby conceivably TM-V relative to TM-III movements inhibits the high constitutive signaling.......Based on a rare, natural Glu for Ala204(C+6) variant located six residues after the conserved Cys residue in extracellular loop 2 (ECL2b) associated with selective elimination of the high constitutive signaling of the ghrelin receptor, this loop was subjected to a detailed structure functional...

  2. Proliferative signaling initiated in ACTH receptors

    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.

  3. A disulfide-bridged mutant of natriuretic peptide receptor-A displays constitutive activity. Role of receptor dimerization in signal transduction.

    Labrecque, J; Mc Nicoll, N; Marquis, M; De Léan, A

    1999-04-01

    Natriuretic peptide receptor-A (NPR-A), a particulate guanylyl cyclase receptor, is composed of an extracellular domain (ECD) with a ligand binding site, a transmembrane spanning, a kinase homology domain (KHD), and a guanylyl cyclase domain. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), the natural agonists, bind and activate the receptor leading to cyclic GMP production. This receptor has been reported to be spontaneously dimeric or oligomeric. In response to agonists, the KHD-mediated guanylate cyclase repression is removed, and it is assumed that ATP binds to the KHD. Since NPR-A displays a pair of juxtamembrane cysteines separated by 8 residues, we hypothesized that the removal of one of those cysteines would leave the other unpaired and reactive, thus susceptible to form an interchain disulfide bridge and to favor the dimeric interactions. Here we show that NPR-AC423S mutant, expressed mainly as a covalent dimer, increases the affinity of pBNP for this receptor by enhancing a high affinity binding component. Dimerization primarily depends on ECD since a secreted NPR-A C423S soluble ectodomain (ECDC423S) also documents a covalent dimer. ANP binding to the unmutated ECD yields up to 80-fold affinity loss as compared with the membrane receptor. However, the ECD C423S mutation restores a high binding affinity. Furthermore, C423S mutation leads to cellular constitutive activation (20-40-fold) of basal catalytic production of cyclic GMP by the full-length mutant. In vitro particulate guanylyl cyclase assays demonstrate that NPR-AC423S displays an increased sensitivity to ATP treatment alone and that the effect of ANP + ATP joint treatment is cumulative instead of synergistic. Finally, the cellular and particulate guanylyl cyclase assays indicate that the receptor is desensitized to agonist stimulation. We conclude the following: 1) dimers are functional units of NPR-A guanylyl cyclase activation; and 2) agonists are inducing dimeric contact

  4. 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.

    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

  5. Corticosterone enhances N-methyl-D-aspartate receptor signaling to promote isolated ventral tegmental area activity in a reconstituted mesolimbic dopamine pathway.

    Berry, Jennifer N; Saunders, Meredith A; Sharrett-Field, Lynda J; Reynolds, Anna R; Bardo, Michael T; Pauly, James R; Prendergast, Mark A

    2016-01-01

    Elevations in circulating corticosteroids during periods of stress may influence activity of the mesolimbic dopamine reward pathway by increasing glutamatergic N-methyl-D-aspartate (NMDA) receptor expression and/or function in a glucocorticoid receptor-dependent manner. The current study employed organotypic co-cultures of the ventral tegmental area (VTA) and nucleus accumbens (NAcc) to examine the effects of corticosterone exposure on NMDA receptor-mediated neuronal viability. Co-cultures were pre-exposed to vehicle or corticosterone (CORT; 1 μM) for 5 days prior to a 24 h co-exposure to NMDA (200 μM). Co-cultures pre-exposed to a non-toxic concentration of corticosterone and subsequently NMDA showed significant neurotoxicity in the VTA only. This was evidenced by increases in propidium iodide uptake as well as decreases in immunoreactivity of the neuronal nuclear protein (NeuN). Co-exposure to the NMDA receptor antagonist 2-amino-7-phosphonovaleric acid (APV; 50 μM) or the glucocorticoid receptor (GR) antagonist mifepristone (10 μM) attenuated neurotoxicity. In contrast, the combination of corticosterone and NMDA did not produce any significant effects on either measure within the NAcc. Cultures of the VTA and NAcc maintained without synaptic contact showed no response to CORT or NMDA. These results demonstrate the ability to functionally reconstitute key regions of the mesolimbic reward pathway ex vivo and to reveal a GR-dependent enhancement of NMDA receptor-dependent signaling in the VTA. PMID:26631585

  6. Epileptiform activity triggers long-term plasticity of GABA(B) receptor signalling in the developing rat hippocampus.

    Tosetti, Patrizia; Ferrand, Nadine; Colin-Le Brun, Isabelle; Gaïarsa, Jean-Luc

    2005-01-01

    International audience GABA(B) receptor (GABA(B)R)-mediated presynaptic inhibition regulates neurotransmitter release from synaptic terminals. In the neonatal hippocampus, GABA(B)R activation reduces GABA release and terminates spontaneous network discharges called giant depolarizing potentials (GDPs). Blocking GABA(B)Rs transforms GDPs into longer epileptiform discharges. Thus, GABA(B)R-mediated presynaptic inhibition of GABA release (GABA auto-inhibition) controls both spontaneous networ...

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

    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

  8. Curcumin attenuates renal ischemia reperfusion injury in mice by inhibiting the activation of toll-like receptor 4 signaling

    Di-ying WU

    2012-07-01

    -L 135.8±15.2, CM-H 125.6±14.6pg/ml; P < 0.05 or P < 0.01. Conclusion Curcumin can ameliorate renal IR injury in mice, which is attributable to inhibition of the activation of Toll-like receptor 4 signaling.

  9. Effects of Somatic Mutations in the C-Terminus of Insulin-Like Growth Factor 1 Receptor on Activity and Signaling

    Barbara P. Craddock

    2012-01-01

    Full Text Available The insulin-like growth factor I receptor (IGF1R is overexpressed in several forms of human cancer, and it has emerged as an important target for anticancer drug design. Cancer genome sequencing efforts have recently identified three somatic mutations in IGF1R: A1374V, a deletion of S1278 in the C-terminal tail region of the receptor, and M1255I in the C-terminal lobe of the kinase catalytic domain. The possible effects of these mutations on IGF1R activity and biological function have not previously been tested. Here, we tested the effects of the mutations on the in vitro biochemical activity of IGF1R and on major IGF1R signaling pathways in mammalian cells. While the mutations do not affect the intrinsic tyrosine kinase activity of the receptor, we demonstrate that the basal (unstimulated levels of MAP kinase and Akt activation are increased in the mutants (relative to wild-type IGF1R. We hypothesize that the enhanced signaling potential of these mutants is due to changes in protein-protein interactions between the IGF1R C-terminus and cellular substrates or modulators.

  10. 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.

    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

  11. Central GLP-1 receptor signalling accelerates plasma clearance of triacylglycerol and glucose by activating brown adipose tissue in mice

    Kooijman, Sander; Wang, Yanan; Parlevliet, Edwin T.; Boon, Mariëtte R.; Edelschaap, David; Snaterse, Gido; Pijl, Hanno; Romijn, Johannes A.; Rensen, Patrick C. N.

    2015-01-01

    Aims/hypothesis Glucagon-like peptide 1 (GLP-1) receptor (GLP-1R) agonism, used in the treatment of type 2 diabetes, has recently been shown to increase thermogenesis via the brain. As brown adipose tissue (BAT) produces heat by burning triacylglycerol (TG) and takes up glucose for de novo lipogenesis, the aim of this study was to evaluate the potential of chronic central GLP-1R activation by exendin-4 to facilitate clearance of lipids and glucose from the circulation by activating BAT. Metho...

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

    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

  13. The aryl hydrocarbon receptor suppresses osteoblast proliferation and differentiation through the activation of the ERK signaling pathway

    Yu, Haitao; Du, Yuxuan; Zhang, Xulong; Sun, Ying; Li, Shentao; Dou, Yunpeng [Department of Immunology, School of Basic Medical Sciences, Capital Medical University, No. 10 Xitoutiao, You An Men, Beijing 100069 (China); Li, Zhanguo [Department of Rheumatology and Immunology, Clinical Immunology Center, Peking University People' s Hospital, No. 11 Xizhimen South Street, Beijing 100044 (China); Yuan, Huihui, E-mail: huihui_yuan@163.com [Department of Immunology, School of Basic Medical Sciences, Capital Medical University, No. 10 Xitoutiao, You An Men, Beijing 100069 (China); Zhao, Wenming, E-mail: zhao-wenming@163.com [Department of Immunology, School of Basic Medical Sciences, Capital Medical University, No. 10 Xitoutiao, You An Men, Beijing 100069 (China)

    2014-11-01

    Ahr activation is known to be associated with synovitis and exacerbated rheumatoid arthritis (RA), but its contributions to bone loss have not been completely elucidated. Osteoblast proliferation and differentiation are abnormal at the erosion site in RA. Here, we reported that the expression of Ahr was increased in the hind paws' bone upon collagen-induced arthritis (CIA) in mice, and the levels of Ahr were negatively correlated with bone mineral density (BMD). In addition, immunofluorescent staining showed that the high expression of Ahr was mainly localized in osteoblasts from the CIA mice compared to normal controls. Moreover, the luciferase intensity of Ahr in the nucleus increased by 12.5% in CIA osteoblasts compared to that in normal controls. In addition, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) activation of the Ahr inhibited pre-osteoblast MC3T3-E1 cellular proliferation and differentiation in a dose-dependent manner. Interestingly, the levels of alkaline phosphatase (ALP) mRNA expression in the osteoblasts of CIA mice were reduced compared to normal controls. In contrast, decreased ALP expression by activated Ahr was completely reversed after pretreatment with an Ahr inhibitor (CH-223191) in MC3T3-E1 cell lines and primary osteoblasts on day 5. Our data further showed that activation of Ahr promoted the phosphorylation of ERK after 5 days. Moreover, Ahr-dependent activation of the ERK signaling pathway decreased the levels of proliferation cells and inhibited ALP activity in MC3T3-E1 cells. These results demonstrated that the high expression of Ahr may suppress osteoblast proliferation and differentiation through activation of the ERK signaling pathway, further enabling bone erosion in CIA mice. - Highlights: • The upregulation of Ahr was localized in osteoblasts of CIA mice. • The overexpression of Ahr suppressed osteoblast development. • The Ahr activated ERK signaling pathway to exacerbate bone erosion.

  14. The aryl hydrocarbon receptor suppresses osteoblast proliferation and differentiation through the activation of the ERK signaling pathway

    Ahr activation is known to be associated with synovitis and exacerbated rheumatoid arthritis (RA), but its contributions to bone loss have not been completely elucidated. Osteoblast proliferation and differentiation are abnormal at the erosion site in RA. Here, we reported that the expression of Ahr was increased in the hind paws' bone upon collagen-induced arthritis (CIA) in mice, and the levels of Ahr were negatively correlated with bone mineral density (BMD). In addition, immunofluorescent staining showed that the high expression of Ahr was mainly localized in osteoblasts from the CIA mice compared to normal controls. Moreover, the luciferase intensity of Ahr in the nucleus increased by 12.5% in CIA osteoblasts compared to that in normal controls. In addition, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) activation of the Ahr inhibited pre-osteoblast MC3T3-E1 cellular proliferation and differentiation in a dose-dependent manner. Interestingly, the levels of alkaline phosphatase (ALP) mRNA expression in the osteoblasts of CIA mice were reduced compared to normal controls. In contrast, decreased ALP expression by activated Ahr was completely reversed after pretreatment with an Ahr inhibitor (CH-223191) in MC3T3-E1 cell lines and primary osteoblasts on day 5. Our data further showed that activation of Ahr promoted the phosphorylation of ERK after 5 days. Moreover, Ahr-dependent activation of the ERK signaling pathway decreased the levels of proliferation cells and inhibited ALP activity in MC3T3-E1 cells. These results demonstrated that the high expression of Ahr may suppress osteoblast proliferation and differentiation through activation of the ERK signaling pathway, further enabling bone erosion in CIA mice. - Highlights: • The upregulation of Ahr was localized in osteoblasts of CIA mice. • The overexpression of Ahr suppressed osteoblast development. • The Ahr activated ERK signaling pathway to exacerbate bone erosion

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

    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...

  16. Morphine Induces Desensitization of Insulin Receptor Signaling

    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...

  17. α-Melanocyte stimulating hormone attenuates dexamethasone-induced osteoblast damages through activating melanocortin receptor 4-SphK1 signaling.

    Guo, Shiguang; Xie, Yue; Fan, Jian-bo; Ji, Feng; Wang, Shouguo; Fei, Haodong

    2016-01-01

    Long-term glucocorticoid (GC) usage may cause non-traumatic femoral head osteonecrosis. Dexamethasone (Dex) is shown to exert potent cytotoxic effect to osteoblasts. Here, we investigated the potential activity of α-melanocyte stimulating hormone (α-MSH) against the process. Our data revealed that pretreatment of α-MSH significantly inhibited Dex-induced apoptosis and necrosis in both osteoblastic-like MC3T3-E1 cells and primary murine osteoblasts. Melanocortin receptor 4 (MC4R) acts as the receptor of α-MSH in mediating its actions in osteoblasts. The MC4R antagonist SHU9119, or shRNA-mediated knockdown of MC4R, almost abolished α-MSH-induced activation of downstream signalings (Akt and Erk1/2) and its pro-survival effect in osteoblasts. Further studies showed that α-MSH activated MC4R downstream sphingosine kinase 1 (SphK1) and increased cellular sphingosine-1-phosphate (S1P) content in MC3T3-E1 cells and primary murine osteoblasts, which were blocked by SHU9119 or MC4R shRNAs. SphK1 inhibition by the its inhibitor N,N-dimethylsphingosine (DMS), or SphK1 knockdown by targeted-shRNAs, largely attenuated α-MSH-mediated osteoblast protection against Dex. Together, these results suggest that α-MSH alleviates Dex-induced damages to cultured osteoblasts through activating MC4R-SphK1 signaling. PMID:26631960

  18. Molecular mechanisms of glucocorticoid receptor signaling

    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.

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

    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

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

    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

  1. The Size of Activating and Inhibitory Killer Ig-like Receptor Nanoclusters Is Controlled by the Transmembrane Sequence and Affects Signaling

    Anna Oszmiana

    2016-05-01

    Full Text Available Super-resolution microscopy has revealed that immune cell receptors are organized in nanoscale clusters at cell surfaces and immune synapses. However, mechanisms and functions for this nanoscale organization remain unclear. Here, we used super-resolution microscopy to compare the surface organization of paired killer Ig-like receptors (KIR, KIR2DL1 and KIR2DS1, on human primary natural killer cells and cell lines. Activating KIR2DS1 assembled in clusters two-fold larger than its inhibitory counterpart KIR2DL1. Site-directed mutagenesis established that the size of nanoclusters is controlled by transmembrane amino acid 233, a lysine in KIR2DS1. Super-resolution microscopy also revealed two ways in which the nanoscale clustering of KIR affects signaling. First, KIR2DS1 and DAP12 nanoclusters are juxtaposed in the resting cell state but coalesce upon receptor ligation. Second, quantitative super-resolution microscopy revealed that phosphorylation of the kinase ZAP-70 or phosphatase SHP-1 is favored in larger KIR nanoclusters. Thus, the size of KIR nanoclusters depends on the transmembrane sequence and affects downstream signaling.

  2. Activation of AMPA receptor promotes TNF-α release via the ROS-cSrc-NFκB signaling cascade in RAW264.7 macrophages

    Cheng, Xiu-Li [Department of Physiology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, Beijing (China); Ding, Fan [Office of Scientific R& D, Tsinghua University, Beijing (China); Li, Hui; Tan, Xiao-Qiu [Department of Physiology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, Beijing (China); Liu, Xiao [Department of Pathophysiology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, Beijing (China); Cao, Ji-Min, E-mail: caojimin@126.com [Department of Physiology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, Beijing (China); Gao, Xue, E-mail: longlongnose@163.com [Department of Pathophysiology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, Beijing (China)

    2015-05-29

    The relationship between glutamate signaling and inflammation has not been well defined. This study aimed to investigate the role of AMPA receptor (AMPAR) in the expression and release of tumor necrosis factor-alpha (TNF-α) from macrophages and the underlying mechanisms. A series of approaches, including confocal microscopy, immunofluorescency, flow cytometry, ELISA and Western blotting, were used to estimate the expression of AMPAR and downstream signaling molecules, TNF-α release and reactive oxygen species (ROS) generation in the macrophage-like RAW264.7 cells. The results demonstrated that AMPAR was expressed in RAW264.7 cells. AMPA significantly enhanced TNF-α release from RAW264.7 cells, and this effect was abolished by CNQX (AMPAR antagonist). AMPA also induced elevation of ROS production, phosphorylation of c-Src and activation of nuclear factor (NF)-κB in RAW264.7 cells. Blocking c-Src by PP2, scavenging ROS by glutathione (GSH) or inhibiting NF-κB activation by pyrrolidine dithiocarbamate (PDTC) decreased TNF-α production from RAW264.7 cells. We concluded that AMPA promotes TNF-α release in RAW264.7 macrophages likely through the following signaling cascade: AMPAR activation → ROS generation → c-Src phosphorylation → NF-κB activation → TNF-α elevation. The study suggests that AMPAR may participate in macrophage activation and inflammation. - Highlights: • AMPAR is expressed in RAW264.7 macrophages and is upregulated by AMPA stimulation. • Activation of AMPAR stimulates TNF-α release in macrophages through the ROS-cSrc-NFκB signaling cascade. • Macrophage AMPAR signaling may play an important role in inflammation.

  3. Activation of AMPA receptor promotes TNF-α release via the ROS-cSrc-NFκB signaling cascade in RAW264.7 macrophages

    The relationship between glutamate signaling and inflammation has not been well defined. This study aimed to investigate the role of AMPA receptor (AMPAR) in the expression and release of tumor necrosis factor-alpha (TNF-α) from macrophages and the underlying mechanisms. A series of approaches, including confocal microscopy, immunofluorescency, flow cytometry, ELISA and Western blotting, were used to estimate the expression of AMPAR and downstream signaling molecules, TNF-α release and reactive oxygen species (ROS) generation in the macrophage-like RAW264.7 cells. The results demonstrated that AMPAR was expressed in RAW264.7 cells. AMPA significantly enhanced TNF-α release from RAW264.7 cells, and this effect was abolished by CNQX (AMPAR antagonist). AMPA also induced elevation of ROS production, phosphorylation of c-Src and activation of nuclear factor (NF)-κB in RAW264.7 cells. Blocking c-Src by PP2, scavenging ROS by glutathione (GSH) or inhibiting NF-κB activation by pyrrolidine dithiocarbamate (PDTC) decreased TNF-α production from RAW264.7 cells. We concluded that AMPA promotes TNF-α release in RAW264.7 macrophages likely through the following signaling cascade: AMPAR activation → ROS generation → c-Src phosphorylation → NF-κB activation → TNF-α elevation. The study suggests that AMPAR may participate in macrophage activation and inflammation. - Highlights: • AMPAR is expressed in RAW264.7 macrophages and is upregulated by AMPA stimulation. • Activation of AMPAR stimulates TNF-α release in macrophages through the ROS-cSrc-NFκB signaling cascade. • Macrophage AMPAR signaling may play an important role in inflammation

  4. Proteolytic activation of proapoptotic kinase protein kinase Cδ by tumor necrosis factor α death receptor signaling in dopaminergic neurons during neuroinflammation

    Gordon Richard

    2012-04-01

    Full Text Available Abstract Background The mechanisms of progressive dopaminergic neuronal loss in Parkinson’s disease (PD remain poorly understood, largely due to the complex etiology and multifactorial nature of disease pathogenesis. Several lines of evidence from human studies and experimental models over the last decade have identified neuroinflammation as a potential pathophysiological mechanism contributing to disease progression. Tumor necrosis factor α (TNF has recently emerged as the primary neuroinflammatory mediator that can elicit dopaminergic cell death in PD. However, the signaling pathways by which TNF mediates dopaminergic cell death have not been completely elucidated. Methods In this study we used a dopaminergic neuronal cell model and recombinant TNF to characterize intracellular signaling pathways activated during TNF-induced dopaminergic neurotoxicity. Etanercept and neutralizing antibodies to tumor necrosis factor receptor 1 (TNFR1 were used to block TNF signaling. We confirmed the results from our mechanistic studies in primary embryonic mesencephalic cultures and in vivo using the stereotaxic lipopolysaccharide (LPS model of nigral dopaminergic degeneration. Results TNF signaling in dopaminergic neuronal cells triggered the activation of protein kinase Cδ (PKCδ, an isoform of the novel PKC family, by caspase-3 and caspase-8 dependent proteolytic cleavage. Both TNFR1 neutralizing antibodies and the soluble TNF receptor Etanercept blocked TNF-induced PKCδ proteolytic activation. Proteolytic activation of PKCδ was accompanied by translocation of the kinase to the nucleus. Notably, inhibition of PKCδ signaling by small interfering (siRNA or overexpression of a PKCδ cleavage-resistant mutant protected against TNF-induced dopaminergic neuronal cell death. Further, primary dopaminergic neurons obtained from PKCδ knockout (−/− mice were resistant to TNF toxicity. The proteolytic activation of PKCδ in the mouse substantia nigra in the

  5. Cannabinoid receptor 2 expression modulates Gβ(1)γ(2) protein interaction with the activator of G protein signalling 2/dynein light chain protein Tctex-1.

    Nagler, Marina; Palkowitsch, Lysann; Rading, Sebastian; Moepps, Barbara; Karsak, Meliha

    2016-01-01

    The activator of G protein signalling AGS2 (Tctex-1) forms protein complexes with Gβγ, and controls cell proliferation by regulating cell cycle progression. A direct interaction of Tctex-1 with various G protein-coupled receptors has been reported. Since the carboxyl terminal portion of CB2 carries a putative Tctex-1 binding motif, we investigated the potential interplay of CB2 and Tctex-1 in the absence and presence of Gβγ. The supposed interaction of cannabinoid receptor CB2 with Tctex-1 and the influence of CB2 on the formation of Tctex-1-Gβγ-complexes were studied by co- and/or immunoprecipitation experiments in transiently transfected HEK293 cells. The analysis on Tctex-1 protein was performed in the absence and presence of the ligands JWH 133, 2-AG, and AM 630, the protein biosynthesis inhibitor cycloheximide or the protein degradation blockers MG132, NH4Cl/leupeptin or bafilomycin. Our results show that CB2 neither directly nor indirectly via Gβγ interacts with Tctex-1, but competes with Tctex-1 in binding to Gβγ. The Tctex-1-Gβγ protein interaction was disrupted by CB2 receptor expression resulting in a release of Tctex-1 from the complex, and its degradation by the proteasome and partly by lysosomes. The decrease in Tctex-1 protein levels is induced by CB2 expression "dose-dependently" and is independent of stimulation by agonist or blocking by an inverse agonist treatment. The results suggest that CB2 receptor expression independent of its activation by agonists is sufficient to competitively disrupt Gβγ-Tctex-1 complexes, and to initiate Tctex-1 degradation. These findings implicate that CB2 receptor expression modifies the stability of intracellular protein complexes by a non-canonical pathway. PMID:26410677

  6. Insulin glulisine: insulin receptor signaling characteristics in vivo.

    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

  7. Gq/5-HT2c receptor signals activate a local GABAergic inhibitory feedback circuit to modulate serotonergic firing and anxiety in mice.

    Spoida, Katharina; Masseck, Olivia A; Deneris, Evan S; Herlitze, Stefan

    2014-04-29

    Serotonin 2c receptors (5-HT2c-Rs) are drug targets for certain mental disorders, including schizophrenia, depression, and anxiety. 5-HT2c-Rs are expressed throughout the brain, making it difficult to link behavioral changes to circuit specific receptor expression. Various 5-HT-Rs, including 5-HT2c-Rs, are found in the dorsal raphe nucleus (DRN); however, the function of 5-HT2c-Rs and their influence on the serotonergic signals mediating mood disorders remain unclear. To investigate the role of 5-HT2c-Rs in the DRN in mice, we developed a melanopsin-based optogenetic probe for activation of Gq signals in cellular domains, where 5-HT2c-Rs are localized. Our results demonstrate that precise temporal control of Gq signals in 5-HT2c-R domains in GABAergic neurons upstream of 5-HT neurons provides negative feedback regulation of serotonergic firing to modulate anxiety-like behavior in mice. PMID:24733892

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

    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

  9. Complexity of Receptor Tyrosine Kinase Signal Processing

    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 ...

  10. Rho-kinase signaling controls nucleocytoplasmic shuttling of class IIa Histone Deacetylase (HDAC7) and transcriptional activation of orphan nuclear receptor NR4A1

    Rho-kinase (ROCK) has been well documented to play a key role in RhoA-induced actin remodeling. ROCK activation results in myosin light chain (MLC) phosphorylation either by direct action on MLC kinase (MLCK) or by inhibition of MLC phosphatase (MLCP), modulating actin–myosin contraction. We found that inhibition of the ROCK pathway in induced pluripotent stem cells, leads to nuclear export of HDAC7 and transcriptional activation of the orphan nuclear receptor NR4A1 while in cells with constitutive ROCK hyperactivity due to loss of function of the RhoGTPase activating protein Oligophrenin-1 (OPHN1), the orphan nuclear receptor NR4A1 is downregulated. Our study identify a new target of ROCK signaling via myosin phosphatase subunit (MYPT1) and Histone Deacetylase (HDAC7) at the nuclear level and provide new insights in the cellular functions of ROCK. - Highlights: • ROCK regulates nucleocytoplasmic shuttling of HDAC7 via phosphorylation of MYPT1. • Nuclear export of HDAC7 and upregulation of NR4A1 occurs with low ROCK activity. • High levels of ROCK activity due to OPHN1 loss of function downregulate NR4A1

  11. Rho-kinase signaling controls nucleocytoplasmic shuttling of class IIa Histone Deacetylase (HDAC7) and transcriptional activation of orphan nuclear receptor NR4A1

    Compagnucci, Claudia; Barresi, Sabina [Unit of Molecular Medicine for Neuromuscular and Neurodegenerative Disorders, Department of Neurosciences, Bambino Gesù Children’s Hospital, IRCCS, Rome (Italy); Petrini, Stefania [Research Laboratories, Confocal Microscopy Core Facility, Bambino Gesù Children’s Hospital, IRCCS, Rome (Italy); Bertini, Enrico [Unit of Molecular Medicine for Neuromuscular and Neurodegenerative Disorders, Department of Neurosciences, Bambino Gesù Children’s Hospital, IRCCS, Rome (Italy); Zanni, Ginevra, E-mail: ginevra.zanni@opbg.net [Unit of Molecular Medicine for Neuromuscular and Neurodegenerative Disorders, Department of Neurosciences, Bambino Gesù Children’s Hospital, IRCCS, Rome (Italy)

    2015-04-03

    Rho-kinase (ROCK) has been well documented to play a key role in RhoA-induced actin remodeling. ROCK activation results in myosin light chain (MLC) phosphorylation either by direct action on MLC kinase (MLCK) or by inhibition of MLC phosphatase (MLCP), modulating actin–myosin contraction. We found that inhibition of the ROCK pathway in induced pluripotent stem cells, leads to nuclear export of HDAC7 and transcriptional activation of the orphan nuclear receptor NR4A1 while in cells with constitutive ROCK hyperactivity due to loss of function of the RhoGTPase activating protein Oligophrenin-1 (OPHN1), the orphan nuclear receptor NR4A1 is downregulated. Our study identify a new target of ROCK signaling via myosin phosphatase subunit (MYPT1) and Histone Deacetylase (HDAC7) at the nuclear level and provide new insights in the cellular functions of ROCK. - Highlights: • ROCK regulates nucleocytoplasmic shuttling of HDAC7 via phosphorylation of MYPT1. • Nuclear export of HDAC7 and upregulation of NR4A1 occurs with low ROCK activity. • High levels of ROCK activity due to OPHN1 loss of function downregulate NR4A1.

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

    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...

  13. Interdependent epidermal growth factor receptor signalling and trafficking.

    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

  14. Optogenetic activation of intracellular adenosine A2A receptor signaling in the hippocampus is sufficient to trigger CREB phosphorylation and impair memory.

    Li, P; Rial, D; Canas, P M; Yoo, J-H; Li, W; Zhou, X; Wang, Y; van Westen, G J P; Payen, M-P; Augusto, E; Gonçalves, N; Tomé, A R; Li, Z; Wu, Z; Hou, X; Zhou, Y; IJzerman, A P; PIJzerman, Ad; Boyden, E S; Cunha, R A; Qu, J; Chen, J-F

    2015-11-01

    Human and animal studies have converged to suggest that caffeine consumption prevents memory deficits in aging and Alzheimer's disease through the antagonism of adenosine A2A receptors (A2ARs). To test if A2AR activation in the hippocampus is actually sufficient to impair memory function and to begin elucidating the intracellular pathways operated by A2AR, we have developed a chimeric rhodopsin-A2AR protein (optoA2AR), which retains the extracellular and transmembrane domains of rhodopsin (conferring light responsiveness and eliminating adenosine-binding pockets) fused to the intracellular loop of A2AR to confer specific A2AR signaling. The specificity of the optoA2AR signaling was confirmed by light-induced selective enhancement of cAMP and phospho-mitogen-activated protein kinase (p-MAPK) (but not cGMP) levels in human embryonic kidney 293 (HEK293) cells, which was abolished by a point mutation at the C terminal of A2AR. Supporting its physiological relevance, optoA2AR activation and the A2AR agonist CGS21680 produced similar activation of cAMP and p-MAPK signaling in HEK293 cells, of p-MAPK in the nucleus accumbens and of c-Fos/phosphorylated-CREB (p-CREB) in the hippocampus, and similarly enhanced long-term potentiation in the hippocampus. Remarkably, optoA2AR activation triggered a preferential p-CREB signaling in the hippocampus and impaired spatial memory performance, while optoA2AR activation in the nucleus accumbens triggered MAPK signaling and modulated locomotor activity. This shows that the recruitment of intracellular A2AR signaling in the hippocampus is sufficient to trigger memory dysfunction. Furthermore, the demonstration that the biased A2AR signaling and functions depend on intracellular A2AR loops prompts the possibility of targeting the intracellular A2AR-interacting partners to selectively control different neuropsychiatric behaviors. PMID:25687775

  15. IL-1RI (interleukin-1 receptor type I signalling is essential for host defence and hemichannel activity during acute central nervous system bacterial infection

    Tammy Kielian

    2012-04-01

    Full Text Available Staphylococcus aureus is a common aetiological agent of bacterial brain abscesses. We have previously established that a considerable IL-1 (interleukin-1 response is elicited immediately following S. aureus infection, where the cytokine can exert pleiotropic effects on glial activation and blood–brain barrier permeability. To assess the combined actions of IL-1α and IL-1β during CNS (central nervous system infection, host defence responses were evaluated in IL-1RI (IL-1 receptor type I KO (knockout animals. IL-1RI KO mice were exquisitely sensitive to intracerebral S. aureus infection, as demonstrated by enhanced mortality rates and bacterial burdens within the first 24 h following pathogen exposure compared with WT (wild-type animals. Loss of IL-1RI signalling also dampened the expression of select cytokines and chemokines, concomitant with significant reductions in neutrophil and macrophage infiltrates into the brain. In addition, the opening of astrocyte hemichannels during acute infection was shown to be dependent on IL-1RI activity. Collectively, these results demonstrate that IL-1RI signalling plays a pivotal role in the genesis of immune responses during the acute stage of brain abscess development through S. aureus containment, inflammatory mediator production, peripheral immune cell recruitment, and regulation of astrocyte hemichannel activity. Taken in the context of previous studies with MyD88 (myeloid differentiation primary response gene 88 and TLR2 (Toll-like receptor 2 KO animals, the current report advances our understanding of MyD88-dependent cascades and implicates IL-1RI signalling as a major antimicrobial effector pathway during acute brain-abscess formation.

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

    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

  17. Molecular mechanisms of alternative estrogen receptor signaling

    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...

  18. SOCS proteins in regulation of receptor tyrosine kinase signaling

    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...

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

    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

  20. Electrospun Gelatin/β-TCP Composite Nanofibers Enhance Osteogenic Differentiation of BMSCs and In Vivo Bone Formation by Activating Ca2+-Sensing Receptor Signaling

    Xuehui Zhang

    2015-01-01

    Full Text Available Calcium phosphate- (CaP- based composite scaffolds have been used extensively for the bone regeneration in bone tissue engineering. Previously, we developed a biomimetic composite nanofibrous membrane of gelatin/β-tricalcium phosphate (TCP and confirmed their biological activity in vitro and bone regeneration in vivo. However, how these composite nanofibers promote the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs is unknown. Here, gelatin/β-TCP composite nanofibers were fabricated by incorporating 20 wt% β-TCP nanoparticles into electrospun gelatin nanofibers. Electron microscopy showed that the composite β-TCP nanofibers had a nonwoven structure with a porous network and a rough surface. Spectral analyses confirmed the presence and chemical stability of the β-TCP and gelatin components. Compared with pure gelatin nanofibers, gelatin/β-TCP composite nanofibers caused increased cell attachment, proliferation, alkaline phosphatase activity, and osteogenic gene expression in rat BMSCs. Interestingly, the expression level of the calcium-sensing receptor (CaSR was significantly higher on the composite nanofibrous scaffolds than on pure gelatin. For rat calvarial critical sized defects, more extensive osteogenesis and neovascularization occurred in the composite scaffolds group compared with the gelatin group. Thus, gelatin/β-TCP composite scaffolds promote osteogenic differentiation of BMSCs in vitro and bone regeneration in vivo by activating Ca2+-sensing receptor signaling.

  1. NOX3 NADPH oxidase couples transient receptor potential vanilloid 1 to signal transducer and activator of transcription 1-mediated inflammation and hearing loss.

    Mukherjea, Debashree; Jajoo, Sarvesh; Sheehan, Kelly; Kaur, Tejbeer; Sheth, Sandeep; Bunch, Jennifer; Perro, Christopher; Rybak, Leonard P; Ramkumar, Vickram

    2011-03-15

    Transient receptor potential vanilloid 1 (TRPV1) is implicated in cisplatin ototoxicity. Activation of this channel by cisplatin increases reactive oxygen species generation, which contribute to loss of outer hair cells in the cochlea. Knockdown of TRPV1 by short interfering RNA protected against cisplatin ototoxicity. In this study, we examined the mechanism underlying TRPV1-mediated ototoxicity using cultured organ of Corti transformed cells (UB/OC-1) and rats. Trans-tympanic injections of capsaicin produced transient hearing loss within 24 h, which recovered by 72 h. In UB/OC-1 cells, capsaicin increased NOX3 NADPH oxidase activity and activation of signal transducer and activator of transcription 1 (STAT1). Intratympanic administration of capsaicin transiently increased STAT1 activity and expression of downstream proinflammatory molecules. Capsaicin produced a transient increase in CD14-positive inflammatory cells into the cochlea, which mimicked the temporal course of STAT1 activation but did not alter the expression of apoptotic genes or damage to outer hair cells. In addition, trans-tympanic administration of STAT1 short interfering RNA protected against capsaicin-induced hearing loss. These data suggest that activation of TRPV1 mediates temporary hearing loss by initiating an inflammatory process in the cochlea via activation of NOX3 and STAT1. Thus, these proteins represent reasonable targets for ameliorating hearing loss. PMID:20712533

  2. Signal Transducer and Activator of Transcription (Stat)-Induced Stat Inhibitor 1 (Ssi-1)/Suppressor of Cytokine Signaling 1 (Socs1) Inhibits Insulin Signal Transduction Pathway through Modulating Insulin Receptor Substrate 1 (Irs-1) Phosphorylation

    Kawazoe, Yoshinori; Naka, Tetsuji; Fujimoto, Minoru; Kohzaki, Hidetsugu; Morita, Yoshiaki; Narazaki, Masashi; Okumura, Kohichi; Saitoh, Hiroshi; Nakagawa, Reiko; Uchiyama, Yasuo; Akira, Shizuo; Kishimoto, Tadamitsu

    2001-01-01

    Signal transducer and activator of transcription (STAT)-induced STAT inhibitor 1 (SSI-1) is known to function as a negative feedback regulator of cytokine signaling, but it is unclear whether it is involved in other biological events. Here, we show that SSI-1 participates and plays an important role in the insulin signal transduction pathway. SSI-1–deficient mice showed a significantly low level of blood sugar. While the forced expression of SSI-1 reduced the phosphorylation level of insulin ...

  3. Nuclear epidermal growth factor receptor (EGFR) interacts with signal transducer and activator of transcription 5 (STAT5) in activating Aurora-A gene expression

    Hung, Liang-Yi; Tseng, Joseph T.; Lee, Yi-Chao; Xia, Weiya; Wang, Ying-Nai; Wu, Min-Li; Chuang, Yu-Hsuan; Lai, Chein-Hsien; Chang, Wen-Chang

    2008-01-01

    Loss of the maintenance of genetic material is a critical step leading to tumorigenesis. It was reported that overexpression of Aurora-A and the constitutive activation of the epidermal growth factor (EGF) receptor (EGFR) are implicated in chromosome instability. In this study, we examined that when cells treated with EGF result in centrosome amplification and microtubule disorder, which are critical for genetic instability. Interestingly, the expression of Aurora-A was also increased by EGF ...

