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Sample records for factor receptor transactivation

  1. Resveratrol prevents angiotensin II-induced hypertrophy of vascular smooth muscle cells through the transactivation of growth factor receptors.

    Science.gov (United States)

    Hossain, Ekhtear; Anand-Srivastava, Madhu B

    2017-08-01

    We previously showed that augmented levels of endogenous angiotensin II (AngII) contribute to vascular smooth muscle cell (VSMC) hypertrophy through the transactivation of growth factor receptors in spontaneously hypertensive rats. Resveratrol (RV), a polyphenolic component of red wine, has also been shown to attenuate AngII-evoked VSMC hypertrophy; however, the molecular mechanism mediating this response is obscure. The present study was therefore undertaken to examine whether RV could prevent AngII-induced VSMC hypertrophy through the transactivation of growth factor receptor and associated signaling pathways. AngII treatment of VSMC enhanced the protein synthesis that was attenuated towards control levels by RV pretreatment as well as by the inhibitors of NADPH oxidase, c-Src, and growth factor receptors. Furthermore, RV pretreatment also inhibited enhanced levels of superoxide anion, NADPH oxidase activity, increased expression of NADPH oxidase subunits, and phosphorylation of c-Src, EGF-R, PDGE-R, ERK1/2, and AKT1/2. In conclusion, these results indicate that RV attenuates AngII-induced VSMC hypertrophy through the inhibition of enhanced oxidative stress and activation of c-Src, growth factor receptors, and MAPK/AKT signaling. We suggest that RV could be used as a therapeutic agent in the treatment of vascular complications associated with hypertension and hypertrophy.

  2. Epidermal Growth Factor Receptor Transactivation Is Required for Mitogen-Activated Protein Kinase Activation by Muscarinic Acetylcholine Receptors in HaCaT Keratinocytes

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    Wymke Ockenga

    2014-11-01

    Full Text Available Non-neuronal acetylcholine plays a substantial role in the human skin by influencing adhesion, migration, proliferation and differentiation of keratinocytes. These processes are regulated by the Mitogen-Activated Protein (MAP kinase cascade. Here we show that in HaCaT keratinocytes all five muscarinic receptor subtypes are expressed, but M1 and M3 are the subtypes involved in mitogenic signaling. Stimulation with the cholinergic agonist carbachol leads to activation of the MAP kinase extracellular signal regulated kinase, together with the protein kinase Akt. The activation is fully dependent on the transactivation of the epidermal growth factor receptor (EGFR, which even appears to be the sole pathway for the muscarinic receptors to facilitate MAP kinase activation in HaCaT cells. The transactivation pathway involves a triple-membrane-passing process, based on activation of matrix metalloproteases, and extracellular ligand release; whereas phosphatidylinositol 3-kinase, Src family kinases or protein kinase C do not appear to be involved in MAP kinase activation. Furthermore, phosphorylation, ubiquitination and endocytosis of the EGF receptor after cholinergic transactivation are different from that induced by a direct stimulation with EGF, suggesting that ligands other than EGF itself mediate the cholinergic transactivation.

  3. Stromal cell-derived factor-1α (SDF-1α/CXCL12) stimulates ovarian cancer cell growth through the EGF receptor transactivation

    International Nuclear Information System (INIS)

    Porcile, Carola; Bajetto, Adriana; Barbieri, Federica; Barbero, Simone; Bonavia, Rudy; Biglieri, Marianna; Pirani, Paolo; Florio, Tullio; Schettini, Gennaro

    2005-01-01

    Ovarian cancer (OC) is the leading cause of death in gynecologic diseases in which there is evidence for a complex chemokine network. Chemokines are a family of proteins that play an important role in tumor progression influencing cell proliferation, angiogenic/angiostatic processes, cell migration and metastasis, and, finally, regulating the immune cells recruitment into the tumor mass. We previously demonstrated that astrocytes and glioblastoma cells express both the chemokine receptor CXCR4 and its ligand stromal cell-derived factor-1 (SDF-1), and that SDF-1α treatment induced cell proliferation, supporting the hypothesis that chemokines may play an important role in tumor cells' growth in vitro. In the present study, we report that CXCR4 and SDF-1 are expressed in OC cell lines. We demonstrate that SDF-1α induces a dose-dependent proliferation in OC cells, by the specific interaction with CXCR4 and a biphasic activation of ERK1/2 and Akt kinases. Our results further indicate that CXCR4 activation induces EGF receptor (EGFR) phosphorylation that in turn was linked to the downstream intracellular kinases activation, ERK1/2 and Akt. In addition, we provide evidence for cytoplasmic tyrosine kinase (c-Src) involvement in the SDF-1/CXCR4-EGFR transactivation. These results suggest a possible important 'cross-talk' between SDF-1/CXCR4 and EGFR intracellular pathways that may link signals of cell proliferation in ovarian cancer

  4. DEPENDENCE OF PPAR LIGAND-INDUCED MAPK SIGNALING ON EPIDERMAL GROWTH FACTOR RECEPTOR TRANSACTIVATION HEPARIN-BINDING EGF CLEAVAGE MEDIATES ZINC-INDUCED EGF RECEPTOR PHOSPHORYLATION

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    Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that function as ligand-activated transcription factors regulating lipid metabolism and homeostasis. In addition to their ability to regulate PPAR-mediated gene transcription, PPARalpha and gamma li...

  5. Endothelin B Receptors on Primary Chicken Müller Cells and the Human MIO-M1 Müller Cell Line Activate ERK Signaling via Transactivation of Epidermal Growth Factor Receptors.

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    Mohammad Harun-Or-Rashid

    Full Text Available Injury to the eye or retina triggers Müller cells, the major glia cell of the retina, to dedifferentiate and proliferate. In some species they attain retinal progenitor properties and have the capacity to generate new neurons. The epidermal growth factor receptor (EGFR system and extracellular signal-regulated kinase (ERK signaling are key regulators of these processes in Müller cells. The extracellular signals that modulate and control these processes are not fully understood. In this work we studied whether endothelin receptor signaling can activate EGFR and ERK signaling in Müller cells. Endothelin expression is robustly upregulated at retinal injury and endothelin receptors have been shown to transactivate EGFRs in other cell types. We analyzed the endothelin signaling system in chicken retina and cultured primary chicken Müller cells as well as the human Müller cell line MIO-M1. The Müller cells were stimulated with receptor agonists and treated with specific blockers to key enzymes in the signaling pathway or with siRNAs. We focused on endothelin receptor mediated transactivation of EGFRs by using western blot analysis, quantitative reverse transcriptase PCR and immunocytochemistry. The results showed that chicken Müller cells and the human Müller cell line MIO-M1 express endothelin receptor B. Stimulation by the endothelin receptor B agonist IRL1620 triggered phosphorylation of ERK1/2 and autophosphorylation of (Y1173 EGFR. The effects could be blocked by Src-kinase inhibitors (PP1, PP2, EGFR-inhibitor (AG1478, EGFR-siRNA and by inhibitors to extracellular matrix metalloproteinases (GM6001, consistent with a Src-kinase mediated endothelin receptor response that engage ligand-dependent and ligand-independent EGFR activation. Our data suggest a mechanism for how injury-induced endothelins, produced in the retina, may modulate the Müller cell responses by Src-mediated transactivation of EGFRs. The data give support to a view in

  6. Inhibitory effects of omega-3 fatty acids on injury-induced epidermal growth factor receptor transactivation contribute to delayed wound healing.

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    Turk, Harmony F; Monk, Jennifer M; Fan, Yang-Yi; Callaway, Evelyn S; Weeks, Brad; Chapkin, Robert S

    2013-05-01

    Epidermal growth factor receptor (EGFR)-mediated signaling is required for optimal intestinal wound healing. Since n-3 polyunsaturated fatty acids (PUFA), specifically docosahexaenoic acid (DHA), alter EGFR signaling and suppress downstream activation of key signaling pathways, we hypothesized that DHA would be detrimental to the process of intestinal wound healing. Using a mouse immortalized colonocyte model, DHA uniquely reduced EGFR ligand-induced receptor activation, whereas DHA and its metabolic precursor eicosapentaenoic acid (EPA) reduced wound-induced EGFR transactivation compared with control (no fatty acid or linoleic acid). Under wounding conditions, the suppression of EGFR activation was associated with a reduction in downstream activation of cytoskeletal remodeling proteins (PLCγ1, Rac1, and Cdc42). Subsequently, DHA and EPA reduced cell migration in response to wounding. Mice were fed a corn oil-, DHA-, or EPA-enriched diet prior to intestinal wounding (2.5% dextran sodium sulfate for 5 days followed by termination after 0, 3, or 6 days of recovery). Mortality was increased in EPA-fed mice and colonic histological injury scores were increased in EPA- and DHA-fed mice compared with corn oil-fed (control) mice. Although kinetics of colonic EGFR activation and downstream signaling (PLCγ1, Rac1, and Cdc42) were delayed by both n-3 PUFA, colonic repair was increased in EPA- relative to DHA-fed mice. These results indicate that, during the early response to intestinal wounding, DHA and EPA uniquely delay the activation of key wound-healing processes in the colon. This effect is mediated, at least in part, via suppression of EGFR-mediated signaling and downstream cytoskeletal remodeling.

  7. The aryl hydrocarbon receptor and estrogen receptor alpha differentially modulate nuclear factor erythroid-2-related factor 2 transactivation in MCF-7 breast cancer cells

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    Lo, Raymond; Matthews, Jason, E-mail: jason.matthews@utoronto.ca

    2013-07-15

    Nuclear factor erythroid-2-related factor 2 (NRF2; NFE2L2) plays an important role in mediating cellular protection against reactive oxygen species. NRF2 signaling is positively modulated by the aryl hydrocarbon receptor (AHR) but inhibited by estrogen receptor alpha (ERα). In this study we investigated the crosstalk among NRF2, AHR and ERα in MCF-7 breast cancer cells treated with the NRF2 activator sulforaphane (SFN), a dual AHR and ERα activator, 3,3′-diindolylmethane (DIM), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or 17β-estradiol (E2). SFN-dependent increases in NADPH-dependent oxidoreductase 1 (NQO1) and heme oxygenase I (HMOX1) mRNA levels were significantly reduced after co-treatment with E2. E2-dependent repression of NQO1 and HMOX1 was associated with increased ERα but reduced p300 recruitment and reduced histone H3 acetylation at both genes. In contrast, DIM + SFN or TCDD + SFN induced NQO1 and HMOX1 mRNA expression to levels higher than SFN alone, which was prevented by RNAi-mediated knockdown of AHR. DIM + SFN but not TCDD + SFN also induced recruitment of ERα to NQO1 and HMOX1. However, the presence of AHR at NQO1 and HMOX1 restored p300 recruitment and histone H3 acetylation, thereby reversing the ERα-dependent repression of NRF2. Taken together, our study provides further evidence of functional interplay among NRF2, AHR and ERα signaling pathways through altered p300 recruitment to NRF2-regulated target genes. - Highlights: • We examined crosstalk among ERα, AHR, and NRF2 in MCF-7 breast cancer cells. • AHR enhanced the mRNA expression levels of two NRF2 target genes – HMOX1 and NQO1. • ERα repressed HMOX1 and NQO1 expression via decreased histone acetylation. • AHR prevented ERα-dependent repression of HMOX1 and NQO1.

  8. Molecular basis for gene-specific transactivation by nuclear receptors

    DEFF Research Database (Denmark)

    Jørgensen, Mads Aagaard; Siersbæk, Rasmus; Mandrup, Susanne

    2010-01-01

    Nuclear receptors (NRs) are key transcriptional regulators of metazoan physiology and metabolism. Different NRs bind to similar or even identical core response elements; however, they regulate transcription in a highly receptor- and gene-specific manner. These differences in gene activation can m...... on the recent advances in the molecular mechanisms responsible for receptor- and gene-specific transcriptional activation. This article is part of a Special Issue entitled: Translating nuclear receptors from health to disease....... most likely be accounted for by mechanisms involving receptor-specific interactions with DNA as well as receptor-specific interactions with protein complexes binding to adjacent and distant DNA sequences. Here, we review key molecular aspects of transactivation by NRs with special emphasis...

  9. Inhibitory effects of omega-3 fatty acids on injury-induced epidermal growth factor receptor transactivation contribute to delayed wound healing

    OpenAIRE

    Turk, Harmony F.; Monk, Jennifer M.; Fan, Yang-Yi; Callaway, Evelyn S.; Weeks, Brad; Chapkin, Robert S.

    2013-01-01

    Epidermal growth factor receptor (EGFR)-mediated signaling is required for optimal intestinal wound healing. Since n-3 polyunsaturated fatty acids (PUFA), specifically docosahexaenoic acid (DHA), alter EGFR signaling and suppress downstream activation of key signaling pathways, we hypothesized that DHA would be detrimental to the process of intestinal wound healing. Using a mouse immortalized colonocyte model, DHA uniquely reduced EGFR ligand-induced receptor activation, whereas DHA and its m...

  10. Phosphorylation of Src by phosphoinositide 3-kinase regulates beta-adrenergic receptor-mediated EGFR transactivation.

    Science.gov (United States)

    Watson, Lewis J; Alexander, Kevin M; Mohan, Maradumane L; Bowman, Amber L; Mangmool, Supachoke; Xiao, Kunhong; Naga Prasad, Sathyamangla V; Rockman, Howard A

    2016-10-01

    β2-Adrenergic receptors (β2AR) transactivate epidermal growth factor receptors (EGFR) through formation of a β2AR-EGFR complex that requires activation of Src to mediate signaling. Here, we show that both lipid and protein kinase activities of the bifunctional phosphoinositide 3-kinase (PI3K) enzyme are required for β2AR-stimulated EGFR transactivation. Mechanistically, the generation of phosphatidylinositol (3,4,5)-tris-phosphate (PIP3) by the lipid kinase function stabilizes β2AR-EGFR complexes while the protein kinase activity of PI3K regulates Src activation by direct phosphorylation. The protein kinase activity of PI3K phosphorylates serine residue 70 on Src to enhance its activity and induce EGFR transactivation following βAR stimulation. This newly identified function for PI3K, whereby Src is a substrate for the protein kinase activity of PI3K, is of importance since Src plays a key role in pathological and physiological signaling. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Cell-Surface Receptors Transactivation Mediated by G Protein-Coupled Receptors

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    Cattaneo, Fabio; Guerra, Germano; Parisi, Melania; De Marinis, Marta; Tafuri, Domenico; Cinelli, Mariapia; Ammendola, Rosario

    2014-01-01

    G protein-coupled receptors (GPCRs) are seven transmembrane-spanning proteins belonging to a large family of cell-surface receptors involved in many intracellular signaling cascades. Despite GPCRs lack intrinsic tyrosine kinase activity, tyrosine phosphorylation of a tyrosine kinase receptor (RTK) occurs in response to binding of specific agonists of several such receptors, triggering intracellular mitogenic cascades. This suggests that the notion that GPCRs are associated with the regulation of post-mitotic cell functions is no longer believable. Crosstalk between GPCR and RTK may occur by different molecular mechanism such as the activation of metalloproteases, which can induce the metalloprotease-dependent release of RTK ligands, or in a ligand-independent manner involving membrane associated non-receptor tyrosine kinases, such as c-Src. Reactive oxygen species (ROS) are also implicated as signaling intermediates in RTKs transactivation. Intracellular concentration of ROS increases transiently in cells stimulated with GPCR agonists and their deliberated and regulated generation is mainly catalyzed by enzymes that belong to nicotinamide adenine dinucleotide phosphate (NADPH) oxidase family. Oxidation and/or reduction of cysteine sulfhydryl groups of phosphatases tightly controls the activity of RTKs and ROS-mediated inhibition of cellular phosphatases results in an equilibrium shift from the non-phosphorylated to the phosphorylated state of RTKs. Many GPCR agonists activate phospholipase C, which catalyze the hydrolysis of phosphatidylinositol 4,5-bis-phosphate to produce inositol 1,4,5-triphosphate and diacylglicerol. The consequent mobilization of Ca2+ from endoplasmic reticulum leads to the activation of protein kinase C (PKC) isoforms. PKCα mediates feedback inhibition of RTK transactivation during GPCR stimulation. Recent data have expanded the coverage of transactivation to include Serine/Threonine kinase receptors and Toll-like receptors. Herein, we

  12. Nuclear Factor Kappa B Activation and Peroxisome Proliferator-activated Receptor Transactivational Effects of Chemical Components of the Roots of Polygonum multiflorum.

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    Sun, Ya Nan; Li, Wei; Song, Seok Bean; Yan, Xi Tao; Yang, Seo Young; Kim, Young Ho

    2016-01-01

    Polygonum multiflorum is well-known as "Heshouwu" in traditional Chinese herbal medicine. In Northeast Asia, it is often used as a tonic to prevent premature aging of the kidney and liver, tendons, and bones and strengthening of the lower back and knees. To research the anti-inflammatory activities of components from P. multiflorum. The compounds were isolated by a combination of silica gel and YMC R-18 column chromatography, and their structures were identified by analysis of spectroscopic data (1D, 2D-nuclear magnetic resonance, and mass spectrometry). The anti-inflammatory activities of the isolated compounds 1-15 were evaluated by luciferase reporter gene assays. Fifteen compounds (1-15) were isolated from the roots of P. multiflorum. Compounds 1-5 and 14-15 significantly inhibited tumor necrosis factor-α-induced nuclear factor kappa B-luciferase activity, with IC50 values of 24.16-37.56 μM. Compounds 1-5 also greatly enhanced peroxisome proliferator-activated receptors transcriptional activity with EC50 values of 18.26-31.45 μM. The anthraquinone derivatives were the active components from the roots of P. multiflorum as an inhibitor on inflammation-related factors in human hepatoma cells. Therefore, we suggest that the roots of P. multiflorum can be used to treat natural inflammatory diseases. This study presented that fifteen compounds (1-15) isolated from the roots of Polygonum multiflrum exert signifiant anti inflmmatory effects by inhibiting TNF α induced NF κB activation and PPARs transcription. Abbreviation used: NF κB: Nuclear factor kappa B, PPARs: Peroxisome proliferator activated receptors, PPREs: Peroxisome proliferator response elements, TNF α: Tumor necrosis factor α, ESI-MS: Electrospray ionization mass spectrometry, HepG2: Human hepatoma cells.

  13. Co-factors necessary for PPAR mediated transactivation of endogenous target genes

    DEFF Research Database (Denmark)

    Grøntved, Lars; Nielsen, Ronni; Stunnenberg, Henk

    of endogenous target gene in different cell types are elusive. To mutually compare the ability of the PPAR subtypes to activate endogenous target genes in a given cell, PPARa, PPARb/d and PPARg2 were HA tagged and rapidly, equally and synchronously expressed using adenoviral delivery. Within a few hours after...... subtype specific activation of target genes. Accumulating evidence suggests that transcriptional co-factors can function as master regulators for nuclear receptors and impose promoter selectivity. To study co-factor necessity for PPAR mediated transactivation of endogenous target genes, specific co...

  14. A lactobacillus rhamnosus GG-derived soluble protein, p40, stimulates ligand release from intestinal epithelial cells to transactivate epidermal growth factor receptor

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    Protein p40, a Lactobacillus rhamnosus GG (LGG)-derived soluble protein, ameliorates intestinal injury and colitis, reduces apoptosis and preserves barrier function by activation of EGF receptor (EGFR) in intestinal epithelial cells. The aim of this study was to determine the mechanisms by which p40...

  15. The proteosome inhibitor MG132 attenuates Retinoic Acid Receptor trans-activation and enhances trans-repression of Nuclear Factor κB. Potential relevance to chemo-preventive interventions with retinoids

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    Rosier Randy N

    2004-03-01

    Full Text Available Abstract Background Nuclear factor kappa B (NFκB is a pro-malignant transcription factor with reciprocal effects on pro-metastatic and anti-metastatic gene expression. Interestingly, NFκB blockade results in the reciprocal induction of retinoic acid receptors (RARs. Given the established property of RARs as negative regulators of malignant progression, we postulated that reciprocal interactions between NFκB and RARs constitute a signaling module in metastatic gene expression and malignant progression. Using Line 1 tumor cells as a model for signal regulation of metastatic gene expression, we investigated the reciprocal interactions between NFκB and RARs in response to the pan-RAR agonist, all-trans retinoic acid (at-RA and the pan-RAR antagonist, AGN193109. Results At-RA [0.1–1 μM] dose-dependently activated RAR and coordinately trans-repressed NFκB, while AGN193109 [1–10 μM] dose-dependently antagonized the effects of at-RA. At-RA and AGN193109 reciprocally regulate pro-metastatic matrix metalloprotease 9 (MMP 9 and its endogenous inhibitor, the tissue inhibitor of metalloprotease 1 (TIMP 1, in a manner consistent with the putative roles of NFκB and RAR in malignant progression. Activation of RAR concurs with its ubiquitination and proteosomal degradation. Accordingly, the proteosome inhibitor, MG132 [5 μM], blocked RAR degradation, quelled RAR trans-activation and enhanced RAR trans-repression of NFκB. Conclusion We conclude that reciprocal interactions between NFκB and RARs constitute a signaling module in metastatic gene expression and malignant progression and propose that the dissociative effect of proteosome inhibitors could be harnessed towards enhancing the anticancer activity of retinoids.

  16. Transactivation of ErbB receptors by leptin in the cardiovascular system: mechanisms, consequences and target for therapy.

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    Bełtowski, Jerzy; Jazmroz-Wiśniewska, Anna

    2014-01-01

    Many experimental and clinical studies have demonstrated that elevated leptin concentration in patients with obesity/metabolic syndrome contributes to the pathogenesis of cardiovascular disorders including arterial hypertension, atherosclerosis, restenosis after coronary angioplasty and myocardial hypertrophy. Receptor tyrosine kinases belonging to the ErbB family, especially ErbB1 (epidermal growth factor receptor) and ErbB2 are abundantly expressed in the blood vessels and the heart. EGFR is activated not only by its multiple peptide ligands but also by many other factors including angiotensin II, endothelin-1, norepinephrine, thrombin and prorenin; the phenomenon referred to as "transactivation". Augmented EGFR signaling contributes to abnormalities of vascular tone and renal sodium handling as well as vascular remodeling and myocardial hypertrophy through various intracellular mechanisms, in particular extracellular signal-regulated kinases (ERK) and phosphoinositide 3-kinase (PI3K). Recent experimental studies indicate that chronically elevated leptin transactivates the EGFR through the mechanisms requiring reactive oxygen species and cytosolic tyrosine kinase, c-Src. In addition, hyperleptinemia increases ErbB2 activity in the arterial wall. Stimulation of EGFR and ErbB2 downstream signaling pathways such as ERK and PI3K in the vascular wall and the kidney may contribute to the increase in vascular tone, enhanced tubular sodium reabsorption as well as vascular and renal lesions in hyperleptinemic obese subjects.

  17. GPCR responses in vascular smooth muscle can occur predominantly through dual transactivation of kinase receptors and not classical Gαq protein signalling pathways.

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    Little, Peter J

    2013-05-30

    GPCR signalling is well known to proceed through several linear pathways involving activation of G proteins and their downstream signalling pathways such as activation of phospholipase C. In addition, GPCRs signal via transactivation of Protein Tyrosine Kinase receptors such as that for Epidermal Growth Factor (EGF) and Platelet-Derived Growth Factor (PDGF) where GPCR agonists mediate increase levels of phosphorylated Erk (pErk) the immediate downstream product of the activation of EGF receptor. It has recently been shown that this paradigm can be extended to include the GPCR transactivation of a Protein Serine/Threonine Kinase receptor, specifically the Transforming Growth Factor β Type I receptor (also known as Alk V) (TβRI) in which case GPCR activation leads to the formation of carboxy terminal polyphosphorylated Smad2 (phosphoSmad2) being the immediate downstream product of the activation of TβRI. Growth factor and hormone regulation of proteoglycan synthesis in vascular smooth muscle cells represent one component of an in vitro model of atherosclerosis because modified proteoglycans show enhanced binding to lipoproteins as the initiating step in atherosclerosis. In the example of proteoglycan synthesis stimulated by GPCR agonists such as thrombin and endothelin-1, the transactivation pathways for the EGF receptor and TβRI are both active and together account for essentially all of the response to the GPCRs. In contrast, signalling downstream of GPCRs such as increased inositol 1,4,5 trisphosphate (IP3) and intracellular calcium do not have any effect on GPCR stimulated proteoglycan synthesis. These data lead to the conclusion that dual transactivation pathways for protein tyrosine and serine/threonine kinase receptors may play a far greater role in GPCR signalling than currently recognised. Copyright © 2013 Elsevier Inc. All rights reserved.

  18. A structural explanation of the effects of dissociated glucocorticoids on glucocorticoid receptor transactivation.

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    Dezitter, Xavier; Fagart, Jérôme; Taront, Solenne; Fay, Michel; Masselot, Bernadette; Hétuin, Dominique; Formstecher, Pierre; Rafestin-Oblin, Marie-Edith; Idziorek, Thierry

    2014-02-01

    There is a therapeutic need for glucocorticoid receptor (GR) ligands that distinguish between the transrepression and transactivation activity of the GR, the later thought to be responsible for side effects. These ligands are known as "dissociated glucocorticoids" (dGCs). The first published dGCs, RU24782 (9α-fluoro-11β-hydroxy-16α-methylpregna-21-thiomethyl-1,4-diene-3,20-dione) and RU24858 (9α-fluoro-11β-hydroxy-16α-methylpregna-21-cyanide-1,4-diene-3,20-dione), do not have the 17α-hydroxyl group that characterizes dexamethasone (Dex; 9α-fluoro-11β,17α,21-trihydroxy-16α-methylpregna-1,4-diene-3,20-dione), and they differ from one another by having C21-thiomethyl and C21-cyanide moieties, respectively. Our aim was therefore to establish the structural basis of their activity. Both RU24782 and RU24858 induced a transactivation activity highly dependent on the GR expression level but always lower than dexamethasone. They also display less ability than dexamethasone to trigger steroid receptor coactivator 1 (SRC-1) recruitment and histone H3 acetylation. Docking studies, validated by mutagenesis experiments, revealed that dGCs are not anchored by Gln642, in contrast to Dex, which is hydrogen bonded to this residue via its 17α-hydroxyl group. This contact is essential for SRC-1 recruitment and subsequent dexamethasone-induced GR transactivation, but not transrepression. The ability of dGCs to make contacts with Ile747, for both RU24858 and RU24782 and with Asn564 for RU24858 are not strong enough to maintain GR in a conformation able to efficiently recruit SRC-1, unless SRC-1 is overexpressed. Overall, our findings provide some structural guidelines for the synthesis of potential new dissociated glucocorticoids with a better therapeutic ratio.

  19. Effects of nuclear receptor transactivation on steroid hormone synthesis and gene expression in porcine Leydig cells.

    Science.gov (United States)

    Gray, Matthew A; Squires, E James

    2013-01-01

    Male pigs are routinely castrated at a young age to prevent the formation of androstenone, a 16-androstene testicular steroid that is a major component of boar taint. The practice of castration has been increasingly viewed as unfavorable, due to both economic considerations and animal welfare concerns. Other means of controlling boar taint, including reducing the synthesis of androstenone in the testes, would eliminate the need for castration. In this study, we determined the effects of transactivation of three nuclear receptors, the constitutive androstane receptor (CAR), pregnane X receptor (PXR), and farnesoid X receptor (FXR), on gene expression and steroid hormone metabolism in primary porcine Leydig cells. Primary cells were isolated from mature boars, and transcript expression levels were assayed using real-time PCR. The transcripts of interest included porcine orthologs of common phase I and phase II metabolic enzymes, enzymes involved in steroidogenesis, and transcripts previously shown to be differentially expressed in boars with high androstenone and boar taint levels. Transactivation of CAR, PXR, or FXR increased the expression of several genes involved in steroidogenesis, including cytochrome B5A (CYB5A) and cytochrome B5 reductase 1 (CYB5R1), as well as hydroxysteroid (17-beta) dehydrogenase 4 (HSD17B4) and retinol dehydrogenase 12 (RDH12). Treatment with (6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde-O-(3,4-dichlorobenzyl)oxime (CITCO), a CAR agonist, or rifampicin (RIF), a PXR agonist, resulted in significantly (pnuclear receptors may lead to increased levels of 16-androstene steroids, likely by altering the activity of CYP17A1 through CYB5A and CYB5R1 to the andien-β synthase reaction and away from the 17α-hydroxylase and C17, 20 lyase reactions. Copyright © 2012 Elsevier Ltd. All rights reserved.

  20. Characterization of the novel progestin gestodene by receptor binding studies and transactivation assays.

    Science.gov (United States)

    Fuhrmann, U; Slater, E P; Fritzemeier, K H

    1995-01-01

    Gestodene is a novel progestin used in oral contraceptives with an increased separation of progestogenic versus androgenic activity and a distinct antimineralocorticoid activity. This specific pharmacological profile of gestodene is defined by its pattern of binding affinities to a variety of steroid hormone receptors. In the present study the affinity of gestodene to the progesterone receptor (PR), the androgen receptor (AR), the glucocorticoid receptor (GR), the mineralocorticoid receptor (MR) and the estrogen receptor (ER) was re-evaluated by steroid binding assays and compared to those obtained for 3-keto-desogestrel and progesterone. The two synthetic progestins displayed identical high affinity to rabbit PR and similar marked binding to rat AR and GR, while progesterone showed high affinity to PR but only low binding to AR and GR. Furthermore, 3-keto-desogestrel exhibited almost no binding to MR, whereas gestodene, similar to progesterone, showed marked affinity to this receptor. In addition to receptor binding studies, transactivation assays were carried out to investigate the effects of gestodene on AR-, GR- and MR-mediated induction of transcription. In contrast to progesterone, which showed antiandrogenic activity, gestodene and 3-keto-desogestrel both exhibited androgenic activity. Furthermore, all three progestins exhibited weak GR-mediated antagonistic activity. In contrast to progesterone, which showed almost no glucocorticoid activity, gestodene and 3-keto-desogestrel showed weak glucocorticoid action. In addition, gestodene inhibited the aldosterone-induced reporter gene transcription, similar to progesterone, whereas unlike progesterone, gestodene did not induce reporter gene transcription. 3-Keto-desogestrel showed neither antimineralocorticoid nor mineralocorticoid action.

  1. Use of cryopreserved transiently transfected cells in high-throughput pregnane X receptor transactivation assay.

    Science.gov (United States)

    Zhu, Zhengrong; Puglisi, Jaime; Connors, David; Stewart, Jeremy; Herbst, John; Marino, Anthony; Sinz, Michael; O'Connell, Jonathan; Banks, Martyn; Dickinson, Kenneth; Cacace, Angela

    2007-03-01

    Cryopreserved, transiently transfected HepG2 cells were compared to freshly transfected HepG2 cells for use in a pregnane X receptor (PXR) transactivation assay. Assay performance was similar for both cell preparations; however, cryopreserved cells demonstrated less interassay variation. Validation with drugs of different PXR activation potencies and efficacies demonstrated an excellent correlation (r(2) > 0.95) between cryopreserved and fresh cells. Cryopreservation did not change the effect of known CYP3A4 inducers that have poor cell permeability, indicating that cryopreservation had little effect on membrane permeability. In addition, cryopreserved HepG2 cells did not exhibit enhanced susceptibility to cytotoxic compounds compared to transiently transfected control cells. The use of cryopreserved cells enables this assay to run with enhanced efficiency.

  2. Antidepressant drugs transactivate TrkB neurotrophin receptors in the adult rodent brain independently of BDNF and monoamine transporter blockade.