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

    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

  5. Evolution of retinoic acid receptors and retinoic acid signaling.

    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. Relaxin Family Peptide Receptors – former orphans reunite with their parent ligands to activate multiple signalling pathways

    Halls, M L; van der Westhuizen, E T; Bathgate, R A D; Summers, R. J.

    2007-01-01

    The relaxin family peptides, although structurally closely related to insulin, act on a group of four G protein-coupled receptors now known as Relaxin Family Peptide (RXFP) Receptors. The leucine-rich repeat containing RXFP1 and RXFP2 and the small peptide-like RXFP3 and RXFP4 are the physiological targets for relaxin, insulin-like (INSL) peptide 3, relaxin-3 and INSL5, respectively. RXFP1 and RXFP2 have at least two binding sites – a high-affinity site in the leucine-rich repeat region of th...

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

    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.

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

    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

  9. Diverse FGF receptor signaling controls astrocyte specification and proliferation

    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.

  10. Diverse FGF receptor signaling controls astrocyte specification and proliferation

    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.

  11. Activation of the SPHK/S1P signalling pathway is coupled to muscarinic receptor-dependent regulation of peripheral airways

    Kummer Wolfgang

    2005-05-01

    Full Text Available Abstract Background In peripheral airways, acetylcholine induces contraction via activation of muscarinic M2-and M3-receptor subtypes (M2R and M3R. Cholinergic hypersensitivity is associated with chronic obstructive pulmonary disease and asthma, and therefore the identification of muscarinic signaling pathways are of great therapeutic interest. A pathway that has been shown to be activated via MR and to increase [Ca2+]i includes the activation of sphingosine kinases (SPHK and the generation of the bioactive sphingolipid sphingosine 1-phosphate (S1P. Whether the SPHK/S1P signaling pathway is integrated in the muscarinic control of peripheral airways is not known. Methods To address this issue, we studied precision cut lung slices derived from FVB and M2R-KO and M3R-KO mice. Results In peripheral airways of FVB, wild-type, and MR-deficient mice, SPHK1 was mainly localized to smooth muscle. Muscarine induced a constriction in all investigated mouse strains which was reduced by inhibition of SPHK using D, L-threo-dihydrosphingosine (DHS and N, N-dimethyl-sphingosine (DMS but not by N-acetylsphingosine (N-AcS, a structurally related agent that does not affect SPHK function. The initial phase of constriction was nearly absent in peripheral airways of M3R-KO mice when SPHK was inhibited by DHS and DMS but was unaffected in M2R-KO mice. Quantitative RT-PCR revealed that the disruption of the M2R and M3R genes had no significant effect on the expression levels of the SPHK1-isoform in peripheral airways. Conclusion These results demonstrate that the SPHK/S1P signaling pathway contributes to cholinergic constriction of murine peripheral airways. In addition, our data strongly suggest that SPHK is activated via the M2R. Given the important role of muscarinic mechanisms in pulmonary disease, these findings should be of considerable therapeutic relevance.

  12. The poly-γ-d-glutamic acid capsule surrogate of the Bacillus anthracis capsule induces nitric oxide production via the platelet activating factor receptor signaling pathway.

    Lee, Hae-Ri; Jeon, Jun Ho; Park, Ok-Kyu; Chun, Jeong-Hoon; Park, Jungchan; Rhie, Gi-Eun

    2015-12-01

    The poly-γ-d-glutamic acid (PGA) capsule, a major virulence factor of Bacillus anthracis, confers protection of the bacillus from phagocytosis and allows its unimpeded growth in the host. PGA capsules released from B. anthracis are associated with lethal toxin in the blood of experimentally infected animals and enhance the cytotoxic effect of lethal toxin on macrophages. In addition, PGA capsule itself activates macrophages and dendritic cells to produce proinflammatory cytokine such as IL-1β, indicating multiple roles of PGA capsule in anthrax pathogenesis. Here we report that PGA capsule of Bacillus licheniformis, a surrogate of B. anthracis capsule, induces production of nitric oxide (NO) in RAW264.7 cells and bone marrow-derived macrophages. NO production was induced by PGA in a dose-dependent manner and was markedly reduced by inhibitors of inducible NO synthase (iNOS), suggesting iNOS-dependent production of NO. Induction of NO production by PGA was not observed in macrophages from TLR2-deficient mice and was also substantially inhibited in RAW264.7 cells by pretreatment of TLR2 blocking antibody. Subsequently, the downstream signaling events such as ERK, JNK and p38 of MAPK pathways as well as NF-κB activation were required for PGA-induced NO production. In addition, the induced NO production was significantly suppressed by treatment with antagonists of platelet activating factor receptor (PAFR) or PAFR siRNA, and mediated through PAFR/Jak2/STAT-1 signaling pathway. These findings suggest that PGA capsule induces NO production in macrophages by triggering both TLR2 and PAFR signaling pathways which lead to activation of NF-kB and STAT-1, respectively. PMID:26350415

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

    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.

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

    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

  15. Cocaine-induced changes in NMDA receptor signaling

    Ortinski, Pavel I.

    2014-01-01

    Addictive states are often thought to rely on lasting modification of signaling at relevant synapses. A long-standing theory posits that activity at N-methyl-D-aspartate receptors (NMDARs) is a critical component of long-term synaptic plasticity in many brain areas. Indeed, NMDAR signaling has been found to play a role in the etiology of addictive states, in particular following cocaine exposure. However, no consensus is apparent with respect to the specific effects of cocaine exposure on NMD...

  16. Follicle-stimulating hormone (FSH activates extracellular signal-regulated kinase phosphorylation independently of beta-arrestin- and dynamin-mediated FSH receptor internalization

    Crepieux Pascale

    2006-06-01

    Full Text Available Abstract Background The follicle-stimulating hormone receptor (FSH-R is a seven transmembrane spanning receptor (7TMR which plays a crucial role in male and female reproduction. Upon FSH stimulation, the FSH-R activates the extracellular signal-regulated kinases (ERK. However, the mechanisms whereby the agonist-stimulated FSH-R activates ERK are poorly understood. In order to activate ERK, some 7 TMRs require beta-arrestin-and dynamin-dependent internalization to occur, whereas some others do not. In the present study, we examined the ability of the FSH-activated FSH-R to induce ERK phosphorylation, in conditions where its beta-arrestin- and dynamin-mediated internalization was impaired. Methods Human embryonic kidney (HEK 293 cells were transiently transfected with the rat FSH-R. Internalization of the FSH-R was manipulated by co-expression of either a beta-arrestin (319–418 dominant negative peptide, either an inactive dynamin K44A mutant or of wild-type beta-arrestin 1 or 2. The outcomes on the FSH-R internalization were assayed by measuring 125I-FSH binding at the cell surface when compared to internalized 125I-FSH binding. The resulting ERK phosphorylation level was visualized by Western blot analysis. Results In HEK 293 cells, FSH stimulated ERK phosphorylation in a dose-dependent manner. Co-transfection of the beta- arrestin (319–418 construct, or of the dynamin K44A mutant reduced FSH-R internalization in response to FSH, without affecting ERK phosphorylation. Likewise, overexpression of wild-type beta-arrestin 1 or 2 significantly increased the FSH-R internalization level in response to FSH, without altering FSH-induced ERK phosphorylation. Conclusion From these results, we conclude that the FSH-R does not require beta-arrestin- nor dynamin-mediated internalization to initiate ERK phosphorylation in response to FSH.

  17. P2X and P2Y receptor signaling in red blood cells

    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

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

    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...

  19. Fatty acids and hypolipidemic drugs regulate peroxisome proliferator-activated receptors α- and γ-mediated gene expression via liver fatty acid binding protein: A signaling path to the nucleus

    Wolfrum, Christian; Borrmann, Carola M.; Börchers, Torsten; Spener, Friedrich

    2001-01-01

    Peroxisome proliferator-activated receptor α (PPARα) is a key regulator of lipid homeostasis in hepatocytes and target for fatty acids and hypolipidemic drugs. How these signaling molecules reach the nuclear receptor is not known; however, similarities in ligand specificity suggest the liver fatty acid binding protein (L-FABP) as a possible candidate. In localization studies using laser-scanning microscopy, we show that L-FABP and PPARα colocalize in the nucleus of...

  20. ERK activation causes epilepsy by stimulating NMDA receptor activity

    Nateri, Abdolrahman S.; Raivich, Gennadij; Gebhardt, Christine; Da Costa, Clive; Naumann, Heike; Vreugdenhil, Martin; Makwana, Milan; Brandner, Sebastian; Adams, Ralf H.; Jefferys, John G. R.; Kann, Oliver; Behrens, Axel

    2007-01-01

    The ERK MAPK signalling pathway is a highly conserved kinase cascade linking transmembrane receptors to downstream effector mechanisms. To investigate the function of ERK in neurons, a constitutively active form of MEK1 (caMEK1) was conditionally expressed in the murine brain, which resulted in ERK activation and caused spontaneous epileptic seizures. ERK activation stimulated phosphorylation of eukaryotic translation initiation factor 4E (eIF4E) and augmented NMDA receptor 2B (NR2B) protein ...

  1. p21-activated kinase group II small compound inhibitor GNE-2861 perturbs estrogen receptor alpha signaling and restores tamoxifen-sensitivity in breast cancer cells

    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 ...

  2. RELAXIN ACTIVATES PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR GAMMA

    Singh, Sudhir; Bennett, Robert G

    2009-01-01

    Relaxin is a polypeptide hormone that triggers multiple signaling pathways through its receptor RXFP1. Many of relaxin’s functions, including vascular and antifibrotic effects, are similar to those induced by activation of PPARγ. In this study, we tested the hypothesis that relaxin signaling through RXFP1 would activate PPARγ activity. In cells overexpressing RXFP1 (HEK-RXFP1), relaxin increased transcriptional activity through a PPAR response element (PPRE) in a concentration-dependent manne...

  3. NMDA receptor activation by spontaneous glutamatergic neurotransmission.

    Espinosa, Felipe; Kavalali, Ege T

    2009-05-01

    Under physiological conditions N-methyl-D-aspartate (NMDA) receptor activation requires coincidence of presynaptic glutamate release and postsynaptic depolarization due to the voltage-dependent block of these receptors by extracellular Mg(2+). Therefore spontaneous neurotransmission in the absence of action potential firing is not expected to lead to significant NMDA receptor activation. Here we tested this assumption in layer IV neurons in neocortex at their resting membrane potential (approximately -67 mV). In long-duration stable recordings, we averaged a large number of miniature excitatory postsynaptic currents (mEPSCs, >100) before or after application of dl-2 amino 5-phosphonovaleric acid, a specific blocker of NMDA receptors. The difference between the two mEPSC waveforms showed that the NMDA current component comprises approximately 20% of the charge transfer during an average mEPSC detected at rest. Importantly, the contribution of the NMDA component was markedly enhanced at membrane potentials expected for the depolarized up states (approximately -50 mV) that cortical neurons show during slow oscillations in vivo. In addition, partial block of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor component of the mEPSCs did not cause a significant reduction in the NMDA component, indicating that potential AMPA receptor-driven local depolarizations did not drive NMDA receptor activity at rest. Collectively these results indicate that NMDA receptors significantly contribute to signaling at rest in the absence of dendritic depolarizations or concomitant AMPA receptor activity. PMID:19261712

  4. Corazonin receptor signaling in ecdysis initiation.

    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

  5. Diphenylarsinic acid, a chemical warfare-related neurotoxicant, promotes liver carcinogenesis via activation of aryl hydrocarbon receptor signaling and consequent induction of oxidative DAN damage in rats

    Wei, Min; Yamada, Takanori; Yamano, Shotaro; Kato, Minoru; Kakehashi, Anna; Fujioka, Masaki; Tago, Yoshiyuki; Kitano, Mistuaki; Wanibuchi, Hideki, E-mail: wani@med.osaka-cu.ac.jp

    2013-11-15

    Diphenylarsinic acid (DPAA), a chemical warfare-related neurotoxic organic arsenical, is present in the groundwater and soil in some regions of Japan due to illegal dumping after World War II. Inorganic arsenic is carcinogenic in humans and its organic arsenic metabolites are carcinogenic in animal studies, raising serious concerns about the carcinogenicity of DPAA. However, the carcinogenic potential of DPAA has not yet been evaluated. In the present study we found that DPAA significantly enhanced the development of diethylnitrosamine-induced preneoplastic lesions in the liver in a medium-term rat liver carcinogenesis assay. Evaluation of the expression of cytochrome P450 (CYP) enzymes in the liver revealed that DPAA induced the expression of CYP1B1, but not any other CYP1, CYP2, or CYP3 enzymes, suggesting that CYP1B1 might be the enzyme responsible for the metabolic activation of DPAA. We also found increased oxidative DNA damage, possibly due to elevated CYP1B1 expression. Induction of CYP1B1 has generally been linked with the activation of AhR, and we found that DPAA activates the aryl hydrocarbon receptor (AhR). Importantly, the promotion effect of DPAA was observed only at a dose that activated the AhR, suggesting that activation of AhR and consequent induction of AhR target genes and oxidative DNA damage plays a vital role in the promotion effects of DPAA. The present study provides, for the first time, evidence regarding the carcinogenicity of DPAA and indicates the necessity of comprehensive evaluation of its carcinogenic potential using long-term carcinogenicity studies. - Highlights: • DPAA, an environmental neurotoxicant, promotes liver carcinogenesis in rats. • DPAA is an activator of AhR signaling pathway. • DPAA promoted oxidative DNA damage in rat livers. • AhR target gene CYP 1B1 might be involved in the metabolism of DPAA.

  6. Diphenylarsinic acid, a chemical warfare-related neurotoxicant, promotes liver carcinogenesis via activation of aryl hydrocarbon receptor signaling and consequent induction of oxidative DAN damage in rats

    Diphenylarsinic acid (DPAA), a chemical warfare-related neurotoxic organic arsenical, is present in the groundwater and soil in some regions of Japan due to illegal dumping after World War II. Inorganic arsenic is carcinogenic in humans and its organic arsenic metabolites are carcinogenic in animal studies, raising serious concerns about the carcinogenicity of DPAA. However, the carcinogenic potential of DPAA has not yet been evaluated. In the present study we found that DPAA significantly enhanced the development of diethylnitrosamine-induced preneoplastic lesions in the liver in a medium-term rat liver carcinogenesis assay. Evaluation of the expression of cytochrome P450 (CYP) enzymes in the liver revealed that DPAA induced the expression of CYP1B1, but not any other CYP1, CYP2, or CYP3 enzymes, suggesting that CYP1B1 might be the enzyme responsible for the metabolic activation of DPAA. We also found increased oxidative DNA damage, possibly due to elevated CYP1B1 expression. Induction of CYP1B1 has generally been linked with the activation of AhR, and we found that DPAA activates the aryl hydrocarbon receptor (AhR). Importantly, the promotion effect of DPAA was observed only at a dose that activated the AhR, suggesting that activation of AhR and consequent induction of AhR target genes and oxidative DNA damage plays a vital role in the promotion effects of DPAA. The present study provides, for the first time, evidence regarding the carcinogenicity of DPAA and indicates the necessity of comprehensive evaluation of its carcinogenic potential using long-term carcinogenicity studies. - Highlights: • DPAA, an environmental neurotoxicant, promotes liver carcinogenesis in rats. • DPAA is an activator of AhR signaling pathway. • DPAA promoted oxidative DNA damage in rat livers. • AhR target gene CYP 1B1 might be involved in the metabolism of DPAA

  7. Androgen receptor signaling and mutations in prostate cancer

    Koochekpour, Shahriar

    2010-01-01

    Normal and neoplastic growth of the prostate gland are dependent on androgen receptor (AR) expression and function. Androgenic activation of the AR, in association with its coregulatory factors, is the classical pathway that leads to transcriptional activity of AR target genes. Alternatively, cytoplasmic signaling crosstalk of AR by growth factors, neurotrophic peptides, cytokines or nonandrogenic hormones may have important roles in prostate carcinogenesis and in metastatic or androgen-indep...

  8. Transport of receptors, receptor signaling complexes and ion channels via neuropeptide-secretory vesicles

    Bo Zhao; Hai-Bo Wang; Ying-Jin Lu; Jian-Wen Hu; Lan Bao; Xu Zhang

    2011-01-01

    Stimulus-induced exocytosis of large dense-core vesicles(LDCVs)leads to discharge of neuropeptides and fusion of LDCV membranes with the plasma membrane. However, the contribution of LDCVs to the properties of the neuronal membrane remains largely unclear. The present study found that LDCVs were associated with multiple receptors, channels and signaling molecules, suggesting that neuronal sensitivity is modulated by an LDCV-mediated mechanism. Liquid chromatography-mass spectrometry combined with immunoblotting of subcellular fractions identified 298 proteins in LDCV membranes purified from the dorsal spinal cord, including Gprotein-coupled receptors, Gproteins and other signaling molecules, ion channels and trafficking-related proteins. Morphological assays showed that δ-opioid receptor 1(DORI), β2 adrenergic receptor(AR), Gα12,voltage-gated calcium channel a2δ1subunit and P2X purinoceptor 2 were localized in substance P(SP)-positive LDCVs in small-diameter dorsal root ganglion neurons, whereas β1 AR, Wnt receptor frizzled 8 and dishevelled 1 were present in SP-negative LDCVs.Furthermore, DOR1/α12/Gβ1γ5/phospholipase C β2 complexes were associated with LDCVs. Blockade of the DOR1/Gαi2 interaction largely abolished the LDCV localization of Gαi2 and impaired stimulation-induced surface expression of Gαi2. Thus, LDCVs serve as carriers of receptors, ion channels and preassembled receptor signaling complexes, enabling a rapid, activity-dependent modulation of neuronal sensitivity.

  9. Interfering with interferon receptor sorting and trafficking: impact on signaling.

    Claudinon, Julie; Monier, Marie-Noëlle; Lamaze, Christophe

    2007-01-01

    Interferons (IFNs) and their receptors (IFN-Rs) play fundamental roles in a multitude of biological functions. Many articles and reviews emphasize that the JAK/STAT machinery is obligatory for relay of the information transmitted by IFNs after binding to their cognate receptors at the plasma membrane. In contrast, very few studies have addressed the endocytosis and the intracellular trafficking of IFN-Rs, the immediate step following IFN binding. However, recent findings have shed light on the importance of IFN-R sorting and trafficking in the control of IFN signaling. Thus, IFN-Rs can be included in the growing family of signaling receptors for which regulation of biological activity critically involves endocytosis and trafficking. PMID:17493737

  10. INHIBITION OF PROTEIN TYROSINE PHOSPHATASE ACTIVITY MEDIATES EPIDERMAL GROWTH FACTOR RECEPTOR SIGNALING IN HUMAN AIRWAY EPITHELIAL CELLS

    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...

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

    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

  12. Distinct Phosphorylation Clusters Determine the Signaling Outcome of Free Fatty Acid Receptor 4/G Protein-Coupled Receptor 120

    Prihandoko, Rudi; Alvarez-Curto, Elisa; Hudson, Brian D; Butcher, Adrian J; Ulven, Trond; Miller, Ashley M; Tobin, Andrew B; Milligan, Graeme

    2016-01-01

    phosphoacceptor sites to alanine completely prevented phosphorylation of mFFA4 but did not limit receptor coupling to extracellular signal regulated protein kinase 1 and 2 (ERK1/2) activation. Rather, an inhibitor of Gq/11proteins completely prevented receptor signaling to ERK1/2. By contrast, the recruitment of......It is established that long-chain free fatty acids includingω-3 fatty acids mediate an array of biologic responses through members of the free fatty acid (FFA) receptor family, which includes FFA4. However, the signaling mechanisms and modes of regulation of this receptor class remain unclear. Here...... arrestin 3, receptor internalization, and activation of Akt were regulated by mFFA4 phosphorylation. The analysis of mFFA4 phosphorylation-dependent signaling was extended further by selective mutations of the phosphoacceptor sites. Mutations within cluster 2 did not affect agonist activation of Akt but...

  13. Prunetin signals via G-protein-coupled receptor, GPR30(GPER1): Stimulation of adenylyl cyclase and cAMP-mediated activation of MAPK signaling induces Runx2 expression in osteoblasts to promote bone regeneration.

    Khan, Kainat; Pal, Subhashis; Yadav, Manisha; Maurya, Rakesh; Trivedi, Arun Kumar; Sanyal, Sabyasachi; Chattopadhyay, Naibedya

    2015-12-01

    Prunetin is found in red clover and fruit of Prunus avium (red cherry). The effect of prunetin on osteoblast function, its mode of action and bone regeneration in vivo were investigated. Cultures of primary osteoblasts, osteoblastic cell line and HEK293T cells were used for various in vitro studies. Adult female rats received drill-hole injury at the femur diaphysis to assess the bone regenerative effect of prunetin. Prunetin at 10nM significantly (a) increased proliferation and differentiation of primary cultures of osteoblasts harvested from rats and (b) promoted formation of mineralized nodules by bone marrow stromal/osteoprogenitor cells. At this concentration, prunetin did not activate any of the two nuclear estrogen receptors (α and β). However, prunetin triggered signaling via a G-protein-coupled receptor, GPR30/GPER1, and enhanced cAMP levels in osteoblasts. G15, a selective GPR30 antagonist, abolished prunetin-induced increases in osteoblast proliferation, differentiation and intracellular cAMP. In osteoblasts, prunetin up-regulated runt-related transcription factor 2 (Runx2) protein through cAMP-dependent Erk/MAP kinase activation that ultimately resulted in the up-regulation of GPR30. Administration of prunetin at 0.25mg/kg given to rats stimulated bone regeneration at the site of drill hole and up-regulated Runx2 expression in the fractured callus and the effect was comparable to human parathyroid hormone, the only clinically used osteogenic therapy. We conclude that prunetin promotes osteoinduction in vivo and the mechanism is defined by signaling through GPR30 resulting in the up-regulation of the key osteogenic gene Runx2 that in turn up-regulates GPR30. PMID:26345541

  14. Characterization of Thyrotropin Receptor Antibody-Induced Signaling Cascades

    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...

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

    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...

  16. Topical treatment with Xiaozheng Zhitong Paste alleviates bone cancer pain by inhibiting proteinase-activated receptor 2 signaling pathway.

    Bao, Yanju; Wang, Gaimei; Gao, Yebo; Du, Maobo; Yang, Liping; Kong, Xiangying; Zheng, Honggang; Hou, Wei; Hua, Baojin

    2015-09-01

    Herbal analgesic Xiaozheng Zhitong Paste (XZP) and related modifications are often used in traditional Chinese medicine to manage cancer pain. However, its underlying mechanism remains unknown. To investigate the effects and mechanism of XZP on bone cancer pain in a rat model of breast cancer-induced bone pain, a bone cancer pain model was established by inoculating Walker 256 cells into Wistar rats. Bone cancer-bearing rats were topically treated with different doses of XZP or injected with 5 mg/kg of osteoprotegerin (OPG) as positive control. Bone destruction, bone mineral content (BMC) and bone mineral density (BMD) were analyzed by radiology. Paw withdrawal threshold (PWT) and paw withdrawal latency (PWL) were examined to determine pain levels. Trypsin, TNF-α and IL-1β serum levels were determined using enzyme-linked immunosorbent assay (ELISA). Central sensitization markers such as c-Fos, GFAP, IBA1 and CGRP, as well as proteinase-activated receptor 2 (PAR2) signaling pathway mediators such as PAR2, PKC-γ, PKA and TRPV1, were determined by quantitative RT-PCR and western blotting assay. XZP treatment significantly mitigated bone cancer-related nociceptive behavior, bone damage, BMC and BMD; and decreased radiological scores in rats. XZP treatment significantly inhibited IBA1, GFAP, c-Fos and CGRP expressions in the spinal cord; and significantly mitigated trypsin, TNF-α and IL-1β serum levels. Furthermore, PAR2, PKC-γ, PKA and TRPV1 relative mRNA levels and protein expression in bone lesions were significantly reduced in rats treated with XZP. XZP significantly alleviates breast cancer-induced bone pain by inhibiting the PAR2 signaling pathway. PMID:26133236

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

    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

  18. Assembly of Oligomeric Death Domain Complexes during Toll Receptor Signaling*

    Moncrieffe, Martin C.; Grossmann, J. Günter; Gay, Nicholas J.

    2008-01-01

    The Drosophila Toll receptor is activated by the endogenous protein ligand Spätzle in response to microbial stimuli in immunity and spatial cues during embryonic development. Downstream signaling is mediated by the adaptor proteins Tube, the kinase Pelle, and the Drosophila homologue of myeloid differentiation primary response protein (dMyD88). Here we have characterized heterodimeric (dMyD88-Tube) and heterotrimeric (dMyD88-Tube-Pelle) death domain complexes. We show ...

  19. Norepinephrine-Induced Adrenergic Activation Strikingly Increased the Atrial Fibrillation Duration through β1- and α1-Adrenergic Receptor-Mediated Signaling in Mice.

    Kenji Suita

    Full Text Available Atrial fibrillation (AF is the most common arrhythmias among old people. It causes serious long-term health problems affecting the quality of life. It has been suggested that the autonomic nervous system is involved in the onset and maintenance of AF in human. However, investigation of its pathogenesis and potential treatment has been hampered by the lack of suitable AF models in experimental animals.Our aim was to establish a long-lasting AF model in mice. We also investigated the role of adrenergic receptor (AR subtypes, which may be involved in the onset and duration of AF.Trans-esophageal atrial burst pacing in mice could induce AF, as previously shown, but with only a short duration (29.0 ± 8.1 sec. We found that adrenergic activation by intraperitoneal norepinephrine (NE injection strikingly increased the AF duration. It increased the duration to more than 10 minutes, i.e., by more than 20-fold (656.2 ± 104.8 sec; P<0.001. In this model, a prior injection of a specific β1-AR blocker metoprolol and an α1-AR blocker prazosin both significantly attenuated NE-induced elongation of AF. To further explore the mechanisms underlying these receptors' effects on AF, we assessed the SR Ca(2+ leak, a major trigger of AF, and consequent spontaneous SR Ca(2+ release (SCR in atrial myocytes. Consistent with the results of our in-vivo experiments, both metoprolol and prazosin significantly inhibited the NE-induced SR Ca(2+ leak and SCR. These findings suggest that both β1-AR and α1-AR may play important roles in the development of AF.We have established a long-lasting AF model in mice induced by adrenergic activation, which will be valuable in future AF study using experimental animals, such as transgenic mice. We also revealed the important role of β1- and α1-AR-mediated signaling in the development of AF through in-vivo and in-vitro experiments.

  20. Quantifying Agonist Activity at G Protein-coupled Receptors

    Ehlert, Frederick J.; Suga, Hinako; Griffin, Michael T.

    2011-01-01

    When an agonist activates a population of G protein-coupled receptors (GPCRs), it elicits a signaling pathway that culminates in the response of the cell or tissue. This process can be analyzed at the level of a single receptor, a population of receptors, or a downstream response. Here we describe how to analyze the downstream response to obtain an estimate of the agonist affinity constant for the active state of single receptors.

  1. ITAM-coupled receptors inhibit IFNAR signaling and alter macrophage responses to TLR4 and Listeria monocytogenes1

    Huynh, Linda; WANG, LU; Shi, Chao; Park-Min, Kyung-Hyun; Ivashkiv, Lionel B.

    2012-01-01

    Immunoreceptor tyrosine based activation motif (ITAM)-coupled receptors play an essential role in regulating macrophage activation and function by cross-regulating signaling from heterologous receptors. We investigated mechanisms by which ITAM-associated receptors inhibit type I interferon (IFN-α/β) signaling in primary human macrophages and tested the effects of simultaneous ligation of ITAM-associated receptors and TLR4 on TLR4-induced Jak-STAT signaling that is mediated by autocrine IFN-β....

  2. Biased signaling through G-protein-coupled PROKR2 receptors harboring missense mutations.

    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

  3. 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.

    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

  4. Human Neuropeptide S Receptor Is Activated via a Gαq Protein-biased Signaling Cascade by a Human Neuropeptide S Analog Lacking the C-terminal 10 Residues.

    Liao, Yuan; Lu, Bin; Ma, Qiang; Wu, Gang; Lai, Xiangru; Zang, Jiashu; Shi, Ying; Liu, Dongxiang; Han, Feng; Zhou, Naiming

    2016-04-01

    Human neuropeptide S (NPS) and its cognate receptor regulate important biological functions in the brain and have emerged as a future therapeutic target for treatment of a variety of neurological and psychiatric diseases. The human NPS (hNPS) receptor has been shown to dually couple to Gαs- and Gαq-dependent signaling pathways. The human NPS analog hNPS-(1-10), lacking 10 residues from the C terminus, has been shown to stimulate Ca(2+)mobilization in a manner comparable with full-length hNPSin vitrobut seems to fail to induce biological activityin vivo Here, results derived from a number of cell-based functional assays, including intracellular cAMP-response element (CRE)-driven luciferase activity, Ca(2+)mobilization, and ERK1/2 phosphorylation, show that hNPS-(1-10) preferentially activates Gαq-dependent Ca(2+)mobilization while exhibiting less activity in triggering Gαs-dependent CRE-driven luciferase activity. We further demonstrate that both Gαq- and Gαs-coupled signaling pathways contribute to full-length hNPS-mediated activation of ERK1/2, whereas hNPS-(1-10)-promoted ERK1/2 activation is completely inhibited by the Gαqinhibitor UBO-QIC but not by the PKA inhibitor H89. Moreover, the results of Ala-scanning mutagenesis of hNPS-(1-13) indicated that residues Lys(11)and Lys(12)are structurally crucial for the hNPS receptor to couple to Gαs-dependent signaling. In conclusion, our findings demonstrate that hNPS-(1-10) is a biased agonist favoring Gαq-dependent signaling. It may represent a valuable chemical probe for further investigation of the therapeutic potential of human NPS receptor-directed signalingin vivo. PMID:26865629

  5. IgE receptor signaling in food allergy pathogenesis.

    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

  6. Sweet taste receptor expressed in pancreatic beta-cells activates the calcium and cyclic AMP signaling systems and stimulates insulin secretion.

    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.

  7. Opioid receptors: Structural and mechanistic insights into pharmacology and signaling.

    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

  8. Analysis of C-type lectin receptor induced NF-kappaB signaling

    Straßer, Andreas Dominikus

    2014-01-01

    Myeloid C-type lectin receptors (CLRs) that signal via Syk and the central Card9-Bcl10-Malt1 (CBM) complex induce the transcription of NF-κB-regulated genes. Activation of those receptors mediates inflammatory reactions and the defense against various pathogens. Despite the non-redundant role of CLRs for the induction of innate immune responses, particularly receptor-proximal events that transduce ligand binding to downstream signaling remain to be defined. This dissertation identifies PKCδ a...

  9. Dopamine D1-D2 receptor heteromer signaling pathway in the brain: emerging physiological relevance

    Hasbi Ahmed

    2011-06-01

    Full Text Available Abstract Dopamine is an important catecholamine neurotransmitter modulating many physiological functions, and is linked to psychopathology of many diseases such as schizophrenia and drug addiction. Dopamine D1 and D2 receptors are the most abundant dopaminergic receptors in the striatum, and although a clear segregation between the pathways expressing these two receptors has been reported in certain subregions, the presence of D1-D2 receptor heteromers within a unique subset of neurons, forming a novel signaling transducing functional entity has been shown. Recently, significant progress has been made in elucidating the signaling pathways activated by the D1-D2 receptor heteromer and their potential physiological relevance.

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

    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. Esculetin attenuates receptor activator of nuclear factor kappa-B ligand-mediated osteoclast differentiation through c-Fos/nuclear factor of activated T-cells c1 signaling pathway

    Baek, Jong Min; Park, Sun-Hyang; Cheon, Yoon-Hee; Ahn, Sung-Jun [Department of Anatomy, School of Medicine, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Lee, Myeung Su [Division of Rheumatology, Department of Internal Medicine, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Imaging Science-based Lung and Bone Diseases Research Center, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Institute for Skeletal Disease, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Oh, Jaemin, E-mail: jmoh@wku.ac.kr [Department of Anatomy, School of Medicine, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Imaging Science-based Lung and Bone Diseases Research Center, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Institute for Skeletal Disease, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Kim, Ju-Young, E-mail: kimjy1014@gmail.com [Imaging Science-based Lung and Bone Diseases Research Center, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of)

    2015-05-29

    Esculetin exerts various biological effects on anti-oxidation, anti-tumors, and anti-inflammation. However, the involvement of esculetin in the bone metabolism process, particularly osteoclast differentiation has not yet been investigated. In the present study, we first confirmed the inhibitory effect of esculetin on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation. We then revealed the relationship between esculetin and the expression of osteoclast-specific molecules to elucidate its underlying mechanisms. Esculetin interfered with the expression of c-Fos and nuclear factor of activated T cell c1 (NFATc1) both at the mRNA and protein level with no involvement in osteoclast-associated early signaling pathways, suppressing the expression of various transcription factors exclusively expressed in osteoclasts such as tartrate-resistant acid phosphatase (Trap), osteoclast-associated receptor (Oscar), dendritic cell-specific transmembrane protein (Dcstamp), osteoclast stimulatory transmembrane protein (Ocstamp), cathepsin K, αvβ3 integrin, and calcitonin receptor (Ctr). Additionally, esculetin inhibited the formation of filamentous actin (F-actin) ring-positive osteoclasts during osteoclast differentiation. However, the development of F-actin structures and subsequent bone resorbing activity of mature osteoclasts, which are observed in osteoclast/osteoblast co-culture systems were not affected by esculetin. Taken together, our results indicate for the first time that esculetin inhibits RANKL-mediated osteoclastogenesis via direct suppression of c-Fos and NFATc1 expression and exerts an inhibitory effect on actin ring formation during osteoclastogenesis. - Highlights: • We first investigated the effects of esculetin on osteoclast differentiation and function. • Our data demonstrate for the first time that esculetin can suppress osteoclastogenesis in vitro. • Esculetin acts as an inhibitor of c-Fos and NFATc1 activation.

  12. Esculetin attenuates receptor activator of nuclear factor kappa-B ligand-mediated osteoclast differentiation through c-Fos/nuclear factor of activated T-cells c1 signaling pathway

    Esculetin exerts various biological effects on anti-oxidation, anti-tumors, and anti-inflammation. However, the involvement of esculetin in the bone metabolism process, particularly osteoclast differentiation has not yet been investigated. In the present study, we first confirmed the inhibitory effect of esculetin on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation. We then revealed the relationship between esculetin and the expression of osteoclast-specific molecules to elucidate its underlying mechanisms. Esculetin interfered with the expression of c-Fos and nuclear factor of activated T cell c1 (NFATc1) both at the mRNA and protein level with no involvement in osteoclast-associated early signaling pathways, suppressing the expression of various transcription factors exclusively expressed in osteoclasts such as tartrate-resistant acid phosphatase (Trap), osteoclast-associated receptor (Oscar), dendritic cell-specific transmembrane protein (Dcstamp), osteoclast stimulatory transmembrane protein (Ocstamp), cathepsin K, αvβ3 integrin, and calcitonin receptor (Ctr). Additionally, esculetin inhibited the formation of filamentous actin (F-actin) ring-positive osteoclasts during osteoclast differentiation. However, the development of F-actin structures and subsequent bone resorbing activity of mature osteoclasts, which are observed in osteoclast/osteoblast co-culture systems were not affected by esculetin. Taken together, our results indicate for the first time that esculetin inhibits RANKL-mediated osteoclastogenesis via direct suppression of c-Fos and NFATc1 expression and exerts an inhibitory effect on actin ring formation during osteoclastogenesis. - Highlights: • We first investigated the effects of esculetin on osteoclast differentiation and function. • Our data demonstrate for the first time that esculetin can suppress osteoclastogenesis in vitro. • Esculetin acts as an inhibitor of c-Fos and NFATc1 activation.

  13. In brown adipocytes, adrenergically induced β1-/β3-(Gs)-, α2-(Gi)- and α1-(Gq)-signalling to Erk1/2 activation is not mediated via EGF receptor transactivation

    Brown adipose tissue is unusual in that the neurotransmitter norepinephrine influences cell destiny in ways generally associated with effects of classical growth factors: regulation of cell proliferation, of apoptosis, and progression of differentiation. The norepinephrine effects are mediated through G-protein-coupled receptors; further mediation of such stimulation to e.g. Erk1/2 activation is in cell biology in general accepted to occur through transactivation of the EGF receptor (by external or internal pathways). We have examined here the significance of such transactivation in brown adipocytes. Stimulation of mature brown adipocytes with cirazoline (α1-adrenoceptor coupled via Gq), clonidine (α2 via Gi) or CL316243 (β3 via Gs) or via β1-receptors significantly activated Erk1/2. Pretreatment with the EGF receptor kinase inhibitor AG1478 had, remarkably, no significant effect on Erk1/2 activation induced by any of these adrenergic agonists (although it fully abolished EGF-induced Erk1/2 activation), demonstrating absence of EGF receptor-mediated transactivation. Results with brown preadipocytes (cells in more proliferative states) were not qualitatively different. Joint stimulation of all adrenoceptors with norepinephrine did not result in synergism on Erk1/2 activation. AG1478 action on EGF-stimulated Erk1/2 phosphorylation showed a sharp concentration–response relationship (IC50 0.3 µM); a minor apparent effect of AG1478 on norepinephrine-stimulated Erk1/2 phosphorylation showed nonspecific kinetics, implying caution in interpretation of partial effects of AG1478 as reported in other systems. Transactivation of the EGF receptor is clearly not a universal prerequisite for coupling of G-protein coupled receptors to Erk1/2 signalling cascades. - Highlights: • In brown adipocytes, norepinephrine regulates proliferation, apoptosis, differentiation. • EGF receptor transactivation is supposed to mediate GPCR-induced Erk1/2 activation. • α1-, α2-, β1

  14. NMDA receptor activity in neuropsychiatric disorders

    ShaheenELakhan

    2013-06-01

    Full Text Available N-Methyl-D-aspartate (NMDA receptors play a variety of physiologic roles and their proper signaling is essential for cellular homeostasis. Any disruption in this pathway, leading to either enhanced or decreased activity, may result in the manifestation of neuropsychiatric pathologies such as schizophrenia, mood disorders, substance induced psychosis, Huntington's disease, Alzheimer's disease, and neuropsychiatric systemic lupus erythematosus. Here, we explore the notion that the overlap in activity of at least one biochemical pathway, the NMDA receptor pathway, may be the link to understanding the overlap in psychotic symptoms between diseases. This review intends to present a broad overview of those neuropsychiatric disorders for which alternations in NMDA receptor activity is prominent thus suggesting that continued direction of pharmaceutical intervention to this pathway may present a viable option for managing symptoms.

  15. Strigolactone and karrikin signal perception: receptors, enzymes, or both?

    Bart Jan Janssen

    2012-12-01

    Full Text Available The signaling molecules strigolactone (SL and karrikin are involved in seed germination, development of axillary meristems, senescence of leaves and interactions with arbuscular mycorrhizal fungi. The signal transduction pathways for both SLs and karrikins require the same F-box protein (MAX2 and closely related α/β hydrolase fold proteins (DAD2 and KAI2. The crystal structure of DAD2 has been solved revealing an α/β hydrolase fold protein with an internal cavity capable of accommodating SLs. DAD2 responds to the SL analog GR24 by changing conformation and binding to MAX2 in a GR24 concentration-dependent manner. DAD2 can also catalyse hydrolysis of GR24. Structure activity relationships of analogs indicate that the butenolide ring common to both SLs and karrikins is essential for biological activity, but the remainder of the molecules can be significantly modified without loss of activity. The combination of data from the study of DAD2, KAI2 and chemical analogs of SLs and karrikins suggests a model for binding that requires nucleophilic attack by the active site serine of the hydrolase at the carbonyl atom of the butenolide ring. A conformational change occurs in the hydrolase that results in interaction with the F-box protein MAX2. Downstream signal transduction is then likely to occur via SCF complex-mediated ubiquitination of target proteins and their subsequent degradation. The role of the catalytic activity of the hydrolase is unclear but it may be integral in binding as well as possibly allowing the signal to be cleared from the receptor. The α/β hydrolase fold family consists mostly of active enzymes, with a few notable exceptions. We suggest that the DAD2 and KAI2 represent an intermediate where some catalytic activity is retained at the same time as a receptor role has evolved.

  16. Metazoan-like signaling in a unicellular receptor tyrosine kinase

    Schultheiss Kira P

    2013-02-01

    Full Text Available Abstract Background Receptor tyrosine kinases (RTKs are crucial components of signal transduction systems in multicellular animals. Surprisingly, numerous RTKs have been identified in the genomes of unicellular choanoflagellates and other protists. Here, we report the first biochemical study of a unicellular RTK, namely RTKB2 from Monosiga brevicollis. Results We cloned, expressed, and purified the RTKB2 kinase, and showed that it is enzymatically active. The activity of RTKB2 is controlled by autophosphorylation, as in metazoan RTKs. RTKB2 possesses six copies of a unique domain (designated RM2 in its C-terminal tail. An isolated RM2 domain (or a synthetic peptide derived from the RM2 sequence served as a substrate for RTKB2 kinase. When phosphorylated, the RM2 domain bound to the Src homology 2 domain of MbSrc1 from M. brevicollis. NMR structural studies of the RM2 domain indicated that it is disordered in solution. Conclusions Our results are consistent with a model in which RTKB2 activation stimulates receptor autophosphorylation within the RM2 domains. This leads to recruitment of Src-like kinases (and potentially other M. brevicollis proteins and further phosphorylation, which may serve to increase or dampen downstream signals. Thus, crucial features of signal transduction circuitry were established prior to the evolution of metazoans from their unicellular ancestors.

  17. Neu1 sialidase and matrix metalloproteinase-9 cross-talk regulates nucleic acid-induced endosomal TOLL-like receptor-7 and -9 activation, cellular signaling and pro-inflammatory responses.

    Abdulkhalek, Samar; Szewczuk, Myron R

    2013-11-01

    The precise mechanism(s) by which intracellular TOLL-like receptors (TLRs) become activated by their ligands remains unclear. Here, we report a molecular organizational G-protein coupled receptor (GPCR) signaling platform to potentiate a novel mammalian neuraminidase-1 (Neu1) and matrix metalloproteinase-9 (MMP-9) cross-talk in alliance with neuromedin B GPCR, all of which form a tripartite complex with TLR-7 and -9. siRNA silencing Neu1, MMP-9 and neuromedin-B GPCR in RAW-blue macrophage cells significantly reduced TLR7 imiquimod- and TLR9 ODN1826-induced NF-κB (NF-κB-pSer(536)) activity. Tamiflu, specific MMP-9 inhibitor, neuromedin B receptor specific antagonist BIM23127, and the selective inhibitor of whole heterotrimeric G-protein complex BIM-46174 significantly block nucleic acid-induced TLR-7 and -9 MyD88 recruitment, NF-κB activation and proinflammatory TNFα and MCP-1 cytokine responses. For the first time, Neu1 clearly plays a central role in mediating nucleic acid-induced intracellular TLR activation, and the interactions involving NMBR-MMP9-Neu1 cross-talk constitute a novel intracellular TLR signaling platform that is essential for NF-κB activation and pro-inflammatory responses. PMID:23827939

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

    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.

  19. Anti-inflammatory activity of Odina wodier Roxb, an Indian folk remedy, through inhibition of toll-like receptor 4 signaling pathway.

    Durbadal Ojha

    Full Text Available Inflammation is part of self-limiting non-specific immune response, which occurs during bodily injury. In some disorders the inflammatory process becomes continuous, leading to the development of chronic inflammatory diseases including cardiovascular diseases, diabetes, cancer etc. Several Indian tribes used the bark of Odina wodier (OWB for treating inflammatory disorders. Thus, we have evaluated the immunotherapeutic potential of OWB methanol extract and its major constituent chlorogenic acid (CA, using three popular in vivo antiinflammatory models: Carrageenan- and Dextran-induced paw edema, Cotton pellet granuloma, and Acetic acid-induced vascular permeability. To elucidate the possible anti-inflammatory mechanism of action we determine the level of major inflammatory mediators (NO, iNOS, COX-2-dependent prostaglandin E2 or PGE2, and pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, and IL-12. Further, we determine the toll-like receptor 4 (TLR4, Myeloid differentiation primary response gene 88 (MyD88, c-Jun N-terminal kinases (JNK, nuclear factor kappa-B cells (NF-κB, and NF-kB inhibitor alpha (IK-Bα by protein and mRNA expression, and Western blot analysis in drug treated LPS-induced murine macrophage model. Moreover, we determined the acute and sub-acute toxicity of OWB extract in BALB/c mice. Our study demonstrated a significant anti-inflammatory activity of OWB extract and CA along with the inhibition of TNF-α, IL-1β, IL-6 and IL-12 expressions. Further, the expression of TLR4, NF-κBp65, MyD88, iNOS and COX-2 molecules were reduced in drug-treated groups, but not in the LPS-stimulated untreated or control groups, Thus, our results collectively indicated that the OWB extract and CA can efficiently inhibit inflammation through the down regulation of TLR4/MyD88/NF-kB signaling pathway.

  20. Biased Signaling of the Angiotensin II Type 1 Receptor Can Be Mediated through Distinct Mechanisms

    Bonde, Marie Mi; Hansen, Jonas Tind; Sanni, Samra Joke; Haunsø, Stig; Gammeltoft, Steen; Lyngsø, Christina; Hansen, Jakob Lerche

    2010-01-01

    Background Seven transmembrane receptors (7TMRs) can adopt different active conformations facilitating a selective activation of either G protein or β-arrestin-dependent signaling pathways. This represents an opportunity for development of novel therapeutics targeting selective biological effects of a given receptor. Several studies on pathway separation have been performed, many of these on the Angiotensin II type 1 receptor (AT1R). It has been shown that certain ligands or mutations facilit...

  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.

    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. Analysis of corkscrew signaling in the Drosophila epidermal growth factor receptor pathway during myogenesis.

    Johnson Hamlet, M R; Perkins, L A

    2001-01-01

    The Drosophila nonreceptor protein tyrosine phosphatase, Corkscrew (Csw), functions positively in multiple receptor tyrosine kinase (RTK) pathways, including signaling by the epidermal growth factor receptor (EGFR). Detailed phenotypic analyses of csw mutations have revealed that Csw activity is required in many of the same developmental processes that require EGFR function. However, it is still unclear where in the signaling hierarchy Csw functions relative to other proteins whose activities...

  3. Nuclear Receptor Signaling Atlas: Opening Access to the Biology of Nuclear Receptor Signaling Pathways.

    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.

  4. Nuclear Receptor Signaling Atlas: Opening Access to the Biology of Nuclear Receptor Signaling Pathways

    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

  5. Self-enforcing Feedback Activation between BCL6 and Pre-B Cell Receptor Signaling Defines a Distinct Subtype of Acute Lymphoblastic Leukemia

    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

  6. 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.

    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

  7. Wnt signaling and the activation of myogenesis in mammals.

    Cossu, G.; Borello, U

    1999-01-01

    In the amniote embryos, specification of skeletal myoblasts occurs in the paraxial mesoderm in response to a number of signaling molecules produced by neighboring tissues such as neural tube, notochord and dorsal ectoderm. Candidate molecules for this complex signaling activity include Sonic hedgehog, Wnts and Noggin as positive activators and BMP4 as a possible inhibitor. Recently, the receptors and the post-receptor pathways for Sonic hedgehog and Wnts have been characterized, and this has ...

  8. Appetitive Cue-Evoked ERK Signaling in the Nucleus Accumbens Requires NMDA and D1 Dopamine Receptor Activation and Regulates CREB Phosphorylation

    Kirschmann, Erin K. Z.; Mauna, Jocelyn C.; Willis, Cory M.; Foster, Rebecca L.; Chipman, Amanda M.; Thiels, Edda

    2014-01-01

    Conditioned stimuli (CS) can modulate reward-seeking behavior. This modulatory effect can be maladaptive and has been implicated in excessive reward seeking and relapse to drug addiction. We previously demonstrated that exposure to an appetitive CS causes an increase in the activation of extracellular signal-regulated kinase (ERK) and cyclic-AMP…

  9. Signal interaction of Hedgehog/GLI and epidermal growth factor receptor signaling in cancer development

    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)

  10. Regulator of G protein signaling 2 (RGS2 and RGS4 form distinct G protein-dependent complexes with protease activated-receptor 1 (PAR1 in live cells.

    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.

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

    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

  12. Serotonin acts as a novel regulator of interleukin-6 secretion in osteocytes through the activation of the 5-HT(2B) receptor and the ERK1/2 signalling pathway.

    Li, Xianxian; Ma, Yuanyuan; Wu, Xiangnan; Hao, Zhichao; Yin, Jian; Shen, Jiefei; Li, Xiaoyu; Zhang, Ping; Wang, Hang

    2013-11-29

    Interleukin-6 (IL-6) is a potent stimulator of osteoclastic bone resorption. Osteocyte secretion of IL-6 plays an important role in bone metabolism. Serotonin (5-HT) has recently been reported to regulate bone metabolism. The aim of this study was to evaluate the effect of serotonin on osteocyte expression of IL-6. The requirement for the 5-HT receptor(s) and the role of the extracellular signal-regulated kinase 1/2 (ERK1/2) in serotonin-induced IL-6 synthesis were examined. In this study, real-time PCR and ELISA were used to analyse IL-6 gene and protein expression in serotonin-stimulated MLO-Y4 cells. ERK1/2 pathway activation was determined by Western blot. We found that serotonin significantly activated the ERK1/2 pathway and induced IL-6 mRNA expression and protein synthesis in cultured MLO-Y4 cells. However, these effects were abolished by pre-treatment of MLO-Y4 cells with a 5-HT2B receptor antagonist, RS127445 or the ERK1/2 inhibitor, PD98059. Our results indicate that serotonin stimulates osteocyte secretion of IL-6 and that this effect is associated with activation of 5-HT2B receptor and the ERK1/2 pathway. These findings provide support for a role of serotonin in bone metabolism by indicating serotonin regulates bone remodelling by mediating an inflammatory cytokine. PMID:24211588

  13. The maintenance of cisplatin- and paclitaxel-induced mechanical and cold allodynia is suppressed by cannabinoid CB2 receptor activation and independent of CXCR4 signaling in models of chemotherapy-induced peripheral neuropathy

    Deng Liting

    2012-09-01

    Full Text Available Abstract Background Chemotherapeutic agents produce dose-limiting peripheral neuropathy through mechanisms that remain poorly understood. We previously showed that AM1710, a cannabilactone CB2 agonist, produces antinociception without producing central nervous system (CNS-associated side effects. The present study was conducted to examine the antinociceptive effect of AM1710 in rodent models of neuropathic pain evoked by diverse chemotherapeutic agents (cisplatin and paclitaxel. A secondary objective was to investigate the potential contribution of alpha-chemokine receptor (CXCR4 signaling to both chemotherapy-induced neuropathy and CB2 agonist efficacy. Results AM1710 (0.1, 1 or 5 mg/kg i.p. suppressed the maintenance of mechanical and cold allodynia in the cisplatin and paclitaxel models. Anti-allodynic effects of AM1710 were blocked by the CB2 antagonist AM630 (3 mg/kg i.p., but not the CB1 antagonist AM251 (3 mg/kg i.p., consistent with a CB2-mediated effect. By contrast, blockade of CXCR4 signaling with its receptor antagonist AMD3100 (10 mg/kg i.p. failed to attenuate mechanical or cold hypersensitivity induced by either cisplatin or paclitaxel. Moreover, blockade of CXCR4 signaling failed to alter the anti-allodynic effects of AM1710 in the paclitaxel model, further suggesting distinct mechanisms of action. Conclusions Our results indicate that activation of cannabinoid CB2 receptors by AM1710 suppresses both mechanical and cold allodynia in two distinct models of chemotherapy-induced neuropathic pain. By contrast, CXCR4 signaling does not contribute to the maintenance of chemotherapy-induced established neuropathy or efficacy of AM1710. Our studies suggest that CB2 receptors represent a promising therapeutic target for the treatment of toxic neuropathies produced by cisplatin and paclitaxel chemotherapeutic agents.

  14. Toll-like receptor signaling in primary immune deficiencies.

    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

  15. Pharmacology of bile acid receptors: Evolution of bile acids from simple detergents to complex signaling molecules.

    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

  16. Noradrenaline represses PPAR (peroxisome-proliferator-activated receptor) gamma2 gene expression in brown adipocytes: intracellular signalling and effects on PPARgamma2 and PPARgamma1 protein levels

    Lindgren, Eva M; Nielsen, Ronni; Petrovic, Natasa;

    2004-01-01

    PPAR (peroxisome-proliferator-activated receptor) gamma is expressed in brown and white adipose tissues and is involved in the control of differentiation and proliferation. Noradrenaline stimulates brown pre-adipocyte proliferation and brown adipocyte differentiation. The aim of the present study...... was thus to investigate the influence of noradrenaline on PPARgamma gene expression in brown adipocytes. In primary cultures of brown adipocytes, PPARgamma2 mRNA levels were 20-fold higher than PPARgamma1 mRNA levels. PPARgamma expression occurred during both the proliferation and the differentiation...

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

    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.

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

    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

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

    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.

  20. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia

    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.

  1. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia

    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

  2. Glutamate Delta-1 Receptor Regulates Metabotropic Glutamate Receptor 5 Signaling in the Hippocampus.

    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

  3. 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

    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

  4. Novel roles of nuclear angiotensin receptors and signaling mechanisms.

    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

  5. A comprehensive map of the toll-like receptor signaling network

    Oda, Kanae; Kitano, Hiroaki

    2006-01-01

    Recognition of pathogen-associated molecular signatures is critically important in proper activation of the immune system. The toll-like receptor (TLR) signaling network is responsible for innate immune response. In mammalians, there are 11 TLRs that recognize a variety of ligands from pathogens to trigger immunological responses. In this paper, we present a comprehensive map of TLRs and interleukin 1 receptor signaling networks based on papers published so far. The map illustrates the possib...

  6. Exercise-induced changes in expression and activity of proteins involved in insulin signal transduction in skeletal muscle: Differential effects on insulin-receptor substrates 1 and 2

    Chibalin, Alexander V; Yu, Mei; Ryder, Jeffrey W.; Song, Xiao Mei; Galuska, Dana; Krook, Anna; Wallberg-Henriksson, Harriet; Juleen R. Zierath

    2000-01-01

    Level of physical activity is linked to improved glucose homeostasis. We determined whether exercise alters the expression and/or activity of proteins involved in insulin-signal transduction in skeletal muscle. Wistar rats swam 6 h per day for 1 or 5 days. Epitrochlearis muscles were excised 16 h after the last exercise bout, and were incubated with or without insulin (120 nM). Insulin-stimulated glucose transport increased 30% and 50% after 1 and 5 days of exercise, respectively. Glycogen co...

  7. Thyrotropin Receptor Stimulates Internalization-Independent Persistent Phosphoinositide Signaling

    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...

  8. Effects of thermal fluctuation and the receptor-receptor interaction in bacterial chemotactic signalling and adaptation

    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...

  9. From pure compounds to complex exposure: Effects of dietary cadmium and lignans on estrogen, epidermal growth factor receptor, and mitogen activated protein kinase signaling in vivo.

    Ali, Imran; Hurmerinta, Teija; Nurmi, Tarja; Berglund, Marika; Rüegg, Joelle; Poutanen, Matti; Halldin, Krister; Mäkelä, Sari; Damdimopoulou, Pauliina

    2016-06-24

    Exposure to environmental endocrine active compounds correlates with altered susceptibility to disease in human populations. Chemical risk assessment is single compound based, although exposure often takes place as heterogeneous mixtures of man-made and natural substances within complex matrices like diet. Here we studied whether the effects of cadmium and enterolactone on endocrine endpoints in dietary exposure can be predicted based on pure compound effects. Ovariectomized estrogen reporter ERE-luciferase (ERE-luc) mice were maintained on diets that intrinsically contain increasing concentrations of cadmium and enterolactone precursors for three and 21 days. The activation of the ERE-luc, epidermal growth factor receptor (EGFR), mitogen activated protein kinase (MAPK)-ERK1/2, and classical estrogen responses were measured. Interactions between the diets and endogenous hormone were evaluated by challenging the animals with 17β-estradiol. Compared to animals on basal purified diet, mice consuming experimental diets were exposed to significantly higher levels of cadmium and enterolactone, yet the exposure remained comparable to typical human dietary intake. Surprisingly, we could not detect effects on endpoints regulated by pure enterolactone, such as ERE-luc activation. However, cadmium accumulation in the liver was accompanied with activation of EGFR and MAPK-ERK1/2 in line with our earlier CdCl2 studies. Further, attenuation of 17β-estradiol-induced ERE-luc response in liver by experimental diets was observed. Our findings indicate that the exposure context can have substantial effects on the activity of endocrine active compounds in vivo. Thus, whenever possible, a context that mimics human exposure should be tested along with pure compounds. PMID:27108949

  10. DMPD: Signal transduction by the lipopolysaccharide receptor, Toll-like receptor-4. [Dynamic Macrophage Pathway CSML Database

    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

  11. Insulin-like growth factor I receptor dose not contribute to heat shock-induced activation of Akt and extracellular signal-regulated kinase (ERK) in mouse embryo fibroblasts

    We have investigated the role of insulin-like growth factor I receptor (IGF-IR) in heat shock-induced activation of the mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3' kinase (PI3-K) pathways. We utilized mouse embryo fibroblasts (MEFs) devoid of endogenous IGF-LR (R-) and MEFs overexpressing human IGF-IR (WT) and examined the activation kinetics of extracellular signal-regulated kinase (ERK) and Akt following heat shock treatment. There were no differences in the kinetics or temperature dependence of activation of either ERK or Akt between the cell lines. As expected, heat shock failed to induce autophosphorylation of IGF-IR overexpressed in WT cells. Surprisingly, the autophosphorylation of endogenous epidermal growth factor receptor (EGFR), which is thought to play an important role in heat shock-induced activation of the MAPK and PI3-K pathways, was not observed in either WT or R-cells. These results suggest that neither IGF-IR nor EGFR contributes to the heat shock-induced activation of ERK and Akt in these cell lines. (author)

  12. Chromatin remodeling by curcumin alters endogenous aryl hydrocarbon receptor signaling.

    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

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

    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

  14. Signal Processing under Active Monitoring

    Mostovyi, Oleksii

    2005-01-01

    This paper describes a method of signal preprocessing under active monitoring. Suppose we want to solve the inverse problem of getting the response of a medium to one powerful signal, which is equivalent to obtaining the transmission function of the medium, but do not have an opportunity to conduct such an experiment (it might be too expensive or harmful for the environment). Practically the problem can be reduced to obtaining the transmission function of the medium. In this case ...

  15. Role of Phospholipase C-L2, a Novel Phospholipase C-Like Protein That Lacks Lipase Activity, in B-Cell Receptor Signaling

    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. ...

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

    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

  17. Cannabinoid receptor-interacting protein Crip1a modulates CB1 receptor signaling in mouse hippocampus.

    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

  18. Inhibition of protein tyrosine phosphatase activity mediates epidermal growth factor receptor signaling in human airway epithelial cells exposed to Zn2+

    Epidemiological studies have implicated zinc (Zn2+) 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 to Src-dependent activation of EGFR signaling in B82 and A431 cells. In order to elucidate the mechanism of Zn2+-induced EGFR activation in HAEC, we treated HAEC with 500 μM ZnSO4 for 5-20 min and measured the state of activation of EGFR, c-Src and PTPs. Western blots revealed that exposure to Zn2+ results in increased phosphorylation at both trans- and autophosphorylation sites in the EGFR. Zn2+-mediated EGFR phosphorylation did not require ligand binding and was ablated by the EGFR kinase inhibitor PD153035, but not by the Src kinase inhibitor PP2. Src activity was inhibited by Zn2+ treatment of HAEC, consistent with Src-independent EGFR transactivation in HAEC exposed to Zn2+. The rate of exogenous EGFR dephosphorylation in lysates of HAEC exposed to Zn2+ or V4+ was significantly diminished. Moreover, exposure of HAEC to Zn2+ also resulted in a significant impairment of dephosphorylation of endogenous EGFR. These data show that Zn2+-induced activation of EGFR in HAEC involves a loss of PTP activities whose function is to dephosphorylate EGFR in opposition to baseline EGFR kinase activity. These findings also suggest that there are marked cell-type-specific differences in the mechanism of EGFR activation induced by Zn2+ exposure

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

    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.

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

    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...

  1. Insulin signaling inhibits the 5-HT2C receptor in choroid plexus via MAP kinase

    Guan Kunliang

    2003-06-01

    Full Text Available Abstract Background G protein-coupled receptors (GPCRs interact with heterotrimeric GTP-binding proteins (G proteins to modulate acute changes in intracellular messenger levels and ion channel activity. In contrast, long-term changes in cellular growth, proliferation and differentiation are often mediated by tyrosine kinase receptors and certain GPCRs by activation of mitogen-activated protein (MAP kinases. Complex interactions occur between these signaling pathways, but the specific mechanisms of such regulatory events are not well-understood. In particular it is not clear whether GPCRs are modulated by tyrosine kinase receptor-MAP kinase pathways. Results Here we describe tyrosine kinase receptor regulation of a GPCR via MAP kinase. Insulin reduced the activity of the 5-HT2C receptor in choroid plexus cells which was blocked by the MAP kinase kinase (MEK inhibitor, PD 098059. We demonstrate that the inhibitory effect of insulin and insulin-like growth factor type 1 (IGF-1 on the 5-HT2C receptor is dependent on tyrosine kinase, RAS and MAP kinase. The effect may be receptor-specific: insulin had no effect on another GPCR that shares the same G protein signaling pathway as the 5-HT2C receptor. This effect is also direct: activated MAP kinase mimicked the effect of insulin, and removing a putative MAP kinase site from the 5-HT2C receptor abolished the effect of insulin. Conclusion These results show that insulin signaling can inhibit 5-HT2C receptor activity and suggest that MAP kinase may play a direct role in regulating the function of a specific GPCR.

  2. Innate immune receptor Toll-like receptor 4 signalling in neuropsychiatric diseases.

    García Bueno, B; Caso, J R; Madrigal, J L M; Leza, J C

    2016-05-01

    The innate immunity is a stereotyped first line of defense against pathogens and unspecified damage signals. One of main actors of innate immunity are the Toll-like receptors (TLRs), and one of the better characterized members of this family is TLR-4, that it is mainly activated by Gram-negative bacteria lipopolysaccharide. In brain, TLR-4 organizes innate immune responses against infections or cellular damage, but also possesses other physiological functions. In the last years, some evidences suggest a role of TLR-4 in stress and stress-related neuropsychiatric diseases. Peripheral and brain TLR-4 activation triggers sickness behavior, and its expression is a risk factor of depression. Some elements of the TLR-4 signaling pathway are up-regulated in peripheral samples and brain post-mortem tissue from depressed and suicidal patients. The "leaky gut" hypothesis of neuropsychiatric diseases is based on the existence of an increase of the intestinal permeability which results in bacterial translocation able to activate TLR-4. Enhanced peripheral TLR-4 expression/activity has been described in subjects diagnosed with schizophrenia, bipolar disorder and in autistic children. A role for TLR-4 in drugs abuse has been also proposed. The therapeutic potential of pharmacological/genetic modulation of TLRs signaling pathways in neuropsychiatry is promising, but a great preclinical/clinical scientific effort is still needed. PMID:26905767

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

    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

  4. Molecular signalling mediating the protective effect of A1 adenosine and mGlu3 metabotropic glutamate receptor activation against apoptosis by oxygen/glucose deprivation in cultured astrocytes.

    Ciccarelli, Renata; D'Alimonte, Iolanda; Ballerini, Patrizia; D'Auro, Mariagrazia; Nargi, Eleonora; Buccella, Silvana; Di Iorio, Patrizia; Bruno, Valeria; Nicoletti, Ferdinando; Caciagli, Francesco

    2007-05-01

    Astrocyte death may occur in neurodegenerative disorders and complicates the outcome of brain ischemia, a condition associated with high extracellular levels of adenosine and glutamate. We show that pharmacological activation of A(1) adenosine and mGlu3 metabotropic glutamate receptors with N(6)-chlorocyclopentyladenosine (CCPA) and (-)2-oxa-4-aminocyclo-[3.1.0]hexane-4,6-dicarboxylic acid (LY379268), respectively, protects cultured astrocytes against apoptosis induced by a 3-h exposure to oxygen/glucose deprivation (OGD). Protection by CCPA and LY379268 was less than additive and was abrogated by receptor blockade with selective competitive antagonists or pertussis toxin. Both in control astrocytes and in astrocytes exposed to OGD, CCPA and LY379268 induced a rapid activation of the phosphatidylinositol-3-kinase (PI3K) and extracellular signal-regulated kinases 1 and 2 (ERK1/2)/mitogen-activated protein kinase (MAPK) pathways, which are known to support cell survival. In cultures exposed to OGD, CCPA and LY379268 reduced the activation of c-Jun N-terminal kinase and p38/MAPK, reduced the levels of the proapoptotic protein Bad, increased the levels of the antiapoptotic protein Bcl-X(L), and were highly protective against apoptotic death, as shown by nuclear 4'-6-diamidino-2-phenylindole staining and measurements of caspase-3 activity. All of these effects were attenuated by treatment with 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene (U0126) and 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride (LY294002), which inhibit the MAPK and the PI3K pathways, respectively. These data suggest that pharmacological activation of A(1) and mGlu3 receptors protects astrocytes against hypoxic/ischemic damage by stimulating the PI3K and ERK1/2 MAPK pathways. PMID:17293559

  5. Ectodysplasin A (EDA) - EDA receptor signalling and its pharmacological modulation.

    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

  6. Retinoid receptor signaling and autophagy in acute promyelocytic leukemia.

    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.

  7. Current Views of Toll-Like Receptor Signaling Pathways

    Masahiro Yamamoto

    2010-01-01

    Full Text Available On microbial invasion, the host immediately evokes innate immune responses. Recent studies have demonstrated that Toll-like receptors (TLRs play crucial roles in innate responses that lead not only to the clearance of pathogens but also to the efficient establishment of acquired immunity by directly detecting molecules from microbes. In terms of intracellular TLR-mediated signaling pathways, cytoplasmic adaptor molecules containing Toll/IL-1R (TIR domains play important roles in inflammatory immune responses through the production of proinflammatory cytokines, nitric oxide, and type I interferon, and upregulation of costimulatory molecules. In this paper, we will describe our current understanding of the relationship between TLRs and their ligands derived from pathogens such as viruses, bacteria, fungi, and parasites. Moreover, we will review the historical and current literature to describe the mechanisms behind TLR-mediated activation of innate immune responses.

  8. CB1 receptor signaling regulates social anxiety and memory.

    Litvin, Y; Phan, A; Hill, M N; Pfaff, D W; McEwen, B S

    2013-07-01

    The endocannabinoid (eCB) system regulates emotion, stress, memory and cognition through the cannabinoid type 1 (CB1 ) receptor. To test the role of CB1 signaling in social anxiety and memory, we utilized a genetic knockout (KO) and a pharmacological approach. Specifically, we assessed the effects of a constitutive KO of CB1 receptors (CB1 KOs) and systemic administration of a CB1 antagonist (AM251; 5 mg/kg) on social anxiety in a social investigation paradigm and social memory in a social discrimination test. Results showed that when compared with wild-type (WT) and vehicle-treated animals, CB1 KOs and WT animals that received an acute dose of AM251 displayed anxiety-like behaviors toward a novel male conspecific. When compared with WT animals, KOs showed both active and passive defensive coping behaviors, i.e. elevated avoidance, freezing and risk-assessment behaviors, all consistent with an anxiety-like profile. Animals that received acute doses of AM251 also showed an anxiety-like profile when compared with vehicle-treated animals, yet did not show an active coping strategy, i.e. changes in risk-assessment behaviors. In the social discrimination test, CB1 KOs and animals that received the CB1 antagonist showed enhanced levels of social memory relative to their respective controls. These results clearly implicate CB1 receptors in the regulation of social anxiety, memory and arousal. The elevated arousal/anxiety resulting from either total CB1 deletion or an acute CB1 blockade may promote enhanced social discrimination/memory. These findings may emphasize the role of the eCB system in anxiety and memory to affect social behavior. PMID:23647582

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

    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...

  10. Inhibition of WNT Signaling by G Protein-Coupled Receptor (GPCR) Kinase 2 (GRK2)

    WANG, LIMING; Gesty-Palmer, Diane; Fields, Timothy A.; Spurney, Robert F.

    2009-01-01

    Activation of Wnt signaling pathways causes release and stabilization of the transcription regulator β-catenin from a destruction complex composed of axin and the adenomatous polyposis coli (APC) protein (canonical signaling pathway). Assembly of this complex is facilitated by a protein-protein interaction between APC and a regulator of G protein signaling (RGS) domain in axin. Because G protein-coupled receptor kinase 2 (GRK2) has a RGS domain that is closely related to the RGS domain in axi...

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

    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...

  12. The molecular mechanisms behind receptor signaling

    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...

  13. Prolactin receptor and signal transduction to milk protein genes

    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.

  14. Non-genomic actions of aldosterone: From receptors and signals to membrane targets.

    Dooley, Ruth

    2011-07-26

    In tissues which express the mineralocorticoid receptor (MR), aldosterone modulates the expression of membrane targets such as the subunits of the epithelial Na(+) channel, in combination with important signalling intermediates such as serum and glucocorticoid-regulated kinase-1. In addition, the rapid \\'non-genomic\\' activation of protein kinases and secondary messenger signalling cascades has also been detected in aldosterone-sensitive tissues of the nephron, distal colon and cardiovascular system. These rapid actions are variously described as being coupled to MR or to an as yet unidentified, membrane-associated aldosterone receptor. The rapidly activated signalling cascades add a level of fine-tuning to the activity of aldosterone-responsive membrane transporters and also modulate the aldosterone-induced changes in gene expression through receptor and transcription factor phosphorylation.

  15. Non-genomic actions of aldosterone: From receptors and signals to membrane targets.

    2012-02-01

    In tissues which express the mineralocorticoid receptor (MR), aldosterone modulates the expression of membrane targets such as the subunits of the epithelial Na(+) channel, in combination with important signalling intermediates such as serum and glucocorticoid-regulated kinase-1. In addition, the rapid \\'non-genomic\\' activation of protein kinases and secondary messenger signalling cascades has also been detected in aldosterone-sensitive tissues of the nephron, distal colon and cardiovascular system. These rapid actions are variously described as being coupled to MR or to an as yet unidentified, membrane-associated aldosterone receptor. The rapidly activated signalling cascades add a level of fine-tuning to the activity of aldosterone-responsive membrane transporters and also modulate the aldosterone-induced changes in gene expression through receptor and transcription factor phosphorylation.

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

    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

  17. Comparison of the activation kinetics of the M3 acetylcholine receptor and a constitutively active mutant receptor in living cells.

    Hoffmann, Carsten; Nuber, Susanne; Zabel, Ulrike; Ziegler, Nicole; Winkler, Christiane; Hein, Peter; Berlot, Catherine H; Bünemann, Moritz; Lohse, Martin J

    2012-08-01

    Activation of G-protein-coupled receptors is the first step of the signaling cascade triggered by binding of an agonist. Here we compare the activation kinetics of the G(q)-coupled M(3) acetylcholine receptor (M(3)-AChR) with that of a constitutively active mutant receptor (M(3)-AChR-N514Y) using M(3)-AChR constructs that report receptor activation by changes in the fluorescence resonance energy transfer (FRET) signal. We observed a leftward shift in the concentration-dependent FRET response for acetylcholine and carbachol with M(3)-AChR-N514Y. Consistent with this result, at submaximal agonist concentrations, the activation kinetics of M(3)-AChR-N514Y were significantly faster, whereas at maximal agonist concentrations the kinetics of receptor activation were identical. Receptor deactivation was significantly faster with carbachol than with acetylcholine and was significantly delayed by the N514Y mutation. Receptor-G-protein interaction was measured by FRET between M(3)-AChR-yellow fluorescent protein (YFP) and cyan fluorescent protein (CFP)-Gγ(2). Agonist-induced receptor-G-protein coupling was of a time scale similar to that of receptor activation. As observed for receptor deactivation, receptor-G-protein dissociation was slower for acetylcholine than that for carbachol. Acetylcholine-stimulated increases in receptor-G-protein coupling of M(3)-AChR-N514Y reached only 12% of that of M(3)-AChR and thus cannot be kinetically analyzed. G-protein activation was measured using YFP-tagged Gα(q) and CFP-tagged Gγ(2). Activation of G(q) was significantly slower than receptor activation and indistinguishable for the two agonists. However, G(q) deactivation was significantly prolonged for acetylcholine compared with that for carbachol. Consistent with decreased agonist-stimulated coupling to G(q), agonist-stimulated G(q) activation by M(3)-AChR-N514Y was not detected. Taken together, these results indicate that the N514Y mutation produces constitutive activation of M(3

  18. Progesterone in pregnancy; receptor-ligand interaction and signaling pathways.

    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. P2X receptor-mediated ATP purinergic signaling in health and disease

    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

  20. Testin, a novel binding partner of the calcium-sensing receptor, enhances receptor-mediated Rho-kinase signalling

    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.