    Directory of Open Access Journals (Sweden)

    Tomi Rantamäki

    Full Text Available BACKGROUND: Antidepressant drugs (ADs have been shown to activate BDNF (brain-derived neurotrophic factor receptor TrkB in the rodent brain but the mechanism underlying this phenomenon remains unclear. ADs act as monoamine reuptake inhibitors and after prolonged treatments regulate brain bdnf mRNA levels indicating that monoamine-BDNF signaling regulate AD-induced TrkB activation in vivo. However, recent findings demonstrate that Trk receptors can be transactivated independently of their neurotrophin ligands. METHODOLOGY: In this study we examined the role of BDNF, TrkB kinase activity and monoamine reuptake in the AD-induced TrkB activation in vivo and in vitro by employing several transgenic mouse models, cultured neurons and TrkB-expressing cell lines. PRINCIPAL FINDINGS: Using a chemical-genetic TrkB(F616A mutant and TrkB overexpressing mice, we demonstrate that ADs specifically activate both the maturely and immaturely glycosylated forms of TrkB receptors in the brain in a TrkB kinase dependent manner. However, the tricyclic AD imipramine readily induced the phosphorylation of TrkB receptors in conditional bdnf⁻/⁻ knock-out mice (132.4±8.5% of control; P = 0.01, indicating that BDNF is not required for the TrkB activation. Moreover, using serotonin transporter (SERT deficient mice and chemical lesions of monoaminergic neurons we show that neither a functional SERT nor monoamines are required for the TrkB phosphorylation response induced by the serotonin selective reuptake inhibitors fluoxetine or citalopram, or norepinephrine selective reuptake inhibitor reboxetine. However, neither ADs nor monoamine transmitters activated TrkB in cultured neurons or cell lines expressing TrkB receptors, arguing that ADs do not directly bind to TrkB. CONCLUSIONS: The present findings suggest that ADs transactivate brain TrkB receptors independently of BDNF and monoamine reuptake blockade and emphasize the need of an intact tissue context for the

  3. 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

    International Nuclear Information System (INIS)

    Wang, Yanling; Fälting, Johanna M.; Mattsson, Charlotte L.; Holmström, Therése E.; Nedergaard, Jan

    2013-01-01

    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 (α 1 -adrenoceptor coupled via G q ), clonidine (α 2 via G i ) or CL316243 (β 3 via G s ) or via β 1 -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 (IC 50 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. •

  4. Nuclear receptor corepressor-dependent repression of peroxisome-proliferator-activated receptor delta-mediated transactivation

    DEFF Research Database (Denmark)

    Krogsdam, Anne-M; Nielsen, Curt A F; Neve, Søren

    2002-01-01

    delta-RXR alpha heterodimer bound to an acyl-CoA oxidase (ACO)-type peroxisome-proliferator response element recruited a glutathione S-transferase-NCoR fusion protein in a ligand-independent manner. Contrasting with most other nuclear receptors, PPAR delta was found to interact equally well......The nuclear receptor corepressor (NCoR) was isolated as a peroxisome-proliferator-activated receptor (PPAR) delta interacting protein using the yeast two-hybrid system. NCoR interacted strongly with the ligand-binding domain of PPAR delta, whereas interactions with the ligand-binding domains...

  5. Fkbp52 Regulates Androgen Receptor Transactivation Activity and Male Urethra Morphogenesis*

    Science.gov (United States)

    Chen, Hanying; Yong, Weidong; Hinds, Terry D.; Yang, Zuocheng; Zhou, Yuhong; Sanchez, Edwin R.; Shou, Weinian

    2010-01-01

    Hypospadias is a common birth defect in humans, yet its etiology and pattern of onset are largely unknown. Recent studies have shown that male mice with targeted ablation of FK506-binding protein-52 (Fkbp52) develop hypospadias, most likely due to actions of Fkbp52 as a molecular co-chaperone of the androgen receptor (AR). Here, we further dissect the developmental and molecular mechanisms that underlie hypospadias in Fkbp52-deficient mice. Scanning electron microscopy revealed a defect in the elevation of prepucial swelling that led to the onset of the ventral penile cleft. Interestingly, expression of Fkbp52 was highest in the ventral aspect of the developing penis that undergoes fusion of the urethral epithelium. Although in situ hybridization and immunohistochemical analyses suggested that Fkbp52 mutants had a normal urethral epithelium signaling center and epithelial differentiation, a reduced apoptotic cell index at ventral epithelial cells at the site of fusion and a defect of genital mesenchymal cell migration were observed. Supplementation of gestating females with excess testosterone partially rescued the hypospadic phenotype in Fkbp52 mutant males, showing that loss of Fkbp52 desensitizes AR to hormonal activation. Direct measurement of AR activity was performed in mouse embryonic fibroblast cells treated with dihydrotestosterone or synthetic agonist R1881. Reduced AR activity at genes controlling sexual dimorphism and cell growth was found in Fkbp52-deficient mouse embryonic fibroblast cells. However, chromatin immunoprecipitation analysis revealed normal occupancy of AR at gene promoters, suggesting that Fkbp52 exerts downstream effects on the transactivation function of AR. Taken together, our data show Fkbp52 to be an important molecular regulator in the androgen-mediated pathway of urethra morphogenesis. PMID:20605780

  6. Fkbp52 regulates androgen receptor transactivation activity and male urethra morphogenesis.

    Science.gov (United States)

    Chen, Hanying; Yong, Weidong; Hinds, Terry D; Yang, Zuocheng; Zhou, Yuhong; Sanchez, Edwin R; Shou, Weinian

    2010-09-03

    Hypospadias is a common birth defect in humans, yet its etiology and pattern of onset are largely unknown. Recent studies have shown that male mice with targeted ablation of FK506-binding protein-52 (Fkbp52) develop hypospadias, most likely due to actions of Fkbp52 as a molecular co-chaperone of the androgen receptor (AR). Here, we further dissect the developmental and molecular mechanisms that underlie hypospadias in Fkbp52-deficient mice. Scanning electron microscopy revealed a defect in the elevation of prepucial swelling that led to the onset of the ventral penile cleft. Interestingly, expression of Fkbp52 was highest in the ventral aspect of the developing penis that undergoes fusion of the urethral epithelium. Although in situ hybridization and immunohistochemical analyses suggested that Fkbp52 mutants had a normal urethral epithelium signaling center and epithelial differentiation, a reduced apoptotic cell index at ventral epithelial cells at the site of fusion and a defect of genital mesenchymal cell migration were observed. Supplementation of gestating females with excess testosterone partially rescued the hypospadic phenotype in Fkbp52 mutant males, showing that loss of Fkbp52 desensitizes AR to hormonal activation. Direct measurement of AR activity was performed in mouse embryonic fibroblast cells treated with dihydrotestosterone or synthetic agonist R1881. Reduced AR activity at genes controlling sexual dimorphism and cell growth was found in Fkbp52-deficient mouse embryonic fibroblast cells. However, chromatin immunoprecipitation analysis revealed normal occupancy of AR at gene promoters, suggesting that Fkbp52 exerts downstream effects on the transactivation function of AR. Taken together, our data show Fkbp52 to be an important molecular regulator in the androgen-mediated pathway of urethra morphogenesis.

  7. Differential transactivation by orphan nuclear receptor NOR1 and its fusion gene product EWS/NOR1: possible involvement of poly(ADP-ribose) polymerase I, PARP-1.

    Science.gov (United States)

    Ohkura, Naganari; Nagamura, Yuko; Tsukada, Toshihiko

    2008-10-15

    In extraskeletal myxoid chondrosarcoma, a chromosomal translocation creates a gene fusion between EWS and an orphan nuclear receptor, NOR1. The resulting fusion protein EWS/NOR1 has been believed to lead to malignant transformation by functioning as a transactivator for NOR1-target genes. By comparing the gene expression profiles of NOR1- and EWS/NOR1-overexpressing cells, we found that they largely shared up-regulated genes, but no significant correlation was observed with respect to the transactivation levels of each gene. In addition, the proteins associated with NOR1 and EWS/NOR1 were mostly the same in these cells. The results suggest that these proteins differentially transactivate overlapping target genes through a similar transcriptional machinery. To clarify the mechanisms underlying the transcriptional divergence between NOR1 and EWS/NOR1, we searched for alternatively associated proteins, and identified poly(ADP-ribose) polymerase I (PARP-1) as an NOR1-specific binding protein. Consistent with its binding properties, PARP-1 acted as a transcriptional repressor of NOR1, but not EWS/NOR1, in a luciferase reporter assay employing PARP-1(-/-) fibroblasts. Interestingly, suppressive activity of PARP-1 was observed in a DNA response element-specific manner, and in a subtype-specific manner toward the NR4A family (Nur77, Nurr1, and NOR1), suggesting that PARP-1 plays a role in the diversity of transcriptional regulation mediated by the NR4A family in normal cells. Altogether, our findings suggest that NOR1 and EWS/NOR1 regulate overlapping target genes differently by utilizing associated proteins, including PARP-1; and that EWS/NOR1 may acquire oncogenic activities by avoiding (or gaining) transcription factor-specific modulation by the associated proteins. (c) 2008 Wiley-Liss, Inc.

  8. SAR of PXR transactivation in benzimidazole-based IGF-1R kinase inhibitors.

    Science.gov (United States)

    Zimmermann, Kurt; Wittman, Mark D; Saulnier, Mark G; Velaparthi, Upender; Sang, Xiaopeng; Frennesson, David B; Struzynski, Charles; Seitz, Steven P; He, Liqi; Carboni, Joan M; Li, Aixin; Greer, Ann F; Gottardis, Marco; Attar, Ricardo M; Yang, Zheng; Balimane, Praveen; Discenza, Lorell N; Lee, Francis Y; Sinz, Michael; Kim, Sean; Vyas, Dolatrai

    2010-03-01

    The SAR of PXR transactivation by 3-(benzimidazol-2-yl)-pyridine-2-one based ATP competitive inhibitors of Insulin-like Growth Factor 1 Receptor kinase (IGF-1R) is discussed. Compounds without PXR transactivation, with in vivo antitumor activity, reduced protein binding and improved oral exposure are presented. Copyright 2010 Elsevier Ltd. All rights reserved.

  9. Mineralocorticoid Receptor (MR) trans-Activation of Inflammatory AP-1 Signaling

    Science.gov (United States)

    Dougherty, Edward J.; Elinoff, Jason M.; Ferreyra, Gabriela A.; Hou, Angela; Cai, Rongman; Sun, Junfeng; Blaine, Kevin P.; Wang, Shuibang; Danner, Robert L.

    2016-01-01

    Glucocorticoids are commonly used to treat inflammatory disorders. The glucocorticoid receptor (GR) can tether to inflammatory transcription factor complexes, such as NFκB and AP-1, and trans-repress the transcription of cytokines, chemokines, and adhesion molecules. In contrast, aldosterone and the mineralocorticoid receptor (MR) primarily promote cardiovascular inflammation by incompletely understood mechanisms. Although MR has been shown to weakly repress NFκB, its role in modulating AP-1 has not been established. Here, the effects of GR and MR on NFκB and AP-1 signaling were directly compared using a variety of ligands, two different AP-1 consensus sequences, GR and MR DNA-binding domain mutants, and siRNA knockdown or overexpression of core AP-1 family members. Both GR and MR repressed an NFκB reporter without influencing p65 or p50 binding to DNA. Likewise, neither GR nor MR affected AP-1 binding, but repression or activation of AP-1 reporters occurred in a ligand-, AP-1 consensus sequence-, and AP-1 family member-specific manner. Notably, aldosterone interactions with both GR and MR demonstrated a potential to activate AP-1. DNA-binding domain mutations that eliminated the ability of GR and MR to cis-activate a hormone response element-driven reporter variably affected the strength and polarity of these responses. Importantly, MR modulation of NFκB and AP-1 signaling was consistent with a trans-mechanism, and AP-1 effects were confirmed for specific gene targets in primary human cells. Steroid nuclear receptor trans-effects on inflammatory signaling are context-dependent and influenced by nuclear receptor conformation, DNA sequence, and the expression of heterologous binding partners. Aldosterone activation of AP-1 may contribute to its proinflammatory effects in the vasculature. PMID:27650495

  10. Cav1.3 channel α1D protein is overexpressed and modulates androgen receptor transactivation in prostate cancers.

    Science.gov (United States)

    Chen, Ruibao; Zeng, Xing; Zhang, Ruitao; Huang, Jiaoti; Kuang, Xiangxing; Yang, Jun; Liu, Jihong; Tawfik, Ossama; Thrasher, James Brantley; Li, Benyi

    2014-07-01

    Widespread use of L-type calcium channel blockers for treating hypertension has led to multiple epidemiologic studies to assess the risk of prostate cancer incidence. These studies revealed a reverse correlation between the likelihood of prostate cancer risk and the use of L-type calcium channel blockers among men without family history but the mechanism was not clear. In this study, we examined the expression profiles of multiple L-type calcium channel genes in prostate cancers and determined their functional roles in androgen receptor (AR) transactivation and cell growth. By reanalyzing the ONCOMINE database, we found that L-type calcium channel CACNA1D gene expression levels in cancer tissues were significantly higher than noncancer tissues in 14 of 15 published complementary deoxyribonucleic acid microarray data sets, of which 9 data sets showed an increase of 2- to 17-folds. Quantitative polymerase chain reaction and immunostaining experiments revealed that CACNA1D gene and its coding protein α1D were highly expressed in prostate cancers, especially in castration-resistant diseases, compared with benign prostate tissues. Consistent with the notion of CACNA1D as an ERG-regulated gene, CACNA1D gene expression levels were significantly higher in prostate cancers with TMPRSS2-ERG gene fusion compared with the cases without this gene fusion. Blocking L-type channel's function or knocking down CACNA1D gene expression significantly suppressed androgen-stimulated Ca(2+) influx, AR transactivation, and cell growth in prostate cancer cells. Taken together, these data suggest that CACNA1D gene overexpression is associated with prostate cancer progression and might play an important role in Ca(2+) influx, AR activation, and cell growth in prostate cancer cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Vitamin D3 transactivates the zinc and manganese transporter SLC30A10 via the Vitamin D receptor.

    Science.gov (United States)

    Claro da Silva, Tatiana; Hiller, Christian; Gai, Zhibo; Kullak-Ublick, Gerd A

    2016-10-01

    Vitamin D3 regulates genes critical for human health and its deficiency is associated with an increased risk for osteoporosis, cancer, diabetes, multiple sclerosis, hypertension, inflammatory and immunological diseases. To study the impact of vitamin D3 on genes relevant for the transport and metabolism of nutrients and drugs, we employed next-generation sequencing (NGS) and analyzed global gene expression of the human-derived Caco-2 cell line treated with 500nM vitamin D3. Genes involved in neuropeptide signaling, inflammation, cell adhesion and morphogenesis were differentially expressed. Notably, genes implicated in zinc, manganese and iron homeostasis were largely increased by vitamin D3 treatment. An ∼10-fold increase in ceruloplasmin and ∼4-fold increase in haptoglobin gene expression suggested a possible association between vitamin D and iron homeostasis. SLC30A10, the gene encoding the zinc and manganese transporter ZnT10, was the chiefly affected transporter, with ∼15-fold increase in expression. SLC30A10 is critical for zinc and manganese homeostasis and mutations in this gene, resulting in impaired ZnT10 function or expression, cause manganese intoxication, with Parkinson-like symptoms. Our NGS results were validated by real-time PCR in Caco-2 cells, as well as in duodenal biopsies taken from healthy human subjects treated with 0.5μg vitamin D3 daily for 10 days. In addition to increasing gene expression of SLC30A10 and the positive control TRPV6, vitamin D3 also increased ZnT10 protein expression, as indicated by Western blot and cytofluorescence. In silico identification of potential vitamin D responsive elements (VDREs) in the 5'-flanking region of the SLC30A10 promoter and dual-luciferase reporter assay showed enhanced promoter activity in the presence of vitamin D receptor (VDR) and retinoid X receptor (RXR) constructs, as well as vitamin D3, but not when one of these factors was absent. Electrophoretic mobility shift assay (EMSA) and

  12. Pax6 represses androgen receptor-mediated transactivation by inhibiting recruitment of the coactivator SPBP.

    Directory of Open Access Journals (Sweden)

    Julianne Elvenes

    Full Text Available The androgen receptor (AR has a central role in development and maintenance of the male reproductive system and in the etiology of prostate cancer. The transcription factor Pax6 has recently been reported to act as a repressor of AR and to be hypermethylated in prostate cancer cells. SPBP is a transcriptional regulator that previously has been shown to enhance the activity of Pax6. In this study we have identified SPBP to act as a transcriptional coactivator of AR. We also show that Pax6 inhibits SPBP-mediated enhancement of AR activity on the AR target gene probasin promoter, a repression that was partly reversed by increased expression of SPBP. Enhanced expression of Pax6 reduced the amount of SPBP associated with the probasin promoter when assayed by ChIP in HeLa cells. We mapped the interaction between both AR and SPBP, and AR and Pax6 to the DNA-binding domains of the involved proteins. Further binding studies revealed that Pax6 and SPBP compete for binding to AR. These results suggest that Pax6 represses AR activity by displacing and/or inhibiting recruitment of coactivators to AR target promoters. Understanding the mechanism for inhibition of AR coactivators can give rise to molecular targeted drugs for treatment of prostate cancer.

  13. In brown adipocytes, adrenergically induced β{sub 1}-/β{sub 3}-(G{sub s})-, α{sub 2}-(G{sub i})- and α{sub 1}-(G{sub q})-signalling to Erk1/2 activation is not mediated via EGF receptor transactivation

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yanling; Fälting, Johanna M.; Mattsson, Charlotte L.; Holmström, Therése E.; Nedergaard, Jan, E-mail: jan@metabol.su.se

    2013-10-15

    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 (α{sub 1}-adrenoceptor coupled via G{sub q}), clonidine (α{sub 2} via G{sub i}) or CL316243 (β{sub 3} via G{sub s}) or via β{sub 1}-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 (IC{sub 50} 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

  14. The activation of G protein-coupled estrogen receptor induces relaxation via cAMP as well as potentiates contraction via EGFR transactivation in porcine coronary arteries.

    Directory of Open Access Journals (Sweden)

    Xuan Yu

    Full Text Available Estrogen exerts protective effects against cardiovascular diseases in premenopausal women, but is associated with an increased risk of both coronary heart disease and stroke in older postmenopausal women. Studies have shown that activation of the G-protein-coupled estrogen receptor 1 (GPER can cause either relaxation or contraction of arteries. It is highly likely that these dual actions of GPER may contribute to the seemingly paradoxical effects of estrogen in regulating coronary artery function. The objective of this study was to test the hypothesis that activation of GPER enhances agonist-stimulated porcine coronary artery contraction via epidermal growth factor receptor (EGFR transactivation and its downstream extracellular signal-regulated kinases (ERK1/2 pathway. Isometric tension studies and western blot were performed to determine the effect of GPER activation on coronary artery contraction. Our findings demonstrated that G-1 caused concentration-dependent relaxation of ET-1-induced contraction, while pretreatment of arterial rings with G-1 significantly enhanced ET-1-induced contraction. GPER antagonist, G-36, significantly inhibited both the G-1-induced relaxation effect and G-1-enhanced ET-1 contraction. Gallein, a Gβγ inhibitor, significantly increased G-1-induced relaxation, yet inhibited G-1-enhanced ET-1-mediated contraction. Similarly, inhibition of EGFR with AG1478 or inhibition of Src with phosphatase 2 further increased G-1-induced relaxation responses in coronary arteries, but decreased G-1-enhanced ET-1-induced contraction. Western blot experiments in porcine coronary artery smooth muscle cells (PCASMC showed that G-1 increased tyrosine phosphorylation of EGFR, which was inhibited by AG-1478. Furthermore, enzyme-linked immunosorbent assays showed that the level of heparin-binding EGF (HB-EGF released by ET-1 treatment increased two-fold; whereas pre-incubation with G-1 further increased ET-1-induced HB-EGF release to four

  15. CysLT1 receptor-induced human airway smooth muscle cells proliferation requires ROS generation, EGF receptor transactivation and ERK1/2 phosphorylation

    Directory of Open Access Journals (Sweden)

    Capra Valérie

    2006-03-01

    Full Text Available Abstract Background Cysteine-containing leukotrienes (cysteinyl-LTs are pivotal inflammatory mediators that play important roles in the pathophysiology of asthma, allergic rhinitis, and other inflammatory conditions. In particular, cysteinyl-LTs exert a variety of effects with relevance to the aetiology of asthma such as smooth muscle contraction, eosinophil recruitment, increased microvascular permeability, enhanced mucus secretion and decreased mucus transport and, finally, airway smooth muscle cells (ASMC proliferation. We used human ASMC (HASMC to identify the signal transduction pathway(s of the leukotriene D4 (LTD4-induced DNA synthesis. Methods Proliferation of primary HASMC was measured by [3H]thymidine incorporation. Phosphorylation of EGF receptor (EGF-R and ERK1/2 was assessed with a polyclonal anti-EGF-R or anti-phosphoERKl/2 monoclonal antibody. A Ras pull-down assay kit was used to evaluate Ras activation. The production of reactive oxygen species (ROS was estimated by measuring dichlorodihydrofluorescein (DCF oxidation. Results We demonstrate that in HASMC LTD4-stimulated thymidine incorporation and potentiation of EGF-induced mitogenic signaling mostly depends upon EGF-R transactivation through the stimulation of CysLT1-R. Accordingly, we found that LTD4 stimulation was able to trigger the increase of Ras-GTP and, in turn, to activate ERK1/2. We show here that EGF-R transactivation was sensitive to pertussis toxin (PTX and phosphoinositide 3-kinase (PI3K inhibitors and that it occurred independently from Src activity, despite the observation of a strong impairment of LTD4-induced DNA synthesis following Src inhibition. More interestingly, CysLT1-R stimulation increased the production of ROS and N-acetylcysteine (NAC abolished LTD4-induced EGF-R phosphorylation and thymidine incorporation. Conclusion Collectively, our data demonstrate that in HASMC LTD4 stimulation of a Gi/o coupled CysLT1-R triggers the transactivation of the EGF

  16. Estrogen receptor transcription and transactivation: Estrogen receptor alpha and estrogen receptor beta - regulation by selective estrogen receptor modulators and importance in breast cancer

    International Nuclear Information System (INIS)

    S Katzenellenbogen, Benita; A Katzenellenbogen, John

    2000-01-01

    Estrogens display intriguing tissue-selective action that is of great biomedical importance in the development of optimal therapeutics for the prevention and treatment of breast cancer, for menopausal hormone replacement, and for fertility regulation. Certain compounds that act through the estrogen receptor (ER), now referred to as selective estrogen receptor modulators (SERMs), can demonstrate remarkable differences in activity in the various estrogen target tissues, functioning as agonists in some tissues but as antagonists in others. Recent advances elucidating the tripartite nature of the biochemical and molecular actions of estrogens provide a good basis for understanding these tissue-selective actions. As discussed in this thematic review, the development of optimal SERMs should now be viewed in the context of two estrogen receptor subtypes, ERα and ERβ, that have differing affinities and responsiveness to various SERMs, and differing tissue distribution and effectiveness at various gene regulatory sites. Cellular, biochemical, and structural approaches have also shown that the nature of the ligand affects the conformation assumed by the ER-ligand complex, thereby regulating its state of phosphorylation and the recruitment of different coregulator proteins. Growth factors and protein kinases that control the phosphorylation state of the complex also regulate the bioactivity of the ER. These interactions and changes determine the magnitude of the transcriptional response and the potency of different SERMs. As these critical components are becoming increasingly well defined, they provide a sound basis for the development of novel SERMs with optimal profiles of tissue selectivity as medical therapeutic agents

  17. Non-canonical Glucocorticoid Receptor Transactivation of gilz by Alcohol Suppresses Cell Inflammatory Response

    Directory of Open Access Journals (Sweden)

    Hang Pong Ng

    2017-06-01

    Full Text Available Acute alcohol exposure suppresses cell inflammatory response. The underlying mechanism has not been fully defined. Here we report that alcohol was able to activate glucocorticoid receptor (GR signaling in the absence of glucocorticoids (GCs and upregulated glucocorticoid-induced leucine zipper (gilz, a prominent GC-responsive gene. Such a non-canonical activation of GR was not blocked by mifepristone, a potent GC competitor. The proximal promoter of gilz, encompassing five GC-responsive elements (GREs, was incorporated and tested in a luciferase reporter system. Deletion and/or mutation of the GREs abrogated the promoter responsiveness to alcohol. Thus, the GR–GRE interaction transduced the alcohol action on gilz. Alcohol induced GR nuclear translocation, which was enhanced by the alcohol dehydrogenase inhibitor fomepizole, suggesting that it was alcohol, not its metabolites, that engendered the effect. Gel mobility shift assay showed that unliganded GR was able to bind GREs and such interaction withstood clinically relevant levels of alcohol. GR knockout via CRISPR/Cas9 gene targeting or GILZ depletion via small RNA interference diminished alcohol suppression of cell inflammatory response to LPS. Thus, a previously unrecognized, non-canonical GR activation of gilz is involved in alcohol modulation of cell immune response.

  18. Overexpression of ERβ is sufficient to inhibit hypoxia-inducible factor-1 transactivation

    Energy Technology Data Exchange (ETDEWEB)

    Park, Choa; Lee, YoungJoo, E-mail: yjlee@sejong.ac.kr

    2014-07-18

    Highlights: • We examined the effect of ERβ specific ligand on HIF-1 inhibition. • DPN down-regulates the ARNT protein levels in PC3 cells. • DPN did not show additional effect in ERβ transfected MCF-7 cells. • Our study shows that unliganded ERβ is sufficient to inhibit HIF-1 in systems of overexpression. - Abstract: Estrogen receptor (ER) β is predicted to play an important role in the prevention of breast cancer development and progression. We have previously shown that ERβ suppresses hypoxia inducible factor (HIF)-1-mediated transcription through aryl hydrocarbon receptor nuclear translocator (ARNT) degradation via ubiquitination processes. In this study, we attempted to examine the effect of ERβ specific ligand on HIF-1 inhibition in ERβ positive PC3 cells and ERβ transfected MCF-7 cells. ERβ specific agonist diarylpropionitrile (DPN) stimulated estrogen response element (ERE)-luciferase activity in a similar fashion to estradiol in PC3 cells. We observed that DPN down-regulates the ARNT protein levels leading to an attenuation of hypoxia-induced hypoxia response element (HRE)-driven luciferase reporter gene activation in PC3 cells. Treatment of DPN reduced vascular endothelial growth factor (VEGF) expression and co-treatment with ERβ specific antagonist PHTPP abrogated the effect in PC3 cells. We then examined the effect of DPN in ERβ transfected MCF-7 cells. HIF-1 transcriptional activity repression by ERβ was not further reduced by DPN, as examined by HRE-driven luciferase assays. Expression of ERβ significantly decreased VEGF secretion and ARNT expression under hypoxic conditions. However, DPN did not additionally affect this suppression in MCF-7 cells transfected with ERβ. This result shows that unliganded ERβ is sufficient to inhibit HIF-1 in systems of overexpression.

  19. The C-terminal domain of the nuclear factor I-B2 isoform is glycosylated and transactivates the WAP gene in the JEG-3 cells

    International Nuclear Information System (INIS)

    Mukhopadhyay, Sudit S.; Rosen, Jeffrey M.

    2007-01-01

    The transcription factor nuclear factor I (NFI) has been shown previously both in vivo and in vitro to be involved in the cooperative regulation of whey acidic protein (WAP) gene transcription along with the glucocorticoid receptor and STAT5. In addition, one of the specific NFI isoforms, NFI-B2, was demonstrated in transient co-transfection experiments in JEG cells, which lack endogenous NFI, to be preferentially involved in the cooperative regulation of WAP gene expression. A comparison of the DNA-binding specificities of the different NFI isoforms only partially explained their differential ability to activate the WAP gene transcription. Here, we analyzed the transactivation regions of two NFI isoforms by making chimeric proteins between the NFI-A and B isoforms. Though, their DNA-binding specificities were not altered as compared to the corresponding wild-type transcription factors, the C-terminal region of the NFI-B isoform was shown to preferentially activate WAP gene transcription in cooperation with GR and STAT5 in transient co-transfection assays in JEG-3 cells. Furthermore, determination of serine and threonine-specific glycosylation (O-linked N-acetylglucosamine) of the C-terminus of the NFI-B isoform suggested that the secondary modification by O-GlcNAc might play a role in the cooperative regulation of WAP gene transcription by NFI-B2 and STAT5

  20. Newly developed quantitative transactivation system shows difference in activation by Vitis CBF transcription factors on DRE/CRT elements.

    Science.gov (United States)

    Nassuth, Annette; Siddiqua, Mahbuba; Xiao, Huogen; Moody, Michelle A; Carlow, Chevonne E

    2014-01-01

    Agroinfiltration-based transactivation systems can determine if a protein functions as a transcription factor, and via which promoter element. However, this activation is not always a yes or no proposition. Normalization for variation in plasmid delivery into plant cells, sample collection and protein extraction is desired to allow for a quantitative comparison between transcription factors or promoter elements. We developed new effector and reporter plasmids which carry additional reporter genes, as well as a procedure to assay all three reporter enzymes from a single extract. The applicability of these plasmids was demonstrated with the analysis of CBF transcription factors and their target promoter sequence, DRE/CRT. Changes in the core DRE/CRT sequence abolished activation by Vitis CBF1 or Vitis CBF4, whereas changes in the surrounding sequence lowered activation by Vitis CBF1 but much less so for Vitis CBF4. The system also detected a reduction in activation due to one amino acid change in Vitis CBF1. The newly developed effector and reporter plasmids improve the ability to quantitatively compare the activation on two different promoter elements by the same transcription factor, or between two different transcription factors on the same promoter element. The quantitative difference in activation by VrCBF1 and VrCBF4 on various DRE/CRT elements support the hypothesis that these transcription factors have unique roles in the cold acclimation process.

  1. The MHC-II transactivator CIITA, a restriction factor against oncogenic HTLV-1 and HTLV-2 retroviruses: similarities and differences in the inhibition of Tax-1 and Tax-2 viral transactivators

    Science.gov (United States)

    Forlani, Greta; Abdallah, Rawan; Accolla, Roberto S.; Tosi, Giovanna

    2013-01-01

    The activation of CD4+ T helper cells is strictly dependent on the presentation of antigenic peptides by MHC class II (MHC-II) molecules. MHC-II expression is primarily regulated at the transcriptional level by the AIR-1 gene product CIITA (class II transactivator). Thus, CIITA plays a pivotal role in the triggering of the adaptive immune response against pathogens. Besides this well known function, we recently found that CIITA acts as an endogenous restriction factor against HTLV-1 (human T cell lymphotropic virus type 1) and HTLV-2 oncogenic retroviruses by targeting their viral transactivators Tax-1 and Tax-2, respectively. Here we review our findings on CIITA-mediated inhibition of viral replication and discuss similarities and differences in the molecular mechanisms by which CIITA specifically counteracts the function of Tax-1 and Tax-2 molecules. The dual function of CIITA as a key regulator of adaptive and intrinsic immunity represents a rather unique example of adaptation of host-derived factors against pathogen infections during evolution. PMID:23986750

  2. Structural characterization of a noncovalent complex between ubiquitin and the transactivation domain of the erythroid-specific factor EKLF.