  1. Anti-inflammatory effects of cordycepin in lipopolysaccharide-stimulated RAW 264.7 macrophages through Toll-like receptor 4-mediated suppression of mitogen-activated protein kinases and NF-κB signaling pathways

    Choi YH

    2014-10-01

    Full Text Available Yung Hyun Choi,1,2 Gi-Young Kim,3 Hye Hyeon Lee4 1Department of Biochemistry, Dongeui University College of Korean Medicine, Busan, 2Anti-Aging Research Center and Blue-Bio Industry RIC, Dongeui University, Busan, 3Laboratory of Immunobiology, Department of Marine Life Sciences, Jeju National University, Jeju, 4Daegu Gyeongbuk Institute of Science and Technology, Daegu, Republic of Korea Abstract: Cordycepin is the main functional component of the Cordyceps species, which has been widely used in traditional Oriental medicine. This compound possesses many pharmacological properties, such as an ability to enhance immune function, as well as antioxidant, antiaging, and anticancer effects. In the present study, we investigated the anti-inflammatory effects of cordycepin using a murine macrophage RAW 264.7 cell model. Our data demonstrated that cordycepin suppressed production of proinflammatory mediators such as nitric oxide (NO and prostaglandin E2 by inhibiting inducible NO synthase and cyclooxygenase-2 gene expression. Cordycepin also inhibited the release of proinflammatory cytokines, including tumor necrosis factor-alpha and interleukin-1-beta, through downregulation of respective mRNA expression. In addition, pretreatment with cordycepin significantly inhibited lipopolysaccharide (LPS-induced phosphorylation of mitogen-activating protein kinases and attenuated nuclear translocation of NF-κB by LPS, which was associated with abrogation of inhibitor kappa B-alpha degradation. Furthermore, cordycepin potently inhibited the binding of LPS to macrophages and LPS-induced Toll-like receptor 4 and myeloid differentiation factor 88 expression. Taken together, the results suggest that the inhibitory effects of cordycepin on LPS-stimulated inflammatory responses in RAW 264.7 macrophages are associated with suppression of mitogen-activating protein kinases and activation of NF-κB by inhibition of the Toll-like receptor 4 signaling pathway. Keywords

  2. Vitamin D Receptor Signaling Inhibits Atherosclerosis in Mice

    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...

  3. Biased signaling of the angiotensin II type 1 receptor can be mediated through distinct mechanisms

    Bonde, Marie Mi; Hansen, Jonas Tind; Sanni, Samra Joke; Gammeltoft, Steen; Haunsø, Stig; Lyngsø, Christina; Hansen, Jakob Lerche

    2010-01-01

    Seven transmembrane receptors (7TMRs) can adopt different active conformations facilitating a selective activation of either G protein or β-arrestin-dependent signaling pathways. This represents an opportunity for development of novel therapeutics targeting selective biological effects of a given...

  4. Biased signaling of the angiotensin II type 1 receptor can be mediated through distinct mechanisms

    Bonde, Marie Mi; Hansen, Jonas Tind; Sanni, Samra Joke; Gammeltoft, Steen; Haunsø, Stig; Lyngsø, Christina; Hansen, Jakob Lerche

    2010-01-01

    Seven transmembrane receptors (7TMRs) can adopt different active conformations facilitating a selective activation of either G protein or ß-arrestin-dependent signaling pathways. This represents an opportunity for development of novel therapeutics targeting selective biological effects of a given...

  5. Shc adaptor proteins are key transducers of mitogenic signaling mediated by the G protein-coupled thrombin receptor

    Chen, Y; Grall, D; Salcini, A E; Pelicci, P G; Pouysségur, J; Van Obberghen-Schilling, E

    1996-01-01

    The serine protease thrombin activates G protein signaling systems that lead to Ras activation and, in certain cells, proliferation. Whereas the steps leading to Ras activation by G protein-coupled receptors are not well defined, the mechanisms of Ras activation by receptor tyrosine kinases have...... kinase activation, gene induction and cell growth. From these data, we conclude that Shc represents a crucial point of convergence between signaling pathways activated by receptor tyrosine kinases and G protein-coupled receptors....... recently been elucidated biochemically and genetically. The present study was undertaken to determine whether common signaling components are used by these two distinct classes of receptors. Here we report that the adaptor protein Shc, is phosphorylated on tyrosine residues following stimulation of the...

  6. Mechanism for the activation of glutamate receptors

    Scientists at the NIH have used a technique called cryo-electron microscopy to determine a molecular mechanism for the activation and desensitization of ionotropic glutamate receptors, a prominent class of neurotransmitter receptors in the brain and spina

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

    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

  8. Trans-Cinnamic Acid Increases Adiponectin and the Phosphorylation of AMP-Activated Protein Kinase through G-Protein-Coupled Receptor Signaling in 3T3-L1 Adipocytes

    Christina Kopp

    2014-02-01

    Full Text Available Adiponectin and intracellular 5'adenosine monophosphate-activated protein kinase (AMPK are important modulators of glucose and fat metabolism. Cinnamon exerts beneficial effects by improving insulin sensitivity and blood lipids, e.g., through increasing adiponectin concentrations and AMPK activation. The underlying mechanism is unknown. The Gi/Go-protein-coupled receptor (GPR 109A stimulates adiponectin secretion after binding its ligand niacin. Trans-cinnamic acid (tCA, a compound of cinnamon is another ligand. We hypothesize whether AMPK activation and adiponectin secretion by tCA is transmitted by GPR signaling. Differentiated 3T3-L1 cells were incubated with pertussis toxin (PTX, an inhibitor of Gi/Go-protein-coupling, and treated with different tCA concentrations. Treatment with tCA increased adiponectin and the pAMPK/AMPK ratio (p ≤ 0.001. PTX incubation abolished the increased pAMPK/AMPK ratio and adiponectin secretion. The latter remained increased compared to controls (p ≤ 0.002. tCA treatment stimulated adiponectin secretion and AMPK activation; the inhibitory effect of PTX suggests GPR is involved in tCA stimulated signaling.

  9. Functionally biased signalling properties of 7TM receptors - opportunities for drug development for the ghrelin receptor

    Sivertsen, B; Holliday, N; Madsen, A N; Holst, B

    2013-01-01

    UNLABELLED: The ghrelin receptor is a 7 transmembrane (7TM) receptor involved in a variety of physiological functions including growth hormone secretion, increased food intake and fat accumulation as well as modulation of reward and cognitive functions. Because of its important role in metabolism...... several different signalling pathways including Gαq , Gαi/o , Gα12/13 and arrestin recruitment. These multiple signalling pathways allow for functionally biased signalling, where one signalling pathway may be favoured over another either by selective ligands or through mutations in the receptor. In the...... on the properties of the agonist. Importantly, ligands with such biased signalling properties may allow the development of drugs that selectively modulate only the therapeutically relevant physiological functions, thereby decreasing the risk of side effects. LINKED ARTICLES: This article is part of a...

  10. Toll-like receptors: cellular signal transducers for exogenous molecular patterns causing immune responses.

    Kirschning, C J; Bauer, S

    2001-09-01

    Innate immunity initiates protection of the host organism against invasion and subsequent multiplication of microbes by specific recognition. Germ line-encoded receptors have been identified for microbial products such as mannan, lipopeptide, peptidoglycan (PGN), lipoteichoic acid (LTA), lipopolysaccharide (LPS), and CpG-DNA. The Drosophila Toll protein has been shown to be involved in innate immune response of the adult fruitfly. Members of the family of Toll-like receptors (TLRs) in vertebrates have been implicated as pattern recognition receptors (PRRs). Ten TLRs are known and six of these have been demonstrated to mediate cellular activation by distinct microbial products. TLR4 has been implicated as activator of adaptive immunity, and analysis of systemic LPS responses in mice led to the identification of LPS-resistant strains instrumental in its identification as a transmembrane LPS signal transducer. Structural similarities between TLRs and receptor molecules involved in immune responses such as CD14 and the IL-1 receptors (IL-1Rs), as well as functional analysis qualified TLR2 as candidate receptor for LPS and other microbial products. Targeted disruption of the TLR9 gene in mice led to identification of TLR9 as CpG-DNA signal transducer. Involvement of TLR5 in cell activation by bacterial flagellin has been demonstrated. Further understanding of recognition and cellular signaling activated through the ancient host defense system represented by Toll will eventually lead to means for its therapeutic modulation. PMID:11680785

  11. ATAR, a novel tumor necrosis factor receptor family member, signals through TRAF2 and TRAF5.

    Hsu, H; Solovyev, I; Colombero, A; Elliott, R; Kelley, M; Boyle, W J

    1997-05-23

    Members of tumor necrosis factor receptor (TNFR) family signal largely through interactions with death domain proteins and TRAF proteins. Here we report the identification of a novel TNFR family member ATAR. Human and mouse ATAR contain 283 and 276 amino acids, respectively, making them the shortest known members of the TNFR superfamily. The receptor is expressed mainly in spleen, thymus, bone marrow, lung, and small intestine. The intracellular domains of human and mouse ATAR share only 25% identity, yet both interact with TRAF5 and TRAF2. This TRAF interaction domain resides at the C-terminal 20 amino acids. Like most other TRAF-interacting receptors, overexpression of ATAR activates the transcription factor NF-kappaB. Co-expression of ATAR with TRAF5, but not TRAF2, results in synergistic activation of NF-kappaB, suggesting potentially different roles for TRAF2 and TRAF5 in post-receptor signaling. PMID:9153189

  12. Mitochondrial H2O2 as an enable signal for triggering autophosphorylation of insulin receptor in neurons

    Persiyantseva, Nadezhda A; Storozhevykh, Tatiana P; Senilova, Yana E; Gorbacheva, Lubov R; Pinelis, Vsevolod G.; Pomytkin, Igor A

    2013-01-01

    Background: Insulin receptors are widely distributed in the brain, where they play roles in synaptic function, memory formation, and neuroprotection. Autophosphorylation of the receptor in response to insulin stimulation is a critical step in receptor activation. In neurons, insulin stimulation leads to a rise in mitochondrial H2O2 production, which plays a role in receptor autophosphorylation. However, the kinetic characteristics of the H2O2 signal and its functional relationships with the i...

  13. Cell death sensitization of leukemia cells by opioid receptor activation

    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

  14. Mislocalized activation of oncogenic RTKs switches downstream signaling outcomes

    Choudhary, Chuna Ram; Olsen, Jesper V; Brandts, Christian;

    2009-01-01

    Inappropriate activation of oncogenic kinases at intracellular locations is frequently observed in human cancers, but its effects on global signaling are incompletely understood. Here, we show that the oncogenic mutant of Flt3 (Flt3-ITD), when localized at the endoplasmic reticulum (ER), aberrant...... patterns of the receptor itself. Thus, intracellular activation of RTKs by oncogenic mutations in the biosynthetic route may exploit cellular architecture to initiate aberrant signaling cascades, thus evading negative regulation....

  15. Nanoconjugation prolongs endosomal signaling of the epidermal growth factor receptor and enhances apoptosis

    Wu, L.; Xu, F.; Reinhard, B. M.

    2016-07-01

    It is becoming increasingly clear that intracellular signaling can be subject to strict spatial control. As the covalent attachment of a signaling ligand to a nanoparticle (NP) impacts ligand-receptor binding, uptake, and trafficking, nanoconjugation provides new opportunities for manipulating intracellular signaling in a controlled fashion. To establish the effect of nanoconjugation on epidermal growth factor (EGF) mediated signaling, we investigate here the intracellular fate of nanoconjugated EGF (NP-EGF) and its bound receptor (EGFR) by quantitative correlated darkfield/fluorescence microscopy and density-based endosomal fractionation. We demonstrate that nanoconjugation prolongs the dwell time of phosphorylated receptors in the early endosomes and that the retention of activated EGFR in the early endosomes is accompanied by an EGF 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.It is becoming increasingly clear that intracellular signaling can be subject to strict spatial control. As the covalent attachment of a signaling ligand to a nanoparticle (NP) impacts ligand-receptor binding, uptake, and trafficking, nanoconjugation provides new opportunities for manipulating intracellular signaling in a controlled fashion. To establish the effect of nanoconjugation on epidermal growth factor (EGF) mediated signaling, we investigate here the intracellular fate of nanoconjugated EGF (NP-EGF) and its bound receptor (EGFR) by quantitative correlated darkfield/fluorescence microscopy and density-based endosomal fractionation. We demonstrate that nanoconjugation prolongs the dwell time of phosphorylated receptors in the early endosomes and that the retention of activated EGFR in the early endosomes is accompanied by an EGF

  16. Inhibition of Nuclear Receptor Signalling by Poly(ADP-Ribose) Polymerase

    Miyamoto, Takahide; Kakizawa, Tomoko; Hashizume, Kiyoshi

    1999-01-01

    Mammalian poly(ADP-ribose) polymerase (PARP) is a nuclear chromatin-associated protein with a molecular mass of 114 kDa that catalyzes the transfer of ADP-ribose units from NAD+ to nuclear proteins that are located within chromatin. We report here the identification of a novel property of PARP as a modulator of nuclear receptor signalling. PARP bound directly to retinoid X receptors (RXR) and repressed ligand-dependent transcriptional activities mediated by heterodimers of RXR and thyroid hor...

  17. Identification of intracellular domains in the growth hormone receptor involved in signal transduction

    Billestrup, N.; Allevato, G.; Moldrup, A. [Hagedorn Research Lab., Gentofte (Denmark)] [and others

    1994-12-31

    The growth hormone (GH) receptor belongs to the GH/prolactin/cytokine super-family of receptors. The signal transduction mechanism utilized by this class of receptors remains largely unknown. In order to identify functional domains in the intracellular region of the GH receptor we generated a number of GH receptor mutants and analyzed their function after transfection into various cell lines. A truncated GH receptor missing 184 amino acids at the C-terminus was unable to medite GH effects on transcription of the Spi 2.1 and insulin genes. However, this mutant was fully active in mediating GH-stimulated metabolic effects such as protein synthesis and lipolysis. Furthermore, this mutant GH receptor internalized rapidly following GH binding. Another truncated GH receptor lacking all but five amino acids of the cytoplasmic domain could not mediate any effects of GH nor did it internalize. Deletion of the proline-rich region or changing the four prolines to alanines also resulted in a GH receptor deficient in signaling. Mutation of phenylalanine 346 to alanine resulted in a GH receptor which did not internalize rapidly; however, this mutant GH receptor was capable of mediating GH-stimulated transcription as well as metabolic effects. These results indicate that the intracellular part of the GH receptor can be divided into at least three functional domains: (1) for transcriptional activity, two domains are involved, one located in the C-terminal 184 amino acids and the other in the proline-rich domain; (2) for metabolic effects, a domain located in or near the proline-rich region is of importance; and (3) for internalization, phenylalanine 346 is necessary. 28 refs., 1 fig.

  18. Mechanism of inhibition of growth hormone receptor signaling by suppressor of cytokine signaling proteins

    Hansen, J A; Lindberg, K; Hilton, D J; Nielsen, Jens Høiriis; Billestrup, N

    1999-01-01

    In this study we have investigated the role of suppressor of cytokine signaling (SOCS) proteins in GH receptor-mediated signaling. GH-induced transcription was inhibited by SOCS-1 and SOCS-3, while SOCS-2 and cytokine inducible SH2-containing protein (CIS) had no effect By using chimeric SOCS pro...

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

    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.

  20. The insulin receptor activation process involves localized conformational changes.

    Baron, V; Kaliman, P; Gautier, N; Van Obberghen, E

    1992-11-15

    The molecular process by which insulin binding to the receptor alpha-subunit induces activation of the receptor beta-subunit with ensuing substrate phosphorylation remains unclear. In this study, we aimed at approaching this molecular mechanism of signal transduction and at delineating the cytoplasmic domains implied in this process. To do this, we used antipeptide antibodies to the following sequences of the receptor beta-subunit: (i) positions 962-972 in the juxtamembrane domain, (ii) positions 1247-1261 at the end of the kinase domain, and (iii) positions 1294-1317 and (iv) positions 1309-1326, both in the receptor C terminus. We have previously shown that insulin binding to its receptor induces a conformational change in the beta-subunit C terminus. Here, we demonstrate that receptor autophosphorylation induces an additional conformational change. This process appears to be distinct from the one produced by ligand binding and can be detected in at least three different beta-subunit regions: the juxtamembrane domain, the kinase domain, and the C terminus. Hence, the cytoplasmic part of the receptor beta-subunit appears to undergo an extended conformational change upon autophosphorylation. By contrast, the insulin-induced change does not affect the juxtamembrane domain 962-972 nor the kinase domain 1247-1261 and may be limited to the receptor C terminus. Further, we show that the hormone-dependent conformational change is maintained in a kinase-deficient receptor due to a mutation at lysine 1018. Therefore, during receptor activation, the ligand-induced change could precede ATP binding and receptor autophosphorylation. We propose that insulin binding leads to a transient receptor form that may allow ATP binding and, subsequently, autophosphorylation. The second conformational change could unmask substrate-binding sites and stabilize the receptor in an active conformation. PMID:1331080

  1. Direct Modulation of Heterotrimeric G Protein-coupled Signaling by a Receptor Kinase Complex.

    Tunc-Ozdemir, Meral; Urano, Daisuke; Jaiswal, Dinesh Kumar; Clouse, Steven D; Jones, Alan M

    2016-07-01

    Plants and some protists have heterotrimeric G protein complexes that activate spontaneously without canonical G protein-coupled receptors (GPCRs). In Arabidopsis, the sole 7-transmembrane regulator of G protein signaling 1 (AtRGS1) modulates the G protein complex by keeping it in the resting state (GDP-bound). However, it remains unknown how a myriad of biological responses is achieved with a single G protein modulator. We propose that in complete contrast to G protein activation in animals, plant leucine-rich repeat receptor-like kinases (LRR RLKs), not GPCRs, provide this discrimination through phosphorylation of AtRGS1 in a ligand-dependent manner. G protein signaling is directly activated by the pathogen-associated molecular pattern flagellin peptide 22 through its LRR RLK, FLS2, and co-receptor BAK1. PMID:27235398

  2. Damage-associated molecular pattern activated Toll-like receptor 4 signalling modulates blood pressure in L-NAME-induced hypertension

    Sollinger, D.; Eissler, R.; Lorenz, S.; Strand, S.; Chmielewski, S.; Aoqui, C.; Schmaderer, Ch.; Bluyssen, H.; Zicha, Josef; Witzke, O.; Scherer, E.; Lutz, J.; Heemann, U.; Baumann, M.

    2014-01-01

    Roč. 101, č. 3 (2014), s. 464-472. ISSN 0008-6363 R&D Projects: GA ČR(CZ) GA305/09/0336 Institutional research plan: CEZ:AV0Z50110509 Institutional support: RVO:67985823 Keywords : innate immunity * Toll-like receptors * reactive oxygen species * vascular contractility Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery Impact factor: 5.940, year: 2014

  3. Ligand-Dependent Actions of the Vitamin D Receptor Are Required for Activation of TGF-β Signaling during the Inflammatory Response to Cutaneous Injury

    Luderer, Hilary F; Nazarian, Rosalynn M.; Zhu, Eric D.; Demay, Marie B.

    2012-01-01

    The vitamin D receptor (VDR) has both 1,25-dihydroxyvitamin D-dependent and -independent actions in the epidermis. Ligand-dependent actions of the VDR have been shown to promote keratinocyte differentiation and to regulate formation of the epidermal barrier. In contrast, the actions of the VDR that regulate postmorphogenic hair cycling do not require 1,25-dihydroxyvitamin D. The VDR also has immunomodulatory actions that are dependent on its ligand, 1,25-dihydroxyvitamin D. To determine wheth...

  4. Wnt signaling and the activation of myogenesis in mammals.

    Cossu, G; Borello, U

    1999-12-15

    In the amniote embryos, specification of skeletal myoblasts occurs in the paraxial mesoderm in response to a number of signaling molecules produced by neighboring tissues such as neural tube, notochord and dorsal ectoderm. Candidate molecules for this complex signaling activity include Sonic hedgehog, Wnts and Noggin as positive activators and BMP4 as a possible inhibitor. Recently, the receptors and the post-receptor pathways for Sonic hedgehog and Wnts have been characterized, and this has opened up the possibility of linking these signaling events to the activation of myogenic regulatory factor genes such as Myf5 and MyoD and functionally related genes such as Pax3. Here we focus on the role of Wnts, their putative receptors Frizzled and the soluble antagonist Frzb1 in regulating mammalian myogenesis. Although it is becoming evident that the signaling downstream of Frizzled receptors is much more complex than anticipated, it is conceivable that it may lead to transcriptional activation of Myf5 and MyoD and to initiation of myogenesis. However, the fact that both Wnts and Sonic hedgehog have a strong effect on cell proliferation and survival suggests that they may contribute to the overall process of myogenesis by a combination of these different biological activities. PMID:10601008

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

    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.

  6. Nuclear receptor-dependent bile acid signaling is required for normal liver regeneration.

    Huang, Wendong; Ma, Ke; Zhang, Jun; Qatanani, Mohammed; Cuvillier, James; Liu, Jun; Dong, Bingning; Huang, Xiongfei; Moore, David D

    2006-04-14

    Liver mass depends on one or more unidentified humoral signals that drive regeneration when liver functional capacity is diminished. Bile acids are important liver products, and their levels are tightly regulated. Here, we identify a role for nuclear receptor-dependent bile acid signaling in normal liver regeneration. Elevated bile acid levels accelerate regeneration, and decreased levels inhibit liver regrowth, as does the absence of the primary nuclear bile acid receptor FXR. We propose that FXR activation by increased bile acid flux is a signal of decreased functional capacity of the liver. FXR, and possibly other nuclear receptors, may promote homeostasis not only by regulating expression of appropriate metabolic target genes but also by driving homeotrophic liver growth. PMID:16614213

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

    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 ...

  8. Alcohol,nutrition and liver cancer:Role of Toll-like receptor signaling

    Samuel; W; French; Joan; Oliva; Barbara; A; French; Fawzia; Bardag-Gorce

    2010-01-01

    This article reviews the evidence that ties the development of hepatocellular carcinoma (HCC) to the natural immune pro-inflammatory response to chronic liver disease, with a focus on the role of Toll-like receptor (TLR) signaling as the mechanism of liver stem cell/progenitor transformation to HCC. Two exemplary models of this phenomenon are reviewed in detail. One model applies chronic ethanol/lipopolysaccharide feeding to the activated TLR4 signaling pathway. The other applies chronic feeding of a carcin...

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

    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

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

    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

  11. NMDA Receptor Activation by Spontaneous Glutamatergic Neurotransmission

    Espinosa, Felipe; Kavalali, Ege T.

    2009-01-01

    Under physiological conditions N-methyl-d-aspartate (NMDA) receptor activation requires coincidence of presynaptic glutamate release and postsynaptic depolarization due to the voltage-dependent block of these receptors by extracellular Mg2+. Therefore spontaneous neurotransmission in the absence of action potential firing is not expected to lead to significant NMDA receptor activation. Here we tested this assumption in layer IV neurons in neocortex at their resting membrane potential (approxi...

  12. Non-Ligand-Induced Dimerization is Sufficient to Initiate the Signalling and Endocytosis of EGF Receptor

    George Kourouniotis

    2016-07-01

    Full Text Available The binding of epidermal growth factor (EGF to EGF receptor (EGFR stimulates cell mitogenesis and survival through various signalling cascades. EGF also stimulates rapid EGFR endocytosis and its eventual degradation in lysosomes. The immediate events induced by ligand binding include receptor dimerization, activation of intrinsic tyrosine kinase and autophosphorylation. However, in spite of intensified efforts, the results regarding the roles of these events in EGFR signalling and internalization is still very controversial. In this study, we constructed a chimeric EGFR by replacing its extracellular domain with leucine zipper (LZ and tagged a green fluorescent protein (GFP at its C-terminus. We showed that the chimeric LZ-EGFR-GFP was constitutively dimerized. The LZ-EGFR-GFP dimer autophosphorylated each of its five well-defined C-terminal tyrosine residues as the ligand-induced EGFR dimer does. Phosphorylated LZ-EGFR-GFP was localized to both the plasma membrane and endosomes, suggesting it is capable of endocytosis. We also showed that LZ-EGFR-GFP activated major signalling proteins including Src homology collagen-like (Shc, extracellular signal-regulated kinase (ERK and Akt. Moreover, LZ-EGFR-GFP was able to stimulate cell proliferation. These results indicate that non-ligand induced dimerization is sufficient to activate EGFR and initiate cell signalling and EGFR endocytosis. We conclude that receptor dimerization is a critical event in EGF-induced cell signalling and EGFR endocytosis.

  13. Non-Ligand-Induced Dimerization is Sufficient to Initiate the Signalling and Endocytosis of EGF Receptor

    Kourouniotis, George; Wang, Yi; Pennock, Steven; Chen, Xinmei; Wang, Zhixiang

    2016-01-01

    The binding of epidermal growth factor (EGF) to EGF receptor (EGFR) stimulates cell mitogenesis and survival through various signalling cascades. EGF also stimulates rapid EGFR endocytosis and its eventual degradation in lysosomes. The immediate events induced by ligand binding include receptor dimerization, activation of intrinsic tyrosine kinase and autophosphorylation. However, in spite of intensified efforts, the results regarding the roles of these events in EGFR signalling and internalization is still very controversial. In this study, we constructed a chimeric EGFR by replacing its extracellular domain with leucine zipper (LZ) and tagged a green fluorescent protein (GFP) at its C-terminus. We showed that the chimeric LZ-EGFR-GFP was constitutively dimerized. The LZ-EGFR-GFP dimer autophosphorylated each of its five well-defined C-terminal tyrosine residues as the ligand-induced EGFR dimer does. Phosphorylated LZ-EGFR-GFP was localized to both the plasma membrane and endosomes, suggesting it is capable of endocytosis. We also showed that LZ-EGFR-GFP activated major signalling proteins including Src homology collagen-like (Shc), extracellular signal-regulated kinase (ERK) and Akt. Moreover, LZ-EGFR-GFP was able to stimulate cell proliferation. These results indicate that non-ligand induced dimerization is sufficient to activate EGFR and initiate cell signalling and EGFR endocytosis. We conclude that receptor dimerization is a critical event in EGF-induced cell signalling and EGFR endocytosis. PMID:27463710

  14. Inhibition of cytokines and JAK-STAT activation by distinct signaling pathways.

    Sengupta, T K; Schmitt, E M; Ivashkiv, L B

    1996-01-01

    An important component of cytokine regulation of cell growth and differentiation is rapid transcriptional activation of genes by the JAK-STAT (signal transducer and activator of transcription) signaling pathway. Ligation of cytokine receptors results in tyrosine phosphorylation and activation of receptor-associated Jak protein tyrosine kinases and cytoplasmic STAT transcription factors, which then translocate to the nucleus. We describe the interruption of cytokine triggered JAK-STAT signals ...

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

    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...

  16. Integrin αvβ3 and CD44 pathways in metastatic prostate cancer cells support osteoclastogenesis via a Runx2/Smad 5/receptor activator of NF-κB ligand signaling axis

    Gupta Aditi

    2012-09-01

    Full Text Available Abstract Background Bone loss and pathological fractures are common skeletal complications associated with androgen deprivation therapy and bone metastases in prostate cancer patients. We have previously demonstrated that prostate cancer cells secrete receptor activator of NF-kB ligand (RANKL, a protein essential for osteoclast differentiation and activation. However, the mechanism(s by which RANKL is produced remains to be determined. The objective of this study is to gain insight into the molecular mechanisms controlling RANKL expression in metastatic prostate cancer cells. Results We show here that phosphorylation of Smad 5 by integrin αvβ3 and RUNX2 by CD44 signaling, respectively, regulates RANKL expression in human-derived PC3 prostate cancer cells isolated from bone metastasis. We found that RUNX2 intranuclear targeting is mediated by phosphorylation of Smad 5. Indeed, Smad5 knock-down via RNA interference and inhibition of Smad 5 phosphorylation by an αv inhibitor reduced RUNX2 nuclear localization and RANKL expression. Similarly, knockdown of CD44 or RUNX2 attenuated the expression of RANKL. As a result, conditioned media from these cells failed to support osteoclast differentiation in vitro. Immunohistochemistry analysis of tissue microarray sections containing primary prostatic tumor (grade2-4 detected predominant localization of RUNX2 and phosphorylated Smad 5 in the nuclei. Immunoblotting analyses of nuclear lysates from prostate tumor tissue corroborate these observations. Conclusions Collectively, we show that CD44 signaling regulates phosphorylation of RUNX2. Localization of RUNX2 in the nucleus requires phosphorylation of Smad-5 by integrin αvβ3 signaling. Our results suggest possible integration of two different pathways in the expression of RANKL. These observations imply a novel mechanistic insight into the role of these proteins in bone loss associated with bone metastases in patients with prostate cancer.

  17. Characterisation of the tissue-specific expression, pharmacology and signalling cascades activated by chicken GnRH receptor subtypes suggested evolutionary specialisation of type III cGnRH receptor function

    Joseph, Nerine Theresa

    2010-01-01

    Variant GnRH ligand and receptor subtypes have been identified in a number of non-mammalian vertebrate species, however research into avian species GnRH systems is lacking. Two isoforms of GnRH are present in the domestic chicken, the evolutionarily conserved GnRH-II and diverged cGnRH-I. The expression of two GnRH ligands parallels the expression of two chicken GnRH receptor subtypes; cGnRH-R-I and the novel cGnRH-R-III. The occurrence of two isoforms of the receptor in the chicken raises qu...

  18. Growth Hormone-induced JAK2 Signaling and GH Receptor Down-regulation: Role of GH Receptor Intracellular Domain Tyrosine Residues

    Deng, Luqin; Jiang, Jing; Frank, Stuart J.

    2012-01-01

    GH receptor (GHR) mediates important somatogenic and metabolic effects of GH. A thorough understanding of GH action requires intimate knowledge of GHR activation mechanisms, as well as determinants of GH-induced receptor down-regulation. We previously demonstrated that a GHR mutant in which all intracellular tyrosine residues were changed to phenylalanine was defective in its ability to activate signal transducer and activator of transcription (STAT)5 and deficient in GH-induced down-regulati...

  19. Abelson tyrosine kinase links PDGFbeta receptor activation to cytoskeletal regulation of NMDA receptors in CA1 hippocampal neurons

    Beazely Michael A

    2008-12-01

    Full Text Available Abstract Background We have previously demonstrated that PDGF receptor activation indirectly inhibits N-methyl-D-aspartate (NMDA currents by modifying the cytoskeleton. PDGF receptor ligand is also neuroprotective in hippocampal slices and cultured neurons. PDGF receptors are tyrosine kinases that control a variety of signal transduction pathways including those mediated by PLCγ. In fibroblasts Src and another non-receptor tyrosine kinase, Abelson kinase (Abl, control PDGF receptor regulation of cytoskeletal dynamics. The mechanism whereby PDGF receptor regulates cytoskeletal dynamics in central neurons remains poorly understood. Results Intracellular applications of active Abl, but not heat-inactivated Abl, decreased NMDA-evoked currents in isolated hippocampal neurons. This mimics the effects of PDGF receptor activation in these neurons. The Abl kinase inhibitor, STI571, blocked the inhibition of NMDA currents by Abl. We demonstrate that PDGF receptors can activate Abl kinase in hippocampal neurons via mechanisms similar to those observed previously in fibroblasts. Furthermore, PDGFβ receptor activation alters the subcellular localization of Abl. Abl kinase is linked to actin cytoskeletal dynamics in many systems. We show that the inhibition of NMDA receptor currents by Abl kinase is blocked by the inclusion of the Rho kinase inhibitor, Y-27632, and that activation of Abl correlates with an increase in ROCK tyrosine phosphorylation. Conclusion This study demonstrates that PDGFβ receptors act via an interaction with Abl kinase and Rho kinase to regulated cytoskeletal regulation of NMDA receptor channels in CA1 pyramidal neurons.

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

    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

  1. Kappa-opioid receptor signaling and brain reward function

    Bruijnzeel, Adrie W.

    2009-01-01

    The dynorphin-like peptides have profound effects on the state of the brain reward system and human and animal behavior. The dynorphin-like peptides affect locomotor activity, food intake, sexual behavior, anxiety-like behavior, and drug intake. Stimulation of kappa-opioid receptors, the endogenous receptor for the dynorphin-like peptides, inhibits dopamine release in the striatum (nucleus accumbens and caudate putamen) and induces a negative mood state in humans and animals. The administrati...

  2. Targeting VEGF signalling via the neuropilin co-receptor.

    Djordjevic, S.; Driscoll, P. C.

    2013-01-01

    The blockade of tumour vascularisation and angiogenesis continues to be a focus for drug development in oncology and other pathologies. Historically, targeting vascular endothelial growth factor (VEGF) activity and its association with VEGF receptors (VEGFRs) has represented the most promising line of attack. More recently, the recognition that VEGFR co-receptors, neuropilin-1 and -2 (NRP1 and NRP2), are also engaged by specific VEGF isoforms in tandem with the VEGFRs has expanded the landsca...

  3. Endosome-mediated retrograde axonal transport of P2X3 receptor signals in primary sensory neurons

    Xu-Qiao Chen; BinWang; Chengbiao Wu; Jin Pan; Bo Yuan; Yuan-Yuan Su; Xing-Yu Jiang; Xu Zhang; Lan Bao

    2012-01-01

    Neurotrophins and their receptors adopt signaling endosomes to transmit retrograde signals.However,the mechanisms of retrograde signaling for other ligand/receptor systems are poorly understood.Here,we report that the signals of the purinergic (P)2X3 receptor,an ATP-gated ion channel are retrogradely transported in dorsal root ganglion (DRG) neuron axons.We found that Rab5,a small GTPase,controls the early sorting of P2X3 receptors into endosomes,while Rab7 mediates the fast retrograde transport of P2X3 receptors.Intraplantar injection and axonal application into the microfluidic chamber of α,β-methylene-ATP (α,β-MeATP),a P2X selective agonist,enhanced the endocytosis and retrograde transport of P2X3 receptors.The α,β-MeATP-induced Ca2+ influx activated a pathway comprised of protein kinase C,rat sarcoma viral oncogene and extracellular signal-regulated protein kinase (ERK),which associated with endocytic P2X3 receptors to form signaling endosomes.Disruption of the lipid rafts abolished the α,β-MeATP-induced ERK phosphorylation,endocytosis and retrograde transport of P2X3 receptors.Furthermore,treatment of peripheral axons with α,β-MeATP increased the activation level of ERK and cAMP response element-binding protein in the cell bodies of DRG neurons and enhanced neuronal excitability.Impairment of either microtubule-based axonal transport in vivo or dynein function in vitro blocked α,β-MeATP-induced retrograde signals.These results indicate that P2X3 receptor-activated signals are transmitted via retrogradely transported endosomes in primary sensory neurons and provide a novel signaling mechanism for ligand-gated channels.

  4. Hormone activation of baculovirus expressed progesterone receptors.

    Elliston, J F; Beekman, J M; Tsai, S Y; O'Malley, B W; Tsai, M J

    1992-03-15

    Human and chicken progesterone receptors (A form) were overproduced in a baculovirus expression system. These recombinant progesterone receptors were full-length bound progesterone specifically and were recognized by monoclonal antibodies, AB52 and PR22, specific for human and chicken progesterone receptor, respectively. In gel retardation studies, binding of recombinant human and chicken progesterone receptors to their progesterone response element (PRE) was specific and was enhanced in the presence of progesterone. Binding of human progesterone receptor to the PRE was also enhanced in the presence of the antiprogestin, RU486, but very little effect was observed in the presence of estradiol, dexamethasone, testosterone, and vitamin D. In our cell-free transcription system, human progesterone receptor induced transcription in a receptor-dependent and hormone-activable manner. Receptor-stimulated transcription required the presence of the PRE in the test template and could be specifically inhibited by excess PRE oligonucleotides. Furthermore, chicken progesterone receptor also induced in vitro transcription in a hormone-activable manner. These results demonstrate that steroid receptors overexpressed in a baculovirus expression system are functional and exhibit steroid-responsive binding and transcription. These observations support our present understanding of the mechanism of steroid receptor-regulated gene expression and provide a technological format for studies of the role of hormone and antihormone in altering gene expression. PMID:1544902

  5. Glucose Enhances Leptin Signaling through Modulation of AMPK Activity

    Haoran Su; Lin Jiang; Christin Carter-Su; Liangyou Rui

    2012-01-01

    Leptin exerts its action by binding to and activating the long form of leptin receptors (LEPRb). LEPRb activates JAK2 that subsequently phosphorylates and activates STAT3. The JAK2/STAT3 pathway is required for leptin control of energy balance and body weight. Defects in leptin signaling lead to leptin resistance, a primary risk factor for obesity. Body weight is also regulated by nutrients, including glucose. Defects in glucose sensing also contribute to obesity. Here we report crosstalk bet...