    Science.gov (United States)

    Raiola, Luca; Lussier-Price, Mathieu; Gagnon, David; Lafrance-Vanasse, Julien; Mascle, Xavier; Arseneault, Genevieve; Legault, Pascale; Archambault, Jacques; Omichinski, James G

    2013-11-05

    Like other acidic transactivation domains (TAD), the minimal TAD from the erythroid-specific transcription factor EKLF (EKLFTAD) has been shown to contribute both to its transcriptional activity as well as to its ubiquitin(UBI)-mediated degradation. In this article, we examine the activation-degradation role of the acidic TAD of EKLF and demonstrate that the first 40 residues (EKLFTAD1) within this region form a noncovalent interaction with UBI. Nuclear magnetic resonance (NMR) structural studies of an EKLFTAD1-UBI complex show that EKLFTAD1 adopts a 14-residue α helix that forms the recognition interface with UBI in a similar manner as the UBI-interacting helix of Rabex5. We also identify a similar interaction between UBI and the activation-degradation region of SREBP1a, but not with the activation-degradation regions of p53, GAL4, and VP16. These results suggest that select activation-degradation regions like the ones found in EKLF and SREBP1a function in part through their ability to form noncovalent interactions with UBI. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Dienogest is a selective progesterone receptor agonist in transactivation analysis with potent oral endometrial activity due to its efficient pharmacokinetic profile.

    Science.gov (United States)

    Sasagawa, Shinichi; Shimizu, Yutaka; Kami, Hideaki; Takeuchi, Takashi; Mita, Shizuka; Imada, Kazunori; Kato, Shigeaki; Mizuguchi, Kiyoshi

    2008-02-01

    Dienogest was introduced as an oral progestin. Yet its strong oral potency on endometrial activity is not clearly explained. To circumvent this situation, steroid hormone receptor profiling using transactivation assay and endometrial activity test in rabbits were carried out with determination of plasma drug concentration. Agonistic/antagonistic activity on human progesterone receptor (PR), androgen receptor (AR), glucocorticoid receptor (GR), mineralocorticoid receptor (MR), estrogen receptor alpha (ERalpha), or estrogen receptor beta (ERbeta) were determined. Dienogest activate PR (EC50=3.4 or 10.5 nmol/l) with antagonistic activity on AR (EC50=420.6 or 775.0 nmol/l) but not agonistic nor antagonistic action on GR, MR (3000 nmol/l). Dienogest activate neither ERalpha nor ERbeta (3000 nmol/l). Progesterone activated PR with antagonistic activity on AR and on MR. Dydrogesterone showed a similar profile to progesterone. Norethisterone activated PR, AR, and ERalpha. Medroxyprogesterone acetate activated PR, AR, and GR. Danazol activated PR and AR. Collectively, dienogest has a good specificity to PR compared with the other drugs. By oral treatment, dienogest showed the strongest endometrial activity (ED50=0.0042 mg/kg) in McPhail test among other progestins (ED50 values for MPA, DYG, NES were 0.074, 1.9, >0.05 mg/kg, respectively). Dienogest showed higher plasma concentrations than those of the other progestins with higher doses. The estimated plasma concentration of dienogest at ED50 (3.66 nmol/l) was close to its EC50 value to activate PR. Thus, the stronger oral activity of dienogest could be explained simply by its in vitro potency on PR and its oral pharmacokinetic profile.

  4. EGFR transactivation: mechanisms, pathophysiology and potential therapies in cardiovascular system

    Science.gov (United States)

    Forrester, Steven J.; Kawai, Tatsuo; Elliott, Katherine J.; O’Brien, Shannon; Thomas, Walter; Harris, Raymond C.; Eguchi, Satoru

    2017-01-01

    Accumulating studies suggest that the epidermal growth factor receptor (EGFR) activation is associated with the physiology and pathophysiology of the cardiovascular system, and inhibition of EGFR activity is emerging as a potential therapeutic strategy to treat diseases, including hypertension, cardiac hypertrophy, renal fibrosis and abdominal aortic aneurysm. The capacity of G protein-coupled receptor (GPCR) agonists, such as angiotensin II (AngII), to promote EGFR signaling is well described – a process termed EGFR “transactivation” – yet delineating the molecular processes and functional relevance of this crosstalk has been challenging. Moreover, these critical findings are dispersed among many different fields. The aim of our review is to highlight the recent advancement of the signaling cascades and downstream consequences of EGFR transactivation within the cardiovascular renal system in vitro and in vivo. We will also focus on linking EGFR transactivation to animal models of the disease as well as the potential therapeutic applications. PMID:26566153

  5. Transcription factor Nrf1 is topologically repartitioned across membranes to enable target gene transactivation through its acidic glucose-responsive domains.

    Science.gov (United States)

    Zhang, Yiguo; Ren, Yonggang; Li, Shaojun; Hayes, John D

    2014-01-01

    The membrane-bound Nrf1 transcription factor regulates critical homeostatic and developmental genes. The conserved N-terminal homology box 1 (NHB1) sequence in Nrf1 targets the cap'n'collar (CNC) basic basic-region leucine zipper (bZIP) factor to the endoplasmic reticulum (ER), but it is unknown how its activity is controlled topologically within membranes. Herein, we report a hitherto unknown mechanism by which the transactivation activity of Nrf1 is controlled through its membrane-topology. Thus after Nrf1 is anchored within ER membranes, its acidic transactivation domains (TADs), including the Asn/Ser/Thr-rich (NST) glycodomain situated between acidic domain 1 (AD1) and AD2, are transiently translocated into the lumen of the ER, where NST is glycosylated in the presence of glucose to yield an inactive 120-kDa Nrf1 glycoprotein. Subsequently, portions of the TADs partially repartition across membranes into the cyto/nucleoplasmic compartments, whereupon an active 95-kDa form of Nrf1 accumulates, a process that is more obvious in glucose-deprived cells and may involve deglycosylation. The repartitioning of Nrf1 out of membranes is monitored within this protein by its acidic-hydrophobic amphipathic glucose-responsive domains, particularly the Neh5L subdomain within AD1. Therefore, the membrane-topological organization of Nrf1 dictates its post-translational modifications (i.e. glycosylation, the putative deglycosylation and selective proteolysis), which together control its ability to transactivate target genes.

  6. E1A-dependent trans-activation of the human MYC promoter is mediated by the E2F factor

    International Nuclear Information System (INIS)

    Hiebert, S.W.; Lipp, M.; Nevins, J.R.

    1989-01-01

    E2F is a cellular transcription factor that binds to two sites in the adenovirus E2 promoter. Previous experiments have implicated E2F in the E1A-dependent transactivation of the E2 gene since levels of active E2F increase markedly during adenovirus infection in parallel with the increase in E2 transcription, and an E2F binding site can confer E1A inducibility to a heterologous promoter. Here the authors show that E2F binds to two sequence elements within the P2 promoter of the human MYC gene which are within a region that is critical for promoter activity. The MYC promoter can be trans-activated in an E1A-dependent manner and site-directed mutagenesis demonstrates that these E2F elements are essential for trans-activation. Finally, they also find that adenovirus infection of quiescent cells results in a stimulation of the endogenous MYC gene. They conclude that the activation of the E2F factor, which is likely responsible for the activation of viral E2 transcription, is also responsible for the E1A-dependent induction of MYC transcription

  7. Cardiac GPCR-Mediated EGFR Transactivation: Impact and Therapeutic Implications.

    Science.gov (United States)

    Grisanti, Laurel A; Guo, Shuchi; Tilley, Douglas G

    2017-07-01

    G protein-coupled receptors (GPCRs) remain primary therapeutic targets for numerous cardiovascular disorders, including heart failure (HF), because of their influence on cardiac remodeling in response to elevated neurohormone signaling. GPCR blockers have proven to be beneficial in the treatment of HF by reducing chronic G protein activation and cardiac remodeling, thereby extending the lifespan of patients with HF. Unfortunately, this effect does not persist indefinitely, thus next-generation therapeutics aim to selectively block harmful GPCR-mediated pathways while simultaneously promoting beneficial signaling. Transactivation of epidermal growth factor receptor (EGFR) has been shown to be mediated by an expanding repertoire of GPCRs in the heart, and promotes cardiomyocyte survival, thus may offer a new avenue of HF therapeutics. However, GPCR-dependent EGFR transactivation has also been shown to regulate cardiac hypertrophy and fibrosis by different GPCRs and through distinct molecular mechanisms. Here, we discuss the mechanisms and impact of GPCR-mediated EGFR transactivation in the heart, focusing on angiotensin II, urotensin II, and β-adrenergic receptor systems, and highlight areas of research that will help us to determine whether this pathway can be engaged as future therapeutic strategy.

  8. Non-transactivational, dual pathways for LPA-induced Erk1/2 activation in primary cultures of brown pre-adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Holmstroem, Therese E.; Mattsson, Charlotte L.; Wang, Yanling; Iakovleva, Irina; Petrovic, Natasa [Department of Physiology, The Wenner-Gren Institute, Stockholm University, SE-106 91 Stockholm (Sweden); Nedergaard, Jan, E-mail: jan@metabol.su.se [Department of Physiology, The Wenner-Gren Institute, Stockholm University, SE-106 91 Stockholm (Sweden)

    2010-10-01

    In many cell types, G-protein-coupled receptor (GPCR)-induced Erk1/2 MAP kinase activation is mediated via receptor tyrosine kinase (RTK) transactivation, in particular via the epidermal growth factor (EGF) receptor. Lysophosphatidic acid (LPA), acting via GPCRs, is a mitogen and MAP kinase activator in many systems, and LPA can regulate adipocyte proliferation. The mechanism by which LPA activates the Erk1/2 MAP kinase is generally accepted to be via EGF receptor transactivation. In primary cultures of brown pre-adipocytes, EGF can induce Erk1/2 activation, which is obligatory and determinant for EGF-induced proliferation of these cells. Therefore, we have here examined whether LPA, via EGF transactivation, can activate Erk1/2 in brown pre-adipocytes. We found that LPA could induce Erk1/2 activation. However, the LPA-induced Erk1/2 activation was independent of transactivation of EGF receptors (or PDGF receptors) in these cells (whereas in transformed HIB-1B brown adipocytes, the LPA-induced Erk1/2 activation indeed proceeded via EGF receptor transactivation). In the brown pre-adipocytes, LPA instead induced Erk1/2 activation via two distinct non-transactivational pathways, one G{sub i}-protein dependent, involving PKC and Src activation, the other, a PTX-insensitive pathway, involving PI3K (but not Akt) activation. Earlier studies showing LPA-induced Erk1/2 activation being fully dependent on RTK transactivation have all been performed in cell lines and transfected cells. The present study implies that in non-transformed systems, RTK transactivation may not be involved in the mediation of GPCR-induced Erk1/2 MAP kinase activation.

  9. Transactivation of the TIEG1 confers growth inhibition of transforming growth factor-β-susceptible hepatocellular carcinoma cells

    Science.gov (United States)

    Jiang, Lei; Lai, Yiu-Kay; Zhang, Jin-Fang; Chan, Chu-Yan; Lu, Gang; Lin, Marie CM; He, Ming-Liang; Li, Ji-Cheng; Kung, Hsiang-Fu

    2012-01-01

    AIM: To investigate the role of transforming growth factor (TGF)-β-inducible early gene 1 (TIEG1) in TGF-β-induced growth inhibition in hepatocellular carcinoma (HCC) cells. METHODS: Human hepatocyte and HCC cell lines with varied susceptibilities to TGF-β1 were tested by methylthiazoletetrazolium (MTT) assay. The expression changes of Smad2, Smad3, Smad4, Smad7, TIEG1 and TIEG2 gene following treatment with TGF-β1 in a TGF-β-sensitive hepatocyte cell line (MIHA), a TGF-β-sensitive hepatoma cell line (Hep3B) and two TGF-β-insensitive hepatoma cell lines (HepG2 and Bel7404) were examined. SiRNA targeting TIEG1 was transfected into Hep3B cells and the sensitivity of cells to TGF-β1 was examined. Overexpression of TIEG1 was induced by lentiviral-mediated transduction in TGF-β1-resistant hepatoma cell lines (Bel7404 and HepG2). MTT assay and 4’,6-Diamidino-2-phenylindole staining were used to identify cell viability and apoptosis, respectively. The expression level of stathmin was measured by reverse transcriptase polymerase chain reaction and Western-blotting analysis, and stathmin promoter activity by TIEG1 was monitored by a luciferase reporter gene system. RESULTS: TIEG1 was significantly upregulated by TGF-β1 in the TGF-β1-sensitive HCC cell line, Hep3B, but not in the resistant cell lines. The suppression of TIEG1 by siRNAs decreased the sensitivity of Hep3B cells to TGF-β1, whereas the overexpression of TIEG1 mediated growth inhibition and apoptosis in TGF-β1-resistant HCC cell lines, which resembled those of TGF-β1-sensitive HCC cells treated with TGF-β1. Our data further suggested that stathmin was a direct target of TIEG1, as stathmin was significantly downregulated by TIEG1 overexpression, and stathmin promoter activity was inhibited by TIEG1 in a dose-dependent manner. CONCLUSION: Our data suggest that transactivation of TIEG1 conferred growth inhibition of TGF-β-susceptible human HCC cells. PMID:22563190

  10. Notch3 Maintains Luminal Phenotype and Suppresses Tumorigenesis and Metastasis of Breast Cancer via Trans-Activating Estrogen Receptor-α.

    Science.gov (United States)

    Dou, Xiao-Wei; Liang, Yuan-Ke; Lin, Hao-Yu; Wei, Xiao-Long; Zhang, Yong-Qu; Bai, Jing-Wen; Chen, Chun-Fa; Chen, Min; Du, Cai-Wen; Li, Yao-Chen; Tian, Jie; Man, Kwan; Zhang, Guo-Jun

    2017-01-01

    The luminal A phenotype is the most common breast cancer subtype and is characterized by estrogen receptor α expression (ERα). Identification of the key regulator that governs the luminal phenotype of breast cancer will clarify the pathogenic mechanism and provide novel therapeutic strategies for this subtype of cancer. ERα signaling pathway sustains the epithelial phenotype and inhibits the epithelial-mesenchymal transition (EMT) of breast cancer. In this study, we demonstrate that Notch3 positively associates with ERα in both breast cancer cell lines and human breast cancer tissues. We found that overexpression of Notch3 intra-cellular domain, a Notch3 active form (N3ICD), in ERα negative breast cancer cells re-activated ERα, while knock-down of Notch3 reduced ERα transcript and proteins, with alteration of down-stream genes, suggesting its ability to regulate ERα. Mechanistically, our results show that Notch3 specifically binds to the CSL binding element of the ERα promoter and activates ERα expression. Moreover, Notch3 suppressed EMT, while suppression of Notch3 promoted EMT in cellular assay. Overexpressing N3ICD in triple-negative breast cancer suppressed tumorigenesis and metastasis in vivo . Conversely, depletion of Notch3 in luminal breast cancer promoted metastasis in vivo . Furthermore, Notch3 transcripts were significantly associated with prolonged relapse-free survival in breast cancer, in particular in ERα positive breast cancer patients. Our observations demonstrate that Notch3 governs the luminal phenotype via trans-activating ERα expression in breast cancer. These findings delineate the role of a Notch3/ERα axis in maintaining the luminal phenotype and inhibiting tumorigenesis and metastasis in breast cancer, providing a novel strategy to re-sensitize ERα negative or low-expressing breast cancers to hormone therapy.

  11. Niacin activates the PI3K/Akt cascade via PKC- and EGFR-transactivation-dependent pathways through hydroxyl-carboxylic acid receptor 2.

    Directory of Open Access Journals (Sweden)

    Huawang Sun

    Full Text Available Niacin has been demonstrated to activate a PI3K/Akt signaling cascade to prevent brain damage after stroke and UV-induced skin damage; however, the underlying molecular mechanisms for HCA2-induced Akt activation remain to be elucidated. Using CHO-K1 cells stably expressing HCA2 and A431 cells, a human epidermoid cell line with high levels of endogenous expression of functional HCA2 receptors, we first demonstrated that niacin induced a robust Akt phosphorylation at both Thr308 and Ser473 in a time-dependent fashion, with a maximal activation at 5 min and a subsequent reduction to baseline by 30 min through HCA2, and that the activation was significantly blocked by pertussis toxin. The HCA2-mediated activation of Akt was also significantly inhibited by the PKC inhibitors GF109203x and Go6983 in both cell lines, by the PDGFR-selective inhibitor tyrphostin A9 in CHO-HCA2 cells and by the MMP inhibitor GM6001 and EGFR-specific inhibitor AG1478 in A431 cells. These results suggest that the PKC pathway and PDGFR/EGFR transactivation pathway play important roles in HCA2-mediated Akt activation. Further investigation indicated that PI3K and the Gβγ subunit were likely to play an essential role in HCA2-induced Akt activation. Moreover, Immunobloting analyses using an antibody that recognizes p70S6K1 phosphorylated at Thr389 showed that niacin evoked p70S6K1 activation via the PI3K/Akt pathway. The results of our study provide new insight into the signaling pathways involved in HCA2 activation.

  12. Src Is a Potential Therapeutic Target in Endocrine-Resistant Breast Cancer Exhibiting Low Estrogen Receptor-Mediated Transactivation.

    Science.gov (United States)

    Guest, Stephanie K; Ribas, Ricardo; Pancholi, Sunil; Nikitorowicz-Buniak, Joanna; Simigdala, Nikiana; Dowsett, Mitch; Johnston, Stephen R; Martin, Lesley-Ann

    2016-01-01

    Despite the effectiveness of endocrine therapies in estrogen receptor positive (ER+) breast cancer, approximately 40% of patients relapse. Previously, we identified the Focal-adhesion kinase canonical pathway as a major contributor of resistance to estrogen deprivation and cellular-sarcoma kinase (c-src) as a dominant gene in this pathway. Dasatinib, a pan-src inhibitor, has recently been used in clinical trials to treat ER+ patients but has shown mixed success. In the following study, using isogenic cell line models, we provide a potential explanation for these findings and suggest a sub-group that may benefit. A panel of isogenic cell lines modelling resistance to aromatase inhibitors (LTED) and tamoxifen (TAMR) were assessed for response to dasatinib ± endocrine therapy. Dasatinib caused a dose-dependent decrease in proliferation in MCF7-TAMR cells and resensitized them to tamoxifen and fulvestrant but not in HCC1428-TAMR. In contrast, in estrogen-deprived conditions, dasatinib increased the proliferation rate of parental-MCF7 cells and had no effect on MCF7-LTED or HCC1428-LTED. Treatment with dasatinib caused a decrease in src-phosphorylation and inhibition of downstream pathways, including AKT and ERK1/2 in all cell lines tested, but only the MCF7-TAMR showed a concomitant decrease in markers of cell cycle progression. Inhibition of src also caused a significant decrease in cell migration in both MCF7-LTED and MCF7-TAMR cells. Finally, we showed that, in MCF7-TAMR cells, in contrast to tamoxifen sensitive cell lines, ER is expressed throughout the cell rather than being restricted to the nucleus and that treatment with dasatinib resulted in nuclear shuttling of ER, which was associated with an increase in ER-mediated transcription. These data suggest that src has differential effects in endocrine-resistant cell lines, particularly in tamoxifen resistant models, with low ER genomic activity, providing further evidence of the importance of patient selection

  13. β-Arrestin Mediates β1-Adrenergic Receptor-Epidermal Growth Factor Receptor Interaction and Downstream Signaling*

    Science.gov (United States)

    Tilley, Douglas G.; Kim, Il-Man; Patel, Priyesh A.; Violin, Jonathan D.; Rockman, Howard A.

    2009-01-01

    β1-Adrenergic receptor (β1AR) stimulation confers cardioprotection via β-arrestin-de pend ent transactivation of epidermal growth factor receptors (EGFRs), however, the precise mechanism for this salutary process is unknown. We tested the hypothesis that the β1AR and EGFR form a complex that differentially directs intracellular signaling pathways. β1AR stimulation and EGF ligand can each induce equivalent EGFR phos pho ryl a tion, internalization, and downstream activation of ERK1/2, but only EGF ligand causes translocation of activated ERK to the nucleus, whereas β1AR-stimulated/EGFR-transactivated ERK is restricted to the cytoplasm. β1AR and EGFR are shown to interact as a receptor complex both in cell culture and endogenously in human heart, an interaction that is selective and undergoes dynamic regulation by ligand stimulation. Although catecholamine stimulation mediates the retention of β1AR-EGFR interaction throughout receptor internalization, direct EGF ligand stimulation initiates the internalization of EGFR alone. Continued interaction of β1AR with EGFR following activation is dependent upon C-terminal tail GRK phos pho ryl a tion sites of the β1AR and recruitment of β-arrestin. These data reveal a new signaling paradigm in which β-arrestin is required for the maintenance of a β1AR-EGFR interaction that can direct cytosolic targeting of ERK in response to catecholamine stimulation. PMID:19509284

  14. Modulation of thyroid hormone receptor transactivation by the early region 1A (E1A-like inhibitor of differentiation 1 (EID1

    Directory of Open Access Journals (Sweden)

    Diana Vargas

    2008-01-01

    Full Text Available Transcriptional activation (TA mediated by the effect of thyroid hormones on target genes requires co-activator proteins such as the early region 1A (E1A associated 300 kDa binding protein (p300 and the cAMP response element binding protein (CREB binding protein (CBP, known as the p300/CBP complex, which acetylate histones 3 and 4 to allow transcriptional machinery access to the target gene promoter. Little is known on the role of p300 in thyroid hormone receptor (TR mediated TA but the E1A-like inhibitor of differentiation 1 (EID1, an inhibitor of p300 histone acetyltransferase (HAT, is a functional homolog of E1A and may inhibit myogenic differentiation factor D (MyoD transcriptional activity and reduces muscle cell differentiation. We evaluated the influence of EID1 on TR-mediated transcriptional activity using transfection and mammalian two-hybrid studies to show that EID1 may partially reduces TA activity of the TR receptor, probably due to p300 blockage since EID1 mutants cannot reduce TR-mediated TA. The EID1 does not affect the function of p160 co-activator proteins (160 kDa proteins of steroid receptor co-activators and is functionally independent of co-repressor proteins or TR binding. Summarizing, EID1 reduces TR-mediated transcriptional activity by blocking p300 and may play an important role in thyroid receptor activity in muscle and other tissues.

  15. Interferon Regulatory Factor 1 Transactivates Expression of Human DNA Polymerase η in Response to Carcinogen N-Methyl-N′-nitro-N-nitrosoguanidine*

    Science.gov (United States)

    Qi, Hongyan; Zhu, Huifang; Lou, Meng; Fan, Yanfeng; Liu, Hong; Shen, Jing; Li, Zhongjie; Lv, Xue; Shan, Jianzhen; Zhu, Lijun; Chin, Y. Eugene; Shao, Jimin

    2012-01-01

    DNA polymerase η (Polη) implements translesion DNA synthesis but has low fidelity in replication. We have previously shown that Polη plays an important role in the genesis of nontargeted mutations at undamaged DNA sites in cells exposed to the carcinogen N-methyl-N′-nitro-N-nitrosoguanidine (MNNG). Here, we report that MNNG-induced Polη expression in an interferon regulatory factor 1 (IRF1)-dependent manner in human cells. Mutagenesis analysis showed that four critical residues (Arg-82, Cys-83, Asn-86, and Ser-87) located in the IRF family conserved DNA binding domain-helix α3 were involved in DNA binding and POLH transactivation by IRF1. Furthermore, Polη up-regulation induced by IRF1 was responsible for the increase of mutation frequency in a SupF shuttle plasmid replicated in the MNNG-exposed cells. Interestingly, IRF1 was acetylated by the histone acetyltransferase CBP in these cells. Lys → Arg substitution revealed that Lys-78 of helix α3 was the major acetylation site, and the IRF1-K78R mutation partially inhibited DNA binding and its transcriptional activity. Thus, we propose that IRF1 activation is responsible for MNNG-induced Polη up-regulation, which contributes to mutagenesis and ultimately carcinogenesis in cells. PMID:22367195

  16. Fibronectin-induced VEGF receptor and calcium channel transactivation stimulate GLUT-1 synthesis and trafficking through PPARγ and TC10 in mouse embryonic stem cells.

    Science.gov (United States)

    Suh, Han Na; Han, Ho Jae

    2013-05-01

    Extracellular matrix (ECM) mediates interactions between integrin and growth factor receptor (GFR) or ion channel. Although this crosstalk promotes integration of the downstream signal pathways and then regulates cellular function, the effect of ECM on glucose transporter (GLUT) in stem cells has not been elucidated. Therefore, we examined the effect of fibronectin on GLUT-1 expression, trafficking, and its related signal pathways in mouse embryonic stem cells (mESCs). Fibronectin increased 2-deoxyglucose (DG) uptake and GLUT-1 protein expression that were blocked by transcription or translation inhibitors. Integrin α5β1-bound fibronectin increased 2-DG uptake through cluster formation with vascular endothelial growth factor receptor (VEGFR) 2, and then activated Ras and PI3K/Akt. In another pathway, integrin α5β1 displayed structural and functional interactions with calcium channels, and stimulated 2-DG uptake through calcium influx and PKC activation. Akt and PKC-induced PPARγ phosphorylation enhanced the decreased expression of PPARγ protein, and subsequently increased GLUT-1 protein synthesis and 2-DG uptake. Fibronectin stimulated TC10 activity and cytoskeleton (F-actin) rearrangement, followed by GLUT-1 trafficking. In conclusion, integrin-bound fibronectin stimulates GLUT-1 synthesis through VEGFR2/Ras/PI3K/Akt and calcium channel/Ca(2+)/PKC, which are merged at PPARγ and GLUT-1 trafficking through TC10 and F-actin. Copyright © 2013 Elsevier B.V. All rights reserved.

  17. Multimodular biosensors reveal a novel platform for activation of G proteins by growth factor receptors.

    Science.gov (United States)

    Midde, Krishna K; Aznar, Nicolas; Laederich, Melanie B; Ma, Gary S; Kunkel, Maya T; Newton, Alexandra C; Ghosh, Pradipta

    2015-03-03

    Environmental cues are transmitted to the interior of the cell via a complex network of signaling hubs. Receptor tyrosine kinases (RTKs) and trimeric G proteins are two such major signaling hubs in eukaryotes. Conventionally, canonical signal transduction via trimeric G proteins is thought to be triggered exclusively by G protein-coupled receptors. Here we used molecular engineering to develop modular fluorescent biosensors that exploit the remarkable specificity of bimolecular recognition, i.e., of both G proteins and RTKs, and reveal the workings of a novel platform for activation of G proteins by RTKs in single living cells. Comprised of the unique modular makeup of guanidine exchange factor Gα-interacting vesicle-associated protein (GIV)/girdin, a guanidine exchange factor that links G proteins to a variety of RTKs, these biosensors provide direct evidence that RTK-GIV-Gαi ternary complexes are formed in living cells and that Gαi is transactivated within minutes after growth factor stimulation at the plasma membrane. Thus, GIV-derived biosensors provide a versatile strategy for visualizing, monitoring, and manipulating the dynamic association of Gαi with RTKs for noncanonical transactivation of G proteins in cells and illuminate a fundamental signaling event regulated by GIV during diverse cellular processes and pathophysiologic states.

  18. Cholinergic Transactivation of the EGFR in HaCaT Keratinocytes Stimulates a Flotillin-1 Dependent MAPK-Mediated Transcriptional Response

    Directory of Open Access Journals (Sweden)

    Sina Kühne

    2015-03-01

    Full Text Available Acetylcholine and its receptors regulate numerous cellular processes in keratinocytes and other non-neuronal cells. Muscarinic acetylcholine receptors are capable of transactivating the epidermal growth factor receptor (EGFR and, downstream thereof, the mitogen-activated protein kinase (MAPK cascade, which in turn regulates transcription of genes involved in cell proliferation and migration. We here show that cholinergic stimulation of human HaCaT keratinocytes results in increased transcription of matrix metalloproteinase MMP-3 as well as several ligands of the epidermal growth factor family. Since both metalloproteinases and the said ligands are involved in the transactivation of the EGFR, this transcriptional upregulation may provide a positive feed-forward loop for EGFR/MAPK activation. We here also show that the cholinergic EGFR and MAPK activation and the upregulation of MMP-3 and EGF-like ligands are dependent on the expression of flotillin-1 which we have previously shown to be a regulator of MAPK signaling.

  19. Cross-talk between constitutive androstane receptor and hypoxia-inducible factor in the regulation of gene expression.

    Science.gov (United States)

    Shizu, Ryota; Shindo, Sawako; Yoshida, Takemi; Numazawa, Satoshi

    2013-05-23

    Hypoxia inducible factor (HIF) and 5'-AMP-activated protein kinase are often activated under similar physiological conditions. Constitutive androstane receptor (CAR) translocates into the nucleus in accordance with 5'-AMP-activated protein kinase and thus confers transactivation. The aim of the present study was to investigate a possible link between CAR and HIFα. Phenobarbital (PB), a typical CAR activator, increased the gene expression of HIF-target genes in the livers of mice, including erythropoietin, heme oxygenase-1 and vascular endothelial growth factor-a. PB induced an accumulation of nuclear HIF-1α and an increase in the HIF-responsive element-mediated transactivation in HepG2 cells. Cobalt chloride, a typical HIF activator, induced the gene expression of CAR-target genes, including cyp2b9 and cyp2b10, an accumulation of nuclear CAR and an increase in the PB-responsive enhancer module-mediated transactivation in the mouse liver. Immunoprecipitation-immunoblot and chromatin immunoprecipitation analyses suggest that CAR binds to the PB-responsive enhancer module with HIF-1α in the liver of untreated mice and that the complex dissociates upon PB treatment. Taken together these results suggest that CAR and HIF-α interact and reciprocally modulate the functions of each other. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  20. Lipid-binding proteins modulate ligand-dependent trans-activation by peroxisome proliferator-activated receptors and localize to the nucleus as well as the cytoplasm

    DEFF Research Database (Denmark)

    Helledie, T; Antonius, M; Sorensen, R V

    2000-01-01

    Peroxisome proliferator-activated receptors (PPARs) are activated by a variety of fatty acids, eicosanoids, and hypolipidemic and insulin-sensitizing drugs. Many of these compounds bind avidly to members of a family of small lipid-binding proteins, the fatty acid-binding proteins (FABPs). Fatty a...

  1. Notch3 Maintains Luminal Phenotype and Suppresses Tumorigenesis and Metastasis of Breast Cancer via Trans-Activating Estrogen Receptor

    NARCIS (Netherlands)

    Dou, Xiao-Wei; Liang, Yuan-Ke; Lin, Hao-Yu; Wei, Xiao-Long; Zhang, Yong-Qu; Bai, Jing-Wen; Chen, Chun-Fa; Chen, Min; Du, Cai-Wen; Li, Yao-Chen; Tian, Jie; Man, Kwan; Zhang, Guo-Jun

    2017-01-01

    The luminal A phenotype is the most common breast cancer subtype and is characterized by estrogen receptor α expression (ERα). Identification of the key regulator that governs the luminal phenotype of breast cancer will clarify the pathogenic mechanism and provide novel therapeutic strategies for

  2. Lipid-binding proteins modulate ligand-dependent trans-activation by peroxisome proliferator-activated receptors and localize to the nucleus as well as the cytoplasm

    DEFF Research Database (Denmark)

    Helledie, T; Antonius, M; Sorensen, R V

    2000-01-01

    Peroxisome proliferator-activated receptors (PPARs) are activated by a variety of fatty acids, eicosanoids, and hypolipidemic and insulin-sensitizing drugs. Many of these compounds bind avidly to members of a family of small lipid-binding proteins, the fatty acid-binding proteins (FABPs). Fatty...

  3. 2,3,7,8-Tetrachlorodibenzo-p-dioxin poly(ADP-ribose) polymerase (TiPARP, ARTD14) is a mono-ADP-ribosyltransferase and repressor of aryl hydrocarbon receptor transactivation.