  6. ABA receptors: The START of a new paradigm in phytohormone signalling

    Klingler, John

    2010-06-03

    The phytohormone abscisic acid (ABA) plays a central role in plant development and in plant adaptation to both biotic and abiotic stressors. In recent years, knowledge of ABA metabolism and signal transduction has advanced rapidly to provide detailed glimpses of the hormone\\'s activities at the molecular level. Despite this progress, many gaps in understanding have remained, particularly at the early stages of ABA perception by the plant cell. The search for an ABA receptor protein has produced multiple candidates, including GCR2, GTG1, and GTG2, and CHLH. In addition to these candidates, in 2009 several research groups converged on a novel family of Arabidopsis proteins that bind ABA, and thereby interact directly with a class of protein phosphatases that are well known as critical players in ABA signal transduction. The PYR/PYL/RCAR receptor family is homologous to the Bet v 1-fold and START domain proteins. It consists of 14 members, nearly all of which appear capable of participating in an ABA receptor-signal complex that responds to the hormone by activating the transcription of ABA-responsive genes. Evidence is provided here that PYR/PYL/RCAR receptors can also drive the phosphorylation of the slow anion channel SLAC1 to provide a fast and timely response to the ABA signal. Crystallographic studies have vividly shown the mechanics of ABA binding to PYR/PYL/RCAR receptors, presenting a model that bears some resemblance to the binding of gibberellins to GID1 receptors. Since this ABA receptor family is highly conserved in crop species, its discovery is likely to usher a new wave of progress in the elucidation and manipulation of plant stress responses in agricultural settings. © 2010 The Author(s).

  7. Intestinal epithelial vitamin D receptor signaling inhibits experimental colitis

    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...

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

    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...

  9. Molecular determinants and feedback circuits regulating type 2 CRH receptor signal integration.

    Markovic, Danijela; Punn, Anu; Lehnert, Hendrik; Grammatopoulos, Dimitris K

    2011-05-01

    In most target tissues, the adenylyl cyclase/cAMP/PKA, the extracellular signal regulated kinase and the protein kinase B/Akt are the main pathways employed by the type 2 corticotropin-releasing hormone receptor to mediate the biological actions of urocortins (Ucns) and CRH. To decipher the molecular determinants of CRH-R2 signaling, we studied the signaling pathways in HEK293 cells overexpressing recombinant human CRH-R2β receptors. Use of specific kinase inhibitors showed that the CRH-R2β cognate agonist, Ucn 2, activated extracellular signal regulated kinase in a phosphoinositide 3-kinase and cyclic adenosine monophosphate/PKA-dependent manner with contribution from Epac activation. Ucn 2 also induced PKA-dependent association between AKAP250 and CRH-R2β that appeared to be necessary for extracellular signal regulated kinase activation. PKB/Akt activation was also mediated via pertussis toxin-sensitive G-proteins and PI3-K activation but did not require cAMP/PKA, Epac or protein kinase C for optimal activation. Potential feedback mechanisms that target the CRH-R2β itself and modulate receptor trafficking and endocytosis were also investigated. Indeed, our results suggested that inhibition of either PKA or extracellular signal regulated kinase pathway accelerates CRH-R2β endocytosis. Furthermore, Ucn 2-activated extracellular signal regulated kinase appeared to target β-arrestin1 and modulate, through phosphorylation at Ser412, β-arrestin1 translocation to the plasma membrane and CRH-R2β internalization kinetics. Loss of this "negative feedback" mechanism through inhibition of the extracellular signal regulated kinase activity resulted in significant attenuation of Ucn 2-induced cAMP response, whereas Akt phosphorylation was not affected by altered receptor endocytosis. These findings reveal a complex interplay between the signaling molecules that allow "fine-tuning" of CRH-R2β functional responses and regulate signal integration. This article is part of

  10. INHIBITION OF PHOSPHATASE ACTIVITY MEDIATES EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR) SIGNALING IN HUMAN AIRWAY EPITHELIAL CELLS (HAEC) EXPOSED TO ZN2+

    A number of studies have implicated zinc in the toxicity of ambient particulate matter (PM) inhalation. We previously showed that exposure to metal-laden particulate matter inhibits protein tyrosine phosphatase activity in HAEC and leads to Src-dependent activation of EGFR sign...

  11. Dual-color dynamic tracking of GM-CSF receptors/JAK2 kinases signaling activation using temporal focusing multiphoton fluorescence excitation and astigmatic imaging.

    Chien, Fan-Ching; Lien, Chi-Hsiang; Dai, Yang-Hong

    2015-11-30

    The dual-color dynamic particle tracking approach that uses temporal focusing multiphoton fluorescence excitation and two-channel astigmatic imaging is utilized to track molecular trajectories in three dimensions to explore molecular interactions. Images of two fluorophores were obtained to extract their positions by optical sectioning excitation using a fast temporal focusing multiphoton excitation microscope (TFMPEM) and by the simultaneous collection of data in two channels. The presented pair of cylindrical lenses, which was used to adjust the astigmatism effect with the minimum shifting of the imaging plane, was more feasible and flexible than single cylindrical lens for aligning two separate detection channels in astigmatic imaging. The lateral and axial positioning resolutions were observed to be approximately 9-13 nm and 23-30 nm respectively, for the two fluorescence channels. The dynamic movement and binding behavior of clusters of GM-CSF receptors and JAK2 kinases in HeLa cells in the presence of GM-CSF ligands were observed. Therefore, the proposed dual-color tracking strategy is useful for the dynamic study of molecular interactions in living specimens with a fast frame rate, less photobleaching, better penetration depth, and minimum optical trapping force. PMID:26698726

  12. DMPD: Toll-like receptor signal transduction. [Dynamic Macrophage Pathway CSML Database

    Full Text Available 17934330 Toll-like receptor signal transduction. Krishnan J, Selvarajoo K, Tsuchiya... M, Lee G, Choi S. Exp Mol Med. 2007 Aug 31;39(4):421-38. (.png) (.svg) (.html) (.csml) Show Toll-like receptor signal tran...sduction. PubmedID 17934330 Title Toll-like receptor signal transduction. Authors Krishnan J,

  13. DMPD: Lysophospholipid receptors: signaling and biology. [Dynamic Macrophage Pathway CSML Database

    Full Text Available 15189145 Lysophospholipid receptors: signaling and biology. Ishii I, Fukushima N, Y...e X, Chun J. Annu Rev Biochem. 2004;73:321-54. (.png) (.svg) (.html) (.csml) Show Lysophospholipid receptors: signaling and bio...logy. PubmedID 15189145 Title Lysophospholipid receptors: signaling and biology. Authors

  14. DMPD: Is HIV infection a TNF receptor signalling-driven disease? [Dynamic Macrophage Pathway CSML Database

    Full Text Available 18178131 Is HIV infection a TNF receptor signalling-driven disease? Herbein G, Khan... KA. Trends Immunol. 2008 Feb;29(2):61-7. (.png) (.svg) (.html) (.csml) Show Is HIV infection a TNF receptor signalling-driven diseas...e? PubmedID 18178131 Title Is HIV infection a TNF receptor signalling-driven disease

  15. PKC and MAPK signalling pathways regulate vascular endothelin receptor expression

    Nilsson, David; Wackenfors, Angelica; Gustafsson, Lotta; Ugander, Martin; Ingemansson, Richard; Edvinsson, Lars; Malmsjö, Malin

    immunofluorescence techniques. Sarafotoxin 6c and endothelin ET-1 were used to examine the endothelin ET(A) and ET(B) receptor effects. The involvement of PKC and MAPK in the receptor regulation was examined by culture in the presence of antagonists. Organ culture resulted in increased sarafotoxin 6c and endothelin......-1 contractions, endothelin ET(A) and ET(B) receptor immunofluorescence staining intensities and endothelin ET(B), but not ET(A), receptor mRNA levels. The general PKC inhibitors, bisindolylmaleimide I (10 microM) or Ro-32-0432 (10 microM), inhibited these effects. Also, the increase in sarafotoxin 6......c contraction, endothelin ET(B) receptor and mRNA levels and endothelin ET(A) and ET(B) immunofluorescence staining intensities were inhibited by MAPK inhibitors for extracellular signal related kinases 1 and 2 (ERK1/2), PD98059 (10 microM), C-jun terminal kinase (JNK), SP600125 (10 microM), but not...

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

    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

  17. Inhibition of Dengue Virus Replication by a Class of Small-Molecule Compounds That Antagonize Dopamine Receptor D4 and Downstream Mitogen-Activated Protein Kinase Signaling

    Smith, Jessica L.; Stein, David A.; Shum, David; Fischer, Matthew A.; Radu, Constantin; Bhinder, Bhavneet; Djaballah, Hakim; Nelson, Jay A.; Früh, Klaus; Hirsch, Alec J.

    2014-01-01

    Dengue viruses (DENV) are endemic pathogens of tropical and subtropical regions that cause significant morbidity and mortality worldwide. To date, no vaccines or antiviral therapeutics have been approved for combating DENV-associated disease. In this paper, we describe a class of tricyclic small-molecule compounds—dihydrodibenzothiepines (DHBTs), identified through high-throughput screening—with potent inhibitory activity against DENV serotype 2. SKI-417616, a highly active representative of ...

  18. Retinoic Acid-mediated Nuclear Receptor Activation and Hepatocyte Proliferation

    Bushue, Nathan; Wan, Yu-Jui Yvonne

    2016-01-01

    Due to their well-known differentiation and apoptosis-inducing abilities, retinoic acid (RA) and its analogs have strong anti-cancer efficacy in human cancers. However, in vivo RA is a liver mitogen. While speculation has persisted that RA-mediated signaling is likely involved in hepatocyte proliferation during liver regeneration, direct evidence is still required. Findings in support of this proposition include observations that a release of retinyl palmitate (the precursor of RA) occurs in liver stellate cells following liver injury. Nevertheless, the biological action of this released vitamin A is virtually unknown. More likely is that the released vitamin A is converted to RA, the biological form, and then bound to a specific receptor (retinoid x receptor; RXRα), which is most abundantly expressed in the liver. Considering the mitogenic effects of RA, the RA-activated RXRα would likely then influence hepatocyte proliferation and liver tissue repair. At present, the mechanism by which RA stimulates hepatocyte proliferation is largely unknown. This review summarizes the activation of nuclear receptors (peroxisome proliferator activated receptor-α, pregnane x receptor, constitutive androstane receptor, and farnesoid x receptor) in an RXRα dependent manner to induce hepatocyte proliferation, providing a link between RA and its proliferative role.

  19. Research Resource: A Reference Transcriptome for Constitutive Androstane Receptor and Pregnane X Receptor Xenobiotic Signaling.

    Ochsner, Scott A; Tsimelzon, Anna; Dong, Jianrong; Coarfa, Cristian; McKenna, Neil J

    2016-08-01

    The pregnane X receptor (PXR) (PXR/NR1I3) and constitutive androstane receptor (CAR) (CAR/NR1I2) members of the nuclear receptor (NR) superfamily of ligand-regulated transcription factors are well-characterized mediators of xenobiotic and endocrine-disrupting chemical signaling. The Nuclear Receptor Signaling Atlas maintains a growing library of transcriptomic datasets involving perturbations of NR signaling pathways, many of which involve perturbations relevant to PXR and CAR xenobiotic signaling. Here, we generated a reference transcriptome based on the frequency of differential expression of genes across 159 experiments compiled from 22 datasets involving perturbations of CAR and PXR signaling pathways. In addition to the anticipated overrepresentation in the reference transcriptome of genes encoding components of the xenobiotic stress response, the ranking of genes involved in carbohydrate metabolism and gonadotropin action sheds mechanistic light on the suspected role of xenobiotics in metabolic syndrome and reproductive disorders. Gene Set Enrichment Analysis showed that although acetaminophen, chlorpromazine, and phenobarbital impacted many similar gene sets, differences in direction of regulation were evident in a variety of processes. Strikingly, gene sets representing genes linked to Parkinson's, Huntington's, and Alzheimer's diseases were enriched in all 3 transcriptomes. The reference xenobiotic transcriptome will be supplemented with additional future datasets to provide the community with a continually updated reference transcriptomic dataset for CAR- and PXR-mediated xenobiotic signaling. Our study demonstrates how aggregating and annotating transcriptomic datasets, and making them available for routine data mining, facilitates research into the mechanisms by which xenobiotics and endocrine-disrupting chemicals subvert conventional NR signaling modalities. PMID:27409825

  20. Bioluminescence imaging of estrogen receptor activity during breast cancer progression.

    Vantaggiato, Cristina; Dell'Omo, Giulia; Ramachandran, Balaji; Manni, Isabella; Radaelli, Enrico; Scanziani, Eugenio; Piaggio, Giulia; Maggi, Adriana; Ciana, Paolo

    2016-01-01

    Estrogen receptors (ER) are known to play an important regulatory role in mammary gland development as well as in its neoplastic transformation. Although several studies highlighted the contribution of ER signaling in the breast transformation, little is known about the dynamics of ER state of activity during carcinogenesis due to the lack of appropriate models for measuring the extent of receptor signaling in time, in the same animal. To this aim, we have developed a reporter mouse model for the non-invasive in vivo imaging of ER activity: the ERE-Luc reporter mouse. ERE-Luc is a transgenic mouse generated with a firefly luciferase (Luc) reporter gene driven by a minimal promoter containing an estrogen responsive element (ERE). This model allows to measure receptor signaling in longitudinal studies by bioluminescence imaging (BLI). Here, we have induced sporadic mammary cancers by treating systemically ERE-Luc reporter mice with DMBA (9,10-dimethyl 1,2-benzanthracene) and measured receptor signaling by in vivo imaging in individual animals from early stage until a clinically palpable tumor appeared in the mouse breast. We showed that DMBA administration induces an increase of bioluminescence in the whole abdominal area 6 h after treatment, the signal rapidly disappears. Several weeks later, strong bioluminescence is observed in the area corresponding to the mammary glands. In vivo and ex vivo imaging analysis demonstrated that this bioluminescent signal is localized in the breast area undergoing neoplastic transformation. We conclude that this non-invasive assay is a novel relevant tool to identify the activation of the ER signaling prior the morphological detection of the neoplastic transformation. PMID:27069764

  1. In vivo characterization of high Basal signaling from the ghrelin receptor

    Petersen, Pia Steen; Woldbye, David P D; Madsen, Andreas Nygaard; Egerod, Kristoffer L; Jin, Chunyu; Lang, Manja; Rasmussen, Maria; Beck-Sickinger, Annette G; Holst, Birgitte

    2009-01-01

    following study, we found that the expression of the ghrelin receptor in the hypothalamus was up-regulated approximately 2-fold in rats both during 48-h fasting and by streptozotocin-induced hyperphagia. In a separate experiment, to probe for the effect of the high basal signaling of the ghrelin receptor in....... Food intake and body weight were significantly decreased in the group of rats treated with the inverse agonist, as compared with the groups treated with the control peptide or the vehicle. In the hypothalamus, the expression of neuropeptide Y and uncoupling protein 2 was decreased by the inverse...... decreased by administration of the inverse agonist. It is suggested that the high constitutive signaling activity is important for the in vivo function of the ghrelin receptor in the control of food intake and body weight....

  2. Organophosphorous pesticide metabolite (DEDTP) induces changes in the activation status of human lymphocytes by modulating the interleukin 2 receptor signal transduction pathway

    Diethyldithiophosphate (DEDTP) is a metabolite formed by biotransformation of organophosphorous (OP) compounds that has a longer half-life than its parental compound. Here we evaluate the effects of DEDTP on human CD4+ T lymphocytes. In vitro exposure to DEDTP (1-50 μM) decreased [3H]thymidine incorporation in resting cells and increased CD25 surface expression without altering cell viability. DEDTP treatment inhibited anti-CD3/anti-CD28 stimulation-induced CD4+ and CD8+ T cell proliferation determined by CFSE dilution. Decreased CD25 expression and intracellular IL-2 levels were correlated with this defect in cell proliferation. IL-2, IFN-γ and IL-10 secretion were also reduced while IL-4 secretion was not altered. Increased phosphorylation of SOCS3 and dephosphorylation of STAT5 were induced by DEDTP after as little as 5 min of exposure. In addition, DEDTP induced phosphorylation of ERK, JNK and p38 and NFAT nuclear translocation. These results suggest that DEDTP can modulate phosphorylation of intracellular proteins such as SOCS3, which functions as a negative regulator of cytokine signalling, and that DEDTP exposure may thus cause T cells to fail to respond to further antigen challenges.

  3. Effect of Spatial Inhomogeneities on the Membrane Surface on Receptor Dimerization and Signal Initiation.

    Kerketta, Romica; Halász, Ádám M; Steinkamp, Mara P; Wilson, Bridget S; Edwards, Jeremy S

    2016-01-01

    Important signal transduction pathways originate on the plasma membrane, where microdomains may transiently entrap diffusing receptors. This results in a non-random distribution of receptors even in the resting state, which can be visualized as "clusters" by high resolution imaging methods. Here, we explore how spatial in-homogeneities in the plasma membrane might influence the dimerization and phosphorylation status of ErbB2 and ErbB3, two receptor tyrosine kinases that preferentially heterodimerize and are often co-expressed in cancer. This theoretical study is based upon spatial stochastic simulations of the two-dimensional membrane landscape, where variables include differential distributions and overlap of transient confinement zones ("domains") for the two receptor species. The in silico model is parameterized and validated using data from single particle tracking experiments. We report key differences in signaling output based on the degree of overlap between domains and the relative retention of receptors in such domains, expressed as escape probability. Results predict that a high overlap of domains, which favors transient co-confinement of both receptor species, will enhance the rate of hetero-interactions. Where domains do not overlap, simulations confirm expectations that homo-interactions are favored. Since ErbB3 is uniquely dependent on ErbB2 interactions for activation of its catalytic activity, variations in domain overlap or escape probability markedly alter the predicted patterns and time course of ErbB3 and ErbB2 phosphorylation. Taken together, these results implicate membrane domain organization as an important modulator of signal initiation, motivating the design of novel experimental approaches to measure these important parameters across a wider range of receptor systems. PMID:27570763

  4. Transcriptional Crosstalk between Nuclear Receptors and Cytokine Signal Transduction Pathways in Immunity

    LihuaWang; XiaohuZhang; WilliamL.Farrar

    2004-01-01

    The nuclear receptor superfamily and the transcriptional factors associated with cytokines are inherently different families of signaling molecules and activate gene transcription by binding to their respective responsive element. However, it has become increasingly clear from our works and others that nuclear receptors are important regulators of cytokine production and function through complex and varied interactions between these distinct transcriptional factors. This review provides a general overview of the mechanism of action of nuclear receptors and their transcriptional crosstalk with transcriptional factors associated with cytokine transduction pathways. One of the most important mechanistic aspects is protein to protein interaction through a direct or co-regulator-mediated indirect manner. Such crosstalk is crucially involved in physiological and therapeutic roles of nuclear receptors and their ligands in immunity,inflammation and cytokine-related tumors. Cellular & Molecular Immunology. 2004;1(6):416-424.

  5. LRP2, an auxiliary receptor that controls sonic hedgehog signaling in development and disease.

    Christ, Annabel; Herzog, Katja; Willnow, Thomas E

    2016-05-01

    To fulfill their multiple roles in organ development and adult tissue homeostasis, hedgehog (HH) morphogens act through their receptor Patched (PTCH) on target cells. However, HH actions also require HH binding proteins, auxiliary cell surface receptors that agonize or antagonize morphogen signaling in a context-dependent manner. Here, we discuss recent findings on the LDL receptor-related protein 2 (LRP2), an exemplary HH binding protein that modulates sonic hedgehog activities in stem and progenitor cell niches in embryonic and adult tissues. LRP2 functions are crucial for developmental processes in a number of tissues, including the brain, the eye, and the heart, and defects in this receptor pathway are the cause of devastating congenital diseases in humans. Developmental Dynamics 245:569-579, 2016. © 2016 Wiley Periodicals, Inc. PMID:26872844

  6. Prediction of compounds activity in nuclear receptor signaling and stress pathway assays using machine learning algorithms and low dimensional molecular descriptors

    Filip eStefaniak

    2015-01-01

    Toxicity evaluation of newly synthesized or used compounds is one of the main challenges during product development in many areas of industry. For example, toxicity is the second reason - after lack of efficacy - for failure in preclinical and clinical studies of drug candidates. To avoid attrition at the late stage of the drug development process, the toxicity analyses are employed at the early stages of a discovery pipeline, along with activity and selectivity enhancing. Although many assa...

  7. Prediction of Compounds Activity in Nuclear Receptor Signaling and Stress Pathway Assays Using Machine Learning Algorithms and Low-Dimensional Molecular Descriptors

    Stefaniak, Filip

    2015-01-01

    Toxicity evaluation of newly synthesized or used compounds is one of the main challenges during product development in many areas of industry. For example, toxicity is the second reason—after lack of efficacy—for failure in preclinical and clinical studies of drug candidates. To avoid attrition at the late stage of the drug development process, the toxicity analyses are employed at the early stages of a discovery pipeline, along with activity and selectivity enhancing. Although many assays fo...

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

    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...

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

    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

  10. Dasatinib inhibits proliferation and activation of CD8+ T-lymphocytes by down-regulation of the T-cell receptor signaling

    Fei, Fei

    2007-01-01

    The novel Src/Abl inhibitor dasatinib (BMS-354825) is a promising therapeutic agent with imatinib-resistant BCR-ABL mutations in patients with chronic myelogenous leukemia (CML) or Ph-positive acute lymphoblastic leukemia (ALL). However, little is known about its effects on T-cell function, especially for patients undergoing allogeneic transplantation for leukemia. Here, we demonstrate that dasatinib at a concentration of 5 nM to 250 nM inhibits the proliferation and activation of CD8+ T-lymp...

  11. Estrogen receptor and PI3K/Akt signaling pathway involvement in S-(-equol-induced activation of Nrf2/ARE in endothelial cells.

    Ting Zhang

    Full Text Available S-(-equol, a natural product of the isoflavone daidzein, has been reported to offer cytoprotective effects with respect to the cardiovascular system, but how this occurs is unclear. Interestingly, S-(-equol is produced by the human gut, suggesting a role in physiological processes. We report that treatment of human umbilical vein endothelial cells and EA.hy926 cells with S-(-equol induces ARE-luciferase reporter gene activity that is dose and time dependent. S-(-equol (10-250 nM increases nuclear factor-erythroid 2-related factor 2 (Nrf2 as well as gene products of Nrf2 target genes heme oxygenase-1 (HO-1 and NAD(PH (nicotinamide-adenine-dinucleotide-phosphate quinone oxidoreductase 1 (NQO1. Endothelial cells transfected with an HA-Nrf2 expression plasmid had elevated HA-Nrf2, HO-1, and NQO1 in response to S-(-equol exposure. S-(-equol treatment affected Nrf2 mRNA only slightly but significantly increased HO-1 and NQO1 mRNA. The pretreatment of cells with specific ER inhibitors or PI3K/Akt (ICI182,780 and LY294002 increased Nrf2, HO-1, and NQO1 protein, impaired nuclear translocation of HA-Nrf2, and decreased ARE-luciferase activity. Identical experiments were conducted with daidzein, which had effects similar to S-(-equol. In addition, DPN treatment (an ERβ agonist induced the ARE-luciferase reporter gene, promoting Nrf2 nuclear translocation. Cell pretreatment with an ERβ antagonist (PHTPP impaired S-(-equol-induced Nrf2 activation. Pre-incubation of cells followed by co-treatment with S-(-equol significantly improved cell survival in response to H2O2 or tBHP and reduced apoptotic and TUNEL-positively-stained cells. Notably, the ability of S-(-equol to protect against H2O2-induced cell apoptosis was attenuated in cells transfected with an siRNA against Nrf2. Thus, beneficial effects of S-(-equol with respect to cytoprotective antioxidant gene activation may represent a novel strategy to prevent and treat cardiovascular diseases.

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

    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.

  13. Activation of estrogen receptor β-dependent nitric oxide signaling mediates the hypotensive effects of estrogen in the rostral ventrolateral medulla of anesthetized rats

    Shih Cheng-Dean

    2009-07-01

    Full Text Available Abstract Background Apart from their well-known peripheral cardiovascular effects, emerging evidence indicates that estrogen acts as a modulator in the brain to regulate cardiovascular functions. The underlying mechanisms of estrogen in central cardiovascular regulation, however, are poorly understood. The present study investigated the cardiovascular effects of 17β-estradiol (E2β in the rostral ventrolateral medulla (RVLM, where sympathetic premotor neurons are located, and delineated the engagement of nitric oxide (NO in E2β-induced cardiovascular responses. Methods In male Sprague-Dawley rats maintained under propofol anesthesia, the changes of blood pressure, heart rate and sympathetic vasomotor tone after microinjection bilaterally into the RVLM of a synthetic estrogen, E2β were examined for at least 120 min. The involvement of ERα and/or ERβ subtypes was determined by microinjection of selective ERα or ERβ agonist into bilateral RVLM. Different NO synthase (NOS inhibitors were used to evaluate the involvement of differential of NOS isoforms in the cardiovascular effects of E2β. Results Bilateral microinjection of E2β (0.5, 1, or 5 pmol into the RVLM dose-dependently decreased systemic arterial pressure (SAP and the power density of the vasomotor components of SAP signals, our experimental index for sympathetic neurogenic vasomotor tone. These cardiovascular depressive effects of E2β (1 pmol were abolished by co-injection of ER antagonist ICI 182780 (0.25 or 0.5 pmol, but not a transcription inhibitor actinomycin D (10 nmol. Like E2β, microinjection bilaterally into the RVLM of a selective ERβ agonist 2,3-bis(4-hydroxyphenyl propionitrile (DPN, 1, 2, or 5 pmol induced significant decreases in these hemodynamic parameters in a dose-dependent manner. In contrast, the selective ERα agonist 1,3,5-tris(4-hydroxyphenyl-4-propyl-1H-pyrazole (5 pmol did not influence the same cardiovascular parameters. Co-administration bilaterally

  14. Anti-cancer activity of an osthole derivative, NBM-T-BMX-OS01: targeting vascular endothelial growth factor receptor signaling and angiogenesis.

    Yang, Hung-Yu; Hsu, Ya-Fen; Chiu, Pei-Ting; Ho, Shiau-Jing; Wang, Chi-Han; Chi, Chih-Chin; Huang, Yu-Han; Lee, Cheng-Feng; Li, Ying-Shiuan; Ou, George; Hsu, Ming-Jen

    2013-01-01

    Angiogenesis occurs during tissue growth, development and wound healing. It is also required for tumor progression and represents a rational target for therapeutic intervention. NBM-T-BMX-OS01 (BMX), derived from the semisynthesis of osthole, an active ingredient isolated from Chinese herb Cnidium monnieri (L.) Cuss., was recently shown to enhance learning and memory in rats. In this study, we characterized the anti-angiogenic activities of NBM-T-BMX-OS01 (BMX) in an effort to develop novel inhibitors to suppress angiogenesis and tumor growth. BMX inhibited vascular endothelial growth factor (VEGF)-induced proliferation, migration and endothelial tube formation in human umbilical endothelial cells (HUVECs). BMX also attenuated VEGF-induced microvessel sprouting from aortic rings ex vivo and reduced HCT116 colorectal cancer cells-induced angiogenesis in vivo. Moreover, BMX inhibited the phosphorylation of VEGFR2, FAK, Akt and ERK in HUVECs exposed to VEGF. BMX was also shown to inhibit HCT116 cell proliferation and to suppress the growth of subcutaneous xenografts of HCT116 cells in vivo. Taken together, this study provides evidence that BMX modulates vascular endothelial cell remodeling and leads to the inhibition of tumor angiogenesis. These results also support the role of BMX as a potential drug candidate and warrant the clinical development in the treatment of cancer. PMID:24312323

  15. Extracts of Feijoa Inhibit Toll-Like Receptor 2 Signaling and Activate Autophagy Implicating a Role in Dietary Control of IBD.

    Noha Ahmed Nasef

    Full Text Available Inflammatory bowel disease (IBD is a heterogeneous chronic inflammatory disease affecting the gut with limited treatment success for its sufferers. This suggests the need for better understanding of the different subtypes of the disease as well as nutritional interventions to compliment current treatments. In this study we assess the ability of a hydrophilic feijoa fraction (F3 to modulate autophagy a process known to regulate inflammation, via TLR2 using IBD cell lines.Mouse embryonic fibroblasts (MEF deleted for ATG5, and two intestinal epithelial cells HCT15 and HCT116, were used to test the anti-inflammatory effect of F3 after stimulating the cells with a TLR2 specific ligand PAM3CSK4.F3 was able to reduce TLR2 specific inflammation and stimulate autophagy in MEFs and HCT15 cells but not in HCT116 cells. The anti-inflammatory effect was reduced in the MEF cells deleted for ATG5. In addition, the activation of autophagy by F3 was enhanced by PAM3CSK4.F3 of feijoa can interact with cells via a TLR2 specific mechanism and reduce Nuclear factor kappa B (NF-κB activation in part due to stimulation of autophagy. These results suggest that there is potential benefit in using feijoa extracts as part of dietary interventions to manage IBD in patients.

  16. Membrane receptor signaling and control of DNA repair after exposure to ionizing radiation

    Accumulated evidence indicates that activation of erbB family of receptors, when mutated or over-expressed, mediates chemo and radiotherapy resistance. In this context signaling pathways down-stream of epidermal growth factor receptor (EGFR), when abnormally activated, invoke cell survival mechanisms, which leads to resistance against radiation. In several reports it has been demonstrated that molecular targeting of EGFR signaling enhances the cytotoxic effects of radiotherapy. The radiosensitizing effects of EGFR antagonists correlate with a suppression of the ability of tumor cells to repair radiation-induced DNA double strand breaks (DNA-DSBs) through non-homologous endjoining repair pathway (NHEJ). The purpose of this review is to highlight the function of EGFR and erbB2 receptors on signaling pathways, i.e. PI3K/Akt activated by ionizing radiation (IR) and involved in repair of DNA-DSB which can explain the radiosensitizing effects of related antagonists. Advances in understanding the mechanism of erbB-signaling in regulating DNA-DSB repair will promote translational approaches to test new strategies for clinically applicable molecular targeting. (orig.)

  17. Toll-like receptors; their physiological role and signal transduction system.

    Takeuchi, O; Akira, S

    2001-04-01

    Drosophila Toll protein is a transmembrane receptor whose function is to recognize the invasion of microorganisms as well as to establish dorso-ventral polarity. Recently, mammalian homologues of Toll, designated as Toll-like receptors (TLRs) have been discovered. So far, six members (TLR1-6) have been reported and two of these, TLR2 and TLR4, have been shown to be essential for the recognition of distinct bacterial cell wall components. TLR2 discriminates peptidoglycan (PGN), lipoprotein, lipoarabinomannan (LAM) and zymosan, whereas TLR4 recognizes lipopolysaccharide (LPS), lipoteichoic acid (LTA) and Taxol. Bacterial components elicit the activation of an intracellular signaling cascade via TLR in a similar way to that occurs upon ligand binding to IL-1 receptor (IL-1R). This signaling pathway leads to the activation of a transcription factor NF-kappaB and c-Jun N-terminal kinase (JNK), which initiate the transcription of proinflammatory cytokine genes. Particularly, analysis of knockout mice revealed a pivotal role for MyD88 in the signaling of the TLR/IL-1R family. Taken together, TLRs and the downstream signaling pathway play a key role in innate immune recognition and in subsequent activation of adaptive immunity. PMID:11357875

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

    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

  19. MicroRNA-27b plays a role in pulmonary arterial hypertension by modulating peroxisome proliferator-activated receptor γ dependent Hsp90-eNOS signaling and nitric oxide production

    Bi, Rui; Bao, Chunrong; Jiang, Lianyong; Liu, Hao; Yang, Yang; Mei, Ju; Ding, Fangbao, E-mail: dbcar126@126.com

    2015-05-01

    Pulmonary artery endothelial dysfunction is associated with pulmonary arterial hypertension (PAH). Based on recent studies showing that microRNA (miR)-27b is aberrantly expressed in PAH, we hypothesized that miR-27b may contribute to pulmonary endothelial dysfunction and vascular remodeling in PAH. The effect of miR-27b on pulmonary endothelial dysfunction and the underlying mechanism were investigated in human pulmonary artery endothelial cells (HPAECs) in vitro and in a monocrotaline (MCT)-induced model of PAH in vivo. miR-27b expression was upregulated in MCT-induced PAH and inversely correlated with the levels of peroxisome proliferator-activated receptor (PPAR)-γ, and miR-27b inhibition attenuated MCT-induced endothelial dysfunction and remodeling and prevented PAH associated right ventricular hypertrophy and systolic pressure in rats. PPARγ was confirmed as a direct target of miR-27b in HPAECs and shown to mediate the effect of miR-27b on the disruption of endothelial nitric oxide synthase (eNOS) coupling to Hsp90 and the suppression of NO production associated with the PAH phenotype. We showed that miR-27b plays a role endothelial function and NO release and elucidated a potential mechanism by which miR-27b regulates Hsp90-eNOS and NO signaling by modulating PPARγ expression, providing potential therapeutic targets for the treatment of PAH. - Highlights: • miR-27b plays a role in endothelial function and NO release. • miR-27b inhibition ameliorates MCT-induced endothelial dysfunction and PAH. • miR-27b targets PPARγ in HPAECs. • miR-27b regulates PPARγ dependent Hsp90-eNOS and NO signaling.

  20. MicroRNA-27b plays a role in pulmonary arterial hypertension by modulating peroxisome proliferator-activated receptor γ dependent Hsp90-eNOS signaling and nitric oxide production

    Pulmonary artery endothelial dysfunction is associated with pulmonary arterial hypertension (PAH). Based on recent studies showing that microRNA (miR)-27b is aberrantly expressed in PAH, we hypothesized that miR-27b may contribute to pulmonary endothelial dysfunction and vascular remodeling in PAH. The effect of miR-27b on pulmonary endothelial dysfunction and the underlying mechanism were investigated in human pulmonary artery endothelial cells (HPAECs) in vitro and in a monocrotaline (MCT)-induced model of PAH in vivo. miR-27b expression was upregulated in MCT-induced PAH and inversely correlated with the levels of peroxisome proliferator-activated receptor (PPAR)-γ, and miR-27b inhibition attenuated MCT-induced endothelial dysfunction and remodeling and prevented PAH associated right ventricular hypertrophy and systolic pressure in rats. PPARγ was confirmed as a direct target of miR-27b in HPAECs and shown to mediate the effect of miR-27b on the disruption of endothelial nitric oxide synthase (eNOS) coupling to Hsp90 and the suppression of NO production associated with the PAH phenotype. We showed that miR-27b plays a role endothelial function and NO release and elucidated a potential mechanism by which miR-27b regulates Hsp90-eNOS and NO signaling by modulating PPARγ expression, providing potential therapeutic targets for the treatment of PAH. - Highlights: • miR-27b plays a role in endothelial function and NO release. • miR-27b inhibition ameliorates MCT-induced endothelial dysfunction and PAH. • miR-27b targets PPARγ in HPAECs. • miR-27b regulates PPARγ dependent Hsp90-eNOS and NO signaling

  1. A peptide tetramer Tk-tPN induces tolerance of cardiac allografting by conversion of type 1 to type 2 immune responses via the Toll-like receptor 2 signal-promoted activation of the MCP1 gene.

    Li, Zuoqing; Yang, Neng; Zhou, Ling; Gu, Peng; Wang, Hui; Zhou, Yun; Zhou, Peijun; Lu, Liming; Chou, Kuang-Yen

    2016-03-01

    The plant protein trichosanthin (Tk) and its derived peptide tetramer Tk-tPN have been shown to stimulate the type 2 immune responses for treating autoimmune disease. This work explores the possibility of using Tk-tPN as a non-toxic immunosuppressant to induce transplantation tolerance using the mechanisms by which T-cell-mediated immune responses are transferred from type 1 to type 2 through innate immunity-related pathways. Immunocytes and cytokine secretions involved in the mouse cardiac allografting model with Tk-tPN treatment were characterized. Identification of critical genes and analysis of their functions through Toll-like receptor (TLR) -initiated signalling and the possible epigenetic changes were performed. Mean survival times of the cardiac allografts were delayed from 7·7 ± 0·3 days (control) to 22·7 ± 3·9 days (P Gata3(+) ), together with a selective expansion of the IL-4/IL-10-producing CD8(+)  CD28(-) regulatory T-cell subset. A TLR2-initiated high expression of chemokine gene MCP1 was detectable simultaneously. Epigenetically Tk/Tk-tPN could also acetylate the histone H3K9 of MCP1 promoter to skew the immunity towards T helper type 2 responses. Tk/Tk-tPN is therefore capable of down-regulating the type 1 response-dominant rejection of cardiac allografts by evoking type 2 immunity through the activation of a TLR2-initiated signalling pathway and MCP1 gene to expand the IL-4/IL-10-secreting CD8(+)  CD28(-) regulatory T cells. Tk-tPN could be a promising novel immunosuppressant to induce tolerance in allotransplantation. PMID:26694804

  2. The Multitarget Ligand 3-Iodothyronamine Modulates β-Adrenergic Receptor 2 Signaling

    Dinter, Juliane; Khajavi, Noushafarin; Mühlhaus, Jessica; Wienchol, Carolin Leonie; Cöster, Maxi; Hermsdorf, Thomas; Stäubert, Claudia; Köhrle, Josef; Schöneberg, Torsten; Kleinau, Gunnar; Mergler, Stefan; Biebermann, Heike

    2015-01-01

    Background 3-Iodothyronamine (3-T1AM), a signaling molecule with structural similarities to thyroid hormones, induces numerous physiological responses including reversible body temperature decline. One target of 3-T1AM is the trace amine-associated receptor 1 (TAAR1), which is a member of the rhodopsin-like family of G protein-coupled receptors (GPCRs). Interestingly, the effects of 3-T1AM remain detectable in TAAR1 knockout mice, suggesting further targets for 3-T1AM such as adrenergic receptors. Therefore, we evaluated whether β-adrenergic receptor 1 (ADRB1) and 2 (ADRB2) signaling is affected by 3-T1AM in HEK293 cells and in human conjunctival epithelial cells (IOBA-NHC), where these receptors are highly expressed endogenously. Methods A label-free EPIC system for prescreening the 3-T1AM-induced effects on ADRB1 and ADRB2 in transfected HEK293 cells was used. In addition, ADRB1 and ADRB2 activation was analyzed using a cyclic AMP assay and a MAPK reporter gene assay. Finally, fluorescence Ca2+ imaging was utilized to delineate 3-T1AM-induced Ca2+ signaling. Results 3-T1AM (10−5−10−10M) enhanced isoprenaline-induced ADRB2-mediated Gs signaling but not that of ADRB1-mediated signaling. MAPK signaling remained unaffected for both receptors. In IOBA-NHC cells, norepinephrine-induced Ca2+ influxes were blocked by the nonselective ADRB blocker timolol (10 µM), indicating that ADRBs are most likely linked with Ca2+ channels. Notably, timolol was also found to block 3-T1AM (10−5M)-induced Ca2+ influx. Conclusions The presented data support that 3-T1AM directly modulates β-adrenergic receptor signaling. The relationship between 3-T1AM and β-adrenergic signaling also reveals a potential therapeutic value for suppressing Ca2+ channel-mediated inflammation processes, occurring in eye diseases such as conjunctivitis. PMID:26601070

  3. The Architecture of the TIR Domain Signalosome in the Toll-like Receptor-4 Signaling Pathway

    Emine Guven-Maiorov; Ozlem Keskin; Attila Gursoy; Carter VanWaes; Zhong Chen; Chung-Jung Tsai; Ruth Nussinov

    2015-01-01

    Scientific Reports | 5:13128 | DOI: 10.1038/srep13128 1 www.nature.com/scientificreports The Architecture of the TIR Domain Signalosome in the Toll-like Receptor-4 Signaling Pathway Emine Guven-Maiorov1,2, Ozlem Keskin1,2, Attila Gursoy2,3, Carter VanWaes4, Zhong Chen4, Chung-Jung Tsai5 & Ruth Nussinov5,6 Activated Toll-like receptors (TLRs) cluster in lipid rafts and induce pro- and anti-tumor responses. The organization of the assembly is critical to the understandin...

  4. Heat dissipation guides activation in signaling proteins.

    Weber, Jeffrey K; Shukla, Diwakar; Pande, Vijay S

    2015-08-18

    Life is fundamentally a nonequilibrium phenomenon. At the expense of dissipated energy, living things perform irreversible processes that allow them to propagate and reproduce. Within cells, evolution has designed nanoscale machines to do meaningful work with energy harnessed from a continuous flux of heat and particles. As dictated by the Second Law of Thermodynamics and its fluctuation theorem corollaries, irreversibility in nonequilibrium processes can be quantified in terms of how much entropy such dynamics produce. In this work, we seek to address a fundamental question linking biology and nonequilibrium physics: can the evolved dissipative pathways that facilitate biomolecular function be identified by their extent of entropy production in general relaxation processes? We here synthesize massive molecular dynamics simulations, Markov state models (MSMs), and nonequilibrium statistical mechanical theory to probe dissipation in two key classes of signaling proteins: kinases and G-protein-coupled receptors (GPCRs). Applying machinery from large deviation theory, we use MSMs constructed from protein simulations to generate dynamics conforming to positive levels of entropy production. We note the emergence of an array of peaks in the dynamical response (transient analogs of phase transitions) that draw the proteins between distinct levels of dissipation, and we see that the binding of ATP and agonist molecules modifies the observed dissipative landscapes. Overall, we find that dissipation is tightly coupled to activation in these signaling systems: dominant entropy-producing trajectories become localized near important barriers along known biological activation pathways. We go on to classify an array of equilibrium and nonequilibrium molecular switches that harmonize to promote functional dynamics. PMID:26240354

  5. TAM receptors affect adult brain neurogenesis by negative regulation of microglial cell activation1

    Ji, Rui; Tian, Shifu; Lu, Helen J.; LU, QINGJUN; Yan ZHENG; Wang, Xiaomin; Ding, Jixiang; Li, Qiutang; Lu, Qingxian

    2013-01-01

    TAM tyrosine kinases play multiple functional roles including regulation of the target genes important in homeostatic regulation of cytokine receptors or Toll-like receptor-mediated signal transduction pathways. Here, we show that TAM receptors affect adult hippocampal neurogenesis and loss of TAM receptors impair hippocampal neurogenesis, largely attributed to exaggerated inflammatory responses by microglia characterized by increased MAP kinase and NF-κB activation and elevated production of...

  6. Prediction of compounds activity in nuclear receptor signaling and stress pathway assays using machine learning algorithms and low dimensional molecular descriptors

    Filip eStefaniak

    2015-12-01

    Full Text Available Toxicity evaluation of newly synthesized or used compounds is one of the main challenges during product development in many areas of industry. For example, toxicity is the second reason - after lack of efficacy - for failure in preclinical and clinical studies of drug candidates. To avoid attrition at the late stage of the drug development process, the toxicity analyses are employed at the early stages of a discovery pipeline, along with activity and selectivity enhancing. Although many assays for screening in vitro toxicity are available, their massive application is not always time and cost effective. Thus the need for fast and reliable in silico tools, which can be used not only for toxicity prediction of existing compounds, but also for prioritization of compounds planned for synthesis or acquisition. Here I present the benchmark results of the combination of various attribute selection methods and machine learning algorithms and their application to the data sets of the Tox21 Data Challenge. The best performing method: Best First for attribute selection with the Rotation Forest/ADTree classifier offers good accuracy for most tested cases. For 11 out of 12 targets, the AUROC value for the final evaluation set was ≥0.72, while for three targets the AUROC value was ≥ 0.80, with the average AUROC being 0.784±0.069. The use of two-dimensional descriptors sets enables fast screening and compound prioritization even for a very large database. Open source tools used in this project make the presented approach widely available and encourage the community to further improve the presented scheme.

  7. Potential protective mechanisms of aryl hydrocarbon receptor (AHR) signaling in benign prostatic hyperplasia.

    Mehta, Vatsal; Vezina, Chad M

    2011-01-01

    The aryl hydrocarbon receptor (AHR) is an evolutionarily conserved ligand activated transcription factor best known for its role in mediating toxic responses to dioxin-like environmental contaminants. However, AHR signaling has also emerged as an active participant in processes of normal development and disease progression. Here, we review the role of AHR signaling in prostate development and disease processes, with a particular emphasis on benign prostatic hyperplasia (BPH). Inappropriate AHR activation has recently been associated with a decreased risk of symptomatic BPH in humans and has been shown to impair prostate development and disrupt endocrine signaling in rodents. We highlight known physiological responses to AHR activation in prostate and other tissues and discuss potential mechanisms by which it may act in adult human prostate to protect against symptomatic BPH. PMID:21684673

  8. Structural motifs of importance for the constitutive activity of the orphan 7TM receptor EBI2: analysis of receptor activation in the absence of an agonist

    Benned-Jensen, Tau; Rosenkilde, Mette M

    2008-01-01

    The Epstein-Barr induced receptor 2 (EBI2) is a lymphocyte-expressed orphan seven transmembrane-spanning (7TM) receptor that signals constitutively through Galphai, as shown, for instance by guanosine 5'-O-(3-thio)triphosphate incorporation. Two regions of importance for the constitutive activity...

  9. Neurotrophin receptors expression and JNK pathway activation in human astrocytomas

    Neurotrophins are growth factors that regulate cell growth, differentiation and apoptosis in the nervous system. Their diverse actions are mediated through two different transmembrane – receptor signaling systems: Trk receptor tyrosine kinases (TrkA, TrkB, TrkC) and p75NTR neurotrophin receptor. Trk receptors promote cell survival and differentiation while p75NTR induces, in most cases, the activity of JNK-p53-Bax apoptosis pathway or suppresses intracellular survival signaling cascades. Robust Trk activation blocks p75NTR -induced apoptosis by suppressing the JNK-p53-Bax pathway. The aim of this exploratory study was to investigate the expression levels of neurotrophin receptors, Trks and p75NTR, and the activation of JNK pathway in human astrocytomas and in adjacent non-neoplastic brain tissue. Formalin-fixed paraffin-embedded serial sections from 33 supratentorial astrocytomas (5 diffuse fibrillary astrocytomas, WHO grade II; 6 anaplastic astrocytomas, WHO grade III; 22 glioblastomas multiforme, WHO grade IV) were immunostained following microwave pretreatment. Polyclonal antibodies against TrkA, TrkB, TrkC and monoclonal antibodies against p75NTR and phosphorylated forms of JNK (pJNK) and c-Jun (pc-Jun) were used. The labeling index (LI), defined as the percentage of positive (labeled) cells out of the total number of tumor cells counted, was determined. Moderate to strong, granular cytoplasmic immunoreactivity for TrkA, TrkB and TrkC receptors was detected in greater than or equal to 10% of tumor cells in the majority of tumors independently of grade; on the contrary, p75NTR receptor expression was found in a small percentage of tumor cells (~1%) in some tumors. The endothelium of tumor capillaries showed conspicuous immunoreactivity for TrkB receptor. Trk immunoreactivity seemed to be localized in some neurons and astrocytes in non-neoplastic tissue. Phosphorylated forms of JNK (pJNK) and c-Jun (pc-Jun) were significantly co-expressed in a tumor grade

  10. Monitoring leptin activity using the chicken leptin receptor.

    Hen, Gideon; Yosefi, Sera; Ronin, Ana; Einat, Paz; Rosenblum, Charles I; Denver, Robert J; Friedman-Einat, Miriam

    2008-05-01

    We report on the construction of a leptin bioassay based on the activation of chicken leptin receptor in cultured cells. A human embryonic kidney (HEK)-293 cell line, stably transfected with the full-length cDNA of chicken leptin receptor together with a STAT3-responsive reporter gene specifically responded to recombinant human and Xenopus leptins. The observed higher sensitivity of chicken leptin receptor to the former is in agreement with the degree of sequence similarity among these species (about 60 and 38% identical amino acids between humans and chickens, and between humans and Xenopus respectively). The specific activation of signal transduction through the chicken leptin receptor, shown here for the first time, suggests that the transition of Gln269 (implicated in the Gln-to-Pro Zucker fatty mutation in rats) to Glu in chickens does not impair its activity. Analysis of leptin-like activity in human serum samples of obese and lean subjects coincided well with leptin levels determined by RIA. Serum samples of pre- and post partum cows showed a tight correlation with the degree of adiposity. However, specific activation of the chicken leptin receptor in this assay was not observed with serum samples from broiler or layer chickens (representing fat and lean phenotypes respectively) or with those from turkey. Similar leptin receptor activation profiles were observed with cells transfected with human leptin receptor. Further work is needed to determine whether the lack of leptin-like activity in the chicken serum samples is due to a lack of leptin in this species or simply to a serum level of leptin that is below the detection threshold. PMID:18434362

  11. Extracellular signal-regulated kinases control expression of G protein-coupled receptor kinase 2 (GRK2)

    Theilade, Juliane; Lerche Hansen, Jakob; Haunsø, Stig;

    2002-01-01

    the extracellular signal-regulated kinase (ERK) cascade to regulate GRK2 cellular levels. ERK activation by receptor stimulation elevated endogenous GRK2 while antagonist treatment decreased cellular GRK2. Activating ERK by overexpressing constitutive active MEK-1 or Ras elevated GRK2 protein levels while blocking...

  12. Mediators, Receptors, and Signalling Pathways in the Anti-Inflammatory and Antihyperalgesic Effects of Acupuncture

    John L. McDonald

    2015-01-01

    Full Text Available Acupuncture has been used for millennia to treat allergic diseases including both intermittent rhinitis and persistent rhinitis. Besides the research on the efficacy and safety of acupuncture treatment for allergic rhinitis, research has also investigated how acupuncture might modulate immune function to exert anti-inflammatory effects. A proposed model has previously hypothesized that acupuncture might downregulate proinflammatory neuropeptides, proinflammatory cytokines, and neurotrophins, modulating transient receptor potential vallinoid (TRPV1, a G-protein coupled receptor which plays a central role in allergic rhinitis. Recent research has been largely supportive of this model. New advances in research include the discovery of a novel cholinergic anti-inflammatory pathway activated by acupuncture. A chemokine-mediated proliferation of opioid-containing macrophages in inflamed tissues, in response to acupuncture, has also been demonstrated for the first time. Further research on the complex cross talk between receptors during inflammation is also helping to elucidate the mediators and signalling pathways activated by acupuncture.

  13. Cytoplasmic truncation of the p55 tumour necrosis factor (TNF) receptor abolishes signalling, but not induced shedding of the receptor

    Brakebusch, C; Nophar, Y; Kemper, O;

    1992-01-01

    The mechanistic relationship between the signalling for the TNF effects by the human p55 TNF receptor (hu-p55-TNF-R) and the formation of a soluble form of the receptor, which is inhibitory to these effects, was explored by examining the function of C-terminally truncated mutants of the receptor,...

  14. Neuroplastin-55 binds to and signals through the fibroblast growth factor receptor

    Owczarek, Sylwia; Kiryushko, Darya; Hald Larsen, Marianne;

    2010-01-01

    3), whereas Np55 lacks the Ig1 module. Of these two isoforms, only Np65 is involved in homophilic interactions resulting in cell adhesion, whereas the role of Np55 is poorly understood. The present study reports for the first time the crystal structure of the ectodomain of Np55 at 1.95-A resolution...... and demonstrates that Np55 binds to and activates the fibroblast growth factor receptor 1 (FGFR1). Furthermore, we identify a sequence motif in the Ig2 module of Np55 interacting with FGFR1 and show that a synthetic peptide encompassing this motif, termed narpin, binds to and activates FGFR1. We show...... Np55-induced signaling may be involved in synaptic plasticity in vivo. Owczarek, S., Kiryushko, D., Larsen, M. H., Kastrup, J. S., Gajhede, M., Sandi, C., Berezin, V., Bock, E., Soroka, V. Neuroplastin-55 binds to and signals through the fibroblast growth factor receptor....

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

    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...

  16. Sphingosine-1-phosphate receptor 1 reporter mice reveal receptor activation sites in vivo

    Kono, Mari; Tucker, Ana E.; Tran, Jennifer; Bergner, Jennifer B.; Turner, Ewa M; Proia, Richard L.

    2014-01-01

    Activation of the GPCR sphingosine-1-phosphate receptor 1 (S1P1) by sphingosine-1-phosphate (S1P) regulates key physiological processes. S1P1 activation also has been implicated in pathologic processes, including autoimmunity and inflammation; however, the in vivo sites of S1P1 activation under normal and disease conditions are unclear. Here, we describe the development of a mouse model that allows in vivo evaluation of S1P1 activation. These mice, known as S1P1 GFP signaling mice, produce a ...

  17. Influence of phasic and tonic dopamine release on receptor activation

    Dreyer, Jakob Kristoffer Kisbye; Herrik, Kjartan F; Berg, Rune W; Hounsgaard, Jørn D

    2010-01-01

    Tonic and phasic dopamine release is implicated in learning, motivation, and motor functions. However, the relationship between spike patterns in dopaminergic neurons, the extracellular concentration of dopamine, and activation of dopamine receptors remains unresolved. In the present study, we...... develop a computational model of dopamine signaling that give insight into the relationship between the dynamics of release and occupancy of D(1) and D(2) receptors. The model is derived from first principles using experimental data. It has no free parameters and offers unbiased estimation of the...

  18. Androgen insensitivity syndrome: gonadal androgen receptor activity

    To determine whether abnormalities of the androgen receptor previously observed in skin fibroblasts from patients with androgen insensitivity syndrome also occur in the gonads of affected individuals, androgen receptor activity in the gonads of a patient with testicular feminization syndrome was investigated. Using conditions for optimal recovery of androgen receptor from human testes established by previous studies, we detected the presence of a high-affinity (dissociation constant . 3.2 X 10(-10) mol/L), low-capacity (4.2 X 10(-12) mol/mg DNA), androgen-binding protein when tritium-labeled R1881 was incubated at 4 degrees C with nuclear extracts from the gonads of control patients or from a patient with testicular feminization syndrome but not when incubated at 37 degrees C. Thus this patient has an androgen receptor with a temperature lability similar to that of receptors from normal persons

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

    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

  20. Vitamin D Receptor Signaling and Pancreatic Cancer Cell EMT

    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

  1. Tyrosine kinase JAK1 is associated with the granulocyte-colony-stimulating factor receptor and both become tyrosine-phosphorylated after receptor activation.

    Nicholson, S. E.; Oates, A. C.; Harpur, A G; Ziemiecki, A; Wilks, A F; Layton, J E

    1994-01-01

    Granulocyte-colony-stimulating factor (G-CSF) stimulates the proliferation and differentiation of cells of the neutrophil lineage by interaction with a specific receptor. Early signal transduction events following G-CSF receptor activation were studied. We detected tyrosine phosphorylation of both the G-CSF receptor and the protein tyrosine kinase JAK1 following G-CSF binding to the human G-CSF receptor. In vitro, the kinase activity of JAK1 was increased by G-CSF stimulation. Coimmunoprecipi...

  2. Novel method for the study of receptor Ca2+ signalling exemplified by the NK1 receptor

    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...

  3. Inhibitory effects of omega-3 fatty acids on early brain injury after subarachnoid hemorrhage in rats: Possible involvement of G protein-coupled receptor 120/β-arrestin2/TGF-β activated kinase-1 binding protein-1 signaling pathway.

    Yin, Jia; Li, Haiying; Meng, Chengjie; Chen, Dongdong; Chen, Zhouqing; Wang, Yibin; Wang, Zhong; Chen, Gang

    2016-06-01

    Omega-3 fatty acids have been reported to improve neuron functions during aging and in patients affected by mild cognitive impairment, and mediate potent anti-inflammatory via G protein-coupled receptor 120 (GPR120) signal pathway. Neuron dysfunction and inflammatory response also contributed to the progression of subarachnoid hemorrhage (SAH)-induced early brain injury (EBI). This study was to examine the effects of omega-3 fatty acids on SAH-induced EBI. Two weeks before SAH, 30% Omega-3 fatty acids was administered by oral gavage at 1g/kg body weight once every 24h. Specific siRNA for GPR120 was exploited. Terminal deoxynucleotidyl transferase dUTP nick end labeling, fluoro-Jade B staining, and neurobehavioral scores and brain water content test showed that omega-3 fatty acids effectively suppressed SAH-induced brain cell apoptosis and neuronal degradation, behavioral impairment, and brain edema. Western blot, immunoprecipitation, and electrophoretic mobility shift assays results showed that omega-3 fatty acids effectively suppressed SAH-induced elevation of inflammatory factors, including cyclooxygenase-2, monocyte chemoattractant protein-1, and inducible nitric oxide synthase. In addition, omega-3 fatty acids could inhibit phosphorylation of transforming growth factor β activated kinase-1 (TAK1), MEK4, c-Jun N-terminal kinase, and IkappaB kinase as well as activation of nuclear factor kappa B through regulating GPR120/β-arrestin2/TAK1 binding protein-1 pathway. Furthermore, siRNA-induced GPR120 silencing blocked the protective effects of omega-3 fatty acids. Here, we show that stimulation of GPR120 with omega-3 fatty acids pretreatment causes anti-apoptosis and anti-inflammatory effects via β-arrestin2/TAK1 binding protein-1/TAK1 pathway in the brains of SAH rats. Fish omega-3 fatty acids as part of a daily diet may reduce EBI in an experimental rat model of SAH. PMID:27000704

  4. Bupleurum Polysaccharides Attenuates Lipopolysaccharide-Induced Inflammation via Modulating Toll-Like Receptor 4 Signaling

    Wu, Jian; Zhang, Yun-Yi; Guo, Li; LI Hong; Chen, Dao-Feng

    2013-01-01

    Background Bupleurum polysaccharides (BPs), isolated from Bupleurum smithii var. parvifolium, possesses immunomodulatory activity, particularly on inflammation. Bacterial endotoxin lipopolysaccharide (LPS) triggers innate immune responses through Toll-like receptor 4 (TLR4) on host cell membrane. The present study was performed to evaluate whether the therapeutic efficacy of BPs on suppression of LPS’s pathogenecity could be associated with the modulating of TLR4 signaling pathway. Methodolog...

  5. Nicotinic acetylcholine receptor-mediated calcium signaling in the nervous system

    Jian-xin SHEN; Jerrel L YAKEL

    2009-01-01

    Based on the composition of the five subunits forming functional neuronal nicotinic acetylcholine receptors (nAChRs), they are grouped into either heteromeric (comprising both α and β subunits) or homomeric (comprising only α subunits) recep-tors. The nAChRs are known to be differentially permeable to calcium ions, with the α7 nAChR subtype having one of the highest permeabilities to calcium. Calcium influx through nAChRs, particularly through the α-bungarotoxin-sensitive α7-containing nAChRs, is a very efficient way to raise cytoplasmic calcium levels. The activation of nAChRs can mediate three types of cytoplasmic calcium signals: (1) direct calcium influx through the nAChRs, (2) indirect calcium influx through voltage-dependent calcium channels (VDCCs) which are activated by the nAChR-mediated depolarization, and (3) calcium-induced calcium release (CICR) (triggered by the first two sources) from the endoplasmic reticulum (ER) through the ryanodine receptors and inositol (1,4,5)-triphosphate receptors (IP3Rs). Downstream signaling events mediated by nAChR-mediated calcium responses can be grouped into instantaneous effects (such as neurotransmitter release, which can occur in milliseconds after nAChR activation), short-term effects (such as the recovery of nAChR desensitization through cellular signaling cascades), and long-term effects (such as neuroprotection via gene expression). In addition, nAChR activity can be regulated by cytoplasmic calcium levels, suggesting a complex reciprocal relationship. Further advances in imaging techniques, animal models, and more potent and subtype-selective ligands for neuronal nAChRs would help in understand-ing the neuronal nAChR-mediated calcium signaling, and lead to the development of improved therapeutic treatments.

  6. Distinct Signaling Cascades Elicited by Different Formyl Peptide Receptor 2 (FPR2 Agonists

    Fabio Cattaneo

    2013-04-01

    Full Text Available The formyl peptide receptor 2 (FPR2 is a remarkably versatile transmembrane protein belonging to the G-protein coupled receptor (GPCR family. FPR2 is activated by an array of ligands, which include structurally unrelated lipids and peptide/proteins agonists, resulting in different intracellular responses in a ligand-specific fashion. In addition to the anti-inflammatory lipid, lipoxin A4, several other endogenous agonists also bind FPR2, including serum amyloid A, glucocorticoid-induced annexin 1, urokinase and its receptor, suggesting that the activation of FPR2 may result in potent pro- or anti-inflammatory responses. Other endogenous ligands, also present in biological samples, include resolvins, amyloidogenic proteins, such as beta amyloid (Aβ-42 and prion protein (Prp106–126, the neuroprotective peptide, humanin, antibacterial peptides, annexin 1-derived peptides, chemokine variants, the neuropeptides, vasoactive intestinal peptide (VIP and pituitary adenylate cyclase activating polypeptide (PACAP-27, and mitochondrial peptides. Upon activation, intracellular domains of FPR2 mediate signaling to G-proteins, which trigger several agonist-dependent signal transduction pathways, including activation of phospholipase C (PLC, protein kinase C (PKC isoforms, the phosphoinositide 3-kinase (PI3K/protein kinase B (Akt pathway, the mitogen-activated protein kinase (MAPK pathway, p38MAPK, as well as the phosphorylation of cytosolic tyrosine kinases, tyrosine kinase receptor transactivation, phosphorylation and nuclear translocation of regulatory transcriptional factors, release of calcium and production of oxidants. FPR2 is an attractive therapeutic target, because of its involvement in a range of normal physiological processes and pathological diseases. Here, we review and discuss the most significant findings on the intracellular pathways and on the cross-communication between FPR2 and tyrosine kinase receptors triggered by different FPR2

  7. Angiotensin II activates different calcium signaling pathways in adipocytes.

    Dolgacheva, Lyudmila P; Turovskaya, Maria V; Dynnik, Vladimir V; Zinchenko, Valery P; Goncharov, Nikolay V; Davletov, Bazbek; Turovsky, Egor A

    2016-03-01

    Angiotensin II (Ang II) is an important mammalian neurohormone involved in reninangiotensin system. Ang II is produced both constitutively and locally by RAS systems, including white fat adipocytes. The influence of Ang II on adipocytes is complex, affecting different systems of signal transduction from early Са(2+) responses to cell proliferation and differentiation, triglyceride accumulation, expression of adipokine-encoding genes and adipokine secretion. It is known that white fat adipocytes express all RAS components and Ang II receptors (АТ1 and АТ2). The current work was carried out with the primary white adipocytes culture, and Са(2+) signaling pathways activated by Ang II were investigated using fluorescent microscopy. Са(2+)-oscillations and transient responses of differentiated adipocytes to Ang II were registered in cells with both small and multiple lipid inclusions. Using inhibitory analysis and selective antagonists, we now show that Ang II initiates periodic Са(2+)-oscillations and transient responses by activating АТ1 and АТ2 receptors and involving branched signaling cascades:In these cascades, AT1 receptors play the leading role. The results of the present work open a perspective of using Ang II for correction of signal resistance of adipocytes often observed during obesity and type 2diabetes. PMID:26850364

  8. Signal transduction by VEGF receptors in regulation of angiogenesis and lymphangiogenesis

    The VEGF/VPF (vascular endothelial growth factor/vascular permeability factor) ligands and receptors are crucial regulators of vasculogenesis, angiogenesis, lymphangiogenesis and vascular permeability in vertebrates. VEGF-A, the prototype VEGF ligand, binds and activates two tyrosine kinase receptors: VEGFR1 (Flt-1) and VEGFR2 (KDR/Flk-1). VEGFR1, which occurs in transmembrane and soluble forms, negatively regulates vasculogenesis and angiogenesis during early embryogenesis, but it also acts as a positive regulator of angiogenesis and inflammatory responses, playing a role in several human diseases such as rheumatoid arthritis and cancer. The soluble VEGFR1 is overexpressed in placenta in preeclampsia patients. VEGFR2 has critical functions in physiological and pathological angiogenesis through distinct signal transduction pathways regulating proliferation and migration of endothelial cells. VEGFR3, a receptor for the lymphatic growth factors VEGF-C and VEGF-D, but not for VEGF-A, regulates vascular and lymphatic endothelial cell function during embryogenesis. Loss-of-function variants of VEGFR3 have been identified in lymphedema. Formation of tumor lymphatics may be stimulated by tumor-produced VEGF-C, allowing increased spread of tumor metastases through the lymphatics. Mapping the signaling system of these important receptors may provide the knowledge necessary to suppress specific signaling pathways in major human diseases

  9. Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.

    Hannah Karlsson

    Full Text Available CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs.

  10. Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.

    Karlsson, Hannah; Svensson, Emma; Gigg, Camilla; Jarvius, Malin; Olsson-Strömberg, Ulla; Savoldo, Barbara; Dotti, Gianpietro; Loskog, Angelica

    2015-01-01

    CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G) CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G) CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs. PMID:26700307

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

    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. ...

  12. Dual intracellular signaling pathways mediated by the human cannabinoid CB1 receptor.

    Calandra, B; Portier, M; Kernéis, A; Delpech, M; Carillon, C; Le Fur, G; Ferrara, P; Shire, D

    1999-06-25

    It has long been established that the cannabinoid CB1 receptor transduces signals through a pertussis toxin-sensitive Gi/Go inhibitory pathway. Although there have been reports that the cannabinoid CB1 receptor can also mediate an increase in cyclic AMP levels, in most cases the presence of an adenylyl cyclase costimulant or the use of very high amounts of agonist was necessary. Here, we present evidence for dual coupling of the cannabinoid CB receptor to the classical pathway and to a pertussis toxin-insensitive adenylyl cyclase stimulatory pathway initiated with low quantities of agonist in the absence of any costimulant. Treatment of Chinese hamster ovary (CHO) cells expressing the cannabinoid CB1 receptor with the cannabinoid CP 55,940, {(-)-cis-3-[2-hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans-4-(3-hyd roxypropyl) cyclohexan-1-ol} resulted in cyclic AMP accumulation in a dose-response manner, an accumulation blocked by the cannabinoid CB1 receptor-specific antagonist SR 141716A, {N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-me thyl-1H-pyrazole-3-carboxamide hydrochloride}. In CHO cells coexpressing the cannabinoid CB1 receptor and a cyclic AMP response element (CRE)-luciferase reporter gene system, CP 55,940 induced luciferase expression by a pathway blocked by the protein kinase A inhibitor N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide hydrochloride (H-89). Under the same conditions the peripheral cannabinoid CB2 receptor proved to be incapable of inducing cAMP accumulation or luciferase activity. This incapacity allowed us to study the luciferase activation mediated by CB /CB2 chimeric constructs, from which we determined that the first and second internal loop regions of the cannabinoid CB1 receptor were involved in transducing the pathway leading to luciferase gene expression. PMID:10422789

  13. Mechanisms Governing the Activation and Trafficking of Yeast G Protein-coupled Receptors

    Stefan, Christopher J.; Overton, Mark C.; Blumer, Kendall J.

    1998-01-01

    We have addressed the mechanisms governing the activation and trafficking of G protein-coupled receptors (GPCRs) by analyzing constitutively active mating pheromone receptors (Ste2p and Ste3p) of the yeast Saccharomyces cerevisiae. Substitution of the highly conserved proline residue in transmembrane segment VI of these receptors causes constitutive signaling. This proline residue may facilitate folding of GPCRs into native, inactive conformations, and/or mediate a...

  14. The effect of CD4 receptor downregulation and its downstream signaling molecules on HIV-1 latency

    Kim, Kyung-Chang [National Institute of Health, Chungbuk (Korea, Republic of); School of Life Science and Biotechnology, Korea University, Seoul (Korea, Republic of); Kim, Hyeon Guk [National Institute of Health, Chungbuk (Korea, Republic of); Roh, Tae-Young [Division of Molecular and Life Science, Pohang University of Science and Technology, Pohang, Gyeongbuk (Korea, Republic of); Division of Integrative Biosciences and Biotechnology, Pohang University of Science and Technology, Pohang, Gyeongbuk (Korea, Republic of); Park, Jihwan [Division of Molecular and Life Science, Pohang University of Science and Technology, Pohang, Gyeongbuk (Korea, Republic of); Jung, Kyung-Min; Lee, Joo-Shil [National Institute of Health, Chungbuk (Korea, Republic of); Choi, Sang-Yun [School of Life Science and Biotechnology, Korea University, Seoul (Korea, Republic of); Kim, Sung Soon [National Institute of Health, Chungbuk (Korea, Republic of); Choi, Byeong-Sun, E-mail: byeongsun@korea.kr [National Institute of Health, Chungbuk (Korea, Republic of)

    2011-01-14

    Research highlights: {yields} CD4 receptors were downregulated on the surface of HIV-1 latently infected cells. {yields} CD4 downstream signaling molecules were suppressed in HIV-1 latently infected cells. {yields} HIV-1 progeny can be reactivated by induction of T-cell activation signal molecules. {yields} H3K4me3 and H3K9ac were highly enriched in CD4 downstream signaling molecules. {yields} HIV-1 latency can be maintained by the reduction of downstream signaling molecules. -- Abstract: HIV-1 can establish a latent infection in memory CD4 + T cells to evade the host immune response. CD4 molecules can act not only as the HIV-1 receptor for entry but also as the trigger in an intracellular signaling cascade for T-cell activation and proliferation via protein tyrosine kinases. Novel chronic HIV-1-infected A3.01-derived (NCHA) cells were used to examine the involvement of CD4 downstream signaling in HIV-1 latency. CD4 receptors in NCHA cells were dramatically downregulated on its surface but were slightly decreased in whole-cell lysates. The expression levels of CD4 downstream signaling molecules, including P56{sup Lck}, ZAP-70, LAT, and c-Jun, were sharply decreased in NCHA cells. The lowered histone modifications of H3K4me3 and H3K9ac correlated with the downregulation of P56{sup Lck}, ZAP-70, and LAT in NCHA cells. AP-1 binding activity was also reduced in NCHA cells. LAT and c-Jun suppressed in NCHA cells were highly induced after PMA treatment. In epigenetic analysis, other signal transduction molecules which are associated with active and/or latent HIV-1 infection showed normal states in HIV-1 latently infected cells compared to A3.01 cells. In conclusion, we demonstrated that the HIV-1 latent state is sustained by the reduction of downstream signaling molecules via the downregulation of CD4 and the attenuated activity of transcription factor as AP-1. The HIV-1 latency model via T-cell deactivation may provide some clues for the development of the new

  15. The effect of CD4 receptor downregulation and its downstream signaling molecules on HIV-1 latency

    Research highlights: → CD4 receptors were downregulated on the surface of HIV-1 latently infected cells. → CD4 downstream signaling molecules were suppressed in HIV-1 latently infected cells. → HIV-1 progeny can be reactivated by induction of T-cell activation signal molecules. → H3K4me3 and H3K9ac were highly enriched in CD4 downstream signaling molecules. → HIV-1 latency can be maintained by the reduction of downstream signaling molecules. -- Abstract: HIV-1 can establish a latent infection in memory CD4 + T cells to evade the host immune response. CD4 molecules can act not only as the HIV-1 receptor for entry but also as the trigger in an intracellular signaling cascade for T-cell activation and proliferation via protein tyrosine kinases. Novel chronic HIV-1-infected A3.01-derived (NCHA) cells were used to examine the involvement of CD4 downstream signaling in HIV-1 latency. CD4 receptors in NCHA cells were dramatically downregulated on its surface but were slightly decreased in whole-cell lysates. The expression levels of CD4 downstream signaling molecules, including P56Lck, ZAP-70, LAT, and c-Jun, were sharply decreased in NCHA cells. The lowered histone modifications of H3K4me3 and H3K9ac correlated with the downregulation of P56Lck, ZAP-70, and LAT in NCHA cells. AP-1 binding activity was also reduced in NCHA cells. LAT and c-Jun suppressed in NCHA cells were highly induced after PMA treatment. In epigenetic analysis, other signal transduction molecules which are associated with active and/or latent HIV-1 infection showed normal states in HIV-1 latently infected cells compared to A3.01 cells. In conclusion, we demonstrated that the HIV-1 latent state is sustained by the reduction of downstream signaling molecules via the downregulation of CD4 and the attenuated activity of transcription factor as AP-1. The HIV-1 latency model via T-cell deactivation may provide some clues for the development of the new antireservoir therapy.

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

    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.

  17. The role of GPCR dimerisation/oligomerisation in receptor signalling.

    Milligan, G; Canals, M; Pediani, J D; Ellis, J; Lopez-Gimenez, J F

    2006-01-01

    A wide range of techniques have been employed to examine the quaternary structure of G-protein-coupled receptors (GPCRs). Although it is well established that homo-dimerisation is common, recent studies have sought to explore the physical basis of these interactions and the role of dimerisation in signal transduction. Growing evidence hints at the existence of higher-order organisation of individual GPCRs and the potential for hetero-dimerisation between pairs of co-expressed GPCRs. Here we consider how both homo-dimerisation/oligomerisation and hetero-dimerisation can regulate signal transduction through GPCRs and the potential consequences of this for function of therapeutic medicines that target GPCRs. Hetero-dimerisation is not the sole means by which co-expressed GPCRs may regulate the function of one another. Heterologous desensitisation may be at least as important and we also consider if this can be the basis for physiological antagonism between pairs of co-expressed GPCRs. Although there may be exceptions (Meyer et al. 2006), a great deal of recent evidence has indicated that most G-protein-coupled receptors (GPCRs) do not exist as monomers but rather as dimers or, potentially, within higher-order oligomers (Milligan 2004b; Park et al. 2004). Support for such models has been provided by a range of studies employing different approaches, including co-immunoprecipitation of differentially epitope-tagged but co-expressed forms of the same GPCR, co-operativity in ligand binding and a variety of resonance energy transfer techniques (Milligan and Bouvier 2005). Only for the photon receptor rhodopsin has the organisational structure of a GPCR been studied in situ. The application of atomic force microscopy to murine rod outer segment discs indicated that rhodopsin is organised in a series of parallel arrays of dimers (Liang et al. 2003) and based on this, molecular models were constructed to try to define and interpret regions of contact between the monomers

  18. Honey bee dopamine and octopamine receptors linked to intracellular calcium signaling have a close phylogenetic and pharmacological relationship.