    Science.gov (United States)

    MacPherson, Laura; Tamblyn, Laura; Rajendra, Sharanya; Bralha, Fernando; McPherson, J Peter; Matthews, Jason

    2013-02-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)-inducible poly(ADP-ribose) polymerase (TiPARP/ARTD14) is a member of the PARP family and is regulated by the aryl hydrocarbon receptor (AHR); however, little is known about TiPARP function. In this study, we examined the catalytic function of TiPARP and determined its role in AHR transactivation. We observed that TiPARP exhibited auto-mono-ADP-ribosyltransferase activity and ribosylated core histones. RNAi-mediated knockdown of TiPARP in T-47D breast cancer and HuH-7 hepatoma cells increased TCDD-dependent cytochrome P450 1A1 (CYP1A1) and CYP1B1 messenger RNA (mRNA) expression levels and recruitment of AHR to both genes. Overexpression of TiPARP reduced AHR-dependent increases in CYP1A1-reporter gene activity, which was restored by overexpression of AHR, but not aryl hydrocarbon receptor nuclear translocator. Deletion and mutagenesis studies showed that TiPARP-mediated inhibition of AHR required the zinc-finger and catalytic domains. TiPARP and AHR co-localized in the nucleus, directly interacted and both were recruited to CYP1A1 in response to TCDD. Overexpression of Tiparp enhanced, whereas RNAi-mediated knockdown of TiPARP reduced TCDD-dependent AHR proteolytic degradation. TCDD-dependent induction of AHR target genes was enhanced in Tiparp(-/-) mouse embryonic fibroblasts compared with wildtype controls. Our findings show that TiPARP is a mono-ADP-ribosyltransferase and a transcriptional repressor of AHR, revealing a novel negative feedback loop in AHR signalling.

  4. Notch3 Maintains Luminal Phenotype and Suppresses Tumorigenesis and Metastasis of Breast Cancer via Trans-Activating Estrogen Receptor

    OpenAIRE

    Dou, Xiao-Wei; Liang, Yuan-Ke; Lin, Hao-Yu; Wei, Xiao-Long; Zhang, Yong-Qu; Bai, Jing-Wen; Chen, Chun-Fa; Chen, Min; Du, Cai-Wen; Li, Yao-Chen; Tian, Jie; Man, Kwan; Zhang, Guo-Jun

    2017-01-01

    The luminal A phenotype is the most common breast cancer subtype and is characterized by estrogen receptor α expression (ERα). Identification of the key regulator that governs the luminal phenotype of breast cancer will clarify the pathogenic mechanism and provide novel therapeutic strategies for this subtype of cancer. ERα signaling pathway sustains the epithelial phenotype and inhibits the epithelial-mesenchymal transition (EMT) of breast cancer. In this study, we demonstrate that Notch3 po...

  5. Cross-talk between the calcium-sensing receptor and the epidermal growth factor receptor in Rat-1 fibroblasts

    International Nuclear Information System (INIS)

    Tomlins, Scott A.; Bolllinger, Nikki; Creim, Jeffrey; Rodland, Karin D.

    2005-01-01

    The calcium-sensing receptor (CaR) is a G-protein-coupled receptor that is activated by extracellular calcium (Ca o 2+ ). Rat-1 fibroblasts have been shown to proliferate and increase ERK activity in response to elevation of [Ca 2+ ] o , and these responses are dependent on functional CaR expression. In this report, we examined the role of cross-talk between the CaR and the epidermal growth factor receptor (EGFR) in mediating these responses in Rat-1 cells. This report shows that AG1478, a specific inhibitor of the EGFR kinase, significantly inhibits the increase in proliferation induced by elevated Ca o 2+ . Furthermore, we show that AG1478 acts downstream or separately from G protein subunit activation of phospholipase C. AG1478 significantly inhibits Ca o 2+ -stimulated ERK phosphorylation and in vitro kinase activity. A similar inhibition of ERK phosphorylation was observed in response to the inhibitor AG494. In addition, treatment with inhibitors of metalloproteases involved in shedding of membrane anchored EGF family ligands substantially inhibited the increase in ERK activation in response to elevated Ca o 2+ . This is consistent with the known expression of TGFα by Rat-1 cells. These results indicate that EGFR transactivation is an important component of the CaR-mediated response to increased Ca o 2+ in Rat-1 fibroblasts and most likely involves CaR-mediated induction of regulated proteolysis and ligand shedding

  6. Catalase reverses tumorigenicity in a malignant cell line by an epidermal growth factor receptor pathway.

    Science.gov (United States)

    Finch, Joanne S; Tome, Margaret E; Kwei, Kevin A; Bowden, G Tim

    2006-03-01

    We have used a keratinocyte in vivo/in vitro cell model to test the hypothesis that hydrogen peroxide acts as a signaling molecule, contributing to proliferation and tumorigenesis. A cell line, 6M90, that produces squamous cell carcinoma (SCC), has high levels of ROS and low levels of catalase. A new cell line, MTOC2, generated from parental 6M90 cells by introduction of a Tet-responsive catalase transgene, effectively expressed higher peroxisomal catalase. Increased catalase expression diminished constitutive ROS and enhanced viability after treatment with hydrogen peroxide. Protein tyrosine phosphatase activity was higher in the MTOC2 cells with high catalase, consistent with detection of a lower level of phosphorylation at tyrosine 1068 of the epidermal growth factor receptor (EGF-R). Transcription of downstream c-fos, AP-1 transactivation and cell proliferation were higher in the low catalase cells. An EGF-R inhibitor, AG1478, blocks the higher AP-1 transactivation and cell proliferation of the low catalase 6M90 cells. Tumorigenesis in SCID mice was greatly diminished in the high catalase cells. Our data suggest that hydrogen peroxide functions as a signaling molecule that can modulate activity of a protein tyrosine phosphatase/(s) resulting in phosphorylation of tryrosine/(s) on the EGF-R. Therefore, catalase acts as a tumor-suppressor gene in part by decreasing EGF-R signaling.

  7. Effects of antiandrogenic progestins, chlormadinone and cyproterone acetate, and the estrogen 17α-ethinylestradiol (EE2), and their mixtures: Transactivation with human and rainbowfish hormone receptors and transcriptional effects in zebrafish (Danio rerio) eleuthero-embryos

    Energy Technology Data Exchange (ETDEWEB)

    Siegenthaler, Patricia Franziska [University of Applied Sciences and Arts Northwestern Switzerland (FHNW), School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz (Switzerland); Bain, Peter [Commonwealth Scientific and Industrial Research Organisation (CSIRO), Land and Water Flagship, PMB2, Glen Osmond, 5064 South Australia (Australia); Riva, Francesco [IRCCS – Istituto di Ricerche Farmacologiche “Mario Negri”, Environmental Biomarkers Unit, Department of Environmental Health Sciences, Via La Masa 19, I-20156 Milan (Italy); Fent, Karl, E-mail: karl.fent@fhnw.ch [University of Applied Sciences and Arts Northwestern Switzerland (FHNW), School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz (Switzerland); Swiss Federal Institute of Technology (ETH Zürich), Institute of Biogeochemistry and Pollution Dynamics, Department of Environmental System Sciences, CH-8092 Zürich (Switzerland)

    2017-01-15

    Highlights: • Agonistic and antagonistic activity of CMA and CPA were assessed in vitro. • CMA and CPA showed different interaction with human and fish receptors. • No progestogenic but antiandrogenic and antiglucocorticoid activity occurred in fish. • CMA and CPA showed transcriptional changes in zebrafish embryos. • Binary mixtures of the progestins with EE2 were assessed in vitro and in vivo. - Abstract: Synthetic progestins act as endocrine disrupters in fish but their risk to the environment is not sufficiently known. Here, we focused on an unexplored antiandrogenic progestin, chlormadinone acetate (CMA), and the antiandrogenic progestin cyproterone acetate (CPA). The aim was to evaluate whether their in vitro interaction with human and rainbowfish (Melanotaenia fluviatilis) sex hormone receptors is similar. Furthermore, we investigated their activity in zebrafish (Danio rerio) eleuthero-embryos. First, we studied agonistic and antagonistic activities of CMA, CPA, and 17α-ethinylestradiol (EE2), in recombinant yeast expressing either the human progesterone (PGR), androgen (AR), or estrogen receptor. The same compounds were also investigated in vitro in a stable transfection cell system expressing rainbowfish nuclear steroid receptors. For human receptors, both progestins exhibited progestogenic, androgenic and antiestrogenic activity with no antiandrogenic or estrogenic activity. In contrast, interactions with rainbowfish receptors showed no progestogenic, but antiandrogenic, antiglucocorticoid, and some antiestrogenic activity. Thus, interaction with and transactivation of human and rainbowfish PGR and AR were distinctly different. Second, we analyzed transcriptional alterations in zebrafish eleuthero‐embryos at 96 and 144 h post fertilization after exposure to CPA, CMA, EE2, and binary mixtures of CMA and CPA with EE2, mimicking the use in oral contraceptives. CMA led to slight down-regulation of the ar transcript, while CPA down-regulated ar

  8. In vitro observations and in silico predictions of xenoestrogen mixture effects in T47D-based receptor transactivation and proliferation assays.

    Science.gov (United States)

    Schlotz, Nina; Kim, Gwang-Jin; Jäger, Stefan; Günther, Stefan; Lamy, Evelyn

    2017-12-01

    Within endocrine disruptor research, evaluation and interpretation of mixture effects and the predictive value for downstream responses still warrant more in-depth investigations. We used an estrogen receptor (ER)-mediated reporter gene assay (ER-CALUX®) and a cell proliferation assay (WST-1 assay), both based on the T47D breast cancer cell line, to test mixtures of heterogeneous xenoestrogens. Observed concentration-response curves were compared to those predicted by the concepts of concentration addition (CA), generalized concentration addition (GCA), and a novel full logistic model (FLM). CA performed better regarding mixture potency (EC50 values), whereas GCA was superior in predicting mixture efficacy (maximal response). In comparison, FLM proved to be highly suitable for in silico mixture effect prediction, combining advantages of both CA and GCA. The inter-assay comparison revealed that ER activation is not necessarily predictive for induction of cell proliferation. The results support the use of models like CA, GCA, or FLM in mixture effect evaluation. However, we conclude that reliable estimations regarding the disruptive potential of mixtures of endocrine active substances require an integrative approach considering more than one assay/endpoint to avoid misinterpretations. The formazan-based WST-1 proliferation assay might be a possible alternative to commonly used proliferation assays in endocrine disrupter research. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. miR-16 targets transcriptional corepressor SMRT and modulates NF-kappaB-regulated transactivation of interleukin-8 gene.

    Directory of Open Access Journals (Sweden)

    Rui Zhou

    Full Text Available The signaling pathways associated with the Toll-like receptors (TLRs and nuclear factor-kappaB (NF-κB are essential to pro-inflammatory cytokine and chemokine expression, as well as initiating innate epithelial immune responses. The TLR/NF-κB signaling pathways must be stringently controlled through an intricate network of positive and negative regulatory elements. MicroRNAs (miRNAs are non-coding small RNAs that regulate the stability and/or translation of protein-coding mRNAs. Herein we report that miR-16 promotes NF-κB-regulated transactivation of the IL-8 gene by suppression of the silencing mediator for retinoid and thyroid hormone receptor (SMRT. LPS stimulation activated miR-16 gene transcription in human monocytes (U937 and biliary epithelial cells (H69 through MAPK-dependent mechanisms. Transfection of cells with the miR-16 precursor promoted LPS-induced production of IL-8, IL-6, and IL-1α, without a significant effect on their RNA stability. Instead, an increase in NF-κB-regulated transactivation of the IL-8 gene was confirmed in cells following transfection of miR-16 precursor. Importantly, miR-16 targeted the 3'-untranslated region of SMRT and caused translational suppression of SMRT. LPS decreased SMRT expression via upregulation of miR-16. Moreover, functional manipulation of SMRT altered NF-κB-regulated transactivation of LPS-induced IL-8 expression. These data suggest that miR-16 targets SMRT and modulates NF-κB-regulated transactivation of the IL-8 gene.

  10. TGFβ induces proHB-EGF shedding and EGFR transactivation through ADAM activation in gastric cancer cells

    International Nuclear Information System (INIS)

    Ebi, Masahide; Kataoka, Hiromi; Shimura, Takaya; Kubota, Eiji; Hirata, Yoshikazu; Mizushima, Takashi; Mizoshita, Tsutomu; Tanaka, Mamoru; Mabuchi, Motoshi; Tsukamoto, Hironobu; Tanida, Satoshi; Kamiya, Takeshi; Higashiyama, Shigeki; Joh, Takashi

    2010-01-01

    Research highlights: → TGFβ induces EGFR transactivation through proHB-EGF shedding by activated ADAM members in gastric cancer cells. → TGFβ induces nuclear translocation of HB-EGF-CTF cleaved by ADAM members. → TGFβ enhances cell growth by EGFR transactivation and HB-EGF-CTF nuclear translocation and ADAM inhibitors block these effects. → Silencing of ADAM17 also blocks EGFR transactivation, HB-EGF-CTF nuclear translocation and cancer cell growth by TGFβ. → ADAM17 may play a crucial role in this TGFβ-HB-EGF signal transduction. -- Abstract: Background and aims: Transforming growth factor-beta (TGFβ) is known to potently inhibit cell growth. Loss of responsiveness to TGFβ inhibition on cell growth is a hallmark of many types of cancer, yet its mechanism is not fully understood. Membrane-anchored heparin-binding EGF-like growth factor (proHB-EGF) ectodomain is cleaved by a disintegrin and metalloproteinase (ADAM) members and is implicated in epidermal growth factor receptor (EGFR) transactivation. Recently, nuclear translocation of the C-terminal fragment (CTF) of pro-HB-EGF was found to induce cell growth. We investigated the association between TGFβ and HB-EGF signal transduction via ADAM activation. Materials and methods: The CCK-8 assay in two gastric cancer cell lines was used to determine the effect for cell growth by TGFβ. The effect of two ADAM inhibitors was also evaluated. Induction of EGFR phosphorylation by TGFβ was analyzed and the effect of the ADAM inhibitors was also examined. Nuclear translocation of HB-EGF-CTF by shedding through ADAM activated by TGFβ was also analyzed. EGFR transactivation, HB-EGF-CTF nuclear translocation, and cell growth were examined under the condition of ADAM17 knockdown. Result: TGFβ-induced EGFR phosphorylation of which ADAM inhibitors were able to inhibit. TGFβ induced shedding of proHB-EGF allowing HB-EGF-CTF to translocate to the nucleus. ADAM inhibitors blocked this nuclear translocation. TGF

  11. TGF{beta} induces proHB-EGF shedding and EGFR transactivation through ADAM activation in gastric cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Ebi, Masahide [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, Nagoya (Japan); Kataoka, Hiromi, E-mail: hkataoka@med.nagoya-cu.ac.jp [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, Nagoya (Japan); Shimura, Takaya; Kubota, Eiji; Hirata, Yoshikazu; Mizushima, Takashi; Mizoshita, Tsutomu; Tanaka, Mamoru; Mabuchi, Motoshi; Tsukamoto, Hironobu; Tanida, Satoshi; Kamiya, Takeshi [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, Nagoya (Japan); Higashiyama, Shigeki [Department of Biochemistry and Molecular Genetics, Ehime University Graduate School of Medicine, Ehime (Japan); Joh, Takashi [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, Nagoya (Japan)

    2010-11-19

    Research highlights: {yields} TGF{beta} induces EGFR transactivation through proHB-EGF shedding by activated ADAM members in gastric cancer cells. {yields} TGF{beta} induces nuclear translocation of HB-EGF-CTF cleaved by ADAM members. {yields} TGF{beta} enhances cell growth by EGFR transactivation and HB-EGF-CTF nuclear translocation and ADAM inhibitors block these effects. {yields} Silencing of ADAM17 also blocks EGFR transactivation, HB-EGF-CTF nuclear translocation and cancer cell growth by TGF{beta}. {yields} ADAM17 may play a crucial role in this TGF{beta}-HB-EGF signal transduction. -- Abstract: Background and aims: Transforming growth factor-beta (TGF{beta}) is known to potently inhibit cell growth. Loss of responsiveness to TGF{beta} inhibition on cell growth is a hallmark of many types of cancer, yet its mechanism is not fully understood. Membrane-anchored heparin-binding EGF-like growth factor (proHB-EGF) ectodomain is cleaved by a disintegrin and metalloproteinase (ADAM) members and is implicated in epidermal growth factor receptor (EGFR) transactivation. Recently, nuclear translocation of the C-terminal fragment (CTF) of pro-HB-EGF was found to induce cell growth. We investigated the association between TGF{beta} and HB-EGF signal transduction via ADAM activation. Materials and methods: The CCK-8 assay in two gastric cancer cell lines was used to determine the effect for cell growth by TGF{beta}. The effect of two ADAM inhibitors was also evaluated. Induction of EGFR phosphorylation by TGF{beta} was analyzed and the effect of the ADAM inhibitors was also examined. Nuclear translocation of HB-EGF-CTF by shedding through ADAM activated by TGF{beta} was also analyzed. EGFR transactivation, HB-EGF-CTF nuclear translocation, and cell growth were examined under the condition of ADAM17 knockdown. Result: TGF{beta}-induced EGFR phosphorylation of which ADAM inhibitors were able to inhibit. TGF{beta} induced shedding of proHB-EGF allowing HB-EGF-CTF to

  12. Quantitative Analysis of NF-κB Transactivation Specificity Using a Yeast-Based Functional Assay.

    Directory of Open Access Journals (Sweden)

    Vasundhara Sharma

    Full Text Available The NF-κB transcription factor family plays a central role in innate immunity and inflammation processes and is frequently dysregulated in cancer. We developed an NF-κB functional assay in yeast to investigate the following issues: transactivation specificity of NF-κB proteins acting as homodimers or heterodimers; correlation between transactivation capacity and in vitro DNA binding measurements; impact of co-expressed interacting proteins or of small molecule inhibitors on NF-κB-dependent transactivation. Full-length p65 and p50 cDNAs were cloned into centromeric expression vectors under inducible GAL1 promoter in order to vary their expression levels. Since p50 lacks a transactivation domain (TAD, a chimeric construct containing the TAD derived from p65 was also generated (p50TAD to address its binding and transactivation potential. The p50TAD and p65 had distinct transactivation specificities towards seventeen different κB response elements (κB-REs where single nucleotide changes could greatly impact transactivation. For four κB-REs, results in yeast were predictive of transactivation potential measured in the human MCF7 cell lines treated with the NF-κB activator TNFα. Transactivation results in yeast correlated only partially with in vitro measured DNA binding affinities, suggesting that features other than strength of interaction with naked DNA affect transactivation, although factors such as chromatin context are kept constant in our isogenic yeast assay. The small molecules BAY11-7082 and ethyl-pyruvate as well as expressed IkBα protein acted as NF-κB inhibitors in yeast, more strongly towards p65. Thus, the yeast-based system can recapitulate NF-κB features found in human cells, thereby providing opportunities to address various NF-κB functions, interactions and chemical modulators.

  13. Multiple phosphorylation events control chicken ovalbumin upstream promoter transcription factor I orphan nuclear receptor activity.

    Science.gov (United States)

    Gay, Frédérique; Baráth, Peter; Desbois-Le Péron, Christine; Métivier, Raphaël; Le Guével, Rémy; Birse, Darcy; Salbert, Gilles

    2002-06-01

    Chicken ovalbumin upstream promoter transcription factor I (COUP-TFI) is an orphan member of the nuclear hormone receptor superfamily that comprises key regulators of many biological functions, such as embryonic development, metabolism, homeostasis, and reproduction. Although COUP-TFI can both actively silence gene transcription and antagonize the functions of various other nuclear receptors, the COUP-TFI orphan receptor also acts as a transcriptional activator in certain contexts. Moreover, COUP-TFI has recently been shown to serve as an accessory factor for some ligand-bound nuclear receptors, suggesting that it may modulate, both negatively and positively, a wide range of hormonal responses. In the absence of any identified cognate ligand, the mechanisms involved in the regulation of COUP-TFI activity remain unclear. The elucidation of several putative phosphorylation sites for MAPKs, PKC, and casein kinase II within the sequence of this orphan receptor led us to investigate phosphorylation events regulating the various COUP-TFI functions. After showing that COUP-TFI is phosphorylated in vivo, we provide evidence that in vivo inhibition of either MAPK or PKC signaling pathway leads to a specific and pronounced decrease in COUP-TFI-dependent transcriptional activation of the vitronectin gene promoter. Focusing on the molecular mechanisms underlying the MAPK- and PKC-mediated regulation of COUP-TFI activity, we show that COUP-TFI can be directly targeted by PKC and MAPK. These phosphorylation events differentially modulate COUP-TFI functions: PKC-mediated phosphorylation enhances COUP-TFI affinity for DNA and MAPK-mediated phosphorylation positively regulates the transactivation function of COUP-TFI, possibly through enhancing specific coactivator recruitment. These data provide evidence that COUP-TFI is likely to integrate distinct signaling pathways and raise the possibility that multiple extracellular signals influence biological processes controlled by COUP-TFI.

  14. Cooperation of tyrosine kinase receptor TrkB and epidermal growth factor receptor signaling enhances migration and dispersal of lung tumor cells.

    Directory of Open Access Journals (Sweden)

    Rudolf Götz

    Full Text Available TrkB mediates the effects of brain-derived neurotrophic factor (BDNF in neuronal and nonnneuronal cells. Based on recent reports that TrkB can also be transactivated through epidermal growth-factor receptor (EGFR signaling and thus regulates migration of early neurons, we investigated the role of TrkB in migration of lung tumor cells. Early metastasis remains a major challenge in the clinical management of non-small cell lung cancer (NSCLC. TrkB receptor signaling is associated with metastasis and poor patient prognosis in NSCLC. Expression of this receptor in A549 cells and in another adenocarcinoma cell line, NCI-H441, promoted enhanced migratory capacity in wound healing assays in the presence of the TrkB ligand BDNF. Furthermore, TrkB expression in A549 cells potentiated the stimulatory effect of EGF in wound healing and in Boyden chamber migration experiments. Consistent with a potential loss of cell polarity upon TrkB expression, cell dispersal and de-clustering was induced in A549 cells independently of exogeneous BDNF. Morphological transformation involved extensive cytoskeletal changes, reduced E-cadherin expression and suppression of E-cadherin expression on the cell surface in TrkB expressing tumor cells. This function depended on MEK and Akt kinase activity but was independent of Src. These data indicate that TrkB expression in lung adenoma cells is an early step in tumor cell dissemination, and thus could represent a target for therapy development.

  15. The nuclear cofactor RAC3/AIB1/SRC-3 enhances Nrf2 signaling by interacting with transactivation domains.

    Science.gov (United States)

    Kim, J-H; Yu, S; Chen, J D; Kong, A N

    2013-01-24

    Nuclear factor erythroid 2-related factor 2 (Nrf2, NM 006164, 605 AA) is essential for the antioxidant responsive element (ARE)-mediated expression of a group of detoxifying antioxidant genes that detoxify carcinogens and protect against oxidative stress. Several proteins have been identified as Nrf2-interacting molecules. In this study, we found that the overexpression of receptor-associated coactivator 3 (RAC3)/AIB-1/steroid receptor coactivator-3, a nuclear coregulator and oncogene frequently amplified in human breast cancers, induced heme oxygenase-1 (HO-1) through Nrf2 transactivation in HeLa cells. Next, we determined the interaction between RAC3 and Nrf2 proteins using a co-immunoprecipitation assay and fluorescence resonance energy transfer analysis. The results showed that RAC3 bound directly to the Nrf2 protein in the nucleus. Subsequently, we identified the interacting domains of Nrf2 and RAC3 using a glutathione S-transferase pull-down assay. The results showed that both the N-terminal RAC3-pasB and C-terminal RAC3-R3B3 domains were tightly bound to the Neh4 and Neh5 transactivation domains. Furthermore, chromatin immunoprecipitation showed that RAC3 bound tightly to the ARE enhancer region of the HO-1 promoter via Nrf2 binding. These data suggest that Nrf2 activation is modulated and directly controlled through interactions with the RAC3 protein in HeLa cells.

  16. Crosstalk between the peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) and the vitamin D receptor (VDR) in human breast cancer cells: PPAR{gamma} binds to VDR and inhibits 1{alpha},25-dihydroxyvitamin D{sub 3} mediated transactivation

    Energy Technology Data Exchange (ETDEWEB)

    Alimirah, Fatouma; Peng, Xinjian; Yuan, Liang; Mehta, Rajeshwari R. [Cancer Biology Division, IIT Research Institute, 10 West 35th Street, Chicago, IL 60616 (United States); Knethen, Andreas von [Institute of Biochemistry, Johann Wolfgang Goethe University, Frankfurt (Germany); Choubey, Divaker [Department of Environmental Health, University of Cincinnati, 3223 Eden Avenue, P.O. Box 670056, Cincinnati, OH 45267 (United States); Mehta, Rajendra G., E-mail: rmehta@iitri.org [Cancer Biology Division, IIT Research Institute, 10 West 35th Street, Chicago, IL 60616 (United States)

    2012-11-15

    Heterodimerization and cross-talk between nuclear hormone receptors often occurs. For example, estrogen receptor alpha (ER{alpha}) physically binds to peroxisome proliferator-activated receptor gamma (PPAR{gamma}) and inhibits its transcriptional activity. The interaction between PPAR{gamma} and the vitamin D receptor (VDR) however, is unknown. Here, we elucidate the molecular mechanisms linking PPAR{gamma} and VDR signaling, and for the first time we show that PPAR{gamma} physically associates with VDR in human breast cancer cells. We found that overexpression of PPAR{gamma} decreased 1{alpha},25-dihydroxyvitamin D{sub 3} (1,25D{sub 3}) mediated transcriptional activity of the vitamin D target gene, CYP24A1, by 49% and the activity of VDRE-luc, a vitamin D responsive reporter, by 75% in T47D human breast cancer cells. Deletion mutation experiments illustrated that helices 1 and 4 of PPAR{gamma}'s hinge and ligand binding domains, respectively, governed this suppressive function. Additionally, abrogation of PPAR{gamma}'s AF2 domain attenuated its repressive action on 1,25D{sub 3} transactivation, indicating that this domain is integral in inhibiting VDR signaling. PPAR{gamma} was also found to compete with VDR for their binding partner retinoid X receptor alpha (RXR{alpha}). Overexpression of RXR{alpha} blocked PPAR{gamma}'s suppressive effect on 1,25D{sub 3} action, enhancing VDR signaling. In conclusion, these observations uncover molecular mechanisms connecting the PPAR{gamma} and VDR pathways. -- Highlights: PPAR{gamma}'s role on 1{alpha},25-dihydroxyvitamin D{sub 3} transcriptional activity is examined. Black-Right-Pointing-Pointer PPAR{gamma} physically binds to VDR and inhibits 1{alpha},25-dihydroxyvitamin D{sub 3} action. Black-Right-Pointing-Pointer PPAR{gamma}'s hinge and ligand binding domains are important for this inhibitory effect. Black-Right-Pointing-Pointer PPAR{gamma} competes with VDR for the availability of their binding

  17. Epidermal growth factor receptor signaling in tissue

    Energy Technology Data Exchange (ETDEWEB)

    Shvartsman, Stanislav; Wiley, H. S.; Lauffenburger, Douglas A.

    2004-08-01

    Abstract: A peptide purified from the salivary gland of a mouse was shown few years ago to accelerate incisor eruption and eyelid opening in newborn mice, and was named epidermal growth factor (EGF). The members of this family of peptide growth factors had been identified in numerous physiological and pathological contexts. EGF binds to a cell surface EGF receptor, which induces a biochemical modification (phosphorylation) of the receptor's cytoplasmic tail. There is a growing consensus in the research community that, in addition to cellular and molecular studies, the dynamics of the EGFR network and its operation must be examined in tissues. A key challenge is to integrate the existing molecular and cellular information into a system-level description of the EGFR network at the tissue and organism level. In this paper, the two examples of EGFR signaling in tissues are described, and the recent efforts to model EGFR autocrine loops, which is a predominant mode of EGFR activation in vivo, are summarized.

  18. The glucocorticoid RU24858 does not distinguish between transrepression and transactivation in primary human eosinophils

    Directory of Open Access Journals (Sweden)

    Zhang Xianzhi

    2006-07-01

    Full Text Available Abstract Background Glucocorticoids are used to treat chronic inflammatory diseases such as asthma. Induction of eosinophil apoptosis is considered to be one of the main mechanisms behind the anti-asthmatic effect of glucocorticoids. Glucocorticoid binding to its receptor (GR can have a dual effect on gene transcription. Activated GR can activate transcription (transactivation, or by interacting with other transcription factors such as NF-κB suppress transcription (transrepression. RU24858 has been reported to transrepress but to have little or no transactivation capability in other cell types. The dissociated properties of RU24858 have not been previously studied in non-malignant human cells. As the eosinophils have a very short lifetime and many of the modern molecular biological methods cannot be used, a "dissociated steroid" would be a valuable tool to evaluate the mechanism of action of glucocorticoids in human eosinophils. The aim of this study was to elucidate the ability of RU24858 to activate and repress gene expression in human eosinophils in order to see whether it is a dissociated steroid in human eosinophils. Methods Human peripheral blood eosinophils were isolated under sterile conditions and cultured in the presence and/or absence RU24858. For comparison, dexamethasone and mometasone were used. We measured chemokine receptor-4 (CXCR4 and Annexin 1 expression by flow cytometry and cytokine production by ELISA. Apoptosis was measured by DNA fragmentation and confirmed by morphological analysis. Results RU24858 (1 μM increased CXCR4 and Annexin 1 expression on eosinophils to a similar extent as mometasone (1 μM and dexamethasone (1 μM. Like dexamethasone and mometasone, RU24858 did suppress IL-8 and MCP-1 production in eosinophils. RU24858 also increased spontaneous eosinophil apoptosis to a similar degree as dexamethasone and mometasone, but unlike dexamethasone and mometasone it did not reverse IL-5- or GM

  19. Identification of transactivation-responsive DNA-binding protein 43 (TARDBP43; TDP-43) as a novel factor for TNF-α expression upon lipopolysaccharide stimulation in human monocytes.

    Science.gov (United States)

    Murata, H; Hattori, T; Maeda, H; Takashiba, S; Takigawa, M; Kido, J; Nagata, T

    2015-08-01

    Tumor necrosis factor alpha (TNF-α) is a major cytokine implicated in various inflammatory diseases. The nature of the nuclear factors associated with human TNF-α gene regulation is not well elucidated. We previously identified a novel region located from -550 to -487 in human TNF-α promoter that did not contain the reported binding sites for nuclear factor kappa B (NF-κB) but showed lipopolysaccharide (LPS)-induced transcriptional activity. The purpose of this study is to identify novel factors that bind to the promoter region and regulate TNF-α expression. To identify DNA-binding proteins that bound to the target region of TNF-α promoter, a cDNA library from LPS-stimulated human monocytic cell line THP-1 was screened using a yeast one-hybrid system. Cellular localizations of the DNA-binding protein in the cells were examined by subcellular immunocytochemistry. Nuclear amounts of the protein in LPS-stimulated THP-1 cells were identified by western blot analysis. Expression of mRNA of the protein in the cells was quantified by real-time polymerase chain reaction. Electrophoretic mobility shift assays were performed to confirm the DNA-binding profile. Overexpression of the protein and knockdown of the gene were also performed to investigate the role for TNF-α expression. Several candidates were identified from the cDNA library and transactivation-responsive DNA-binding protein 43 (TARDBP43; TDP-43) was focused on. Western blot analysis revealed that nuclear TDP-43 protein was increased in the LPS-stimulated THP-1 cells. Expression of TDP-43 mRNA was already enhanced before TNF-α induction by LPS. Electrophoretic mobility shift assay analysis showed that nuclear extracts obtained by overexpressing FLAG-tagged TDP-43 bound to the -550 to -487 TNF-α promoter fragments. Overexpression of TDP-43 in THP-1 cells resulted in an increase of TNF-α expression. Knockdown of TDP-43 in THP-1 cells downregulated TNF-α expression. We identified TDP-43 as one of the novel

  20. Genetic dissection of epidermal growth factor receptor signaling during luteinizing hormone-induced oocyte maturation.

    Directory of Open Access Journals (Sweden)

    Minnie Hsieh

    Full Text Available Recent evidence that luteinizing hormone (LH stimulation of ovulatory follicles causes transactivation of the epidermal growth factor receptor (EGFR has provided insights into the mechanisms of ovulation. However, the complete array of signals that promote oocyte reentry into the meiotic cell cycle in the follicle are still incompletely understood. To elucidate the signaling downstream of EGFR involved in oocyte maturation, we have investigated the LH responses in granulosa cells with targeted ablation of EGFR. Oocyte maturation and ovulation is disrupted when EGFR expression is progressively reduced. In granulosa cells from mice with either global or granulosa cell-specific disruption of EGFR signaling, LH-induced phosphorylation of MAPK3/1, p38MAPK, and connexin-43 is impaired. Although the LH-induced decrease in cGMP is EGFR-dependent in wild type follicles, LH still induces a decrease in cGMP in Egfr(delta/f Cyp19-Cre follicles. Thus compensatory mechanisms appear activated in the mutant. Spatial propagation of the LH signal in the follicle also is dependent on the EGF network, and likely is important for the control of signaling to the oocyte. Thus, multiple signals and redundant pathways contribute to regulating oocyte reentry into the cell cycle.

  1. Pattern of hormone receptors and human epidermal growth factor ...

    African Journals Online (AJOL)

    Introduction: Breast cancer is the most common cancer among women globally. With immunohistochemistry (IHC), breast cancer is classified into four groups based on IHC profile of estrogen receptor (ER)/progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2/neu) expression, positive (+) and/or ...

  2. Transcriptional regulation of the human Liver X Receptor α gene by Hepatocyte Nuclear Factor

    Energy Technology Data Exchange (ETDEWEB)

    Theofilatos, Dimitris; Anestis, Aristomenis [University of Crete Medical School and Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology of Hellas, Heraklion, 71003, Crete (Greece); Hashimoto, Koshi [Department of Preemptive Medicine and Metabolism, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-city, Tokyo, 113-8510 (Japan); Kardassis, Dimitris, E-mail: kardasis@imbb.forth.gr [University of Crete Medical School and Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology of Hellas, Heraklion, 71003, Crete (Greece)

    2016-01-15

    Liver X Receptors (LXRs) are sterol-activated transcription factors that play major roles in cellular cholesterol homeostasis, HDL biogenesis and reverse cholesterol transport. The aim of the present study was to investigate the mechanisms that control the expression of the human LXRα gene in hepatic cells. A series of reporter plasmids containing consecutive 5′ deletions of the hLXRα promoter upstream of the luciferase gene were constructed and the activity of each construct was measured in HepG2 cells. This analysis showed that the activity of the human LXRα promoter was significantly reduced by deleting the −111 to −42 region suggesting the presence of positive regulatory elements in this short proximal fragment. Bioinformatics data including motif search and ChIP-Seq revealed the presence of a potential binding motif for Hepatocyte Nuclear Factor 4 α (HNF-4α) in this area. Overexpression of HNF-4α in HEK 293T cells increased the expression of all LXRα promoter constructs except −42/+384. In line, silencing the expression of endogenous HNF-4α in HepG2 cells was associated with reduced LXRα protein levels and reduced activity of the −111/+384 LXRα promoter but not of the −42/+384 promoter. Using ChiP assays in HepG2 cells combined with DNAP assays we mapped the novel HNF-4α specific binding motif (H4-SBM) in the −50 to −40 region of the human LXRα promoter. A triple mutation in this H4-SBM abolished HNF-4α binding and reduced the activity of the promoter to 65% relative to the wild type. Furthermore, the mutant promoter could not be transactivated by HNF-4α. In conclusion, our data indicate that HNF-4α may have a wider role in cell and plasma cholesterol homeostasis by controlling the expression of LXRα in hepatic cells. - Highlights: • The human LXRα promoter contains a HNF-4α specific binding motif in the proximal −50/−40 region. • Mutations in this motif abolished HNF4α binding and transactivation of the h

  3. Transcriptional regulation of the human Liver X Receptor α gene by Hepatocyte Nuclear Factor

    International Nuclear Information System (INIS)

    Theofilatos, Dimitris; Anestis, Aristomenis; Hashimoto, Koshi; Kardassis, Dimitris

    2016-01-01

    Liver X Receptors (LXRs) are sterol-activated transcription factors that play major roles in cellular cholesterol homeostasis, HDL biogenesis and reverse cholesterol transport. The aim of the present study was to investigate the mechanisms that control the expression of the human LXRα gene in hepatic cells. A series of reporter plasmids containing consecutive 5′ deletions of the hLXRα promoter upstream of the luciferase gene were constructed and the activity of each construct was measured in HepG2 cells. This analysis showed that the activity of the human LXRα promoter was significantly reduced by deleting the −111 to −42 region suggesting the presence of positive regulatory elements in this short proximal fragment. Bioinformatics data including motif search and ChIP-Seq revealed the presence of a potential binding motif for Hepatocyte Nuclear Factor 4 α (HNF-4α) in this area. Overexpression of HNF-4α in HEK 293T cells increased the expression of all LXRα promoter constructs except −42/+384. In line, silencing the expression of endogenous HNF-4α in HepG2 cells was associated with reduced LXRα protein levels and reduced activity of the −111/+384 LXRα promoter but not of the −42/+384 promoter. Using ChiP assays in HepG2 cells combined with DNAP assays we mapped the novel HNF-4α specific binding motif (H4-SBM) in the −50 to −40 region of the human LXRα promoter. A triple mutation in this H4-SBM abolished HNF-4α binding and reduced the activity of the promoter to 65% relative to the wild type. Furthermore, the mutant promoter could not be transactivated by HNF-4α. In conclusion, our data indicate that HNF-4α may have a wider role in cell and plasma cholesterol homeostasis by controlling the expression of LXRα in hepatic cells. - Highlights: • The human LXRα promoter contains a HNF-4α specific binding motif in the proximal −50/−40 region. • Mutations in this motif abolished HNF4α binding and transactivation of the h

  4. Molecular analysis of the nerve growth factor receptor

    International Nuclear Information System (INIS)

    Hempstead, B.; Patil, N.; Olson, K.; Chao, M.

    1988-01-01

    An essential molecule in the translocation of information by nerve growth factor (NGF) to responsive cells is the cell-surface receptor for NGF. This paper presents information on the genomic structure of the NGF receptor gene, NGF receptor models, and transfection of NGF receptors. Equilibrium binding of [ 125 I]NGF to cells reveals two distinct affinity states for the NGF receptor. The human NGF receptor gene is a single-copy gene, consisting of six exons that span 23 kb. The receptor gene is capable of being transferred to fibroblast cells from human genomic DNA and expressed at high levels. The constitutive nature of the receptor promoter sequence is a partial explanation of why this tissue-specific gene is expressed efficiently in a variety of nonneuronal cells after genomic gene transfer. The two kinetic forms of the NGF receptor appear to be encoded by the same protein, which is the product of a single gene

  5. Interaction of epidermal growth factor receptors with the cytoskeleton is related to receptor clustering

    NARCIS (Netherlands)

    van Belzen, N.; Spaargaren, M.; Verkleij, A. J.; Boonstra, J.

    1990-01-01

    Recently it has been established that cytoskeleton-associated epidermal growth factor (EGF) receptors are predominantly of the high-affinity class and that EGF induces a recruitment of low-affinity receptors to the cytoskeleton. The nature of this EGF-induced receptor-cytoskeleton interaction,

  6. New factors influencing G protein coupled receptors' system functions

    African Journals Online (AJOL)

    New factors such as the G protein coupled receptor (GPCR) surrounding's chemical environment, cell membrane constituents, the existent gap junction, endogenous receptor affinity status and animal species have been shown to influence the GPCR physiology and variations of those factors can modify the functions of the ...

  7. Apolipoprotein E isoform dependently affects Tat-mediated HIV-1 LTR transactivation.

    Science.gov (United States)

    Khan, Nabab; Datta, Gaurav; Geiger, Jonathan D; Chen, Xuesong

    2018-03-20

    Apolipoprotein E (ApoE) is the major carrier protein that mediates the transport and delivery of cholesterol and other lipids in the brain. Three isoforms of ApoE (ApoE2, ApoE3, ApoE4) exist in humans, and their relative expression levels impact HIV-1 infection, HIV-1/AIDS disease progression, and cognitive decline associated with HIV-1-associated neurocognitive disorder. Because HIV-1 Tat, a viral protein essential for HIV-1 replication, can bind to low-density lipoprotein receptor-related protein 1 (LRP1) that controls ApoE uptake in the brain, we determined the extent to which different isoforms of ApoE affected Tat-mediated HIV-1 LTR transactivation. Using U87MG glioblastoma cells expressing LTR-driven luciferase, we determined the extent to which LRP1 as well as ApoE2, ApoE3, and ApoE4 affected Tat-mediated HIV-1 LTR transactivation. A specific LRP1 antagonist and siRNA knockdown of LRP1 both restricted significantly Tat-mediated LTR transactivation. Of the three ApoEs, ApoE4 was the least potent and effective at preventing HIV-1 Tat internalization and at decreasing Tat-mediated HIV-1 LTR transactivation. Further, Tat-mediated LTR transactivation was attenuated by an ApoE mimetic peptide, and ApoE4-induced restriction of Tat-mediated LTR transactivation was potentiated by an ApoE4 structure modulator that changes ApoE4 into an ApoE3-like phenotype. These findings help explain observed differential effects of ApoEs on HIV-1 infectivity and the prevalence of HAND in people living with HIV-1 infection and suggest that ApoE mimetic peptides and ApoE4 structure modulator might be used as a therapeutic strategy against HIV-1 infection and associated neurocognitive disorders.

  8. Normal epidermal growth factor receptor signaling is dispensable for bone anabolic effects of parathyroid hormone.

    Science.gov (United States)

    Schneider, Marlon R; Dahlhoff, Maik; Andrukhova, Olena; Grill, Jessica; Glösmann, Martin; Schüler, Christiane; Weber, Karin; Wolf, Eckhard; Erben, Reinhold G

    2012-01-01

    Although the bone anabolic properties of intermittent parathyroid hormone (PTH) have long been employed in the treatment of osteoporosis, the molecular mechanisms behind this action remain largely unknown. Previous studies showed that PTH increases the expression and the activity of epidermal growth factor receptor (EGFR) in osteoblasts, and activation of ERK1/2 by PTH in osteoblasts was demonstrated to induce the proteolytical release of EGFR ligands and EGFR transactivation. However, conclusive evidence for an important role of the EGFR system in mediating the anabolic actions of intermittent PTH on bone in vivo is lacking. Here, we evaluated the effects of intermittent PTH on bone in Waved-5 (Wa5) mice which carry an antimorphic Egfr allele whose product acts as a dominant negative receptor. Heterozygous Wa5 females and control littermates received a subcutaneous injection of PTH (80 μg/kg) or buffer on 5 days per week for 4 weeks. Wa5 mice had slightly lower total bone mineral density (BMD), but normal cancellous bone volume and turnover in the distal femoral metaphysis. The presence of the antimorphic Egfr allele neither influenced the PTH-induced increase in serum osteocalcin nor the increases in distal femoral BMD, cortical thickness, cancellous bone volume, and cancellous bone formation rate. Similarly, the PTH-induced rise in lumbar vertebral BMD was unchanged in Wa5 relative to wild-type mice. Wa5-derived osteoblasts showed considerably lower basal extracellular signal-regulated kinase 1/2 (ERK1/2) activation as compared to control osteoblasts. Whereas activation of ERK1/2 by the EGFR ligand amphiregulin was largely blocked in Wa5 osteoblasts, treatment with PTH induced ERK1/2 activation comparable to that observed in control osteoblasts, relative to baseline levels. Our data indicate that impairment of EGFR signaling does not affect the anabolic action of intermittent PTH on cancellous and cortical bone. Copyright © 2011. Published by Elsevier Inc.

  9. Inhibition of E2F-1 transactivation by direct binding of the retinoblastoma protein

    DEFF Research Database (Denmark)

    Helin, K; Harlow, E; Fattaey, A

    1993-01-01

    to transcription factor E2F has provided a model for the mechanism of pRB-mediated growth regulation. Since adenovirus E1A proteins dissociate the pRB-E2F complexes and stimulate E2F-dependent transcription, it has been suggested that pRB inhibits E2F transactivation. Although some evidence for this hypothesis has...... been provided, it has not been possible to determine the mechanism of pRB-mediated inhibition of E2F transactivation. In this study, we constructed mutants of E2F-1 that do not bind to pRB yet retain the ability to transactivate the adenovirus E2 promoter through E2F DNA-binding sites. We demonstrated...

  10. NLRC5/MHC class I transactivator is a target for immune evasion in cancer

    OpenAIRE

    Yoshihama, Sayuri; Roszik, Jason; Downs, Isaac; Meissner, Torsten B.; Vijayan, Saptha; Chapuy, Bjoern; Sidiq, Tabasum; Shipp, Margaret A.; Lizee, Gregory A.; Kobayashi, Koichi S.

    2016-01-01

    Tumor antigen presentation to CD8+ T cells by MHC class I molecules is crucial for immune responses against cancers, whereas the loss of MHC class I is a common immune evasion strategy used by cancers. However, the molecular mechanisms leading to MHC class I deficiency have remained poorly defined. We demonstrate here that MHC class I transactivator (CITA)/NOD-like receptor (NLR) family, caspase recruitment (CARD) domain containing 5 (NLRC5) is a major target for cancer immune evasion. Reduce...

  11. Hepatitis B virus DNA integration and transactivation of cellular genes

    Directory of Open Access Journals (Sweden)

    Vijay Kumar

    2007-02-01

    transactivator can stimulate a wide range of cellular genes and displays oncogenic potential in cell culture as well as in a transgenic environment.

    The HBs transactivators are encoded by the preS/S region of S gene and may involve carboxy terminal truncation to gain transactivation function. Expression of host genes by viral transactivators is mediated by regulatory elements of the cellular transcription factors like c-fos, c-myc, NF-kappa B, SRE and Sp1. Thus, during hepatitis B infection, the tendency of rearrangement of hepatocyte chromosomes is combined with the forcible turnover of cells. This is a constantly operating system for the selection of cells that grow better than normal cells, possibly involving important steps in multi-staged hepatocarcinogeneses. Gene expression profiling and proteomic techniques may help to characterize the molecular mechanisms driving HBV-associated carcinogenesis, and thus potentially identify new strategies in diagnosis and therapy.

     

    REFERENCES

    1. Kekule AS, Lauer U, Meyer M, Caselmann WH, Hofschneider PH, Koshy R. (1990 The preS2/S region of integrated hepatitis B virus DNA encodes a transcriptional transactivator. Nature 343, 457-461.

    2. Caselmann WH. (1996 Trans-activation of cellular genes by hepatitis B virus proteins: a possible mechanism of hepatocarcinogenesis. Adv Virus Res 47, 253-302.

    3. Matsubara K, Tokino T. (1990 Integration of hepatitis B virus DNA and its implications for hepatocarcinogenesis. Mol Biol

  12. Andrographolide regulates epidermal growth factor receptor and transferrin receptor trafficking in epidermoid carcinoma (A-431) cells

    Science.gov (United States)

    Tan, Y; Chiow, KH; Huang, D; Wong, SH

    2010-01-01

    Background and purpose: Andrographolide is the active component of Andrographis paniculata, a plant used in both Indian and Chinese traditional medicine, and it has been demonstrated to induce apoptosis in different cancer cell lines. However, not much is known about how it may affect the key receptors implicated in cancer. Knowledge of how andrographolide affects receptor trafficking will allow us to better understand new mechanisms by which andrographolide may cause death in cancer cells. Experimental approach: We utilized the well-characterized epidermal growth factor receptor (EGFR) and transferrin receptor (TfR) expressed in epidermoid carcinoma (A-431) cells as a model to study the effect of andrographolide on receptor trafficking. Receptor distribution, the total number of receptors and surface receptors were analysed by immunofluorescence, Western blot as well as flow-cytometry respectively. Key results: Andrographolide treatment inhibited cell growth, down-regulated EGFRs on the cell surface and affected the degradation of EGFRs and TfRs. The EGFR was internalized into the cell at an increased rate, and accumulated in a compartment that co-localizes with the lysosomal-associated membrane protein in the late endosomes. Conclusion and implications: This study sheds light on how andrographolide may affect receptor trafficking by inhibiting receptor movement from the late endosomes to lysosomes. The down-regulation of EGFR from the cell surface also indicates a new mechanism by which andrographolide may induce cancer cell death. PMID:20233216

  13. Andrographolide regulates epidermal growth factor receptor and transferrin receptor trafficking in epidermoid carcinoma (A-431) cells.

    Science.gov (United States)

    Tan, Y; Chiow, K H; Huang, D; Wong, S H

    2010-04-01

    Andrographolide is the active component of Andrographis paniculata, a plant used in both Indian and Chinese traditional medicine, and it has been demonstrated to induce apoptosis in different cancer cell lines. However, not much is known about how it may affect the key receptors implicated in cancer. Knowledge of how andrographolide affects receptor trafficking will allow us to better understand new mechanisms by which andrographolide may cause death in cancer cells. We utilized the well-characterized epidermal growth factor receptor (EGFR) and transferrin receptor (TfR) expressed in epidermoid carcinoma (A-431) cells as a model to study the effect of andrographolide on receptor trafficking. Receptor distribution, the total number of receptors and surface receptors were analysed by immunofluorescence, Western blot as well as flow-cytometry respectively. Andrographolide treatment inhibited cell growth, down-regulated EGFRs on the cell surface and affected the degradation of EGFRs and TfRs. The EGFR was internalized into the cell at an increased rate, and accumulated in a compartment that co-localizes with the lysosomal-associated membrane protein in the late endosomes. This study sheds light on how andrographolide may affect receptor trafficking by inhibiting receptor movement from the late endosomes to lysosomes. The down-regulation of EGFR from the cell surface also indicates a new mechanism by which andrographolide may induce cancer cell death.

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

    International Nuclear Information System (INIS)

    Williams, L.T.; Escobedo, J.A.; Keating, M.T.; Coughlin, S.R.

    1988-01-01

    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

  15. Cyclin D1 represses p300 transactivation through a cyclin-dependent kinase-independent mechanism.

    Science.gov (United States)

    Fu, Maofu; Wang, Chenguang; Rao, Mahadev; Wu, Xiaofang; Bouras, Toula; Zhang, Xueping; Li, Zhiping; Jiao, Xuanmao; Yang, Jianguo; Li, Anping; Perkins, Neil D; Thimmapaya, Bayar; Kung, Andrew L; Munoz, Alberto; Giordano, Antonio; Lisanti, Michael P; Pestell, Richard G

    2005-08-19

    Cyclin D1 encodes a regulatory subunit, which with its cyclin-dependent kinase (Cdk)-binding partner forms a holoenzyme that phosphorylates and inactivates the retinoblastoma protein. In addition to its Cdk binding-dependent functions, cyclin D1 regulates cellular differentiation in part by modifying several transcription factors and nuclear receptors. The molecular mechanism through which cyclin D1 regulates the function of transcription factors involved in cellular differentiation remains to be clarified. The histone acetyltransferase protein p300 is a co-integrator required for regulation of multiple transcription factors. Here we show that cyclin D1 physically interacts with p300 and represses p300 transactivation. We demonstrated further that the interaction of the two proteins occurs at the peroxisome proliferator-activated receptor gamma-responsive element of the lipoprotein lipase promoter in the context of the local chromatin structure. We have mapped the domains in p300 and cyclin D1 involved in this interaction. The bromo domain and cysteine- and histidine-rich domains of p300 were required for repression by cyclin D1. Cyclin D1 repression of p300 was independent of the Cdk- and retinoblastoma protein-binding domains of cyclin D1. Cyclin D1 inhibits histone acetyltransferase activity of p300 in vitro. Microarray analysis identified a signature of genes repressed by cyclin D1 and induced by p300 that promotes cellular differentiation and induces cell cycle arrest. Together, our results suggest that cyclin D1 plays an important role in cellular proliferation and differentiation through regulation of p300.

  16. Proteomic analysis of the herpes simplex virus 1 virion protein 16 transactivator protein in infected cells.

    Science.gov (United States)

    Suk, Hyung; Knipe, David M

    2015-06-01

    The herpes simplex virus 1 virion protein 16 (VP16) tegument protein forms a transactivation complex with the cellular proteins host cell factor 1 (HCF-1) and octamer-binding transcription factor 1 (Oct-1) upon entry into the host cell. VP16 has also been shown to interact with a number of virion tegument proteins and viral glycoprotein H to promote viral assembly, but no comprehensive study of the VP16 proteome has been performed at early times postinfection. We therefore performed a proteomic analysis of VP16-interacting proteins at 3 h postinfection. We confirmed the interaction of VP16 with HCF-1 and a large number of cellular Mediator complex proteins, but most surprisingly, we found that the major viral protein associating with VP16 is the infected cell protein 4 (ICP4) immediate-early (IE) transactivator protein. These results raise the potential for a new function for VP16 in associating with the IE ICP4 and playing a role in transactivation of early and late gene expression, in addition to its well-documented function in transactivation of IE gene expression. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Novel Drosophila receptor that binds multiple growth factors

    International Nuclear Information System (INIS)

    Rosner, M.R.; Thompson, K.L.; Garcia, V.; Decker, S.J.

    1986-01-01

    The authors have recently reported the identification of a novel growth factor receptor from Drosophila cell cultures that has dual binding specificity for both insulin and epidermal growth factor (EGF). This 100 kDa protein is also antigenically related to the cytoplasmic region of the mammalian EGF receptor-tyrosine kinase. They now report that this protein binds to mammalian nerve growth factor and human transforming growth factor alpha as well as insulin and EGF with apparent dissociation constants ranging from 10 -6 to 10 -8 M. The 100 kDa protein can be affinity-labeled with these 125 I-labeled growth factors after immunoprecipitation with anti-EGF receptor antiserum. These four growth factors appear to share a common binding site, as evidenced by their ability to block affinity labelling by 125 I-insulin. No significant binding to the 100 kDa protein was observed with platelet-derived growth factor, transforming growth factor beta, or glucagon. The 100 kDa Drosophila protein has a unique ligand-binding spectrum with no direct counterpart in mammalian cells and may represent an evolutionary precursor of the mammalian receptors for these growth factors

  18. Astrocyte Mitogen Inhibitor Related to Epidermal Growth Factor Receptor

    Science.gov (United States)

    Nieto-Sampedro, Manuel

    1988-06-01

    Epidermal growth factor (EGF) is a well-characterized polypeptide hormone with diverse biological activities, including stimulation of astrocyte division. A soluble astrocyte mitogen inhibitor, immunologically related to the EGF receptor, is present in rat brain. Injury to the brain causes a time-dependent reduction in the levels of this inhibitor and the concomitant appearance of EGF receptor on the astrocyte surface. Intracerebral injection of antibody capable of binding the inhibitor caused the appearance of numerous reactive astrocytes. EGF receptor-related inhibitors may play a key role in the control of glial cell division in both normal and injured brain.

  19. Transactivation of EGFR by LPS induces COX-2 expression in enterocytes.

    Directory of Open Access Journals (Sweden)

    Steven J McElroy

    Full Text Available Necrotizing enterocolitis (NEC is the leading cause of gastrointestinal morbidity and mortality in preterm infants. NEC is characterized by an exaggerated inflammatory response to bacterial flora leading to bowel necrosis. Bacterial lipopolysaccharide (LPS mediates inflammation through TLR4 activation and is a key molecule in the pathogenesis of NEC. However, LPS also induces cyclooxygenase-2 (COX-2, which promotes intestinal barrier restitution through stimulation of intestinal cell survival, proliferation, and migration. Epidermal growth factor receptor (EGFR activation prevents experimental NEC and may play a critical role in LPS-stimulated COX-2 production. We hypothesized that EGFR is required for LPS induction of COX-2 expression. Our data show that inhibiting EGFR kinase activity blocks LPS-induced COX-2 expression in small intestinal epithelial cells. LPS induction of COX-2 requires Src-family kinase signaling while LPS transactivation of EGFR requires matrix metalloprotease (MMP activity. EGFR tyrosine kinase inhibitors block LPS stimulation of mitogen-activated protein kinase ERK, suggesting an important role of the MAPK/ERK pathway in EGFR-mediated COX-2 expression. LPS stimulates proliferation of IEC-6 cells, but this stimulation is inhibited with either the EGFR kinase inhibitor AG1478, or the selective COX-2 inhibitor Celecoxib. Taken together, these data show that EGFR plays an important role in LPS-induction of COX-2 expression in enterocytes, which may be one mechanism for EGF in inhibition of NEC.

  20. AKT increases VEGF expression in tumor cells by transactivating the proximal VEGF promoter

    International Nuclear Information System (INIS)

    Pore, N.; Bernhard, E.J.; Shu, H.-K.; Li, B.; O'Rourke, D.M.; Maity, A.; Haas-Kogan, D.

    2003-01-01

    Vascular endothelial growth factor (VEGF) is overexpressed in many cancers including glioblastomas and may contribute to their growth. Epidermal growth factor receptor (EGFR) amplification and loss of PTEN, commonly found in glioblastomas leading to increase phosphatidylinositol-3-kinase (PI3K) activity and VEGF expression. In the current study we show that AKT, which is downstream of PI3K, regulates VEGF expression. U87MG human glioblastoma cells lack wildtype PTEN and express high levels of phosphorylated AKT. Over expression of AKT either by stable expression in immortalized human astrocytes or by transduction with adenovirus containing activated myristoylated AKT in SF188 glioblastoma cells increases VEGF expression. Moreover the elevation of angiogenesis by constitutively expressed AKT is further confirmed by in vivo matrigel plug assay in nude mice. The upregulation of VEGF by AKT is mediated through a region in the proximal promoter located between -88 and -70 (+1 is transcription start site). In transient transfection activity of a luciferase reporter containing the -88/+54 region of the VEGF promoter is increased by cotransfection with myristoylated AKT and downregulated by a dominant negative AKT expression vector. Mutation of the putative Sp1 binding sites located in the -88/-70 region we show that AKT acts through Sp1 to transactivate the VEGF promoter. Cotransfection of the VEGF promoter reporter with both Sp1 and myristoylated AKT expression vectors increases promoter activity to a greater extent than either Sp1 or Akt by itself. In vivo phosphate labeling of proteins reveals that AKT leads to increased Sp1 phosphorylation. Gel shift assays using a radio labeled probe corresponding to nucleotides -88 through -66 in the promoter show increased binding with nuclear extracts from cells transduced with adenovirus expressing myristoylated AKT. In conclusion, our results suggest that loss of PTEN leads to increased VEGF expression by increasing AKT

  1. Mutations that abrogate transactivational activity of the feline leukemia virus long terminal repeat do not affect virus replication

    International Nuclear Information System (INIS)

    Abujamra, Ana L.; Faller, Douglas V.; Ghosh, Sajal K.

    2003-01-01

    The U3 region of the LTR of oncogenic Moloney murine leukemia virus (Mo-MuLV) and feline leukemia viruses (FeLV) have been previously reported to activate expression of specific cellular genes in trans, such as MHC class I, collagenase IV, and MCP-1, in an integration-independent manner. It has been suggested that transactivation of these specific cellular genes by leukemia virus U3-LTR may contribute to the multistage process of leukemogenesis. The U3-LTR region, necessary for gene transactivational activity, also contains multiple transcription factor-binding sites that are essential for normal virus replication. To dissect the promoter activity and the gene transactivational activity of the U3-LTR, we conducted mutational analysis of the U3-LTR region of FeLV-A molecular clone 61E. We identified minimal nucleotide substitution mutants on the U3 LTR that did not disturb transcription factor-binding sites but abrogated its ability to transactivate the collagenase gene promoter. To determine if these mutations actually have altered any uncharacterized important transcription factor-binding site, we introduced these U3-LTR mutations into the full-length infectious molecular clone 61E. We demonstrate that the mutant virus was replication competent but could not transactivate cellular gene expression. These results thus suggest that the gene transactivational activity is a distinct property of the LTR and possibly not related to its promoter activity. The cellular gene transactivational activity-deficient mutant FeLV generated in this study may also serve as a valuable reagent for testing the biological significance of LTR-mediated cellular gene activation in the tumorigenesis caused by leukemia viruses

  2. Prolactin modulates phosphorylation, signaling and trafficking of epidermal growth factor receptor in human T47D breast cancer cells.

    Science.gov (United States)

    Huang, Y; Li, X; Jiang, J; Frank, S J

    2006-12-07

    Prolactin (PRL) is a polypeptide hormone produced by the anterior pituitary gland and other sites that acts both systemically and locally to cause lactation and other biological effects by interacting with the PRL receptor, a Janus kinase (JAK)2-coupled cytokine receptor family member, and activating downstream signal pathways. Recent evidence suggests PRL is a player in the pathogenesis and progression of breast cancer. Epidermal growth factor (EGF) also has effects on breast tissue, working through its receptors, epidermal growth factor receptor (EGFR) and ErbB-2 (c-neu, HER2), both intrinsic tyrosine kinase growth factor receptors. EGFR promotes pubertal breast ductal morphogenesis in mice, and both EGFR and ErbB-2 are relevant in pathogenesis and behavior of breast and other human cancers. Previous studies showed that PRL and EGF synergize to enhance motility in the human breast cancer cell line, T47D. In this study, we explored crosstalk between the PRL and EGF signaling pathways in T47D cells, with an ultimate aim of understanding how these two important factors might work together in vivo to affect breast cancer behavior. Both PRL and EGF caused robust signaling in T47D cells; PRL acutely activated JAK2, signal transducer and activator of transcription-5 (STAT5), and extracellular signal-regulated kinase-1 and -2 (ERK1 and ERK2), whereas EGF caused EGFR activation and consequent src homology collagen (SHC) activation and ERK activation. Notably, PRL also caused phosphorylation of the EGFR and ErbB-2 at sites detected by PTP101, an antibody that recognizes threonine phosphorylation at consensus motifs for ERK-induced phosphorylation. PRL-induced PTP101-reactive phosphorylation was prevented by pretreatment with PD98059, an ERK pathway inhibitor. Furthermore, PRL synergized with EGF in activating SHC and ERK and transactivating a luciferase reporter driven by c-fos gene enhancer elements, suggesting that PRL allowed markedly enhanced EGF signaling. This was

  3. Topical administration of adrenergic receptor pharmaceutics and nerve growth factor

    Directory of Open Access Journals (Sweden)

    Jena J Steinle

    2010-06-01

    Full Text Available Jena J SteinleDepartments of Ophthalmology and Anatomy and Neurobiology, Hamilton Eye Institute, University of Tennessee Health Science Center, Memphis, TN 38163, USAAbstract: Topical application of nerve growth factor (NGF and adrenergic receptor pharmaceutics are currently in use for corneal ulcers and glaucoma. A recent interest in the neuroprotective abilities of NGF has led to a renewed interest in NGF as a therapeutic for retinal and choroidal diseases. NGF can promote cell proliferation through actions of the TrkA receptor or promote apoptosis through receptor p75NTR. This understanding has led to novel interest in the role of NGF for diseases of the posterior eye. The role of β-adrenergic receptor agonists and antagonists for treatments of glaucoma, diabetic retinopathy, and their potential mechanisms of action, are still under investigation. This review discusses the current knowledge and applications of topical NGF and adrenergic receptor drugs for ocular disease.Keywords: NGF, β-adrenergic receptor agents, α-adrenergic receptor agents, retina, cornea, glaucoma

  4. Vascular endothelial growth factor ( VEGF ) receptor expression ...

    African Journals Online (AJOL)

    Background: Colorectal carcinoma (CRC) is the seventh-most common malignancy and is the main cause of death in Iraq. The incidence of this cancer has increased sharply after the invasion of Iraq in 2003. Aim: To estimate immunohistochemical expression of vascular endothelial growth factor (VEGF) in CRC in relation ...