    Kyle T Beggs

    Full Text Available BACKGROUND: Three dopamine receptor genes have been identified that are highly conserved among arthropod species. One of these genes, referred to in honey bees as Amdop2, shows a close phylogenetic relationship to the a-adrenergic-like octopamine receptor family. In this study we examined in parallel the functional and pharmacological properties of AmDOP2 and the honey bee octopamine receptor, AmOA1. For comparison, pharmacological properties of the honey bee dopamine receptors AmDOP1 and AmDOP3, and the tyramine receptor AmTYR1, were also examined. METHODOLOGY/PRINCIPAL FINDINGS: Using HEK293 cells heterologously expressing honey bee biogenic amine receptors, we found that activation of AmDOP2 receptors, like AmOA1 receptors, initiates a rapid increase in intracellular calcium levels. We found no evidence of calcium signaling via AmDOP1, AmDOP3 or AmTYR1 receptors. AmDOP2- and AmOA1-mediated increases in intracellular calcium were inhibited by 10 µM edelfosine indicating a requirement for phospholipase C-β activity in this signaling pathway. Edelfosine treatment had no effect on AmDOP2- or AmOA1-mediated increases in intracellular cAMP. The synthetic compounds mianserin and epinastine, like cis-(Z-flupentixol and spiperone, were found to have significant antagonist activity on AmDOP2 receptors. All 4 compounds were effective antagonists also on AmOA1 receptors. Analysis of putative ligand binding sites offers a possible explanation for why epinastine acts as an antagonist at AmDOP2 receptors, but fails to block responses mediated via AmDOP1. CONCLUSIONS/SIGNIFICANCE: Our results indicate that AmDOP2, like AmOA1, is coupled not only to cAMP, but also to calcium-signalling and moreover, that the two signalling pathways are independent upstream of phospholipase C-β activity. The striking similarity between the pharmacological properties of these 2 receptors suggests an underlying conservation of structural properties related to receptor

  19. Heat dissipation guides activation in signaling proteins

    Weber, Jeffrey K.; Shukla, Diwakar; Pande, Vijay S.

    2015-01-01

    As with their macroscopic counterparts, the moving parts of nanoscale protein machines grow hot while in operation. A portion of the energy biomolecules harness to perform meaningful work is always dissipated as heat into the surroundings. Here, we feature a methodology by which dominant dissipative trajectories can be extracted from detailed models of protein dynamics. In two important classes of signaling proteins [kinases and G-protein–coupled receptors (GPCRs)], we find that the regions o...

  20. Constitutive Activity among Orphan Class-A G Protein Coupled Receptors

    Martin, Adam L.; Michael A Steurer; Aronstam, Robert S.

    2015-01-01

    The purpose of this study was to evaluate the extent of constitutive activity among orphan class-A G protein coupled receptors within the cAMP signaling pathway. Constitutive signaling was revealed by changes in gene expression under control of the cAMP response element. Gene expression was measured in Chinese hamster ovary cells transiently co-transfected with plasmids containing a luciferase reporter and orphan receptor. Criteria adopted for defining constitutive activation were: 1) 200% el...

  1. Functional requirements for inhibitory signal transmission by the immunomodulatory receptor CD300a

    DeBell Karen E

    2012-04-01

    Full Text Available Abstract Background Activation signals can be negatively regulated by cell surface receptors bearing immunoreceptor tyrosine-based inhibitory motifs (ITIMs. CD300a, an ITIM bearing type I transmembrane protein, is expressed on many hematopoietic cells, including subsets of lymphocytes. Results We have taken two approaches to further define the mechanism by which CD300a acts as an inhibitor of immune cell receptor signaling. First, we have expressed in Jurkat T cells a chimeric receptor consisting of the extracellular domains of killer-cell immunoglobulin-like receptor (KIR2DL2 fused to the transmembrane and cytoplasmic segments of CD300a (KIR-CD300a to explore surrogate ligand-stimulated inhibition of superantigen stimulated T cell receptor (TCR mediated cell signaling. We found that intact CD300a ITIMs were essential for inhibition and that the tyrosine phosphorylation of these ITIMs required the src tyrosine kinase Lck. Tyrosine phosphorylation of the CD300a ITIMs created docking sites for both src homology 2 domain containing protein tyrosine phosphatase (SHP-1 and SHP-2. Suppression of SHP-1 and SHP-2 expression in KIR-CD300a Jurkat T cells with siRNA and the use of DT40 chicken B cell lines expressing CD300a and deficient in several phosphatases revealed that SHP-1, but not SHP-2 or the src homology 2 domain containing inositol 5’ phosphatase SHIP, was utilized by CD300a for its inhibitory activity. Conclusion These studies provide new insights into the function of CD300a in tuning T and B cell responses.

  2. Exercício de força ativa a via AKT/mTor pelo receptor de angiotensina II tipo I no músculo cardíaco de ratos Activation of AKT-mTor signaling pathways by angiotensin II receptor type 1 after a session of strength exercise in cardiac muscle of rats

    Stéphano Freitas Soares Melo

    2011-09-01

    Full Text Available O receptor de angiotensina II tipo I (AT1 tem uma importante participação no desenvolvimento da hipertrofia cardíaca. Em um trabalho publicado anteriormente, por nosso grupo, demonstramos que o bloqueio do receptor AT1 durante o treinamento de força inibiu a hipertrofia cardíaca em ratos. Por isso, o objetivo deste trabalho foi estudar a participação do receptor AT1 na ativação de vias de sinalização intracelular relacionadas com o aumento da síntese de proteína em ratos submetidos a uma sessão de exercício de força. Para isso, realizamos um experimento com seis grupos de animais (n = 6; cada: controle (Con, exercitado e sacrificado cinco minutos após o exercício (Exe 5, exercitado e sacrificado 30 minutos após o exercício (Exe 30, controle tratado com losartan (Con Los, tratado com losartan, exercitado e sacrificado cinco minutos após o exercício (Exe 5 Los, tratado com losartan, exercitado e sacrificado 30 minutos após o exercício (Exe 30 Los. Os resultados mostram que no grupo Exe 5 e Exe 30 ocorreu um aumento de 63% (P The angiotensin II type I (AT1 receptor has an important participation in the development of cardiac hypertrophy. Previously, we have shown that AT1 receptor participates in the cardiac hypertrophy induced by resistance training in rats. Here, we studied the involvement of AT1 receptor in the activation of intracellular signaling pathways related to the concentric HC in rats submitted to a session of strength exercise. Male Wistar rats were divided into 6 groups (n= 6 each: control (Con; exercised and killed 5 minutes after exercise (Exe 5; exercised and killed 30 minutes after exercise (Exe 30; control treated with Losartan (Con Los; treated with Losartan, exercised and killed 5 minutes after the exercise (Exe Los 5; treated with Losartan, exercised and killed 30 minutes after training (Exe Los 30. The results show that phosphorylation activity of AKT in group Exe 5 and Exe 30 increased 63% (P < 0

  3. Dopamine signaling in food addiction: role of dopamine D2 receptors

    Ja-Hyun Baik

    2013-11-01

    Full Text Available Dopamine (DA regulates emotional and motivationalbehavior through the mesolimbic dopaminergic pathway.Changes in DA signaling in mesolimbic neurotransmission arewidely believed to modify reward-related behaviors and aretherefore closely associated with drug addiction. Recentevidence now suggests that as with drug addiction, obesitywith compulsive eating behaviors involves reward circuitry ofthe brain, particularly the circuitry involving dopaminergicneural substrates. Increasing amounts of data from humanimaging studies, together with genetic analysis, havedemonstrated that obese people and drug addicts tend to showaltered expression of DA D2 receptors in specific brain areas,and that similar brain areas are activated by food-related anddrug-related cues. This review focuses on the functions of theDA system, with specific focus on the physiological interpretationand the role of DA D2 receptor signaling in foodaddiction. [BMB Reports 2013; 46(11: 519-526

  4. Enhancement of arachidonic acid signaling pathway by nicotinic acid receptor HM74A

    HM74A is a G protein-coupled receptor for nicotinic acid (niacin), which has been used clinically to treat dyslipidemia for decades. The molecular mechanisms whereby niacin exerts its pleiotropic effects on lipid metabolism remain largely unknown. In addition, the most common side effect in niacin therapy is skin flushing that is caused by prostaglandin release, suggesting that the phospholipase A2 (PLA2)/arachidonic acid (AA) pathway is involved. Various eicosanoids have been shown to activate peroxisome-proliferator activated receptors (PPAR) that play a diverse array of roles in lipid metabolism. To further elucidate the potential roles of HM74A in mediating the therapeutic effects and/or side effects of niacin, we sought to explore the signaling events upon HM74A activation. Here we demonstrated that HM74A synergistically enhanced UTP- and bradykinin-mediated AA release in a pertussis toxin-sensitive manner in A431 cells. Activation of HM74A also led to Ca2+-mobilization and enhanced bradykinin-promoted Ca2+-mobilization through Gi protein. While HM74A increased ERK1/2 activation by the bradykinin receptor, it had no effects on UTP-promoted ERK1/2 activation.Furthermore, UTP- and bradykinin-mediated AA release was significantly decreased in the presence of both MAPK kinase inhibitor PD 098059 and PKC inhibitor GF 109203X. However, the synergistic effects of HM74A were not dramatically affected by co-treatment with both inhibitors, indicating the cross-talk occurred at the receptor level. Finally, stimulation of A431 cells transiently transfected with PPRE-luciferase with AA significantly induced luciferase activity, mimicking the effects of PPARγ agonist rosiglitazone, suggesting that alteration of AA signaling pathway can regulate gene expression via endogenous PPARs

  5. Mechanism of FGF receptor dimerization and activation

    Sarabipour, Sarvenaz; Hristova, Kalina

    2016-01-01

    Fibroblast growth factors (fgfs) are widely believed to activate their receptors by mediating receptor dimerization. Here we show, however, that the FGF receptors form dimers in the absence of ligand, and that these unliganded dimers are phosphorylated. We further show that ligand binding triggers structural changes in the FGFR dimers, which increase FGFR phosphorylation. The observed effects due to the ligands fgf1 and fgf2 are very different. The fgf2-bound dimer structure ensures the smallest separation between the transmembrane (TM) domains and the highest possible phosphorylation, a conclusion that is supported by a strong correlation between TM helix separation in the dimer and kinase phosphorylation. The pathogenic A391E mutation in FGFR3 TM domain emulates the action of fgf2, trapping the FGFR3 dimer in its most active state. This study establishes the existence of multiple active ligand-bound states, and uncovers a novel molecular mechanism through which FGFR-linked pathologies can arise.

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

    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.

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

    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...

  8. A Perspective on Studying G-Protein-Coupled Receptor Signaling with Resonance Energy Transfer Biosensors in Living Organisms

    Unen, J. van; Woolard, J.; Rinken, A.; Hoffmann, C.; Hill, S.J.; Goedhart, J.; Bruchas, M.R.; Bouvier, M.; Adjobo-Hermans, M.J.W.

    2015-01-01

    The last frontier for a complete understanding of G-protein-coupled receptor (GPCR) biology is to be able to assess GPCR activity, interactions, and signaling in vivo, in real time within biologically intact systems. This includes the ability to detect GPCR activity, trafficking, dimerization, prote

  9. Activation of Wnt signaling bypasses the requirement for RTK/Ras signaling during C. elegans vulval induction

    Gleason, Julie E.; Korswagen, Hendrik C.; Eisenmann, David M

    2002-01-01

    During Caenorhabditis elegans vulval development, activation of receptor tyrosine kinase/Ras and Notch signaling pathways causes three vulval precursor cells (VPCs) to adopt induced cell fates. A Wnt signaling pathway also acts in cell fate specification by the VPCs, via regulation of the Hox gene lin-39. We show here that either mutation of pry-1 or expression of an activated BAR-1 β-catenin protein causes an Overinduced phenotype, in which greater than three VPCs adopt induced cell fates. T...

  10. ATP release and purinergic signaling in NLRP3 inflammasome activation

    Isabelle eCOUILLIN

    2013-01-01

    Full Text Available The NLRP3 inflammasome is a protein complex involved in IL-1β and IL-18 processing that senses pathogen- and danger-associated molecular patterns. One step- or two step- models have been proposed to explain the tight regulation of IL-1β production during inflammation. Moreover, cellular stimulation triggers ATP release and subsequent activation of purinergic receptors at the cell surface. Importantly some studies have reported roles for extracellular ATP (eATP, in NLRP3 inflammasome activation in response to PAMPs and DAMPs. In this mini review, we will discuss the link between active ATP release, purinergic signaling and NLRP3 inflammasome activation. We will focus on the role of autocrine or paracrine ATP export in particle-induced NLRP3 inflammasome activation and discuss how particle activators are competent to induce maturation and secretion of IL-1β through a process that involves, as a first event, extracellular release of endogenous ATP through hemichannel opening, and as a second event, signaling through purinergic receptors that trigger NLRP3 inflammasome activation. Finally, we will review the evidence for ATP as a key proinflammatory mediator released by dying cells. In particular we will discuss how cancer cells dying via autophagy trigger ATP-dependent NLRP3 inflammasome activation in the macrophages engulfing them, eliciting an immunogenic response against tumors.

  11. Potentiation of nerve growth factor-induced neurite outgrowth by fluvoxamine: role of sigma-1 receptors, IP3 receptors and cellular signaling pathways.

    Tomoko Nishimura

    Full Text Available BACKGROUND: Selective serotonin reuptake inhibitors (SSRIs have been widely used and are a major therapeutic advance in psychopharmacology. However, their pharmacology is quite heterogeneous. The SSRI fluvoxamine, with sigma-1 receptor agonism, is shown to potentiate nerve-growth factor (NGF-induced neurite outgrowth in PC 12 cells. However, the precise cellular and molecular mechanisms underlying potentiation by fluvoxamine are not fully understood. In this study, we examined the roles of cellular signaling pathways in the potentiation of NGF-induced neurite outgrowth by fluvoxamine and sigma-1 receptor agonists. METHODS AND FINDINGS: The effects of three SSRIs (fluvoxamine, sertraline, paroxetine and three sigma-1 receptor agonists (SA4503, 4-phenyl-1-(4-phenylbutyl piperidine (PPBP, and dehydroepiandrosterone (DHEA-sulfate on NGF-induced neurite outgrowth in PC12 cells were examined. Also examined were the effects of the sigma-1 receptor antagonist NE-100, inositol 1,4,5-triphosphate (IP(3 receptor antagonist, and specific inhibitors of signaling pathways in the potentiation of NGF-induced neurite outgrowth by selective sigma-1 receptor agonist SA4503. Fluvoxamine (but not sertraline or paroxetine and the sigma-1 receptor agonists SA4503, PPBP, and DHEA-sulfate significantly potentiated NGF-induced neurite outgrowth in PC12 cells in a concentration-dependent manner. The potentiation by fluvoxamine and the three sigma-1 receptor agonists was blocked by co-administration of the selective sigma-1 receptor antagonist NE-100, suggesting that sigma-1 receptors play a role in blocking the enhancement of NGF-induced neurite outgrowth. Moreover, the potentiation by SA4503 was blocked by co-administration of the IP(3 receptor antagonist xestospongin C. In addition, the specific inhibitors of phospholipase C (PLC-gamma, phosphatidylinositol 3-kinase (PI3K, p38MAPK, c-Jun N-terminal kinase (JNK, and the Ras/Raf/mitogen-activated protein kinase (MAPK

  12. Anxiolytic activity of adenosine receptor activation in mice.

    Jain, N; Kemp, N; Adeyemo, O; Buchanan, P; Stone, T W

    1995-10-01

    1. Purine analogues have been examined for anxiolytic- and anxiogenic-like activity in mice, by use of the elevated plus-maze. 2. The selective A1 receptor agonist, N6-cyclopentyladenosine (CPA) had marked anxiolytic-like activity at 10 and 50 microg kg(-1), with no effect on locomotor performance at these doses. 3. The A1 selective adenosine receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine (CPX) had no significant effect on anxiety-related measures or locomotor behaviour, but blocked the anxiolytic-like activity of CPA. The hydrophilic xanthine, 8-(p-sulphophenyl) theophylline did not prevent anxiolysis by CPA. 4. Caffeine had anxiogenic-like activity at 30 mg kg(-1) which was prevented by CPA at 50 micro kg(-1). 5. The A2 receptor agonist, N6-[2-(3,5-dimethoxyphenyl)-2(2-methylphenyl)-ethyl]adenosine (DPMA) had no effect on anxiety behaviour but depressed locomotor activity at the highest dose tested of 1 mg kg(-1). The A2 receptor antagonist, 1,3-dimethyl-l-propargylxanthine (DMPX) had no effect on anxiety-related measures or locomotion and did not modify the anxiolytic-like activity of CPA. 6. Administration of DPMA in combination with anxiolytic doses of CPA prevented the anxiolytic-like activity of the latter. 7. The results suggest that the selective activation of central A1 adenosine receptors induces anxiolytic-like behaviour, while the activation of A2 sites causes locomotor depression and reduces the effects of A1 receptor activation. The absence of any effect of CPX alone suggests that the receptors involved in modulating behaviour in the elevated plus-maze in mice are not activated tonically by endogenous adenosine. PMID:8640355

  13. The sensing of bacteria: emerging principles for the detection of signal sequences by formyl peptide receptors.

    Bufe, Bernd; Zufall, Frank

    2016-06-01

    The ability to detect specific chemical signatures released by bacteria and other microorganisms is a fundamental feature of immune defense against pathogens. There is increasing evidence that chemodetection of such microorganism-associated molecular patterns (MAMPs) occurs at many places in the body including specific sets of chemosensory neurons in the mammalian nose. Formyl peptide receptors (FPRs) are a unique family of G protein-coupled receptors (GPCRs) that can detect the presence of bacteria and function as chemotactic receptors. Here, we highlight the recent discovery of a vast family of natural FPR agonists, the bacterial signal peptides (or signal sequences), thus providing new insight into the molecular mechanisms of bacterial sensing by human and mouse FPRs. Signal peptides in bacteria are formylated, N-terminal protein signatures required for directing the transfer of proteins through the plasma membrane. After their cleavage and release, signal peptides are available for FPR detection and thus provide a previously unrecognized MAMP. With over 170 000 predicted sequences, bacterial signal peptides represent one of the largest families of GPCR ligands and one of the most complex classes of natural activators of the innate immune system. By recognizing a conserved three-dimensional peptide motif, FPRs employ an unusual detection mechanism that combines structural promiscuity with high specificity and sensitivity, thus solving the problem of detecting thousands of distinct sequences yet maintaining selectivity. How signal peptides are released by bacteria and sensed by GPCRs and how these processes shape the responses of other cells and whole organisms represents an important topic for future research. PMID:27305707

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

    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.

  15. Purinergic receptor-induced Ca2+ signaling in the neuroepithelium of the vomeronasal organ of larval Xenopus laevis

    Dittrich, Katarina; Sansone, Alfredo; Hassenklöver, Thomas; Manzini, Ivan

    2013-01-01

    Purinergic signaling has considerable impact on the functioning of the nervous system, including the special senses. Purinergic receptors are expressed in various cell types in the retina, cochlea, taste buds, and the olfactory epithelium. The activation of these receptors by nucleotides, particularly adenosine-5'-triphosphate (ATP) and its breakdown products, has been shown to tune sensory information coding to control the homeostasis and to regulate the cell turnover in these organs. While ...

  16. Apo-ghrelin receptor (apo-GHSR1a Regulates Dopamine Signaling in the Brain

    Andras eKern

    2014-08-01

    Full Text Available The orexigenic peptide hormone ghrelin is synthesized in the stomach and its receptor growth hormone secretagogue receptor (GHSR1a is expressed mainly in the central nervous system (CNS. In this review we confine our discussion to the physiological role of GHSR1a in the brain. Paradoxically, despite broad expression of GHSR1a in the CNS, other than trace amounts in the hypothalamus, ghrelin is undetectable in the brain. In our efforts to elucidate the function of the ligand-free ghrelin receptor (apo-GHSR1a we identified subsets of neurons that co-express GHSR1a and dopamine receptors. In this review we focus on interactions between apo-GHSR1a and dopamine-2 receptor (DRD2 and formation of GHSR1a:DRD2 heteromers in hypothalamic neurons that regulate appetite, and discuss implications for the treatment of Prader-Willi syndrome. GHSR1a antagonists of distinct chemical structures, a quinazolinone and a triazole, respectively enhance and inhibit dopamine signaling through GHSR1a:DRD2 heteromers by an allosteric mechanism. This finding illustrates a potential strategy for designing the next generation of drugs for treating eating disorders as well as psychiatric disorders caused by abnormal dopamine signaling. Treatment with a GHSR1a antagonist that enhances dopamine/DRD2 activity in GHSR1a:DRD2 expressing hypothalamic neurons has the potential to inhibit the uncontrollable hyperphagia associated with Prader-Willi syndrome. DRD2 antagonists are prescribed for treating schizophrenia, but these block dopamine signaling in all DRD2 expressing neurons and are associated with adverse side effects, including enhanced appetite and excessive weight gain. A GHSR1a antagonist of structural class that allosterically blocks dopamine/DRD2 action in GHSR1a:DRD2 expressing neurons would have no effect on neurons expressing DRD2 alone; therefore, the side effects of DRD2 antagonists would potentially be reduced thereby enhancing patient compliance.

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

    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.

  18. Pleiotropic effects of Blastocystis spp. Subtypes 4 and 7 on ligand-specific toll-like receptor signaling and NF-κB activation in a human monocyte cell line.

    Joshua D W Teo

    Full Text Available Blastocystis spp. is a common enteric stramenopile parasite that colonizes the colon of hosts of a diverse array of species, including humans. It has been shown to compromise intestinal epithelial cell barrier integrity and mediate the production of pro-inflammatory cytokines and chemokines. Mucosal epithelial surfaces, including the intestinal epithelium, are increasingly recognized to perform a vital surveillance role in the context of innate immunity, through the expression of pathogen recognition receptors, such as Toll-like receptors (TLRs. In this study, we use the human TLR reporter monocytic cell line, THP1-Blue, which expresses all human TLRs, to investigate effects of Blastocystis on TLR activation, more specifically the activation of TLR-2, -4 and -5. We have observed that live Blastocystis spp. parasites and whole cell lysate (WCL alone do not activate TLRs in THP1-Blue. Live ST4-WR1 parasites inhibited LPS-mediated NF-κB activation in THP1-Blue. In contrast, ST7-B WCL and ST4-WR1 WCL induced pleiotropic modulation of ligand-specific TLR-2 and TLR-4 activation, with no significant effects on flagellin-mediated TLR-5 activation. Real time-qPCR analysis on SEAP reporter gene confirmed the augmenting effect of ST7-B on LPS-mediated NF-κB activation in THP1-Blue. Taken together, this is the first study to characterize interactions between Blastocystis spp. and host TLR activation using an in vitro reporter model.

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

    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.

  20. A gate-latch-lock mechanism for hormone signalling by abscisic acid receptors

    Melcher, Karsten; Ng, Ley-Moy; Zhou, X Edward; Soon, Fen-Fen; Xu, Yong; Suino-Powell, Kelly M; Park, Sang-Youl; Weiner, Joshua J; Fujii, Hiroaki; Chinnusamy, Viswanathan; Kovach, Amanda; Li, Jun; Wang, Yonghong; Li, Jiayang; Peterson, Francis C; Jensen, Davin R; Yong, Eu-Leong; Volkman, Brian F; Cutler, Sean R; Zhu, Jian-Kang; Xu, H Eric; (NU Sinapore); (Van Andel); (MCW); (UCR); (Chinese Aca. Sci.)

    2010-01-12

    Abscisic acid (ABA) is a ubiquitous hormone that regulates plant growth, development and responses to environmental stresses. Its action is mediated by the PYR/PYL/RCAR family of START proteins, but it remains unclear how these receptors bind ABA and, in turn, how hormone binding leads to inhibition of the downstream type 2C protein phosphatase (PP2C) effectors. Here we report crystal structures of apo and ABA-bound receptors as well as a ternary PYL2-ABA-PP2C complex. The apo receptors contain an open ligand-binding pocket flanked by a gate that closes in response to ABA by way of conformational changes in two highly conserved β-loops that serve as a gate and latch. Moreover, ABA-induced closure of the gate creates a surface that enables the receptor to dock into and competitively inhibit the PP2C active site. A conserved tryptophan in the PP2C inserts directly between the gate and latch, which functions to further lock the receptor in a closed conformation. Together, our results identify a conserved gate-latch-lock mechanism underlying ABA signalling.

  1. A gate-latch-lock mechanism for hormone signalling by abscisic acid receptors

    Melcher, Karsten

    2009-12-03

    Abscisic acid (ABA) is a ubiquitous hormone that regulates plant growth, development and responses to environmental stresses. Its action is mediated by the PYR/PYL/RCAR family of START proteins, but it remains unclear how these receptors bind ABA and, in turn, how hormone binding leads to inhibition of the downstream type 2C protein phosphatase (PP2C) effectors. Here we report crystal structures of apo and ABA-bound receptors as well as a ternary PYL2-ABA-PP2C complex. The apo receptors contain an open ligand-binding pocket flanked by a gate that closes in response to ABA by way of conformational changes in two highly conserved ?-loops that serve as a gate and latch. Moreover, ABA-induced closure of the gate creates a surface that enables the receptor to dock into and competitively inhibit the PP2C active site. A conserved tryptophan in the PP2C inserts directly between the gate and latch, which functions to further lock the receptor in a closed conformation. Together, our results identify a conserved gate-latch-lock mechanism underlying ABA signalling. © 2009 Macmillan Publishers Limited. All rights reserved.

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

    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...

  3. Gonadotropin-inhibitory hormone receptor signaling and its impact on reproduction in chickens.

    Bédécarrats, Grégoy Y; McFarlane, Heather; Maddineni, Sreenivasa R; Ramachandran, Ramesh

    2009-09-01

    In birds, as in other vertebrates, reproduction is controlled by the hypothalamo-pituitary-gonadal axis with each component secreting specific neuropeptides or hormones. Until recently, it was believed this axis is exclusively under the stimulatory control of hypothalamic gonadotropin-releasing hormone I (GnRH-I) which in turn, stimulates luteinizing hormone (LH) and follicle stimulating hormone (FSH) secretion from the pituitary gland. However, the discovery of a novel inhibitory hypothalamic peptide able to reduce LH secretion (gonadotropin-inhibitory hormone: GnIH) challenged this dogma. Furthermore, with the characterization of its specific receptor (GnIHR), progress has been made to clarify the physiological relevance of GnIH in birds. This short review discusses the recent advances in GnIHR signaling at the level of the pituitary gland and the gonads. GnIHR is a member of the G-protein coupled receptor (GPCR) family which couples to G(alphai) and, upon activation inhibits adenylyl cyclase (AC) activity, thus reducing intracellular cAMP levels. This implies that GnIH interferes with signaling of any GPCR coupled to G(alphas), including GnRH, LH and FSH receptors. In the chicken pituitary gland, the GnRHR-II/GnIHR ratio changes during sexual maturation in favor of GnRHR-II that appears to result in hypothalamic control of gonadotropin secretion shifting from inhibitory to stimulatory, with corresponding changes in GnRH-induced cAMP levels. Within the gonads, GnIH and its receptor may act in an autocrine/paracrine manner and may interfere with LH and FSH signaling to influence ovarian follicular maturation and recruitment, as well as spermatogenesis. PMID:19332068

  4. Medial nucleus tractus solitarius oxytocin receptor signaling and food intake control: the role of gastrointestinal satiation signal processing

    Ong, Zhi Yi; Alhadeff, Amber L; Grill, Harvey J.

    2015-01-01

    Central oxytocin (OT) administration reduces food intake and its effects are mediated, in part, by hindbrain oxytocin receptor (OT-R) signaling. The neural substrate and mechanisms mediating the intake inhibitory effects of hindbrain OT-R signaling are undefined. We examined the hypothesis that hindbrain OT-R-mediated feeding inhibition results from an interaction between medial nucleus tractus solitarius (mNTS) OT-R signaling and the processing of gastrointestinal (GI) satiation signals by n...

  5. CERAPP: Collaborative estrogen receptor activity prediction project

    Mansouri, Kamel; Abdelaziz, Ahmed; Rybacka, Aleksandra;

    2016-01-01

    Background: Humans are exposed to thousands of man-made chemicals in the environment. Some chemicals mimic natural endocrine hormones and, thus, have the potential to be endocrine disruptors. Most of these chemicals have never been tested for their ability to interact with the estrogen receptor (ER......). Risk assessors need tools to prioritize chemicals for evaluation in costly in vivo tests, for instance, within the U.S. EPA Endocrine Disruptor Screening Program. oBjectives: We describe a large-scale modeling project called CERAPP (Collaborative Estrogen Receptor Activity Prediction Project) and...

  6. Spongian diterpenoids inhibit androgen receptor activity

    Yang, Yu Chi; Labros G Meimetis; Tien, Amy H; Mawji, Nasrin R.; Carr, Gavin; Wang, Jun; Andersen, Raymond J.; Sadar, Marianne D.

    2013-01-01

    Androgen receptor (AR) is a ligand-activated transcription factor and a validated drug target for all stages of prostate cancer. Antiandrogens compete with physiological ligands for AR ligand-binding domain (LBD). High-throughput screening of a marine natural product library for small molecules that inhibit AR transcriptional activity yielded the furanoditerpenoid spongia-13(16),-14-dien-19-oic acid, designated terpene 1 (T1). Characterization of T1 and the structurally related semi-synthetic...

  7. Identification of Essential Cannabinoid-binding Domains: STRUCTURAL INSIGHTS INTO EARLY DYNAMIC EVENTS IN RECEPTOR ACTIVATION*

    Shim, Joong-Youn; Bertalovitz, Alexander C.; Kendall, Debra A.

    2011-01-01

    The classical cannabinoid agonist HU210, a structural analog of (−)-Δ9-tetrahydrocannabinol, binds to brain cannabinoid (CB1) receptors and activates signal transduction pathways. To date, an exact molecular description of the CB1 receptor is not yet available. Utilizing the minor binding pocket of the CB1 receptor as the primary ligand interaction site, we explored HU210 binding using lipid bilayer molecular dynamics (MD) simulations. Among the potential ligand contact residues, we identifie...

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

    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

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

    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

  10. Endocytosis of pro-inflammatory cytokine receptors and its relevance for signal transduction.

    Hermanns, Heike M; Wohlfahrt, Julia; Mais, Christine; Hergovits, Sabine; Jahn, Daniel; Geier, Andreas

    2016-08-01

    The pro-inflammatory cytokines tumor necrosis factor (TNF), interleukin-1 (IL-1) and interleukin-6 (IL-6) are key players of the innate and adaptive immunity. Their activity needs to be tightly controlled to allow the initiation of an appropriate immune response as defense mechanism against pathogens or tissue injury. Excessive or sustained signaling of either of these cytokines leads to severe diseases, including rheumatoid arthritis, inflammatory bowel diseases (Crohn's disease, ulcerative colitis), steatohepatitis, periodic fevers and even cancer. Studies carried out in the last 30 years have emphasized that an elaborate control system for each of these cytokines exists. Here, we summarize what is currently known about the involvement of receptor endocytosis in the regulation of these pro-inflammatory cytokines' signaling cascades. Particularly in the last few years it was shown that this cellular process is far more than a mere feedback mechanism to clear cytokines from the circulation and to shut off their signal transduction. PMID:27071147

  11. TGF-β-induced profibrotic signaling is regulated in part by the WNT receptor Frizzled-8

    Spanjer, Anita I R; Baarsma, Hoeke A; Oostenbrink, Lisette M; Jansen, Sepp R; Kuipers, Christine C; Lindner, Michael; Postma, Dirkje S; Meurs, Herman; Heijink, Irene H; Gosens, Reinoud; Königshoff, Melanie

    2016-01-01

    TGF-β is important in lung injury and remodeling processes. TGF-β and Wingless/integrase-1 (WNT) signaling are interconnected; however, the WNT ligand-receptor complexes involved are unknown. Thus, we aimed to identify Frizzled (FZD) receptors that mediate TGF-β-induced profibrotic signaling. MRC-5

  12. Fatty acids activate a chimera of the clofibric acid-activated receptor and the glucocorticoid receptor.

    Göttlicher, M; Widmark, E; Q. Li; Gustafsson, J.A.

    1992-01-01

    Peroxisome proliferators such as clofibric acid, nafenopin, and WY-14,643 have been shown to activate PPAR (peroxisome proliferator-activated receptor), a member of the steroid nuclear receptor superfamily. We have cloned the cDNA from the rat that is homologous to that from the mouse [Issemann, I. & Green, S. (1990) Nature (London) 347, 645-650], which encodes a 97% similar protein with a particularly well-conserved putative ligand-binding domain. To search for physiologically occurring acti...

  13. β-Arrestin biosensors reveal a rapid, receptor-dependent activation/deactivation cycle.

    Nuber, Susanne; Zabel, Ulrike; Lorenz, Kristina; Nuber, Andreas; Milligan, Graeme; Tobin, Andrew B; Lohse, Martin J; Hoffmann, Carsten

    2016-03-31

    (β-)Arrestins are important regulators of G-protein-coupled receptors (GPCRs). They bind to active, phosphorylated GPCRs and thereby shut off 'classical' signalling to G proteins, trigger internalization of GPCRs via interaction with the clathrin machinery and mediate signalling via 'non-classical' pathways. In addition to two visual arrestins that bind to rod and cone photoreceptors (termed arrestin1 and arrestin4), there are only two (non-visual) β-arrestin proteins (β-arrestin1 and β-arrestin2, also termed arrestin2 and arrestin3), which regulate hundreds of different (non-visual) GPCRs. Binding of these proteins to GPCRs usually requires the active form of the receptors plus their phosphorylation by G-protein-coupled receptor kinases (GRKs). The binding of receptors or their carboxy terminus as well as certain truncations induce active conformations of (β-)arrestins that have recently been solved by X-ray crystallography. Here we investigate both the interaction of β-arrestin with GPCRs, and the β-arrestin conformational changes in real time and in living human cells, using a series of fluorescence resonance energy transfer (FRET)-based β-arrestin2 biosensors. We observe receptor-specific patterns of conformational changes in β-arrestin2 that occur rapidly after the receptor-β-arrestin2 interaction. After agonist removal, these changes persist for longer than the direct receptor interaction. Our data indicate a rapid, receptor-type-specific, two-step binding and activation process between GPCRs and β-arrestins. They further indicate that β-arrestins remain active after dissociation from receptors, allowing them to remain at the cell surface and presumably signal independently. Thus, GPCRs trigger a rapid, receptor-specific activation/deactivation cycle of β-arrestins, which permits their active signalling. PMID:27007855

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

    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.

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

    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.

  16. Human Receptor Activation by Aroclor 1260, a Polychlorinated Biphenyl Mixture

    Wahlang, Banrida; Falkner, K. Cameron; Clair, Heather B.; Al-Eryani, Laila; Prough, Russell A.; States, J. Christopher; Coslo, Denise M.; Omiecinski, Curtis J.; Cave, Matthew C.

    2014-01-01

    Polychlorinated biphenyls (PCBs) are persistent environmental toxicants, present in 100% of U.S. adults and dose-dependently associated with obesity and non-alcoholic fatty liver disease (NAFLD). PCBs are predicted to interact with receptors previously implicated in xenobiotic/energy metabolism and NAFLD. These receptors include the aryl hydrocarbon receptor (AhR), pregnane xenobiotic receptor (PXR), constitutive androstane receptor (CAR), peroxisome proliferator-activated receptors (PPARs), ...

  17. The glucagon-like peptide 1 receptor agonist enhances intrinsic peroxisome proliferator-activated receptor γ activity in endothelial cells

    Onuma, Hirohisa; Inukai, Kouichi, E-mail: kinukai@ks.kyorin-u.ac.jp; Kitahara, Atsuko; Moriya, Rie; Nishida, Susumu; Tanaka, Toshiaki; Katsuta, Hidenori; Takahashi, Kazuto; Sumitani, Yoshikazu; Hosaka, Toshio; Ishida, Hitoshi

    2014-08-22

    Highlights: • PPARγ activation was involved in the GLP-1-mediated anti-inflammatory action. • Exendin-4 enhanced endogenous PPARγ transcriptional activity in HUVECs. • H89, a PKA inhibitor, abolished GLP-1-induced PPARγ enhancement. • The anti-inflammatory effects of GLP-1 may be explained by PPARγ activation. - Abstract: Recent studies have suggested glucagon-like peptide-1 (GLP-1) signaling to exert anti-inflammatory effects on endothelial cells, although the precise underlying mechanism remains to be elucidated. In the present study, we investigated whether PPARγ activation is involved in the GLP-1-mediated anti-inflammatory action on endothelial cells. When we treated HUVEC cells with 0.2 ng/ml exendin-4, a GLP-1 receptor agonist, endogenous PPARγ transcriptional activity was significantly elevated, by approximately 20%, as compared with control cells. The maximum PPARγ activity enhancing effect of exendin-4 was observed 12 h after the initiation of incubation with exendin-4. As H89, a PKA inhibitor, abolished GLP-1-induced PPARγ enhancement, the signaling downstream from GLP-1 cross-talk must have been involved in PPARγ activation. In conclusion, our results suggest that GLP-1 has the potential to induce PPARγ activity, partially explaining the anti-inflammatory effects of GLP-1 on endothelial cells. Cross-talk between GLP-1 signaling and PPARγ activation would have major impacts on treatments for patients at high risk for cardiovascular disease.