  5. Delineation of atypical insulin receptors from classical insulin and type I insulin-like growth factor receptors in human placenta.

    OpenAIRE

    Jonas, H A; Cox, A J; Harrison, L C

    1989-01-01

    Insulin-like growth factor (IGF)-binding sites copurifying with human placental insulin receptors during insulin-affinity chromatography consist of two immunologically distinct populations. One reacts with monoclonal antibody alpha IR-3, but not with antibodies to the insulin receptor, and represents Type I IGF receptors; the other reacts only with antibodies to the insulin receptor and is precipitated with a polyclonal receptor antibody (B-10) after labelling with 125I-multiplication-stimula...

  6. Diametrically opposed effects of hypoxia and oxidative stress on two viral transactivators

    Directory of Open Access Journals (Sweden)

    Aiyar Ashok

    2010-05-01

    Full Text Available Abstract Background Many pathogens exist in multiple physiological niches within the host. Differences between aerobic and anaerobic conditions are known to alter the expression of bacterial virulence factors, typically through the conditional activity of transactivators that modulate their expression. More recently, changes in physiological niches have been shown to affect the expression of viral genes. For many viruses, differences in oxygen tension between hypoxia and normoxia alter gene expression or function. Oxygen tension also affects many mammalian transactivators including AP-1, NFkB, and p53 by affecting the reduced state of critical cysteines in these proteins. We have recently determined that an essential cys-x-x-cys motif in the EBNA1 transactivator of Epstein-Barr virus is redox-regulated, such that transactivation is favoured under reducing conditions. The crucial Tat transactivator of human immunodeficiency virus (HIV has an essential cysteine-rich region, and is also regulated by redox. Contrary to EBNA1, it is reported that Tat's activity is increased by oxidative stress. Here we have compared the effects of hypoxia, oxidative stress, and cellular redox modulators on EBNA1 and Tat. Results Our results indicate that unlike EBNA1, Tat is less active during hypoxia. Agents that generate hydroxyl and superoxide radicals reduce EBNA1's activity but increase transactivation by Tat. The cellular redox modulator, APE1/Ref-1, increases EBNA1's activity, without any effect on Tat. Conversely, thioredoxin reductase 1 (TRR1 reduces Tat's function without any effect on EBNA1. Conclusions We conclude that oxygen partial pressure and oxidative stress affects the functions of EBNA1 and Tat in a dramatically opposed fashion. Tat is more active during oxidative stress, whereas EBNA1's activity is compromised under these conditions. The two proteins respond to differing cellular redox modulators, suggesting that the oxidized cysteine adduct is a

  7. Orphan Nuclear Receptor Small Heterodimer Partner Negatively Regulates Growth Hormone-mediated Induction of Hepatic Gluconeogenesis through Inhibition of Signal Transducer and Activator of Transcription 5 (STAT5) Transactivation*

    Science.gov (United States)

    Kim, Yong Deuk; Li, Tiangang; Ahn, Seung-Won; Kim, Don-Kyu; Lee, Ji-Min; Hwang, Seung-Lark; Kim, Yong-Hoon; Lee, Chul-Ho; Lee, In-Kyu; Chiang, John Y. L.; Choi, Hueng-Sik

    2012-01-01

    Growth hormone (GH) is a key metabolic regulator mediating glucose and lipid metabolism. Ataxia telangiectasia mutated (ATM) is a member of the phosphatidylinositol 3-kinase superfamily and regulates cell cycle progression. The orphan nuclear receptor small heterodimer partner (SHP: NR0B2) plays a pivotal role in regulating metabolic processes. Here, we studied the role of ATM on GH-dependent regulation of hepatic gluconeogenesis in the liver. GH induced phosphoenolpyruvate carboxykinase (PEPCK) and glucose 6-phosphatase gene expression in primary hepatocytes. GH treatment and adenovirus-mediated STAT5 overexpression in hepatocytes increased glucose production, which was blocked by a JAK2 inhibitor, AG490, dominant negative STAT5, and STAT5 knockdown. We identified a STAT5 binding site on the PEPCK gene promoter using reporter assays and point mutation analysis. Up-regulation of SHP by metformin-mediated activation of the ATM-AMP-activated protein kinase pathway led to inhibition of GH-mediated induction of hepatic gluconeogenesis, which was abolished by an ATM inhibitor, KU-55933. Immunoprecipitation studies showed that SHP physically interacted with STAT5 and inhibited STAT5 recruitment on the PEPCK gene promoter. GH-induced hepatic gluconeogenesis was decreased by either metformin or Ad-SHP, whereas the inhibition by metformin was abolished by SHP knockdown. Finally, the increase of hepatic gluconeogenesis following GH treatment was significantly higher in the liver of SHP null mice compared with that of wild-type mice. Overall, our results suggest that the ATM-AMP-activated protein kinase-SHP network, as a novel mechanism for regulating hepatic glucose homeostasis via a GH-dependent pathway, may be a potential therapeutic target for insulin resistance. PMID:22977252

  8. The epidermal growth factor receptor pathway in chronic kidney diseases

    NARCIS (Netherlands)

    Harskamp, Laura R.; Gansevoort, Ron T.; Goor, van Harry; Meijer, Esther

    The epidermal growth factor receptor (EGFR) pathway has a critical role in renal development, tissue repair and electrolyte handling. Numerous studies have reported an association between dysregulation of this pathway and the initiation and progression of various chronic kidney diseases such as

  9. Epidermal growth factor receptor: Target for delivery and silencing

    NARCIS (Netherlands)

    Santos Oliveira, S.|info:eu-repo/dai/nl/304841455

    2008-01-01

    Epidermal growth factor receptor in cancer therapy Recently, cancer research has been able to identify molecular targets that are specific for (or highly expressed by) cancer cells. These molecular targets serve as models for the development of rationally designed anticancer drugs that target

  10. Morphine potentiates neurodegenerative effects of HIV-1 Tat through actions at μ-opioid receptor-expressing glia.

    Science.gov (United States)

    Zou, Shiping; Fitting, Sylvia; Hahn, Yun-Kyung; Welch, Sandra P; El-Hage, Nazira; Hauser, Kurt F; Knapp, Pamela E

    2011-12-01

    soluble factors. Similar morphine-transactivator of transcription synergy was also observed in studies with the clade C sequence of HIV-1 transactivator of transcription, which did not cause neuron death unless morphine was present. Several paradoxical observations related to opiate effects were noted when µ-opioid receptors were specifically ablated from either glia or neurons. This suggests that µ-opioid receptor loss in isolated cell types can fundamentally distort cell-to-cell signalling, revealing opponent processes that may exist in individual cell types. Our findings show the critical role of glia in orchestrating neurotoxic interactions of morphine and transactivator of transcription, and support the emerging concept that combined exposure to opiates and human immunodeficiency virus drives enhanced pathology within the central nervous system.

  11. Insulin and insulin-like growth factor receptors and responses

    International Nuclear Information System (INIS)

    Roth, R.A.; Steele-Perkins, G.; Hari, J.; Stover, C.; Pierce, S.; Turner, J.; Edman, J.C.; Rutter, W.J.

    1988-01-01

    Insulin is a member of a family of structurally related hormones with diverse physiological functions. In humans, the best-characterized members of this family include insulin, insulin-like growth factor (IGF)-I, and IGF-II. Each of these three polypeptide hormones has its own distinct receptor. The structures of each of these receptors have now been deduced from analyses of isolated cDNA clones. To study further the responses mediated through these three different receptors, the authors have been studying cells expressing the proteins encoded by these three cDNAs. The isolated cDNAs have been transfected into Chinese hamster ovary (CHO) cells, and the resulting transfected cell lines have been characterized as to the ligand-binding activities and signal-transducing activities of the expressed proteins

  12. Ciliary neurotrophic factor receptor regulation of adult forebrain neurogenesis.

    Science.gov (United States)

    Lee, Nancy; Batt, Myra K; Cronier, Brigitte A; Jackson, Michele C; Bruno Garza, Jennifer L; Trinh, Dennis S; Mason, Carter O; Spearry, Rachel P; Bhattacharya, Shayon; Robitz, Rachel; Nakafuku, Masato; MacLennan, A John

    2013-01-16

    Appropriately targeted manipulation of endogenous neural stem progenitor (NSP) cells may contribute to therapies for trauma, stroke, and neurodegenerative disease. A prerequisite to such therapies is a better understanding of the mechanisms regulating adult NSP cells in vivo. Indirect data suggest that endogenous ciliary neurotrophic factor (CNTF) receptor signaling may inhibit neuronal differentiation of NSP cells. We challenged subventricular zone (SVZ) cells in vivo with low concentrations of CNTF to anatomically characterize cells containing functional CNTF receptors. We found that type B "stem" cells are highly responsive, whereas type C "transit-amplifying" cells and type A neuroblasts are remarkably unresponsive, as are GFAP(+) astrocytes found outside the SVZ. CNTF was identified in a subset of type B cells that label with acute BrdU administration. Disruption of in vivo CNTF receptor signaling in SVZ NSP cells, with a "floxed" CNTF receptor α (CNTFRα) mouse line and a gene construct driving Cre recombinase (Cre) expression in NSP cells, led to increases in SVZ-associated neuroblasts and new olfactory bulb neurons, as well as a neuron subtype-specific, adult-onset increase in olfactory bulb neuron populations. Adult-onset receptor disruption in SVZ NSP cells with a recombinant adeno-associated virus (AAV-Cre) also led to increased neurogenesis. However, the maintenance of type B cell populations was apparently unaffected by the receptor disruption. Together, the data suggest that endogenous CNTF receptor signaling in type B stem cells inhibits adult neurogenesis, and further suggest that the regulation may occur in a neuron subtype-specific manner.

  13. Inhibition of Nuclear Transcription Factor-κB and Activation of Peroxisome Proliferator-Activated Receptors in HepG2 Cells by Cucurbitane-Type Triterpene Glycosides from Momordica charantia

    Science.gov (United States)

    Nhiem, Nguyen Xuan; Yen, Pham Hai; Ngan, Nguyen Thi Thanh; Quang, Tran Hong; Kiem, Phan Van; Minh, Chau Van; Tai, Bui Huu; Cuong, Nguyen Xuan; Song, Seok Bean

    2012-01-01

    Abstract Momordica charantia: is used to treat various diseases, including inflammatory conditions. Previous reports indicated that the extract of this plant inhibits activation of nuclear transcription factor-κB (NF-κB) but activates peroxisome proliferator-activated receptor (PPAR). Additionally, cucurbitane-type triterpene glycosides are the main bioactive components of the fruit of M. charantia. Therefore, we investigated the anti-inflammatory activity of 17 cucurbitane-type triterpene glycosides (1–17) isolated from this plant. Their inhibition of NF-κB and activation of PPAR activities in HepG2 cells were measured using luciferase reporter and PPAR subtype transactivation assays. Compounds 6 and 8 were found to inhibit NF-κB activation stimulated by tumor necrosis factor-α (TNFα) in a dose-dependent manner. With 50% inhibition concentration (IC50) values of 0.4 μM, compounds 6 and 8 were more potent inhibitors than the positive control, sulfasalazine (IC50=0.9 μM). Compounds 4, 6, and 8 also inhibited TNFα-induced expressions of inducible nitric oxide synthase and cyclooxygenase-2 mRNA. However, only compound 13 significantly increased PPARγ transactivation. PMID:22248180

  14. TMPRSS2-ERG fusion protein regulates insulin-like growth factor-1 receptor (IGF1R) gene expression in prostate cancer: involvement of transcription factor Sp1.

    Science.gov (United States)

    Meisel Sharon, Shilhav; Pozniak, Yair; Geiger, Tamar; Werner, Haim

    2016-08-09

    Prostate cancer is a major health issue in the Western world. The most common gene rearrangement in prostate cancer is the TMPRSS2-ERG fusion, which results in aberrant expression of the transcription factor ERG. The insulin-like growth factor-1 receptor (IGF1R) plays a key role in cell growth and tumorigenesis, and is overexpressed in most malignancies, including prostate cancer. In this study we show that TMPRSS2-ERG mediates its tumorigenic effects through regulation of IGF1R gene expression. Silencing of T-ERG in VCaP cells resulted in downregulation of both IGF1R and Sp1, a critical IGF1R regulator. Co-immunoprecipitation assays revealed a physical interaction between transcription factors ERG and Sp1, with potential relevance in IGF1R gene regulation. In addition, transactivation of the IGF1R gene by ERG was mediated at the level of transcription, as indicated by results of promoter assays. To identify new co-activators of the TMPRSS2-ERG fusion protein we performed mass spectrometry-based proteomic analyses. Among other interactors, we identified AP-2 complex subunit mu (AP2M1) and caveolin-1 (CAV1) in association with ERG in cell nuclei. These proteins play a mechanistic role in IGF1R internalization. Our analyses are consistent with a potential novel function of TMPRSS2-ERG as a major regulator of IGF1R gene expression. Results may impinge upon ongoing efforts to target the IGF1R in the clinics.

  15. Epidermal growth factor receptor expression in urinary bladder cancer

    Directory of Open Access Journals (Sweden)

    Dayalu S.L. Naik

    2011-01-01

    Full Text Available Objective : To evaluate the expression pattern of epidermal growth factor receptor (EGFR in urinary bladder cancer and its association with human epidermal growth factor receptor 2 (HER2, epidermal growth factor (EGF, interleukin-6 (IL-6, and high risk human papilloma virus (HPV types 16 and 18. Materials and Methods : Thirty cases of urothelial carcinoma were analyzed. EGFR, HER2, EGF, and IL-6 expressions in the tissue were evaluated by immunohistochemical staining. For HPV, DNA from tissue samples was extracted and detection of HPV was done by PCR technique. Furthermore, evaluation of different intracellular molecules associated with EGFR signaling pathways was performed by the western blot method using lysates from various cells and tissues. Results : In this study, the frequencies of immunopositivity for EGFR, HER2, EGF, and IL-6 were 23%, 60%, 47%, and 80%, respectively. No cases were positive for HPV-18, whereas HPV-16 was detected in 10% cases. Overall, expression of EGFR did not show any statistically significant association with the studied parameters. However, among male patients, a significant association was found only between EGFR and HER2. Conclusions : Overexpression of EGFR and/or HER2, two important members of the same family of growth factor receptors, was observed in a considerable proportion of cases. Precise knowledge in this subject would be helpful to formulate a rational treatment strategy in patients with urinary bladder cancer.

  16. Prognostic factors and status of hormone receptors and angiogenic factors in uterine carcinosarcoma.

    Science.gov (United States)

    Etoh, Tomomaro; Nakai, Hidekatsu

    2014-03-01

    To determine novel prognostic factors and treatment modalities for uterine carcinosarcoma (UCS). We performed immunohistochemical staining of estrogen receptor (ER)-α, ER-β, progesterone receptor, gonadotropin-releasing hormone receptor, vascular endothelial growth factor (VEGF), platelet-derived endothelial cell growth factor (PD-ECGF) and platelet-derived growth factor receptor (PDGFR)-β in a clinicopathological study of 15 UCS patients. No significant differences were found between the sarcomatous and carcinomatous components with respect to expression of ER-α, ER-β and progesterone receptor. However, VEGF was significantly more frequently expressed in the carcinomatous component, while PD-ECGF and PDGFR-β were significantly more frequently expressed in the sarcomatous component. Only one patient showed gonadotropin-releasing hormone receptor expression in the sarcomatous component. Moreover, ER-β expression in resected specimens, increased serum levels of carbohydrate antigen (CA)-125 and C-reactive protein (CRP), and thrombocytosis were determined as significant UCS prognostic factors. Combination of anti-VEGF therapy and anti-PD-ECGF or anti-PDGFR-β therapy would be expected in advanced or recurrent UCS. Furthermore, careful monitoring for early detection of recurrence should be performed when UCS patients showed preoperative increase in serum CA-125 levels, CRP and platelet counts, and ER-β expression in biopsied or surgically resected specimens. © 2013 The Authors. Journal of Obstetrics and Gynaecology Research © 2013 Japan Society of Obstetrics and Gynecology.

  17. Decreased expression of serum and microvascular vascular endothelial growth factor receptor-2 in meningococcal sepsis*.

    NARCIS (Netherlands)

    Flier, M. van der; Baerveldt, E.M.; Miedema, A.; Hartwig, N.G.; Hazelzet, J.A.; Emonts, M.; Groot, R. de; Prens, E.P.; Vught, A.J. van; Jansen, N.J.

    2013-01-01

    OBJECTIVES: To determine the skin microvessel expression of vascular endothelial growth factor receptor 2 and serum-soluble vascular endothelial growth factor receptor 2 levels in children with meningococcal sepsis. DESIGN: Observational study. SETTING: Two tertiary academic children hospital PICUs.

  18. Brief report: Glucocorticoid receptor polymorphism affects transrepression but not transactivation

    NARCIS (Netherlands)

    E.L.T. van den Akker (Erica); H. Russcher (Henk); E.F.C. van Rossum (Liesbeth); A.O. Brinkmann (Albert); F.H. de Jong (Frank); A.C.S. Hokken-Koelega (Anita); H.A.P. Pols (Huib); J.W. Koper (Jan); S.W.J. Lamberts (Steven)

    2006-01-01

    textabstractContext: Glucocorticoids (GCs) are extensively used in the treatment of inflammatory and autoimmune diseases. Their beneficial effects are thought to be mediated by GC transrepression on gene expression. However, their use is limited by serious adverse effects, presumably mediated by GC

  19. Cheiradone: a vascular endothelial cell growth factor receptor antagonist

    Directory of Open Access Journals (Sweden)

    Ahmed Nessar

    2008-01-01

    Full Text Available Abstract Background Angiogenesis, the growth of new blood vessels from the pre-existing vasculature is associated with physiological (for example wound healing and pathological conditions (tumour development. Vascular endothelial growth factor (VEGF, fibroblast growth factor-2 (FGF-2 and epidermal growth factor (EGF are the major angiogenic regulators. We have identified a natural product (cheiradone isolated from a Euphorbia species which inhibited in vivo and in vitro VEGF- stimulated angiogenesis but had no effect on FGF-2 or EGF activity. Two primary cultures, bovine aortic and human dermal endothelial cells were used in in vitro (proliferation, wound healing, invasion in Matrigel and tube formation and in vivo (the chick chorioallantoic membrane models of angiogenesis in the presence of growth factors and cheiradone. In all cases, the concentration of cheiradone which caused 50% inhibition (IC50 was determined. The effect of cheiradone on the binding of growth factors to their receptors was also investigated. Results Cheiradone inhibited all stages of VEGF-induced angiogenesis with IC50 values in the range 5.20–7.50 μM but did not inhibit FGF-2 or EGF-induced angiogenesis. It also inhibited VEGF binding to VEGF receptor-1 and 2 with IC50 values of 2.9 and 0.61 μM respectively. Conclusion Cheiradone inhibited VEGF-induced angiogenesis by binding to VEGF receptors -1 and -2 and may be a useful investigative tool to study the specific contribution of VEGF to angiogenesis and may have therapeutic potential.

  20. The F-BAR Protein PACSIN2 Regulates Epidermal Growth Factor Receptor Internalization

    NARCIS (Netherlands)

    de Kreuk, Bart-Jan; Anthony, Eloise C.; Geerts, Dirk; Hordijk, Peter L.

    2012-01-01

    Signaling via growth factor receptors, including the epidermal growth factor (EGF) receptor, is key to various cellular processes, such as proliferation, cell survival, and cell migration. In a variety of human diseases such as cancer, aberrant expression and activation of growth factor receptors

  1. Upregulation of epidermal growth factor receptor 4 in oral leukoplakia

    Science.gov (United States)

    Kobayashi, Hiroshi; Kumagai, Kenichi; Gotoh, Akito; Eguchi, Takanori; Yamada, Hiroyuki; Hamada, Yoshiki; Suzuki, Satsuki; Suzuki, Ryuji

    2013-01-01

    In the present study, we investigate the expression profile of the epidermal growth factor receptor family, which comprises EGFR/ErbB1, HER2/ErbB2, HER3/ErbB3 and HER4/ErbB4 in oral leukoplakia (LP). The expression of four epidermal growth factor receptor (EGFR) family genes and their ligands were measured in LP tissues from 14 patients and compared with levels in 10 patients with oral lichen planus (OLP) and normal oral mucosa (NOM) from 14 healthy donors by real-time polymerase chain reaction (PCR) and immunohistochemistry. Synchronous mRNA coexpression of ErbB1, ErbB2, ErbB3 and ErbB4 was detected in LP lesions. Out of the receptors, only ErbB4 mRNA and protein was more highly expressed in LP compared with NOM tissues. These were strongly expressed by epithelial keratinocytes in LP lesions, as shown by immunohistochemistry. Regarding the ligands, the mRNA of Neuregulin2 and 4 were more highly expressed in OLP compared with NOM tissues. Therefore, enhanced ErbB4 on the keratinocytes and synchronous modulation of EGFR family genes may contribute to the pathogenesis and carcinogenesis of LP. PMID:23492901

  2. Targeting the epidermal growth factor receptor in solid tumor malignancies

    DEFF Research Database (Denmark)

    Nedergaard, Mette K; Hedegaard, Chris J; Poulsen, Hans S

    2012-01-01

    to the extracellular part of EGFR, blocking the binding sites for the EGFR ligands, and intracellular tyrosine kinase inhibitors (TKIs) that block the ATP binding site of the tyrosine kinase domain. Besides an EGFRvIII-targeted vaccine, conjugated anti-EGFR mAbs have been used in different settings to deliver lethal......The epidermal growth factor receptor (EGFR) is over-expressed, as well as mutated, in many types of cancers. In particular, the EGFR variant type III mutant (EGFRvIII) has attracted much attention as it is frequently and exclusively found on many tumor cells, and hence both EGFR and EGFRvIII have...... been proposed as valid targets in many cancer therapy settings. Different strategies have been developed in order to either inhibit EGFR/EGFRvIII activity or to ablate EGFR/EGFRvIII-positive tumor cells. Drugs that inhibit these receptors include monoclonal antibodies (mAbs) that bind...

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

    DEFF Research Database (Denmark)

    Dengjel, Joern; Akimov, Vyacheslav; Blagoev, Blagoy

    2007-01-01

    -out by 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......Phosphorylation-based signaling events happening within the first minute of receptor stimulation have so far only been analyzed by classical cell biological approaches like live-cell microscopy. The development of a quench flow system with a time resolution of one second coupled to a read...... of stimulation. This new technique opens the perspectives for accurate analysis of rapid cellular processes and will help to establish models describing signal initiation at the plasma membrane....

  4. Tyrosine 769 of the keratinocyte growth factor receptor is required for receptor signaling but not endocytosis

    International Nuclear Information System (INIS)

    Ceridono, Mara; Belleudi, Francesca; Ceccarelli, Simona; Torrisi, Maria Rosaria

    2005-01-01

    Keratinocyte growth factor receptor (KGFR) is a receptor tyrosine kinase expressed on epithelial cells which belongs to the family of fibroblast growth factor receptors (FGFRs). Following ligand binding, KGFR is rapidly autophosphorylated on specific tyrosine residues in the intracellular domain, recruits substrate proteins, and is rapidly internalized by clathrin-mediated endocytosis. The role of different autophosphorylation sites in FGFRs, and in particular the role of the tyrosine 766 in FGFR1, first identified as PLCγ binding site, has been extensively studied. We analyzed here the possible role of the tyrosine 769 in KGFR, corresponding to tyrosine 766 in FGFR1, in the regulation of KGFR signal transduction and MAPK activation as well as in the control of the endocytic process of KGFR. A mutant KGFR in which tyrosine 769 was substituted by phenylalanine was generated and transfected in NIH3T3 and HeLa cells. Our results indicate that tyrosine 769 is required for the binding to KGFR and tyrosine phosphorylation of PLCγ as well as for the full activation of MAPKs and for cell proliferation through the regulation of FRS2 tyrosine phosphorylation, suggesting that this residue represents a key regulator of KGFR signal transduction. Our data also show that tyrosine 769 is not involved in the regulation of the endocytic process of KGFR

  5. Dynamic Arginine Methylation of Tumor Necrosis Factor (TNF) Receptor-associated Factor 6 Regulates Toll-like Receptor Signaling.

    Science.gov (United States)

    Tikhanovich, Irina; Kuravi, Sudhakiranmayi; Artigues, Antonio; Villar, Maria T; Dorko, Kenneth; Nawabi, Atta; Roberts, Benjamin; Weinman, Steven A

    2015-09-04

    Arginine methylation is a common post-translational modification, but its role in regulating protein function is poorly understood. This study demonstrates that, TNF receptor-associated factor 6 (TRAF6), an E3 ubiquitin ligase involved in innate immune signaling, is regulated by reversible arginine methylation in a range of primary and cultured cells. Under basal conditions, TRAF6 is methylated by the methyltransferase PRMT1, and this inhibits its ubiquitin ligase activity, reducing activation of toll-like receptor signaling. In response to toll-like receptor ligands, TRAF6 is demethylated by the Jumonji domain protein JMJD6. Demethylation is required for maximal activation of NF-κB. Loss of JMJD6 leads to reduced response, and loss of PRMT1 leads to basal pathway activation with subsequent desensitization to ligands. In human primary cells, variations in the PRMT1/JMJD6 ratio significantly correlate with TRAF6 methylation, basal activation of NF-κB, and magnitude of response to LPS. Reversible arginine methylation of TRAF6 by the opposing effects of PRMT1 and JMJD6 is, therefore, a novel mechanism for regulation of innate immune pathways. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. NLRC5: a newly discovered MHC class I transactivator (CITA)

    OpenAIRE

    Meissner, Torsten B.; Li, Amy; Kobayashi, Koichi S.

    2011-01-01

    Major histocompatibility complex (MHC) class I and class II are crucial for the function of the human adaptive immune system. An NLR protein, CIITA (MHC class II transactivator), is a master regulator of MHC class II gene expression as well as of some of the genes involved in MHC class II antigen presentation. It has recently been discovered that another member of the NLR protein family, NLRC5, transcriptionally activates MHC class I genes, and thus acts as “CITA” (MHC class I transactivator)...

  7. EXPRESSION OF GROWTH-FACTORS AND GROWTH-FACTOR RECEPTORS IN NORMAL AND TUMOROUS HUMAN THYROID TISSUES

    NARCIS (Netherlands)

    van der Laan, B.F.A.M.; FREEMAN, JL; ASA, SL

    A number of growth factors have been implicated as stimuli of thyroid cell proliferation; overexpression of these growth factors and/or their receptors may play a role in the growth of thyroid tumors. To determine if immunohistochemical detection of growth factors and/or their receptors correlates

  8. Changing the insulin receptor to possess insulin-like growth factor I ligand specificity

    International Nuclear Information System (INIS)

    Andersen, A.S.; Kjeldsen, T.; Wiberg, F.C.; Christensen, P.M.; Rasmussen, J.S.; Norris, K.; Moeller, K.B.; Moeller, N.P.H.

    1990-01-01

    To examine the role of the N-terminal part of the insulin-like growth factor I (IGF-I) receptor and insulin receptor in determining ligand specificity, the authors prepared an expression vector encoding a hybrid receptor where exon 1 (encoding the signal peptide and seven amino acids of the α-subunit), exon 2, and exon 3 of the insulin receptor were replaced with the corresponding IGF-I receptor cDNA (938 nucleotides). To allow direct quantitative comparison of the binding capabilities of this hybrid receptor with those of the human IGF-I receptor and the insulin receptor, all three receptors were expressed in baby hamster kidney (BHK) cells as soluble molecules and partially purified before characterization. The hybrid IGF-I/insulin receptor bound IGF-I with an affinity comparable to that of the wild-type IGF-I receptor. In contrast, the hybrid receptor no longer displayed high-affinity binding of insulin. These results directly demonstrate that it is possible to change the specificity of the insulin receptor to that of the IGF-I receptor and, furthermore, that the binding specificity for IGF-I is encoded within the nucleotide sequence from 135 to 938 of the IGF-I receptor cDNA. Since the hybrid receptor only bound insulin with low affinity, the insulin binding region is likely to be located within exons 2 and 3 of the insulin receptor

  9. Changing the insulin receptor to possess insulin-like growth factor I ligand specificity

    Energy Technology Data Exchange (ETDEWEB)

    Andersen, A.S.; Kjeldsen, T.; Wiberg, F.C.; Christensen, P.M.; Rasmussen, J.S.; Norris, K.; Moeller, K.B.; Moeller, N.P.H. (Biopharmaceuticals Div., Bagsvaerd (Denmark))

    1990-08-14

    To examine the role of the N-terminal part of the insulin-like growth factor I (IGF-I) receptor and insulin receptor in determining ligand specificity, the authors prepared an expression vector encoding a hybrid receptor where exon 1 (encoding the signal peptide and seven amino acids of the {alpha}-subunit), exon 2, and exon 3 of the insulin receptor were replaced with the corresponding IGF-I receptor cDNA (938 nucleotides). To allow direct quantitative comparison of the binding capabilities of this hybrid receptor with those of the human IGF-I receptor and the insulin receptor, all three receptors were expressed in baby hamster kidney (BHK) cells as soluble molecules and partially purified before characterization. The hybrid IGF-I/insulin receptor bound IGF-I with an affinity comparable to that of the wild-type IGF-I receptor. In contrast, the hybrid receptor no longer displayed high-affinity binding of insulin. These results directly demonstrate that it is possible to change the specificity of the insulin receptor to that of the IGF-I receptor and, furthermore, that the binding specificity for IGF-I is encoded within the nucleotide sequence from 135 to 938 of the IGF-I receptor cDNA. Since the hybrid receptor only bound insulin with low affinity, the insulin binding region is likely to be located within exons 2 and 3 of the insulin receptor.

  10. Argos inhibits epidermal growth factor receptor signalling by ligand sequestration.

    Science.gov (United States)

    Klein, Daryl E; Nappi, Valerie M; Reeves, Gregory T; Shvartsman, Stanislav Y; Lemmon, Mark A

    2004-08-26

    The epidermal growth factor receptor (EGFR) has critical functions in development and in many human cancers. During development, the spatial extent of EGFR signalling is regulated by feedback loops comprising both well-understood activators and less well-characterized inhibitors. In Drosophila melanogaster the secreted protein Argos functions as the only known extracellular inhibitor of EGFR, with clearly identified roles in multiple stages of development. Argos is only expressed when the Drosophila EGFR (DER) is activated at high levels, and downregulates further DER signalling. Although there is ample genetic evidence that Argos inhibits DER activation, the biochemical mechanism has not been established. Here we show that Argos inhibits DER signalling without interacting directly with the receptor, but instead by sequestering the DER-activating ligand Spitz. Argos binds tightly to the EGF motif of Spitz and forms a 1:1 (Spitz:Argos) complex that does not bind DER in vitro or at the cell surface. Our results provide an insight into the mechanism of Argos function, and suggest new strategies for EGFR inhibitor design.

  11. Nuclear tristetraprolin acts as a corepressor of multiple steroid nuclear receptors in breast cancer cells

    Directory of Open Access Journals (Sweden)

    Tonatiuh Barrios-García

    2016-06-01

    Full Text Available Tristetraprolin (TTP is a 34-kDa, zinc finger-containing factor that in mammalian cells acts as a tumor suppressor protein through two different mechanisms. In the cytoplasm TTP promotes the decay of hundreds of mRNAs encoding cell factors involved in inflammation, tissue invasion, and metastasis. In the cell nucleus TTP has been identified as a transcriptional corepressor of the estrogen receptor alpha (ERα, which has been associated to the development and progression of the majority of breast cancer tumors. In this work we report that nuclear TTP modulates the transactivation activity of progesterone receptor (PR, glucocorticoid receptor (GR and androgen receptor (AR. In recent years these steroid nuclear receptors have been shown to be of clinical and therapeutical relevance in breast cancer. The functional association between TTP and steroid nuclear receptors is supported by the finding that TTP physically interacts with ERα, PR, GR and AR in vivo. We also show that TTP overexpression attenuates the transactivation of all the steroid nuclear receptors tested. In contrast, siRNA-mediated reduction of endogenous TTP expression in MCF-7 cells produced an increase in the transcriptional activities of ERα, PR, GR and AR. Taken together, these results suggest that the function of nuclear TTP in breast cancer cells is to act as a corepressor of ERα, PR, GR and AR. We propose that the reduction of TTP expression observed in different types of breast cancer tumors may contribute to the development of this disease by producing a dysregulation of the transactivation activity of multiple steroid nuclear receptors.