  18. Exercise modulates postreceptor insulin signaling and glucose transport in muscle-specific insulin receptor knockout mice

    Wojtaszewski, Jørgen F. P.; Higaki, Yasuki; Hirshman, Michael F.; Michael, M. Dodson; Dufresne, Scott D.; Kahn, C. Ronald; Goodyear, Laurie J.

    1999-01-01

    Physical exercise promotes glucose uptake into skeletal muscle and makes the working muscles more sensitive to insulin. To understand the role of insulin receptor (IR) signaling in these responses, we studied the effects of exercise and insulin on skeletal muscle glucose metabolism and insulin signaling in mice lacking insulin receptors specifically in muscle. Muscle-specific insulin receptor knockout (MIRKO) mice had normal resting 2-deoxy-glucose (2DG) uptake in soleus muscles but had no si...

  19. The Role of Cgrp-Receptor Component Protein (Rcp in Cgrp-Mediated Signal Transduction

    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.

  20. Agonism, Antagonism, and Inverse Agonism Bias at the Ghrelin Receptor Signaling.

    M'Kadmi, Céline; Leyris, Jean-Philippe; Onfroy, Lauriane; Galés, Céline; Saulière, Aude; Gagne, Didier; Damian, Marjorie; Mary, Sophie; Maingot, Mathieu; Denoyelle, Séverine; Verdié, Pascal; Fehrentz, Jean-Alain; Martinez, Jean; Banères, Jean-Louis; Marie, Jacky

    2015-11-01

    The G protein-coupled receptor GHS-R1a mediates ghrelin-induced growth hormone secretion, food intake, and reward-seeking behaviors. GHS-R1a signals through Gq, Gi/o, G13, and arrestin. Biasing GHS-R1a signaling with specific ligands may lead to the development of more selective drugs to treat obesity or addiction with minimal side effects. To delineate ligand selectivity at GHS-R1a signaling, we analyzed in detail the efficacy of a panel of synthetic ligands activating the different pathways associated with GHS-R1a in HEK293T cells. Besides β-arrestin2 recruitment and ERK1/2 phosphorylation, we monitored activation of a large panel of G protein subtypes using a bioluminescence resonance energy transfer-based assay with G protein-activation biosensors. We first found that unlike full agonists, Gq partial agonists were unable to trigger β-arrestin2 recruitment and ERK1/2 phosphorylation. Using G protein-activation biosensors, we then demonstrated that ghrelin promoted activation of Gq, Gi1, Gi2, Gi3, Goa, Gob, and G13 but not Gs and G12. Besides, we identified some GHS-R1a ligands that preferentially activated Gq and antagonized ghrelin-mediated Gi/Go activation. Finally, we unambiguously demonstrated that in addition to Gq, GHS-R1a also promoted constitutive activation of G13. Importantly, we identified some ligands that were selective inverse agonists toward Gq but not of G13. This demonstrates that bias at GHS-R1a signaling can occur not only with regard to agonism but also to inverse agonism. Our data, combined with other in vivo studies, may facilitate the design of drugs selectively targeting individual signaling pathways to treat only the therapeutically relevant function. PMID:26363071

  1. Active transport improves the precision of linear long distance molecular signalling

    Godec, Aljaz; Metzler, Ralf

    2016-01-01

    Molecular signalling in living cells occurs at low copy numbers and is thereby inherently limited by the noise imposed by thermal diffusion. The precision at which biochemical receptors can count signalling molecules is intimately related to the noise correlation time. In addition to passive thermal diffusion, messenger RNA and vesicle-engulfed signalling molecules can transiently bind to molecular motors and are actively transported across biological cells. Active transport is most beneficia...

  2. Modulation of visceral pain and inflammation by protease-activated receptors

    Vergnolle, Nathalie

    2004-01-01

    The gastrointestinal (GI) tract is exposed to a large array of proteases, under both physiological and pathophysiological conditions. The discovery of G protein-coupled receptors activated by proteases, the protease-activated receptors (PARs), has highlighted new signaling functions for proteases in the GI tract, particularly in the domains of inflammation and pain mechanisms. Activation of PARs by selective peptidic agonists in the intestine or the pancreas leads to inflammatory events and c...

  3. Quercetin Directly Interacts with Vitamin D Receptor (VDR): Structural Implication of VDR Activation by Quercetin

    Lee, Ki-Young; Choi, Hye-Seung; Choi, Ho-Sung; Chung, Ka Young; Lee, Bong-Jin; Maeng, Han-Joo; Seo, Min-Duk

    2016-01-01

    The vitamin D receptor (VDR) is a member of the nuclear receptor (NR) superfamily. The VDR binds to active vitamin D3 metabolites, which stimulates downstream transduction signaling involved in various physiological activities such as calcium homeostasis, bone mineralization, and cell differentiation. Quercetin is a widely distributed flavonoid in nature that is known to enhance transactivation of VDR target genes. However, the detailed molecular mechanism underlying VDR activation by quercet...

  4. Enzymatic Regulation of Steroidogenesis and Nuclear Receptor Activation : Special Focus on Vitamin D and Sex Hormones

    LUNDQVIST, JOHAN

    2011-01-01

    Enzyme-catalyzed reactions are important to regulate steroidogenesis and nuclear receptor activation. The present investigation examines the role of steroid metabolism catalyzed by CYP7B1 for regulation of hormone receptor activation and the effects of vitamin D on enzymatic regulation of steroidogenesis. The study reports data indicating that CYP7B1 can regulate estrogenic signaling by converting estrogens into inactive or less active metabolites. Similar results were obtained for CYP7B1-med...

  5. Interaction with both ZNRF3 and LGR4 is required for the signalling activity of R-spondin

    Xie, Yang; Zamponi, Raffaella; Charlat, Olga; Ramones, Melissa; Swalley, Susanne; Jiang, Xiaomo; Rivera, Daniel; Tschantz, William; Lu, Bo; Quinn, Lisa; Dimitri, Chris; Parker, Jefferson; Jeffery, Doug; Wilcox, Sheri K; Watrobka, Mike

    2013-01-01

    This study shows that both ZNRF3- and LGR4-binding motifs of R-spondin are required for its Wnt-promoting activity. These results support a dual receptor model of R-spondin signalling, where LGR4 serves as the engagement receptor while ZNRF3 functions as the effector receptor.

  6. The Structure of the GM-CSF Receptor Complex Reveals a Distinct Mode of Cytokine Receptor Activation

    Hansen, Guido; Hercus, Timothy R.; McClure, Barbara J.; Stomski, Frank C.; Dottore, Mara; Powell, Jason; Ramshaw, Hayley; Woodcock, Joanna M.; Xu, Yibin; Guthridge, Mark; McKinstry, William J.; Lopez, Angel F.; Parker, Michael W. (SVIMR-A); (Hanson)

    2008-08-11

    Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic cytokine that controls the production and function of blood cells, is deregulated in clinical conditions such as rheumatoid arthritis and leukemia, yet offers therapeutic value for other diseases. Its receptors are heterodimers consisting of a ligand-specific {alpha} subunit and a {beta}c subunit that is shared with the interleukin (IL)-3 and IL-5 receptors. How signaling is initiated remains an enigma. We report here the crystal structure of the human GM-CSF/GM-CSF receptor ternary complex and its assembly into an unexpected dodecamer or higher-order complex. Importantly, mutagenesis of the GM-CSF receptor at the dodecamer interface and functional studies reveal that dodecamer formation is required for receptor activation and signaling. This unusual form of receptor assembly likely applies also to IL-3 and IL-5 receptors, providing a structural basis for understanding their mechanism of activation and for the development of therapeutics.

  7. Aripiprazole Increases the PKA Signalling and Expression of the GABAA Receptor and CREB1 in the Nucleus Accumbens of Rats.

    Pan, Bo; Lian, Jiamei; Huang, Xu-Feng; Deng, Chao

    2016-05-01

    The GABAA receptor is implicated in the pathophysiology of schizophrenia and regulated by PKA signalling. Current antipsychotics bind with D2-like receptors, but not the GABAA receptor. The cAMP-responsive element-binding protein 1 (CREB1) is also associated with PKA signalling and may be related to the positive symptoms of schizophrenia. This study investigated the effects of antipsychotics in modulating D2-mediated PKA signalling and its downstream GABAA receptors and CREB1. Rats were treated orally with aripiprazole (0.75 mg/kg, ter in die (t.i.d.)), bifeprunox (0.8 mg/kg, t.i.d.), haloperidol (0.1 mg/kg, t.i.d.) or vehicle for 1 week. The levels of PKA-Cα and p-PKA in the prefrontal cortex (PFC), nucleus accumbens (NAc) and caudate putamen (CPu) were detected by Western blots. The mRNA levels of Gabrb1, Gabrb2, Gabrb3 and Creb1, and their protein expression were measured by qRT-PCR and Western blots, respectively. Aripiprazole elevated the levels of p-PKA and the ratio of p-PKA/PKA in the NAc, but not the PFC and CPu. Correlated with this elevated PKA signalling, aripiprazole elevated the mRNA and protein expression of the GABAA (β-1) receptor and CREB1 in the NAc. While haloperidol elevated the levels of p-PKA and the ratio of p-PKA/PKA in both NAc and CPu, it only tended to increase the expression of the GABAA (β-1) receptor and CREB1 in the NAc, but not the CPu. Bifeprunox had no effects on PKA signalling in these brain regions. These results suggest that aripiprazole has selective effects on upregulating the GABAA (β-1) receptor and CREB1 in the NAc, probably via activating PKA signalling. PMID:26894264

  8. Interactions between polymorphisms in the aryl hydrocarbon receptor signalling pathway and exposure to persistent organochlorine pollutants affect human semen quality

    Brokken, L J S; Lundberg, P J; Spanò, M;

    2014-01-01

    Persistent organic pollutants (POPs) may affect male reproductive function. Many dioxin-like POPs exert their effects by activation of the aryl hydrocarbon receptor (AHR) signalling pathway. We analysed whether gene-environment interactions between polymorphisms in AHR (R554K) and AHR repressor (...

  9. A Perspective on Studying G-Protein-Coupled Receptor Signaling with Resonance Energy Transfer Biosensors in Living Organisms

    J. van Unen; J. Woolard; A. Rinken; C. Hoffmann; S.J. Hill; J. Goedhart; M.R. Bruchas; M. Bouvier; M.J.W. Adjobo-Hermans

    2015-01-01

    The last frontier for a complete understanding of G-protein-coupled receptor (GPCR) biology is to be able to assess GPCR activity, interactions, and signaling in vivo, in real time within biologically intact systems. This includes the ability to detect GPCR trafficking, dimerization, protein-protein

  10. Engineering the melanocortin-4 receptor to control constitutive and ligand-mediated G(S signaling in vivo.

    Supriya Srinivasan

    Full Text Available The molecular and functional diversity of G protein-coupled receptors is essential to many physiological processes. However, this diversity presents a significant challenge to understanding the G protein-mediated signaling events that underlie a specific physiological response. To increase our understanding of these processes, we sought to gain control of the timing and specificity of G(s signaling in vivo. We used naturally occurring human mutations to develop two G(s-coupled engineered receptors that respond solely to a synthetic ligand (RASSLs. Our G(s-coupled RASSLs are based on the melanocortin-4 receptor, a centrally expressed receptor that plays an important role in the regulation of body weight. These RASSLs are not activated by the endogenous hormone alpha-melanocyte-stimulating hormone but respond potently to a selective synthetic ligand, tetrahydroisoquinoline. The RASSL variants reported here differ in their intrinsic basal activities, allowing the separation of the effects of basal signaling from ligand-mediated activation of the G(s pathway in vivo. These RASSLs can be used to activate G(s signaling in any tissue, but would be particularly useful for analyzing downstream events that mediate body weight regulation in mice. Our study also demonstrates the use of human genetic variation for protein engineering.

  11. PRMT1-mediated methylation of the EGF receptor regulates signaling and cetuximab response

    Liao, Hsin-Wei

    2015-11-16

    Posttranslational modifications to the intracellular domain of the EGFR are known to regulate EGFR functions; however, modifications to the extracellular domain and their effects remain relatively unexplored. Here, we determined that methylation at R198 and R200 of the EGFR extracellular domain by protein arginine methyltransferase 1 (PRMT1) enhances binding to EGF and subsequent receptor dimerization and signaling activation. In a mouse orthotopic colorectal cancer xenograft model, expression of a methylation-defective EGFR reduced tumor growth. Moreover, increased EGFR methylation sustained signaling activation and cell proliferation in the presence of the therapeutic EGFR monoclonal antibody cetuximab. In colorectal cancer patients, EGFR methylation level also correlated with a higher recurrence rate after cetuximab treatment and reduced overall survival. Together, these data indicate that R198/R200 methylation of the EGFR plays an important role in regulating EGFR functionality and resistance to cetuximab treatment.

  12. Multi-organ Site Metastatic Reactivation Mediated by Non-canonical Discoidin Domain Receptor 1 Signaling.

    Gao, Hua; Chakraborty, Goutam; Zhang, Zhanguo; Akalay, Intissar; Gadiya, Mayur; Gao, Yaquan; Sinha, Surajit; Hu, Jian; Jiang, Cizhong; Akram, Muzaffar; Brogi, Edi; Leitinger, Birgit; Giancotti, Filippo G

    2016-06-30

    Genetic screening identifies the atypical tetraspanin TM4SF1 as a strong mediator of metastatic reactivation of breast cancer. Intriguingly, TM4SF1 couples the collagen receptor tyrosine kinase DDR1 to the cortical adaptor syntenin 2 and, hence, to PKCα. The latter kinase phosphorylates and activates JAK2, leading to the activation of STAT3. This non-canonical mechanism of signaling induces the expression of SOX2 and NANOG; sustains the manifestation of cancer stem cell traits; and drives metastatic reactivation in the lung, bone, and brain. Bioinformatic analyses and pathological studies corroborate the clinical relevance of these findings. We conclude that non-canonical DDR1 signaling enables breast cancer cells to exploit the ubiquitous interstitial matrix component collagen I to undergo metastatic reactivation in multiple target organs. PMID:27368100

  13. Mu and delta opioid receptors oppositely regulate motor impulsivity in the signaled nose poke task.

    Mary C Olmstead

    Full Text Available Impulsivity is a primary feature of many psychiatric disorders, most notably attention deficit hyperactivity disorder and drug addiction. Impulsivity includes a number of processes such as the inability to delay gratification, the inability to withhold a motor response, or acting before all of the relevant information is available. These processes are mediated by neural systems that include dopamine, serotonin, norepinephrine, glutamate and cannabinoids. We examine, for the first time, the role of opioid systems in impulsivity by testing whether inactivation of the mu- (Oprm1 or delta- (Oprd1 opioid receptor gene alters motor impulsivity in mice. Wild-type and knockout mice were examined on either a pure C57BL6/J (BL6 or a hybrid 50% C57Bl/6J-50% 129Sv/pas (HYB background. Mice were trained to respond for sucrose in a signaled nose poke task that provides independent measures of associative learning (responses to the reward-paired cue and motor impulsivity (premature responses. Oprm1 knockout mice displayed a remarkable decrease in motor impulsivity. This was observed on the two genetic backgrounds and did not result from impaired associative learning, as responses to the cue signaling reward did not differ across genotypes. Furthermore, mutant mice were insensitive to the effects of ethanol, which increased disinhibition and decreased conditioned responding in wild-type mice. In sharp contrast, mice lacking the Oprd1 gene were more impulsive than controls. Again, mutant animals showed no deficit in associative learning. Ethanol completely disrupted performance in these animals. Together, our results suggest that mu-opioid receptors enhance, whereas delta-opioid receptors inhibit, motor impulsivity. This reveals an unanticipated contribution of endogenous opioid receptor activity to disinhibition. In a broader context, these data suggest that alterations in mu- or delta-opioid receptor function may contribute to impulse control disorders.

  14. Activity of recombinant trypsin isoforms on human proteinase-activated receptors (PAR): mesotrypsin cannot activate epithelial PAR-1, -2, but weakly activates brain PAR-1

    Grishina, Zoryana; Ostrowska, Ewa; Halangk, Walter; Sahin-Tóth, Miklós; Reiser, Georg

    2005-01-01

    Trypsin-like serine proteinases trigger signal transduction pathways through proteolytic cleavage of proteinase-activated receptors (PARs) in many tissues. Three members, PAR-1, PAR-2 and PAR-4, are trypsin substrates, as trypsinolytic cleavage of the extracellular N terminus produces receptor activation. Here, the ability of the three human pancreatic trypsin isoforms (cationic trypsin, anionic trypsin and mesotrypsin (trypsin IV)) as recombinant proteins was tested on PARs.Using fura 2 [Ca2...

  15. Signaling pathways involved in the inhibition of epidermal growth factor receptor by erlotinib in hepatocellular cancer

    Alexander Huether; Michael H(o)pfner; Andreas P Sutter; Viola Baradari; Detlef Schuppan; Hans Scherübl

    2006-01-01

    AIM: To examine the underlying mechanisms of erlotinib-induced growth inhibition in hepatocellular carcinoma (HCC).METHODS: Erlotinib-induced alterations in gene expression were evaluated using cDNA array technology;changes in protein expression and/or protein activation due to erlotinib treatment as well as IGF-1-induced EGFR transactivation were investigated using Western blotting. RESULTS: Erlotinib treatment inhibited the mitogen activated protein (MAP)-kinase pathway and signal transducer of activation and transcription (STAT)mediated signaling which led to an altered expression of apoptosis and cell cycle regulating genes as demonstrated by cDNA array technology. Overexpression of proapoptotic factors like caspases and gadds associated with a down-regulation of antiapoptoticfactors like Bcl-2, Bcl-XL or jun D accounted for erlotinib's potency to induce apoptosis. Downregulation of cell cycle regulators promoting the G1/S-transition and overexpression of cyclin-dependent kinase inhibitors and gadds contributed to the induction of a G1/Go-arrest in response to erlotinib. Furthermore, we displayed the transactivation of EGFR-mediated signaling by the IGF-1-receptor and showed erlotinib's inhibitory effects on the receptor-receptor cross talk. CONCLUSION: Our study sheds light on the understanding of the mechanisms of action of EGFR-TKinhibition in HCC-cells and thus might facilitate the design of combination therapies that act additively or synergistically. Moreover, our data on the pathways responding to erlotinib treatment could be helpful in predicting the responsiveness of tumors to EGFR-TKIs in the future.

  16. The plant natriuretic peptide receptor is a guanylyl cyclase and enables cGMP-dependent signaling.

    Turek, Ilona; Gehring, Chris

    2016-06-01

    The functional homologues of vertebrate natriuretic peptides (NPs), the plant natriuretic peptides (PNPs), are a novel class of peptidic hormones that signal via guanosine 3',5'-cyclic monophosphate (cGMP) and systemically affect plant salt and water balance and responses to biotrophic plant pathogens. Although there is increasing understanding of the complex roles of PNPs in plant responses at the systems level, little is known about the underlying signaling mechanisms. Here we report isolation and identification of a novel Leucine-Rich Repeat (LRR) protein that directly interacts with A. thaliana PNP, AtPNP-A. In vitro binding studies revealed that the Arabidopsis AtPNP-A binds specifically to the LRR protein, termed AtPNP-R1, and the active region of AtPNP-A is sufficient for the interaction to occur. Importantly, the cytosolic part of the AtPNP-R1, much like in some vertebrate NP receptors, harbors a catalytic center diagnostic for guanylyl cyclases and the recombinant AtPNP-R1 is capable of catalyzing the conversion of guanosine triphosphate to cGMP. In addition, we show that AtPNP-A causes rapid increases of cGMP levels in wild type (WT) leaf tissue while this response is significantly reduced in the atpnp-r1 mutants. AtPNP-A also causes cGMP-dependent net water uptake into WT protoplasts, and hence volume increases, whereas responses of the protoplasts from the receptor mutant are impaired. Taken together, our results suggest that the identified LRR protein is an AtPNP-A receptor essential for the PNP-dependent regulation of ion and water homeostasis in plants and that PNP- and vertebrate NP-receptors and their signaling mechanisms share surprising similarities. PMID:26945740

  17. New insights of P2X7 receptor signaling pathway in alveolar functions

    Mishra, Amarjit

    2013-01-01

    Purinergic P2X7 receptor (P2X7R), an ATP-gated cation channel, is unique among all other family members because of its ability to respond to various stimuli and to modulate pro-inflammatory signaling. The activation of P2X7R in immune cells is absolutely required for mature interleukin -1beta (IL-1beta) and IL-18 production and release. Lung alveoli are lined by the structural alveolar epithelial type I (AEC I) and alveolar epithelial type II cells (AEC II). AEC I plays important roles in alv...

  18. Toll Receptor-Mediated Hippo Signaling Controls Innate Immunity in Drosophila.

    Liu, Bo; Zheng, Yonggang; Yin, Feng; Yu, Jianzhong; Silverman, Neal; Pan, Duojia

    2016-01-28

    The Hippo signaling pathway functions through Yorkie to control tissue growth and homeostasis. How this pathway regulates non-developmental processes remains largely unexplored. Here, we report an essential role for Hippo signaling in innate immunity whereby Yorkie directly regulates the transcription of the Drosophila IκB homolog, Cactus, in Toll receptor-mediated antimicrobial response. Loss of Hippo pathway tumor suppressors or activation of Yorkie in fat bodies, the Drosophila immune organ, leads to elevated cactus mRNA levels, decreased expression of antimicrobial peptides, and vulnerability to infection by Gram-positive bacteria. Furthermore, Gram-positive bacteria acutely activate Hippo-Yorkie signaling in fat bodies via the Toll-Myd88-Pelle cascade through Pelle-mediated phosphorylation and degradation of the Cka subunit of the Hippo-inhibitory STRIPAK PP2A complex. Our studies elucidate a Toll-mediated Hippo signaling pathway in antimicrobial response, highlight the importance of regulating IκB/Cactus transcription in innate immunity, and identify Gram-positive bacteria as extracellular stimuli of Hippo signaling under physiological settings. PMID:26824654

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

    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

  20. New insights of P2X7 receptor signaling pathway in alveolar functions.

    Mishra, Amarjit

    2013-01-01

    Purinergic P2X7 receptor (P2X7R), an ATP-gated cation channel, is unique among all other family members because of its ability to respond to various stimuli and to modulate pro-inflammatory signaling. The activation of P2X7R in immune cells is absolutely required for mature interleukin -1beta (IL-1beta) and IL-18 production and release. Lung alveoli are lined by the structural alveolar epithelial type I (AEC I) and alveolar epithelial type II cells (AEC II). AEC I plays important roles in alveolar barrier protection and fluid homeostasis whereas AEC II synthesizes and secrete surfactant and prevents alveoli from collapse. Earlier studies indicated that purinergic P2X7 receptors were specifically expressed in AEC I. However, their implication in alveolar functions has not been explored. This paper reviews two important signaling pathways of P2X7 receptors in surfactant homeostatsis and Acute Lung Injury (ALI). Thus, P2X7R resides at the critical nexus of alveolar pathophysiology. PMID:23634990

  1. Moonlighting kinases with guanylate cyclase activity can tune regulatory signal networks

    Irving, Helen R.

    2012-02-01

    Guanylate cyclase (GC) catalyzes the formation of cGMP and it is only recently that such enzymes have been characterized in plants. One family of plant GCs contains the GC catalytic center encapsulated within the intracellular kinase domain of leucine rich repeat receptor like kinases such as the phytosulfokine and brassinosteroid receptors. In vitro studies show that both the kinase and GC domain have catalytic activity indicating that these kinase-GCs are examples of moonlighting proteins with dual catalytic function. The natural ligands for both receptors increase intracellular cGMP levels in isolated mesophyll protoplast assays suggesting that the GC activity is functionally relevant. cGMP production may have an autoregulatory role on receptor kinase activity and/or contribute to downstream cell expansion responses. We postulate that the receptors are members of a novel class of receptor kinases that contain functional moonlighting GC domains essential for complex signaling roles.

  2. Remote activation of the Wnt/β-catenin signalling pathway using functionalised magnetic particles.

    Michael Rotherham

    Full Text Available Wnt signalling pathways play crucial roles in developmental biology, stem cell fate and tissue patterning and have become an attractive therapeutic target in the fields of tissue engineering and regenerative medicine. Wnt signalling has also been shown to play a role in human Mesenchymal Stem Cell (hMSC fate, which have shown potential as a cell therapy in bone and cartilage tissue engineering. Previous work has shown that biocompatible magnetic nanoparticles (MNP can be used to stimulate specific mechanosensitive membrane receptors and ion channels in vitro and in vivo. Using this strategy, we determined the effects of mechano-stimulation of the Wnt Frizzled receptor on Wnt pathway activation in hMSC. Frizzled receptors were tagged using anti-Frizzled functionalised MNP (Fz-MNP. A commercially available oscillating magnetic bioreactor (MICA Biosystems was used to mechanically stimulate Frizzled receptors remotely. Our results demonstrate that Fz-MNP can activate Wnt/β-catenin signalling at key checkpoints in the signalling pathway. Immunocytochemistry indicated nuclear localisation of the Wnt intracellular messenger β-catenin after treatment with Fz-MNP. A Wnt signalling TCF/LEF responsive luciferase reporter transfected into hMSC was used to assess terminal signal activation at the nucleus. We observed an increase in reporter activity after treatment with Fz-MNP and this effect was enhanced after mechano-stimulation using the magnetic array. Western blot analysis was used to probe the mechanism of signalling activation and indicated that Fz-MNP signal through an LRP independent mechanism. Finally, the gene expression profiles of stress response genes were found to be similar when cells were treated with recombinant Wnt-3A or Fz-MNP. This study provides proof of principle that Wnt signalling and Frizzled receptors are mechanosensitive and can be remotely activated in vitro. Using magnetic nanoparticle technology it may be possible to modulate

  3. Chlorogenic acid improves late diabetes through adiponectin receptor signaling pathways in db/db mice.

    Shasha Jin

    Full Text Available The aim of this study was to examine the effects of chlorogenic acid (CGA on glucose and lipid metabolism in late diabetic db/db mice, as well as on adiponectin receptors and their signaling molecules, to provide evidence for CGA in the prevention of type 2 diabetes. We randomly divided 16 female db/db mice into db/db-CGA and db/db-control (CON groups equally; db/m mice were used as control mice. The mice in both the db/db-CGA and db/m-CGA groups were administered 80 mg/kg/d CGA by lavage for 12 weeks, whereas the mice in both CON groups were given equal volumes of phosphate-buffered saline (PBS by lavage. At the end of the intervention, we assessed body fat and the parameters of glucose and lipid metabolism in the plasma, liver and skeletal muscle tissues as well as the levels of aldose reductase (AR and transforming growth factor-β1 (TGF-β1 in the kidneys and measured adiponectin receptors and the protein expression of their signaling molecules in liver and muscle tissues. After 12 weeks of intervention, compared with the db/db-CON group, the percentage of body fat, fasting plasma glucose (FPG and glycosylated hemoglobin (HbA1c in the db/db-CGA group were all significantly decreased; TGF-β1 protein expression and AR activity in the kidney were both decreased; and the adiponectin level in visceral adipose was increased. The protein expression of adiponectin receptors (ADPNRs, the phosphorylation of AMP-activated protein kinase (AMPK in the liver and muscle, and the mRNA and protein levels of peroxisome proliferator-activated receptor alpha (PPAR-α in the liver were all significantly greater. CGA could lower the levels of fasting plasma glucose and HbA1c during late diabetes and improve kidney fibrosis to some extent through the modulation of adiponectin receptor signaling pathways in db/db mice.

  4. Specification, annotation, visualization and simulation of a large rule-based model for ERBB receptor signaling

    Creamer Matthew S

    2012-08-01

    Full Text Available Abstract Background Mathematical/computational models are needed to understand cell signaling networks, which are complex. Signaling proteins contain multiple functional components and multiple sites of post-translational modification. The multiplicity of components and sites of modification ensures that interactions among signaling proteins have the potential to generate myriad protein complexes and post-translational modification states. As a result, the number of chemical species that can be populated in a cell signaling network, and hence the number of equations in an ordinary differential equation model required to capture the dynamics of these species, is prohibitively large. To overcome this problem, the rule-based modeling approach has been developed for representing interactions within signaling networks efficiently and compactly through coarse-graining of the chemical kinetics of molecular interactions. Results Here, we provide a demonstration that the rule-based modeling approach can be used to specify and simulate a large model for ERBB receptor signaling that accounts for site-specific details of protein-protein interactions. The model is considered large because it corresponds to a reaction network containing more reactions than can be practically enumerated. The model encompasses activation of ERK and Akt, and it can be simulated using a network-free simulator, such as NFsim, to generate time courses of phosphorylation for 55 individual serine, threonine, and tyrosine residues. The model is annotated and visualized in the form of an extended contact map. Conclusions With the development of software that implements novel computational methods for calculating the dynamics of large-scale rule-based representations of cellular signaling networks, it is now possible to build and analyze models that include a significant fraction of the protein interactions that comprise a signaling network, with incorporation of the site

  5. Anxiolytic activity of adenosine receptor activation in mice.

    Jain, N; Kemp, N; Adeyemo, O; Buchanan, P.; Stone, T W

    1995-01-01

    1. Purine analogues have been examined for anxiolytic- and anxiogenic-like activity in mice, by use of the elevated plus-maze. 2. The selective A1 receptor agonist, N6-cyclopentyladenosine (CPA) had marked anxiolytic-like activity at 10 and 50 microg kg(-1), with no effect on locomotor performance at these doses. 3. The A1 selective adenosine receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine (CPX) had no significant effect on anxiety-related measures or locomotor behaviour, but blocked ...

  6. Full and partial agonists of thromboxane prostanoid receptor unveil fine tuning of receptor superactive conformation and G protein activation.

    Valérie Capra

    Full Text Available The intrahelical salt bridge between E/D(3.49 and R(3.50 within the E/DRY motif on helix 3 (H3 and the interhelical hydrogen bonding between the E/DRY and residues on H6 are thought to be critical in stabilizing the class A G protein-coupled receptors in their inactive state. Removal of these interactions is expected to generate constitutively active receptors. This study examines how neutralization of E(3.49/6.30 in the thromboxane prostanoid (TP receptor alters ligand binding, basal, and agonist-induced activity and investigates the molecular mechanisms of G protein activation. We demonstrate here that a panel of full and partial agonists showed an increase in affinity and potency for E129V and E240V mutants. Yet, even augmenting the sensitivity to detect constitutive activity (CA with overexpression of the receptor or the G protein revealed resistance to an increase in basal activity, while retaining fully the ability to cause agonist-induced signaling. However, direct G protein activation measured through bioluminescence resonance energy transfer (BRET indicates that these mutants more efficiently communicate and/or activate their cognate G proteins. These results suggest the existence of additional constrains governing the shift of TP receptor to its active state, together with an increase propensity of these mutants to agonist-induced signaling, corroborating their definition as superactive mutants. The particular nature of the TP receptor as somehow "resistant" to CA should be examined in the context of its pathophysiological role in the cardiovascular system. Evolutionary forces may have favored regulation mechanisms leading to low basal activity and selected against more highly active phenotypes.

  7. The viral G protein-coupled receptor ORF74 unmasks phospholipase C signaling of the receptor tyrosine kinase IGF-1R.

    de Munnik, Sabrina M; van der Lee, Rosan; Velders, Daniëlle M; van Offenbeek, Jody; Smits-de Vries, Laura; Leurs, Rob; Smit, Martine J; Vischer, Henry F

    2016-06-01

    Kaposi's sarcoma-associated herpesvirus (KSHV) encodes the constitutively active G protein-coupled receptor ORF74, which is expressed on the surface of infected host cells and has been linked to the development of the angioproliferative tumor Kaposi's sarcoma. Furthermore, the insulin-like growth factor (IGF)-1 receptor, a receptor tyrosine kinase, also plays an essential role in Kaposi's sarcoma growth and survival. In this study we examined the effect of the constitutively active viral receptor ORF74 on human IGF-1R signaling. Constitutive and CXCL1-induced ORF74 signaling did not transactivate IGF-1R. In contrast, IGF-1 stimulated phospholipase C (PLC) activation in an ORF74-dependent manner without affecting chemokine binding to ORF74. Inhibition of constitutive ORF74 activity by mutagenesis or the inverse agonist CXCL10, or neutralizing IGF-1R with an antibody or silencing IGF-1R expression using siRNA inhibited PLC activation by IGF-1. Transactivation of ORF74 in response to IGF-1 occurred independently of Src, PI3K, and secreted ORF74 ligands. Furthermore, tyrosine residues in the carboxyl-terminus and intracellular loop 2 of ORF74 are not essential for IGF-1-induced PLC activation. Interestingly, PLC activation in response to IGF-1 is specific for ORF74 as IGF-1 was unable to activate PLC in cells expressing the constitutively active human cytomegalovirus (HCMV)-encoded GPCR US28. Interestingly, IGF-1 does not induce β-arrestin recruitment to ORF74. The proximity ligation assay revealed close proximity between ORF74 and IGF-1R on the cell surface, but a physical interaction was not confirmed by co-immunoprecipitation. Unmasking IGF-1R signaling to PLC in response to IGF-1 is a previously unrecognized action of ORF74. PMID:26931381

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

    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

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

    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

  10. Neurohumoral activation in heart failure: the role of adrenergic receptors

    Patricia C. Brum

    2006-09-01

    Full Text Available Heart failure (HF is a common endpoint for many forms of cardiovascular disease and a significant cause of morbidity and mortality. The development of end-stage HF often involves an initial insult to the myocardium that reduces cardiac output and leads to a compensatory increase in sympathetic nervous system activity. Acutely, the sympathetic hyperactivity through the activation of beta-adrenergic receptors increases heart rate and cardiac contractility, which compensate for decreased cardiac output. However, chronic exposure of the heart to elevated levels of catecholamines released from sympathetic nerve terminals and the adrenal gland may lead to further pathologic changes in the heart, resulting in continued elevation of sympathetic tone and a progressive deterioration in cardiac function. On a molecular level, altered beta-adrenergic receptor signaling plays a pivotal role in the genesis and progression of HF. beta-adrenergic receptor number and function are decreased, and downstream mechanisms are altered. In this review we will present an overview of the normal beta-adrenergic receptor pathway in the heart and the consequences of sustained adrenergic activation in HF. The myopathic potential of individual components of the adrenergic signaling will be discussed through the results of research performed in genetic modified animals. Finally, we will discuss the potential clinical impact of beta-adrenergic receptor gene polymorphisms for better understanding the progression of HF.A insuficiência cardíaca (IC é a via final comum da maioria das doenças cardiovasculares e uma das maiores causas de morbi-mortalidade. O desenvolvimento do estágio final da IC freqüentemente envolve um insulto inicial do miocárdio, reduzindo o débito cardíaco e levando ao aumento compensatório da atividade do sistema nervoso simpático (SNS. Existem evidências de que apesar da exposição aguda ser benéfica, exposições crônicas a elevadas concentra

  11. Ric-8A, a Gα protein guanine nucleotide exchange factor potentiates taste receptor signaling

    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.

  12. Real-time trafficking and signaling of the glucagon-like peptide-1 receptor

    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...

  13. Relaxin Family Peptide Receptor 1 (RXFP1) Activation Stimulates the Peroxisome Proliferator-Activated Receptor Gamma

    Singh, Sudhir; Bennett, Robert G

    2009-01-01

    Relaxin (Rlx) has antifibrotic effects in a number of tissues. Many of these effects are similar to those induced by the activators of peroxisome proliferator-activated receptor γ (PPARγ), raising the possibility that a mechanism for Rlx’s antifibrotic effects may involve activation of the PPARγ pathway. This study investigates the effect of Rlx on PPARs and their mechanism of upregulation. It shows that Rlx stimulates ligand-independent PPAR activation in a dose-dependent manner. The combine...

  14. Activation of the unliganded estrogen receptor by EGF involves the MAP kinase pathway and direct phosphorylation.

    Bunone, G; Briand, P A; Miksicek, R J; Picard, D.

    1996-01-01

    The estrogen receptor (ER) can be activated as a transcription factor either by binding of cognate estrogenic ligand or, indirectly, by a variety of other extracellular signals. As a first step towards elucidating the mechanism of 'steroid-independent activation' of the ER by the epidermal growth factor (EGF), we have mapped the ER target domain and determined the signaling pathway. We show that the N-terminal transcriptional