  12. NLRC5/MHC class I transactivator is a target for immune evasion in cancer.

    Science.gov (United States)

    Yoshihama, Sayuri; Roszik, Jason; Downs, Isaac; Meissner, Torsten B; Vijayan, Saptha; Chapuy, Bjoern; Sidiq, Tabasum; Shipp, Margaret A; Lizee, Gregory A; Kobayashi, Koichi S

    2016-05-24

    Cancer cells develop under immune surveillance, thus necessitating immune escape for successful growth. Loss of MHC class I expression provides a key immune evasion strategy in many cancers, although the molecular mechanisms remain elusive. MHC class I transactivator (CITA), known as "NLRC5" [NOD-like receptor (NLR) family, caspase recruitment (CARD) domain containing 5], has recently been identified as a critical transcriptional coactivator of MHC class I gene expression. Here we show that the MHC class I transactivation pathway mediated by CITA/NLRC5 constitutes a target for cancer immune evasion. In all the 21 tumor types we examined, NLRC5 expression was highly correlated with the expression of MHC class I, with cytotoxic T-cell markers, and with genes in the MHC class I antigen-presentation pathway, including LMP2/LMP7, TAP1, and β2-microglobulin. Epigenetic and genetic alterations in cancers, including promoter methylation, copy number loss, and somatic mutations, were most prevalent in NLRC5 among all MHC class I-related genes and were associated with the impaired expression of components of the MHC class I pathway. Strikingly, NLRC5 expression was significantly associated with the activation of CD8(+) cytotoxic T cells and patient survival in multiple cancer types. Thus, NLRC5 constitutes a novel prognostic biomarker and potential therapeutic target of cancers.

  13. NLRC5/MHC class I transactivator is a target for immune evasion in cancer

    Science.gov (United States)

    Yoshihama, Sayuri; Roszik, Jason; Downs, Isaac; Meissner, Torsten B.; Vijayan, Saptha; Chapuy, Bjoern; Sidiq, Tabasum; Shipp, Margaret A.; Lizee, Gregory A.; Kobayashi, Koichi S.

    2016-01-01

    Cancer cells develop under immune surveillance, thus necessitating immune escape for successful growth. Loss of MHC class I expression provides a key immune evasion strategy in many cancers, although the molecular mechanisms remain elusive. MHC class I transactivator (CITA), known as “NLRC5” [NOD-like receptor (NLR) family, caspase recruitment (CARD) domain containing 5], has recently been identified as a critical transcriptional coactivator of MHC class I gene expression. Here we show that the MHC class I transactivation pathway mediated by CITA/NLRC5 constitutes a target for cancer immune evasion. In all the 21 tumor types we examined, NLRC5 expression was highly correlated with the expression of MHC class I, with cytotoxic T-cell markers, and with genes in the MHC class I antigen-presentation pathway, including LMP2/LMP7, TAP1, and β2-microglobulin. Epigenetic and genetic alterations in cancers, including promoter methylation, copy number loss, and somatic mutations, were most prevalent in NLRC5 among all MHC class I-related genes and were associated with the impaired expression of components of the MHC class I pathway. Strikingly, NLRC5 expression was significantly associated with the activation of CD8+ cytotoxic T cells and patient survival in multiple cancer types. Thus, NLRC5 constitutes a novel prognostic biomarker and potential therapeutic target of cancers. PMID:27162338

  14. Studies on Platelet-Derived Growth Factor Beta-Receptor and Hepatocyte Growth Factor Receptor c-met in Paracrine Interactions in Human Breast Cancer

    Science.gov (United States)

    1996-09-01

    specific expression, and transforming submandibular gland . Growth Factors 1994, 10:145- activity of the mouse met proto-oncogene. Cell Growth 151...Derived Growth Factor Beta-Receptor and Hepatocyte Growth Factor Receptor c-met in Paracrine Interactions in Human Breast Cancer PRINCIPAL INVESTIGATOR...Interactions in Human Breast DAMDI7-94-J-4407 Cancer 6. AUTHOR(S) Bruce E. Elliott, Ph.D. 7. PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) 8. PERFORMING

  15. Personalized Radiation Oncology: Epidermal Growth Factor Receptor and Other Receptor Tyrosine Kinase Inhibitors.

    Science.gov (United States)

    Higgins, Geoff S; Krause, Mechthild; McKenna, W Gillies; Baumann, Michael

    Molecular biomarkers are currently evaluated in preclinical and clinical studies in order to establish predictors for treatment decisions in radiation oncology. The receptor tyrosine kinases (RTK) are described in the following text. Among them, the most data are available for the epidermal growth factor receptor (EGFR) that plays a major role for prognosis of patients after radiotherapy, but seems also to be involved in mechanisms of radioresistance, specifically in repopulation of tumour cells between radiotherapy fractions. Monoclonal antibodies against the EGFR improve locoregional tumour control and survival when applied during radiotherapy, however, the effects are heterogeneous and biomarkers for patient selection are warranted. Also other RTK´s such as c-Met and IGF-1R seem to play important roles in tumour radioresistance. Beside the potential to select patients for molecular targeting approaches combined with radiotherapy, studies are also needed to evluate radiotherapy adaptation approaches for selected patients, i.e. adaptation of radiation dose, or, more sophisticated, of target volumes.

  16. Full trans-activation mediated by the immediate-early protein of equine herpesvirus 1 requires a consensus TATA box, but not its cognate binding sequence.

    Science.gov (United States)

    Kim, Seong K; Shakya, Akhalesh K; O'Callaghan, Dennis J

    2016-01-04

    The immediate-early protein (IEP) of equine herpesvirus 1 (EHV-1) has extensive homology to the IEP of alphaherpesviruses and possesses domains essential for trans-activation, including an acidic trans-activation domain (TAD) and binding domains for DNA, TFIIB, and TBP. Our data showed that the IEP directly interacted with transcription factor TFIIA, which is known to stabilize the binding of TBP and TFIID to the TATA box of core promoters. When the TATA box of the EICP0 promoter was mutated to a nonfunctional TATA box, IEP-mediated trans-activation was reduced from 22-fold to 7-fold. The IEP trans-activated the viral promoters in a TATA motif-dependent manner. Our previous data showed that the IEP is able to repress its own promoter when the IEP-binding sequence (IEBS) is located within 26-bp from the TATA box. When the IEBS was located at 100 bp upstream of the TATA box, IEP-mediated trans-activation was very similar to that of the minimal IE(nt -89 to +73) promoter lacking the IEBS. As the distance from the IEBS to the TATA box decreased, IEP-mediated trans-activation progressively decreased, indicating that the IEBS located within 100 bp from the TATA box sequence functions as a distance-dependent repressive element. These results indicated that IEP-mediated full trans-activation requires a consensus TATA box of core promoters, but not its binding to the cognate sequence (IEBS). Copyright © 2015 Elsevier B.V. All rights reserved.

  17. The F-BAR protein PACSIN2 regulates epidermal growth factor receptor internalization

    NARCIS (Netherlands)

    B.J. de Kreuk (Bart-Jan); E.C. Anthony (Eloise); D. Geertss (Dirk); P.L. Hordijk (Peter )

    2012-01-01

    textabstractSignaling via growth factor receptors, including the epidermal growth factor (EGF) receptor, is key to various cellular processes, such as proliferation, cell survival, and cell migration. In a variety of human diseases such as cancer, aberrant expression and activation of growth factor

  18. Nod factor receptors form heteromeric complexes and are essential for intracellular infection in Medicago nodules

    NARCIS (Netherlands)

    Moling, S.; Pietraszewska-Bogiel, A.; Postma, M.; Fedorova, E.E.; Hink, M.A.; Limpens, E.H.M.; Gadella, T.W.J.; Bisseling, T.

    2014-01-01

    Rhizobial Nod factors are the key signaling molecules in the legume-rhizobium nodule symbiosis. In this study, the role of the Nod factor receptors NOD FACTOR PERCEPTION (NFP) and LYSIN MOTIF RECEPTOR-LIKE KINASE3 (LYK3) in establishing the symbiotic interface in root nodules was investigated. It

  19. Nod factor receptors form heteromeric complexes and are essential for intracellular infection in medicago nodules

    NARCIS (Netherlands)

    Moling, S.; Pietraszewska-Bogiel, A.; Postma, M.; Fedorova, E.; Hink, M.A.; Limpens, E.; Gadella, T.W.J.; Bisseling, T.

    2014-01-01

    Rhizobial Nod factors are the key signaling molecules in the legume-rhizobium nodule symbiosis. In this study, the role of the Nod factor receptors NOD FACTOR PERCEPTION (NFP) and LYSIN MOTIF RECEPTOR-LIKE KINASE3 (LYK3) in establishing the symbiotic interface in root nodules was investigated. It

  20. Arctigenin induced gallbladder cancer senescence through modulating epidermal growth factor receptor pathway.

    Science.gov (United States)

    Zhang, Mingdi; Cai, Shizhong; Zuo, Bin; Gong, Wei; Tang, Zhaohui; Zhou, Di; Weng, Mingzhe; Qin, Yiyu; Wang, Shouhua; Liu, Jun; Ma, Fei; Quan, Zhiwei

    2017-05-01

    Gallbladder cancer has poor prognosis and limited therapeutic options. Arctigenin, a representative dibenzylbutyrolactone lignan, occurs in a variety of plants. However, the molecular mechanisms involved in the antitumor effect of arctigenin on gallbladder cancer have not been fully elucidated. The expression levels of epidermal growth factor receptor were examined in 100 matched pairs of gallbladder cancer tissues. A positive correlation between high epidermal growth factor receptor expression levels and poor prognosis was observed in gallbladder cancer tissues. Pharmacological inhibition or inhibition via RNA interference of epidermal growth factor receptor induced cellular senescence in gallbladder cancer cells. The antitumor effect of arctigenin on gallbladder cancer cells was primarily achieved by inducing cellular senescence. In gallbladder cancer cells treated with arctigenin, the expression level of epidermal growth factor receptor significantly decreased. The analysis of the activity of the kinases downstream of epidermal growth factor receptor revealed that the RAF-MEK-ERK signaling pathway was significantly inhibited. Furthermore, the cellular senescence induced by arctigenin could be reverted by pcDNA-epidermal growth factor receptor. Arctigenin also potently inhibited the growth of tumor xenografts, which was accompanied by the downregulation of epidermal growth factor receptor and induction of senescence. This study demonstrates arctigenin could induce cellular senescence in gallbladder cancer through the modulation of epidermal growth factor receptor pathway. These data identify epidermal growth factor receptor as a key regulator in arctigenin-induced gallbladder cancer senescence.

  1. Analysis of receptor signaling pathways by mass spectrometry: identification of vav-2 as a substrate of the epidermal and platelet-derived growth factor receptors

    DEFF Research Database (Denmark)

    Pandey, A; Podtelejnikov, A V; Blagoev, B

    2000-01-01

    Oligomerization of receptor protein tyrosine kinases such as the epidermal growth factor receptor (EGFR) by their cognate ligands leads to activation of the receptor. Transphosphorylation of the receptor subunits is followed by the recruitment of signaling molecules containing src homology 2 (SH2...

  2. Development of real-time reverse transcription polymerase chain reaction assays to quantify insulin-like growth factor receptor and insulin receptor expression in equine tissue

    Directory of Open Access Journals (Sweden)

    Stephen B. Hughes

    2013-08-01

    Full Text Available The insulin-like growth factor system (insulin-like growth factor 1, insulin-like growth factor 2, insulin-like growth factor 1 receptor, insulin-like growth factor 2 receptor and six insulin-like growth factor-binding proteins and insulin are essential to muscle metabolism and most aspects of male and female reproduction. Insulin-like growth factor and insulin play important roles in the regulation of cell growth, differentiation and the maintenance of cell differentiation in mammals. In order to better understand the local factors that regulate equine physiology, such as muscle metabolism and reproduction (e.g., germ cell development and fertilisation, real-time reverse transcription polymerase chain reaction assays for quantification of equine insulin-like growth factor 1 receptor and insulin receptor messenger ribonucleic acid were developed. The assays were sensitive: 192 copies/µLand 891 copies/µL for insulin-like growth factor 1 receptor, messenger ribonucleic acid and insulin receptor respectively (95%limit of detection, and efficient: 1.01 for the insulin-like growth factor 1 receptor assay and 0.95 for the insulin receptor assay. The assays had a broad linear range of detection (seven logs for insulin-like growth factor 1 receptor and six logs for insulin receptor. This allowed for analysis of very small amounts of messenger ribonucleic acid. Low concentrations of both insulin-like growth factor 1 receptor and insulin receptor messenger ribonucleic acid were detected in endometrium, lung and spleen samples, whilst high concentrations were detected in heart, muscle and kidney samples, this was most likely due to the high level of glucose metabolism and glucose utilisation by these tissues. The assays developed for insulin-like growth factor 1 receptor and insulin receptor messenger ribonucleic acid expression have been shown to work on equine tissue and will contribute to the understanding of insulin and insulin-like growth factor 1

  3. HTLV-1 subgroups associated with the risk of HAM/TSP are related to viral and host gene expression in peripheral blood mononuclear cells, independent of the transactivation functions of the viral factors.

    Science.gov (United States)

    Yasuma, Keiko; Matsuzaki, Toshio; Yamano, Yoshihisa; Takashima, Hiroshi; Matsuoka, Masao; Saito, Mineki

    2016-08-01

    Among human T cell leukemia virus type 1 (HTLV-1)-infected individuals, the risk of developing HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) across lifetime differs between ethnic groups. There is an association between HTLV-1 tax gene subgroups (subgroup-A or subgroup-B) and the risk of HAM/TSP in the Japanese population. In this study, we investigated the full-length proviral genome sequences of various HTLV-1-infected cell lines and patient samples. The functional differences in the viral transcriptional regulators Tax and HTLV-1 bZIP factor (HBZ) between each subgroup and the relationships between subgroups and the clinical and laboratory characteristics of HAM/TSP patients were evaluated. The results of these analyses indicated the following: (1) distinct nucleotide substitutions corresponding to each subgroup were associated with nucleotide substitutions in viral structural, regulatory, and accessory genes; (2) the HBZ messenger RNA (mRNA) expression in HTLV-1-infected cells was significantly higher in HAM/TSP patients with subgroup-B than in those with subgroup-A; (3) a positive correlation was observed between the expression of HBZ mRNA and its target Foxp3 mRNA in HAM/TSP patients with subgroup-B, but not in patients with subgroup-A; (4) no clear differences were noted in clinical and laboratory characteristics between HAM/TSP patients with subgroup-A and subgroup-B; and (5) no functional differences were observed in Tax and HBZ between each subgroup based on reporter gene assays. Our results indicate that although different HTLV-1 subgroups are characterized by different patterns of viral and host gene expression in HAM/TSP patients via independent mechanisms of direct transcriptional regulation, these differences do not significantly affect the clinical and laboratory characteristics of HAM/TSP patients.

  4. The MHC-II transactivator CIITA inhibits Tat function and HIV-1 replication in human myeloid cells.

    Science.gov (United States)

    Forlani, Greta; Turrini, Filippo; Ghezzi, Silvia; Tedeschi, Alessandra; Poli, Guido; Accolla, Roberto S; Tosi, Giovanna

    2016-04-18

    We previously demonstrated that the HLA class II transactivator CIITA inhibits HIV-1 replication in T cells by competing with the viral transactivator Tat for the binding to Cyclin T1 subunit of the P-TEFb complex. Here, we analyzed the anti-viral function of CIITA in myeloid cells, another relevant HIV-1 target cell type. We sinvestigated clones of the U937 promonocytic cell line, either permissive (Plus) or non-permissive (Minus) to HIV-1 replication. This different phenotype has been associated with the expression of TRIM22 in U937 Minus but not in Plus cells. U937 Plus cells stably expressing CIITA were generated and HLA-II positive clones were selected by cell sorting and cloning. HLA and CIITA proteins were analyzed by cytofluorometry and western blotting, respectively. HLA-II DR and CIITA mRNAs were quantified by qRT-PCR. Tat-dependent transactivation was assessed by performing the HIV-1 LTR luciferase gene reporter assay. Cells were infected with HIV-1 and viral replication was evaluated by measuring the RT activity in culture supernatants. CIITA was expressed only in HLA-II-positive U937 Minus cells, and this was strictly correlated with inhibition of Tat-dependent HIV-1 LTR transactivation in Minus but not in Plus cells. Overexpression of CIITA in Plus cells restored the suppression of Tat transactivation, confirming the inhibitory role of CIITA. Importantly, HIV-1 replication was significantly reduced in Plus-CIITA cells with respect to Plus parental cells. This effect was independent of TRIM22 as CIITA did not induce TRIM22 expression in Plus-CIITA cells. U937 Plus and Minus cells represent an interesting model to study the role of CIITA in HIV-1 restriction in the monocytic/macrophage cell lineage. The differential expression of CIITA in CIITA-negative Plus and CIITA-positive Minus cells correlated with their capacity to support or not HIV-1 replication, respectively. In Minus cells CIITA targeted the viral transactivator Tat to inhibit HIV-1

  5. Neural cell adhesion molecule differentially interacts with isoforms of the fibroblast growth factor receptor

    DEFF Research Database (Denmark)

    Christensen, Claus; Berezin, Vladimir; Bock, Elisabeth

    2011-01-01

    The fibroblast growth factor receptor (FGFR) can be activated through direct interactions with various fibroblast growth factors or through a number of cell adhesion molecules, including the neural cell adhesion molecule (NCAM). We produced recombinant proteins comprising the ligand...

  6. The Pseudo signal peptide of the corticotropin-releasing factor receptor type 2A prevents receptor oligomerization.

    Science.gov (United States)

    Teichmann, Anke; Rutz, Claudia; Kreuchwig, Annika; Krause, Gerd; Wiesner, Burkhard; Schülein, Ralf

    2012-08-03

    N-terminal signal peptides mediate the interaction of native proteins with the translocon complex of the endoplasmic reticulum membrane and are cleaved off during early protein biogenesis. The corticotropin-releasing factor receptor type 2a (CRF(2(a))R) possesses an N-terminal pseudo signal peptide, which represents a so far unique domain within the large protein family of G protein-coupled receptors (GPCRs). In contrast to a conventional signal peptide, the pseudo signal peptide remains uncleaved and consequently forms a hydrophobic extension at the N terminus of the receptor. The functional consequence of the presence of the pseudo signal peptide is not understood. Here, we have analyzed the significance of this domain for receptor dimerization/oligomerization in detail. To this end, we took the CRF(2(a))R and the homologous corticotropin-releasing factor receptor type 1 (CRF(1)R) possessing a conventional cleaved signal peptide and conducted signal peptide exchange experiments. Using single cell and single molecule imaging methods (fluorescence resonance energy transfer and fluorescence cross-correlation spectroscopy, respectively) as well as biochemical experiments, we obtained two novel findings; we could show that (i) the CRF(2(a))R is expressed exclusively as a monomer, and (ii) the presence of the pseudo signal peptide prevents its oligomerization. Thus, we have identified a novel functional domain within the GPCR protein family, which plays a role in receptor oligomerization and which may be useful to study the functional significance of this process in general.

  7. Host factors that modify Plasmodium falciparum adhesion to endothelial receptors.

    Science.gov (United States)

    Mahamar, Almahamoudou; Attaher, Oumar; Swihart, Bruce; Barry, Amadou; Diarra, Bacary S; Kanoute, Moussa B; Cisse, Kadidia B; Dembele, Adama B; Keita, Sekouba; Gamain, Benoît; Gaoussou, Santara; Issiaka, Djibrilla; Dicko, Alassane; Duffy, Patrick E; Fried, Michal

    2017-10-24

    P. falciparum virulence is related to adhesion and sequestration of infected erythrocytes (IE) in deep vascular beds, but the endothelial receptors involved in severe malaria remain unclear. In the largest ever study of clinical isolates, we surveyed adhesion of freshly collected IE from children under 5 years of age in Mali to identify novel vascular receptors, and examined the effects of host age, hemoglobin type, blood group and severe malaria on levels of IE adhesion to a panel of endothelial receptors. Several novel molecules, including integrin α3β1, VE-cadherin, ICAM-2, junctional adhesion molecule-B (JAM-B), laminin, and cellular fibronectin, supported binding of IE from children. Severe malaria was not significantly associated with levels of IE adhesion to any of the 19 receptors. Hemoglobin AC, which reduces severe malaria risk, reduced IE binding to the receptors CD36 and integrin α5β1, while hemoglobin AS did not modify IE adhesion to any receptors. Blood groups A, AB and B significantly reduced IE binding to ICAM-1. Severe malaria risk varies with age, but age significantly impacted the level of IE binding to only a few receptors: IE binding to JAM-B decreased with age, while binding to CD36 and integrin α5β1 significantly increased with age.

  8. Physical and functional interactions between nuclear receptor LXRα and the forkhead box transcription factor FOXA2 regulate the response of the human lipoprotein lipase gene to oxysterols in hepatic cells.

    Science.gov (United States)

    Kanaki, Maria; Tiniakou, Ioanna; Thymiakou, Efstathia; Kardassis, Dimitris

    2017-08-01

    Lipoprotein lipase (LPL) catalyzes the hydrolysis of triglycerides from triglyceride-rich lipoproteins such as VLDL and chylomicrons in the circulation. Mutations in LPL or its activator apolipoprotein C-II cause hypertriglyceridemia in humans and animal models. The levels of LPL in the liver are low but they can be strongly induced by a high cholesterol diet or by synthetic ligands of Liver X Receptors (LXRs). However, the mechanism by which LXRs activate the human LPL gene is unknown. In the present study we show that LXR agonists increased the mRNA and protein levels as well as the promoter activity of human LPL in HepG2 cells. A promoter deletion analysis defined the proximal -109/-28 region, which contains a functional FOXA2 element, as essential for transactivation by ligand-activated LXRα/RXRα heterodimers. Silencing of endogenous FOXA2 in HepG2 cells by siRNAs or by treatment with insulin compromised the induction of the LPL gene by LXR agonists whereas mutations in the FOXA2 site abolished the synergistic transactivation of the LPL promoter by LXRα/RXRα and FOXA2. Physical and functional interactions between LXRα and FOXA2 were established in vitro and ex vivo. In summary, the present study revealed a novel mechanism of human LPL gene induction by oxysterols in the liver with is based on physical and functional interactions between transcription factors LXRα and FOXA2. This mechanism, which may not be restricted to the LPL gene, is critically important for a better understanding of the regulation of cholesterol and triglyceride metabolism in the liver under healthy or pathological states. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Muscarinic Receptor Signaling in Colon Cancer

    Energy Technology Data Exchange (ETDEWEB)

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

    2011-03-02

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

  10. Muscarinic Receptor Signaling in Colon Cancer

    International Nuclear Information System (INIS)

    Rosenvinge, Erik C. von; Raufman, Jean-Pierre

    2011-01-01

    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

  11. Muscarinic Receptor Signaling in Colon Cancer

    Directory of Open Access Journals (Sweden)

    Jean-Pierre Raufman

    2011-03-01

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

  12. Human corpus luteum: presence of epidermal growth factor receptors and binding characteristics

    International Nuclear Information System (INIS)

    Ayyagari, R.R.; Khan-Dawood, F.S.

    1987-01-01

    Epidermal growth factor receptors are present in many reproductive tissues but have not been demonstrated in the human corpus luteum. To determine the presence of epidermal growth factor receptors and its binding characteristics, we carried out studies on the plasma cell membrane fraction of seven human corpora lutea (days 16 to 25) of the menstrual cycle. Specific epidermal growth factor receptors were present in human corpus luteum. Insulin, nerve growth factor, and human chorionic gonadotropin did not competitively displace epidermal growth factor binding. The optimal conditions for corpus luteum-epidermal growth factor receptor binding were found to be incubation for 2 hours at 4 degrees C with 500 micrograms plasma membrane protein and 140 femtomol 125 I-epidermal growth factor per incubate. The number (mean +/- SEM) of epidermal growth factor binding sites was 12.34 +/- 2.99 X 10(-19) mol/micrograms protein; the dissociation constant was 2.26 +/- 0.56 X 10(-9) mol/L; the association constant was 0.59 +/- 0.12 X 10(9) L/mol. In two regressing corpora lutea obtained on days 2 and 3 of the menstrual cycle, there was no detectable specific epidermal growth factor receptor binding activity. Similarly no epidermal growth factor receptor binding activity could be detected in ovarian stromal tissue. Our findings demonstrate that specific receptors for epidermal growth factor are present in the human corpus luteum. The physiologic significance of epidermal growth factor receptors in human corpus luteum is unknown, but epidermal growth factor may be involved in intragonadal regulation of luteal function

  13. Interaction of the αβ dimers of the insulin-like growth factor I receptor required for receptor autophosphorylation

    International Nuclear Information System (INIS)

    Tollesfsen, S.E.; Stoszek, R.M.; Thompson, K.

    1991-01-01

    The authors have recently found that association of the two αβ dimers of the insulin-like growth factor I (IGF I) receptor is required for formation of a high-affinity binding site for IGF I. To determine the structural requirements for IGF I activated kinase activity, they have examined the effect of dissociation of the two αβ dimers of the IGF I receptor on β subunit autophosphorylation. The αβ dimers formed after treatment with 2 mM dithiothreitol (DTT) at pH 8.75 for 5 min were separated from IGF I receptor remaining as tetramers after DTT treatment by fast protein liquid chromatography on a Superose 6 gel filtration column. Purification of the αβ dimers was confirmed by Western blot analysis using 125 I-labeled αIR-3, a monoclonal antibody to the IGF I receptor. Autophosphorylation of the IGF I receptor (αβ) 2 tetramer, treated without DTT or remaining after DTT treatment, is stimulated 1.6-2.9-fold by IGF I. In contrast, autophosporylation of the αβ dimers incubated in the presence or absence of IGF I (100 ng/mL) does not occur. Both IGF I receptor dimers and tetramers exhibit similar kinase activities using the synthetic substrate Arg-Arg-Leu-Ile-Glu-Asp-Ala-Glu-Tyr-Ala-Ala-Arg-Gly, indicating that the failure to detect autophosphorylation of the IGF I receptor dimers does not result from inactivation of the kinase by DTT treatment. They conclude that autophosphorylation of the IGF I receptor depends upon the interaction of the two αβ dimers

  14. Neural stem cells express melatonin receptors and neurotrophic factors: colocalization of the MT1 receptor with neuronal and glial markers

    Directory of Open Access Journals (Sweden)

    McMillan Catherine R

    2004-10-01

    Full Text Available Abstract Background In order to optimize the potential benefits of neural stem cell (NSC transplantation for the treatment of neurodegenerative disorders, it is necessary to understand their biological characteristics. Although neurotrophin transduction strategies are promising, alternative approaches such as the modulation of intrinsic neurotrophin expression by NSCs, could also be beneficial. Therefore, utilizing the C17.2 neural stem cell line, we have examined the expression of selected neurotrophic factors under different in vitro conditions. In view of recent evidence suggesting a role for the pineal hormone melatonin in vertebrate development, it was also of interest to determine whether its G protein-coupled MT1 and MT2 receptors are expressed in NSCs. Results RT-PCR analysis revealed robust expression of glial cell-line derived neurotrophic factor (GDNF, brain-derived neurotrophic factor (BDNF and nerve growth factor (NGF in undifferentiated cells maintained for two days in culture. After one week, differentiating cells continued to exhibit high expression of BDNF and NGF, but GDNF expression was lower or absent, depending on the culture conditions utilized. Melatonin MT1 receptor mRNA was detected in NSCs maintained for two days in culture, but the MT2 receptor was not seen. An immature MT1 receptor of about 30 kDa was detected by western blotting in NSCs cultured for two days, whereas a mature receptor of about 40 – 45 kDa was present in cells maintained for longer periods. Immunocytochemical studies demonstrated that the MT1 receptor is expressed in both neural (β-tubulin III positive and glial (GFAP positive progenitor cells. An examination of the effects of melatonin on neurotrophin expression revealed that low physiological concentrations of this hormone caused a significant induction of GDNF mRNA expression in NSCs following treatment for 24 hours. Conclusions The phenotypic characteristics of C17.2 cells suggest that they are

  15. Hepatocyte and keratinocyte growth factors and their receptors in human lung emphysema

    Directory of Open Access Journals (Sweden)

    Marchal Joëlle

    2005-10-01

    Full Text Available Abstract Background Hepatocyte and keratinocyte growth factors are key growth factors in the process of alveolar repair. We hypothesized that excessive alveolar destruction observed in lung emphysema involves impaired expression of hepatocyte and keratinocyte growth factors or their respective receptors, c-met and keratinocyte growth factor receptor. The aim of our study was to compare the expression of hepatocyte and keratinocyte growth factors and their receptors in lung samples from 3 groups of patients: emphysema; smokers without emphysema and non-smokers without emphysema. Methods Hepatocyte and keratinocyte growth factor proteins were analysed by immunoassay and western blot; mRNA expression was measured by real time quantitative polymerase chain reaction. Results Hepatocyte and keratinocyte growth factors, c-met and keratinocyte growth factor receptor mRNA levels were similar in emphysema and non-emphysema patients. Hepatocyte growth factor mRNA correlated negatively with FEV1 and the FEV1/FVC ratio both in emphysema patients and in smokers with or without emphysema. Hepatocyte and keratinocyte growth factor protein concentrations were similar in all patients' groups. Conclusion The expression of hepatocyte and keratinocyte growth factors and their receptors is preserved in patients with lung emphysema as compared to patients without emphysema. Hepatocyte growth factor mRNA correlates with the severity of airflow obstruction in smokers.

  16. Multistep change in epidermal growth factor receptors during spontaneous neoplastic progression in Chinese hamster embryo fibroblasts

    International Nuclear Information System (INIS)

    Wakshull, E.; Kraemer, P.M.; Wharton, W.

    1985-01-01

    Whole Chinese hamster embryo lineages have been shown to undergo multistep spontaneous neoplastic progression during serial passage in culture. The authors have studied the binding, internalization, and degradation of 125 I-labeled epidermal growth factor at four different stages of transformation. The whole Chinese hamster embryo cells lost cell surface epidermal growth factor receptors gradually during the course of neoplastic progression until only 10% of the receptor number present in the early-passage cells (precrisis) were retained in the late-passage cells (tumorigenic). No differences in internalization rates, chloroquine sensitivity, or ability to degrade hormone between the various passage levels were seen. No evidence for the presence in conditioned medium of transforming growth factors which might mask or down-regulate epidermal growth factor receptor was obtained. These results suggest that a reduction in cell surface epidermal growth factor receptor might be an early event during spontaneous transformation in whole Chinese hamster embryo cells

  17. Association of coatomer proteins with the beta-receptor for platelet-derived growth factor

    DEFF Research Database (Denmark)

    Hansen, Klaus; Rönnstrand, L; Rorsman, C

    1997-01-01

    The nonreceptor tyrosine kinase Src binds to and is activated by the beta-receptor for platelet-derived growth factor (PDGF). The interaction leads to Src phosphorylation of Tyr934 in the kinase domain of the receptor. In the course of the functional characterization of this phosphorylation, we...

  18. A receptor model for urban aerosols based on oblique factor analysis

    DEFF Research Database (Denmark)

    Keiding, Kristian; Sørensen, Morten S.; Pind, Niels

    1987-01-01

    A procedure is outlined for the construction of receptor models of urban aerosols, based on factor analysis. The advantage of the procedure is that the covariation of source impacts is included in the construction of the models. The results are compared with results obtained by other receptor...

  19. A dual immunocytochemical assay for oestrogen and epidermal growth factor receptors in tumour cell lines

    NARCIS (Netherlands)

    A.K. Sharma (Anisha K.); J.H. Horgan; R.L. McClelland (Robyn); A.G. Douglas-Jones (A.); T. van Agthoven (Ton); L.C.J. Dorssers (Lambert); R.I. Nicholson (R.)

    1994-01-01

    textabstractA new dual immunocytochemical assay for oestrogen receptor (ER) and epidermal growth factor receptor (EGFR) has been developed. It has been tested in a variety of conditions using cell culture lines and the results correlate well with those obtained from single immunocytochemical assays.

  20. Expression of growth factor receptors and targeting of EGFR in cholangiocarcinoma cell lines

    International Nuclear Information System (INIS)

    Xu, Ling; Hausmann, Martin; Dietmaier, Wolfgang; Kellermeier, Silvia; Pesch, Theresa; Stieber-Gunckel, Manuela; Lippert, Elisabeth; Klebl, Frank; Rogler, Gerhard

    2010-01-01

    Cholangiocarcinoma (CC) is a malignant neoplasm of the bile ducts or the gallbladder. Targeting of growth factor receptors showed therapeutic potential in palliative settings for many solid tumors. The aim of this study was to determine the expression of seven growth factor receptors in CC cell lines and to assess the effect of blocking the EGFR receptor in vitro. Expression of EGFR (epithelial growth factor receptor), HGFR (hepatocyte growth factor receptor) IGF1R (insulin-like growth factor 1 receptor), IGF2R (insulin-like growth factor 2 receptor) and VEGFR1-3 (vascular endothelial growth factor receptor 1-3) were examined in four human CC cell lines (EGI-1, HuH28, OZ and TFK-1). The effect of the anti-EGFR-antibody cetuximab on cell growth and apoptosis was studied and cell lines were examined for KRAS mutations. EGFR, HGFR and IGFR1 were present in all four cell lines tested. IGFR2 expression was confirmed in EGI-1 and TFK-1. No growth-inhibitory effect was found in EGI-1 cells after incubation with cetuximab. Cetuximab dose-dependently inhibited growth in TFK-1. Increased apoptosis was only seen in TFK-1 cells at the highest cetuximab dose tested (1 mg/ml), with no dose-response-relationship at lower concentrations. In EGI-1 a heterozygous KRAS mutation was found in codon 12 (c.35G>A; p.G12D). HuH28, OZ and TFK-1 lacked KRAS mutation. CC cell lines express a pattern of different growth receptors in vitro. Growth factor inhibitor treatment could be affected from the KRAS genotype in CC. The expression of EGFR itself does not allow prognoses on growth inhibition by cetuximab

  1. Activation of the epidermal growth factor receptor (EGFR) by a novel metalloprotease pathway.

    LENUS (Irish Health Repository)

    Bergin, David A

    2008-11-14

    Neutrophil Elastase (NE) is a pro-inflammatory protease present at higher than normal levels in the lung during inflammatory disease. NE regulates IL-8 production from airway epithelial cells and can activate both EGFR and TLR4. TACE\\/ADAM17 has been reported to trans-activate EGFR in response to NE. Here, using 16HBE14o-human bronchial epithelial cells we demonstrate a new mechanism by which NE regulates both of these events. A high molecular weight soluble metalloprotease activity detectable only in supernatants from NE-treated cells by gelatin and casein zymography was confirmed to be meprin alpha by Western immunoblotting. In vitro studies demonstrated the ability of NE to activate meprin alpha, which in turn could release soluble TGFalpha and induce IL-8 production from 16HBE14o- cells. These effects were abrogated by actinonin, a specific meprin inhibitor. NE-induced IL-8 expression was also inhibited by meprin alpha siRNA. Immunoprecipitation studies detected EGFR\\/TLR4 complexes in NE-stimulated cells overexpressing these receptors. Confocal studies confirmed colocalization of EGFR and TLR4 in 16HBE14o- cells stimulated with meprin alpha. NFkappaB was also activated via MyD88 in these cells by meprin alpha. In bronchoalveolar lavage fluid from NE knock-out mice infected intra-tracheally with Pseudomonas aeruginosa meprin alpha was significantly decreased compared with control mice, and was significantly increased and correlated with NE activity, in bronchoalveolar lavage fluid from individuals with cystic fibrosis but not healthy controls. The data describe a previously unidentified lung metalloprotease meprin alpha, and its role in NE-induced EGFR and TLR4 activation and IL-8 production.

  2. Human epidermal growth factor receptor (HER 2)/neu expression ...

    African Journals Online (AJOL)

    use

    2011-11-23

    Nov 23, 2011 ... receptor (HER 2)/neu and its clinical significance in colorectal cancer, in this study, clinicopathological data and paraffin-embedded specimen .... Correlation between the HER 2/neu protein expression, amplication and clinicopathologic feature in 192 colorectal cancer. Variable. HER2 protein expression. P.

  3. Expression of the epidermal growth factor receptor in human small cell lung cancer cell lines

    DEFF Research Database (Denmark)

    Damstrup, L; Rygaard, K; Spang-Thomsen, M

    1992-01-01

    Epidermal growth factor (EGF) receptor expression was evaluated in a panel of 21 small cell lung cancer cell lines with radioreceptor assay, affinity labeling, and Northern blotting. We found high-affinity receptors to be expressed in 10 cell lines. Scatchard analysis of the binding data...... lung cancer cell lines express the EGF receptor....... of EGF receptor mRNA in all 10 cell lines that were found to be EGF receptor-positive and in one cell line that was found to be EGF receptor-negative in the radioreceptor assay and affinity labeling. Our results provide, for the first time, evidence that a large proportion of a broad panel of small cell...

  4. Agonists of fibroblast growth factor receptor induce neurite outgrowth and survival of cerebellar granule neurons

    DEFF Research Database (Denmark)

    Li, Shizhong; Christensen, Claus; Køhler, Lene B

    2009-01-01

    Fibroblast growth factor receptor (FGFR) signaling is pivotal in the regulation of neurogenesis, neuronal differentiation and survival, and synaptic plasticity both during development and in adulthood. In order to develop low molecular weight agonists of FGFR, seven peptides, termed hexafins...

  5. Tumor necrosis factor receptor- associated factor 6 (TRAF6) regulation of development, function, and homeostasis of the immune system.

    Science.gov (United States)

    Walsh, Matthew C; Lee, JangEun; Choi, Yongwon

    2015-07-01

    Tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) is an adapter protein that mediates a wide array of protein-protein interactions via its TRAF domain and a RING finger domain that possesses non-conventional E3 ubiquitin ligase activity. First identified nearly two decades ago as a mediator of interleukin-1 receptor (IL-1R)-mediated activation of NFκB, TRAF6 has since been identified as an actor downstream of multiple receptor families with immunoregulatory functions, including members of the TNFR superfamily, the Toll-like receptor (TLR) family, tumor growth factorreceptors (TGFβR), and T-cell receptor (TCR). In addition to NFκB, TRAF6 may also direct activation of mitogen-activated protein kinase (MAPK), phosphoinositide 3-kinase (PI3K), and interferon regulatory factor pathways. In the context of the immune system, TRAF6-mediated signals have proven critical for the development, homeostasis, and/or activation of B cells, T cells, and myeloid cells, including macrophages, dendritic cells, and osteoclasts, as well as for organogenesis of thymic and secondary lymphoid tissues. In multiple cellular contexts, TRAF6 function is essential not only for proper activation of the immune system but also for maintaining immune tolerance, and more recent work has begun to identify mechanisms of contextual specificity for TRAF6, involving both regulatory protein interactions, and messenger RNA regulation by microRNAs. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  6. Tumor necrosis factor receptor associated factor 6 (TRAF6) regulation of development, function, and homeostasis of the immune system

    Science.gov (United States)

    Walsh, Matthew C.; Lee, JangEun; Choi, Yongwon

    2016-01-01

    Summary Tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) is an adaptor protein that mediates a wide array of protein-protein interactions via its TRAF domain and a RING finger domain that possesses non-conventional E3 ubiquitin ligase activity. First identified nearly two decades ago as a mediator of IL-1 receptor (IL-1R)-mediated activation of NFκB, TRAF6 has since been identified as an actor downstream of multiple receptor families with immunoregulatory functions, including members of the TNFR superfamily, the toll-like receptor (TLR) family, tumor growth factorreceptors (TGFβR), and T cell receptor (TCR). In addition to NFκB, TRAF6 may also direct activation of mitogen-activated protein kinase (MAPK), phosphoinositide 3-kinase (PI3K), and interferon regulatory factor (IRF) pathways. In the context of the immune system, TRAF6-mediated signals have proven critical for the development, homeostasis, and/or activation of B cells, T cells, and myeloid cells, including macrophages, dendritic cells, and osteoclasts, as well as for organogenesis of thymic and secondary lymphoid tissues. In multiple cellular contexts, TRAF6 function is essential not only for proper activation of the immune system, but also for maintaining immune tolerance, and more recent works have begun to identify mechanisms of contextual specificity for TRAF6, involving both regulatory protein interactions, and messenger RNA regulation by microRNAs. PMID:26085208

  7. Molecular and biochemical analyses of platelet-derived growth factor receptor (PDGFR) B, PDGFRA, and KIT receptors in chordomas.

    Science.gov (United States)

    Tamborini, Elena; Miselli, Francesca; Negri, Tiziana; Lagonigro, M Stefania; Staurengo, Samantha; Dagrada, Gian Paolo; Stacchiotti, Silvia; Pastore, Elisa; Gronchi, Alessandro; Perrone, Federica; Carbone, Antonino; Pierotti, Marco A; Casali, Paolo G; Pilotti, Silvana

    2006-12-01

    We have previously shown the presence of an activated platelet-derived growth factor (PDGF) receptor (PDGFR) B and its ligand PDGFB in a limited number of patients with clinical and radiological responses to imatinib mesylate treatment. This article describes the results of comprehensive molecular/biochemical analyses of the three receptors targeted by the drug (PDGFRB, PDGFRA, and KIT) in a series of 31 chordoma patients. The presence and activation status of PDGFRB, PDGFRA, and KIT receptors were investigated by means of immunoprecipitation and Western blot analyses complemented by immunohistochemistry, their expression level was analyzed by means of real-time PCR, and the occurrence of activating point mutations was investigated by means of cDNA sequencing. The PDGFB, PDGFA, and stem cell factor cognate ligands were investigated by reverse transcription-PCR, and gene status was assessed by fluorescence in situ hybridization. The results show that PDGFRB was highly expressed and phosphorylated, whereas PDGFRA and KIT were less expressed but phosphorylated and thus activated. These findings, together with the absence of gain-of-function mutations and the presence of the cognate ligands, strongly support the hypothesis that the activation mechanism is the autocrine/paracrine loop. No role seems to be played by gene amplification. In the light of our findings, the clinical benefit observed in chordoma patients treated with imatinib seems to be attributable to the switching off of all three receptors.

  8. The nerve growth factor and its receptors in airway inflammatory diseases.

    Science.gov (United States)

    Freund-Michel, V; Frossard, N

    2008-01-01

    The nerve growth factor (NGF) belongs to the neurotrophin family and induces its effects through activation of 2 distinct receptor types: the tropomyosin-related kinase A (TrkA) receptor, carrying an intrinsic tyrosine kinase activity in its intracellular domain, and the receptor p75 for neurotrophins (p75NTR), belonging to the death receptor family. Through activation of its TrkA receptor, NGF activates signalling pathways, including phospholipase Cgamma (PLCgamma), phosphatidyl-inositol 3-kinase (PI3K), the small G protein Ras, and mitogen-activated protein kinases (MAPK). Through its p75NTR receptor, NGF activates proapoptotic signalling pathways including the MAPK c-Jun N-terminal kinase (JNK), ceramides, and the small G protein Rac, but also activates pathways promoting cell survival through the transcription factor nuclear factor-kappaB (NF-kappaB). NGF was first described by Rita Levi-Montalcini and collaborators as an important factor involved in nerve differentiation and survival. Another role for NGF has since been established in inflammation, in particular of the airways, with increased NGF levels in chronic inflammatory diseases. In this review, we will first describe NGF structure and synthesis and NGF receptors and their signalling pathways. We will then provide information about NGF in the airways, describing its expression and regulation, as well as pointing out its potential role in inflammation, hyperresponsiveness, and remodelling process observed in airway inflammatory diseases, in particular in asthma.

  9. Increased concentrations of tumour necrosis factor (TNF) and soluble TNF receptors in biliary obstruction in mice; soluble TNF receptors as prognostic factors for mortality

    NARCIS (Netherlands)

    Bemelmans, M. H.; Greve, J. W.; Gouma, D. J.; Buurman, W. A.

    1996-01-01

    Systemic tumour necrosis factor (TNF) is present in jaundiced mice. Two soluble TNF receptors, sTNFr-P55 and sTNFr-P75 are reported to play a part in the natural defence against TNF. This study investigated the properties of circulating TNF and sTNFr in jaundiced mice. The data show that TNF in

  10. Bevacizumab reduces tumor targeting of antiepidermal growth factor and anti-insulin-like growth factor 1 receptor antibodies

    NARCIS (Netherlands)

    Heskamp, Sandra; Boerman, Otto C.; Molkenboer-Kuenen, Janneke D. M.; Oyen, Wim J. G.; van der Graaf, Winette T. A.; van Laarhoven, Hanneke W. M.

    2013-01-01

    Bevacizumab (antivascular endothelial growth factor [anti-VEGF]) and cetuximab (antiepidermal growth factor receptor [anti-EGFR]) are approved antibodies for treatment of cancer. However, in advanced colorectal cancer, the combination fails to improve survival. As the reason for the lack of activity

  11. Essential role of TEA domain transcription factors in the negative regulation of the MYH 7 gene by thyroid hormone and its receptors.

    Directory of Open Access Journals (Sweden)

    Hiroyuki Iwaki

    Full Text Available MYH7 (also referred to as cardiac myosin heavy chain β gene expression is known to be repressed by thyroid hormone (T3. However, the molecular mechanism by which T3 inhibits the transcription of its target genes (negative regulation remains to be clarified, whereas those of transcriptional activation by T3 (positive regulation have been elucidated in detail. Two MCAT (muscle C, A, and T sites and an A/T-rich region in the MYH7 gene have been shown to play a critical role in the expression of this gene and are known to be recognized by the TEAD/TEF family of transcription factors (TEADs. Using a reconstitution system with CV-1 cells, which has been utilized in the analysis of positive as well as negative regulation, we demonstrate that both T3 receptor (TR β1 and α1 inhibit TEAD-dependent activation of the MYH7 promoter in a T3 dose-dependent manner. TRβ1 bound with GC-1, a TRβ-selective T3 analog, also repressed TEAD-induced activity. Although T3-dependent inhibition required the DNA-binding domain (DBD of TRβ1, it remained after the putative negative T3-responsive elements were mutated. A co-immunoprecipitation study demonstrated the in vivo association of TRβ1 with TEAD-1, and the interaction surfaces were mapped to the DBD of the TRβ1 and TEA domains of TEAD-1, both of which are highly conserved among TRs and TEADs, respectively. The importance of TEADs in MYH7 expression was also validated with RNA interference using rat embryonic cardiomyocyte H9c2 cells. These results indicate that T3-bound TRs interfere with transactivation by TEADs via protein-protein interactions, resulting in the negative regulation of MYH7 promoter activity.

  12. Essential role of TEA domain transcription factors in the negative regulation of the MYH 7 gene by thyroid hormone and its receptors.

    Science.gov (United States)

    Iwaki, Hiroyuki; Sasaki, Shigekazu; Matsushita, Akio; Ohba, Kenji; Matsunaga, Hideyuki; Misawa, Hiroko; Oki, Yutaka; Ishizuka, Keiko; Nakamura, Hirotoshi; Suda, Takafumi

    2014-01-01

    MYH7 (also referred to as cardiac myosin heavy chain β) gene expression is known to be repressed by thyroid hormone (T3). However, the molecular mechanism by which T3 inhibits the transcription of its target genes (negative regulation) remains to be clarified, whereas those of transcriptional activation by T3 (positive regulation) have been elucidated in detail. Two MCAT (muscle C, A, and T) sites and an A/T-rich region in the MYH7 gene have been shown to play a critical role in the expression of this gene and are known to be recognized by the TEAD/TEF family of transcription factors (TEADs). Using a reconstitution system with CV-1 cells, which has been utilized in the analysis of positive as well as negative regulation, we demonstrate that both T3 receptor (TR) β1 and α1 inhibit TEAD-dependent activation of the MYH7 promoter in a T3 dose-dependent manner. TRβ1 bound with GC-1, a TRβ-selective T3 analog, also repressed TEAD-induced activity. Although T3-dependent inhibition required the DNA-binding domain (DBD) of TRβ1, it remained after the putative negative T3-responsive elements were mutated. A co-immunoprecipitation study demonstrated the in vivo association of TRβ1 with TEAD-1, and the interaction surfaces were mapped to the DBD of the TRβ1 and TEA domains of TEAD-1, both of which are highly conserved among TRs and TEADs, respectively. The importance of TEADs in MYH7 expression was also validated with RNA interference using rat embryonic cardiomyocyte H9c2 cells. These results indicate that T3-bound TRs interfere with transactivation by TEADs via protein-protein interactions, resulting in the negative regulation of MYH7 promoter activity.

  13. Altered [125I]epidermal growth factor binding and receptor distribution in psoriasis

    International Nuclear Information System (INIS)

    Nanney, L.B.; Stoscheck, C.M.; Magid, M.; King, L.E. Jr.

    1986-01-01

    Stimulation of growth and differentiation of human epidermis by epidermal growth factor (EGF) is mediated by its binding to specific receptors. Whether EGF receptors primarily mediate cell division or differentiation in hyperproliferative disease such as psoriasis vulgaris is unclear. To study the pathogenesis of psoriasis, 4-mm2 punch biopsy specimens of normal, uninvolved, and involved psoriatic skin were assayed for EGF receptors by autoradiographic, immunohistochemical, and biochemical methods. Using autoradiographic and immunohistochemical methods, basal keratinocytes were found to contain the greatest number of EGF binding sites and immunoreactive receptors as compared to the upper layers of the epidermis in both normal epidermis and psoriatic skin. No EGF receptor differences between normal and psoriatic epidermis were observed in this layer. In the upper layers of the epidermis, a 2-fold increase in EGF binding capacity was observed in psoriatic skin as compared with normal thin or thick skin. Biochemical methods indicated that [ 125 I]EGF binding was increased in psoriatic epidermis as compared with similar thickness normal epidermis when measured on a protein basis. Epidermal growth factor was shown to increase phosphorylation of the EGF receptor in skin. EGF receptors retained in the nonmitotic stratum spinosum and parakeratotic stratum corneum may reflect the incomplete, abnormal differentiation that occurs in active psoriatic lesions. Alternatively, retained EGF receptors may play a direct role in inhibiting cellular differentiation in the suprabasal layers

  14. Positron emission mammography: correlation of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 status and 18F-FDG.

    Science.gov (United States)

    Wang, Carolyn L; MacDonald, Lawrence R; Rogers, James V; Aravkin, Aleksandr; Haseley, David R; Beatty, J David

    2011-08-01

    The study objective was to assess the correlation between (18)F-FDG uptake values on positron emission mammography (PEM), expressed as maximum uptake value and lesion-to-background ratio, and receptor status (i.e., estrogen receptor [ER], progesterone receptor [PR], and human epidermal growth factor receptor 2 [HER2]), tumor histology, and tumor grade. We also evaluated for the correlation between maximum uptake value on PEM and maximum uptake value on a whole-body PET/CT. We retrospectively reviewed our database for patients with newly diagnosed breast cancer who were referred for PEM between June 2007 and September 2009. A subset of patients also underwent a whole-body PET/CT scan. The original pathology reports were reviewed to establish the histologic type, grade, and receptor status. The study involved 98 patients with 100 lesions. ER-negative tumors and PR-negative tumors had significantly higher mean lesion-to-background ratio than did their respective receptor-positive tumors (p = 0.02). Triple-negative tumors (i.e., ER-negative, PR-negative, and HER2-negative tumors) had statistically higher mean lesion-to-background ratio than did ER-positive PR-positive HER2-negative tumors (p = 0.04). Infiltrating ductal carcinomas had significantly higher PEM FDG uptake values than did infiltrating lobular carcinomas (p = 0.02-0.04). Breast tumors with higher histologic grade also had significantly higher PEM FDG uptake values than did those with lower grade (p = 0.03 and p PEM uptake values. This study shows a correlation between PEM FDG uptake values and the prognostic factors that have been shown to predict breast cancer survival.

  15. Insulin-like growth factor-II receptors in cultured rat hepatocytes: regulation by cell density

    International Nuclear Information System (INIS)

    Scott, C.D.; Baxter, R.C.

    1987-01-01

    Insulin-like growth factor-II (IGF-II) receptors in primary cultures of adult rat hepatocytes were characterized and their regulation by cell density examined. In hepatocytes cultured at 5 X 10(5) cells per 3.8 cm2 plate [ 125 I]IGF-II bound to specific, high affinity receptors (Ka = 4.4 +/- 0.5 X 10(9) l/mol). Less than 1% cross-reactivity by IGF-I and no cross-reactivity by insulin were observed. IGF-II binding increased when cells were permeabilized with 0.01% digitonin, suggesting the presence of an intracellular receptor pool. Determined by Scatchard analysis and by polyacrylamide gel electrophoresis after affinity labeling, the higher binding was due solely to an increase in binding sites present on 220 kDa type II IGF receptors. In hepatocytes cultured at low densities, the number of cell surface receptors increased markedly, from 10-20,000 receptors per cell at a culture density of 6 X 10(5) cells/well to 70-80,000 receptors per cell at 0.38 X 10(5) cells/well. The increase was not due simply to the exposure of receptors from the intracellular pool, as a density-related increase in receptors was also seen in cells permeabilized with digitonin. There was no evidence that IGF binding proteins, either secreted by hepatocytes or present in fetal calf serum, had any effect on the measurement of receptor concentration or affinity. We conclude that rat hepatocytes in primary culture contain specific IGF-II receptors and that both cell surface and intracellular receptors are regulated by cell density

  16. Computed Radiography Photostimulable Phosphor Plate Receptors: Effect on Grid Conversion Factors.

    Science.gov (United States)

    Noble, Lauren B; Culp, Melissa P

    2017-11-01

    Grid conversion factors were established for using grids with varying ratios for film-screen systems. Evidence from the physics community suggests that the relationship between grid conversion factors and exposure to the receptor does not apply with different types of digital imaging receptors. To evaluate the relationship among analog grid conversion factors when using various grid ratios with a photostimulable phosphor plate (PSP) receptor system. Three experiments were conducted using a range of grid ratios (6:1, 8:1, 12:1) to assess the effect of conversion factors on exposure indicators with a computed radiography PSP receptor. When using the analog grid conversion factors established for varying grid ratios with a PSP receptor and constant peak kilovoltage (kVp), the plate was overexposed with an exposure indicator outside the acceptable limits. The discussion of antiscatter grids in radiologic technologist literature should remain in the context of receptor type and digital imaging system being used so that technologists can make appropriate decisions about the use of grids while minimizing patient radiation dose. Calibrated grid conversions based on the digital systems in use and the radiologist's image quality preference could be created within radiology departments. ©2017 American Society of Radiologic Technologists.

  17. Molecular and functional characterization of pigeon (Columba livia) tumor necrosis factor receptor-associated factor 3.

    Science.gov (United States)

    Zhou, Yingying; Kang, Xilong; Xiong, Dan; Zhu, Shanshan; Zheng, Huijuan; Xu, Ying; Guo, Yaxin; Pan, Zhiming; Jiao, Xinan

    2017-04-01

    Tumor necrosis factor receptor-associated factor 3 (TRAF3) plays a key antiviral role by promoting type I interferon production. We cloned the pigeon TRAF3 gene (PiTRAF3) according to its predicted mRNA sequence to investigate its function. The 1704-bp full-length open reading frame encodes a 567-amino acid protein. One Ring finger, two TRAF-type Zinc fingers, one Coiled coil, and one MATH domain were inferred. RT-PCR showed that PiTRAF3 was expressed in all tissues, with relatively weak expression in the heart and liver. In HEK293T cells, over-expression of wild-type, △Ring, △Zinc finger, and △Coiled coil PiTRAF3, but not a △MATH form, significantly increased IFN-β promoter activity. Zinc finger and Coiled coil domains were essential for NF-κB activation. In chicken HD11 cells, PiTRAF3 increased IFN-β promoter activity and four domains were all contributing. R848 stimulation of pigeon peripheral blood mononuclear cells and splenocytes significantly increased expression of PiTRAF3 and the inflammatory cytokine genes CCL5, IL-8, and IL-10. These data demonstrate TRAF3's innate immune function and improve understanding of its involvement in poultry antiviral defense. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Distinct phosphotyrosines on a growth factor receptor bind to specific molecules that mediate different signaling pathways.

    Science.gov (United States)

    Fantl, W J; Escobedo, J A; Martin, G A; Turck, C W; del Rosario, M; McCormick, F; Williams, L T

    1992-05-01

    The receptor for platelet-derived growth factor (PDGF) binds two proteins containing SH2 domains, GTPase activating protein (GAP) and phosphatidylinositol 3-kinase (PI3-kinase). The sites on the receptor that mediate this interaction were identified by using phosphotyrosine-containing peptides representing receptor sequences to block specifically binding of either PI3-kinase or GAP. These results suggested that PI3-kinase binds two phosphotyrosine residues, each located in a 5 aa motif with an essential methionine at the fourth position C-terminal to the tyrosine. Point mutations at these sites caused a selective elimination of PI3-kinase binding and loss of PDGF-stimulated DNA synthesis. Mutation of the binding site for GAP prevented the receptor from associating with or phosphorylating GAP, but had no effect on PI3-kinase binding and little effect on DNA synthesis. Therefore, GAP and PI3-kinase interact with the receptor by binding to different phosphotyrosine-containing sequence motifs.

  19. Distribution and number of epidermal growth factor receptors in skin is related to epithelial cell growth

    DEFF Research Database (Denmark)

    Green, M R; Basketter, D A; Couchman, J R

    1983-01-01

    markedly with age. This decrease in receptor number is similar in trend to the known drop in basal cell [3H]thymidine labelling index which occurs over the same time period. The data suggest that the distribution of EGF receptors and EGF cell surface receptor number in skin are important in the spatial......, and keratinisation when injected into neonatal mice (S. Cohen and G.A. Elliott, 1963, J. Invest. Dermatol, 40, 1-5). We have determined the distribution of the available receptors for epidermal growth factor in rat skin using autoradiography following incubation of explants with 125I-labelled mouse EGF. EGF...... receptors are detected on the epithelial cells overlying the basement membranes of the epidermis, sebaceous gland, and regions of the hair follicle all of which have proliferative capacity. In marked contrast, tissues which have started to differentiate and lost their growth potential, carry either...

  20. Internalization Mechanisms of the Epidermal Growth Factor Receptor after Activation with Different Ligands

    Science.gov (United States)

    Henriksen, Lasse; Grandal, Michael Vibo; Knudsen, Stine Louise Jeppe; van Deurs, Bo; Grøvdal, Lene Melsæther

    2013-01-01

    The epidermal growth factor receptor (EGFR) regulates normal growth and differentiation, but dysregulation of the receptor or one of the EGFR ligands is involved in the pathogenesis of many cancers. There are eight ligands for EGFR, however most of the research into trafficking of the receptor after ligand activation focuses on the effect of epidermal growth factor (EGF) and transforming growth factor-α (TGF-α). For a long time it was believed that clathrin-mediated endocytosis was the major pathway for internalization of the receptor, but recent work suggests that different pathways exist. Here we show that clathrin ablation completely inhibits internalization of EGF- and TGF-α-stimulated receptor, however the inhibition of receptor internalization in cells treated with heparin-binding EGF-like growth factor (HB-EGF) or betacellulin (BTC) was only partial. In contrast, clathrin knockdown fully inhibits EGFR degradation after all ligands tested. Furthermore, inhibition of dynamin function blocked EGFR internalization after stimulation with all ligands. Knocking out a number of clathrin-independent dynamin-dependent pathways of internalization had no effect on the ligand-induced endocytosis of the EGFR. We suggest that EGF and TGF-α lead to EGFR endocytosis mainly via the clathrin-mediated pathway. Furthermore, we suggest that HB-EGF and BTC also lead to EGFR endocytosis via a clathrin-mediated pathway, but can additionally use an unidentified internalization pathway or better recruit the small amount of clathrin remaining after clathrin knockdown. PMID:23472148

  1. Internalization mechanisms of the epidermal growth factor receptor after activation with different ligands.

    Directory of Open Access Journals (Sweden)

    Lasse Henriksen

    Full Text Available The epidermal growth factor receptor (EGFR regulates normal growth and differentiation, but dysregulation of the receptor or one of the EGFR ligands is involved in the pathogenesis of many cancers. There are eight ligands for EGFR, however most of the research into trafficking of the receptor after ligand activation focuses on the effect of epidermal growth factor (EGF and transforming growth factor-α (TGF-α. For a long time it was believed that clathrin-mediated endocytosis was the major pathway for internalization of the receptor, but recent work suggests that different pathways exist. Here we show that clathrin ablation completely inhibits internalization of EGF- and TGF-α-stimulated receptor, however the inhibition of receptor internalization in cells treated with heparin-binding EGF-like growth factor (HB-EGF or betacellulin (BTC was only partial. In contrast, clathrin knockdown fully inhibits EGFR degradation after all ligands tested. Furthermore, inhibition of dynamin function blocked EGFR internalization after stimulation with all ligands. Knocking out a number of clathrin-independent dynamin-dependent pathways of internalization had no effect on the ligand-induced endocytosis of the EGFR. We suggest that EGF and TGF-α lead to EGFR endocytosis mainly via the clathrin-mediated pathway. Furthermore, we suggest that HB-EGF and BTC also lead to EGFR endocytosis via a clathrin-mediated pathway, but can additionally use an unidentified internalization pathway or better recruit the small amount of clathrin remaining after clathrin knockdown.

  2. Epidermal growth factor receptor: an independent predictor of survival in astrocytic tumors given definitive irradiation

    International Nuclear Information System (INIS)

    An Zhu; Shaeffer, James; Leslie, Susan; Kolm, Paul; El-Mahdi, Anas M.

    1996-01-01

    Purpose: To determine whether the expression of epidermal growth factor receptor (EGFR) protein was predictive of patient survival independently of other prognostic factors in astrocytic tumors. Methods and Materials: Epidermal growth factor receptor protein expression was investigated immunohistochemically in formalin-fixed, paraffin-embedded surgical specimens of 55 glioblastoma multiforme, 14 anaplastic astrocytoma, and 2 astrocytomas given definitive irradiation. We evaluated the relationship of EGFR protein expression and tumor grade, histologic features, age at diagnosis, sex, patient survival, and recurrence-free survival. Results: The percentage of tumor cells which were EGFR positive related to reduced survival by Cox regression analysis in both univariate (p = 0.0424) and multivariate analysis (p = 0.0016). Epidermal growth factor receptor positivity was the only 1 of 11 clinical and histological variables associated with decreased recurrence-free survival by either univariate (p = 0.0353) or multivariate (p = 0.0182) analysis. Epidermal growth factor receptor protein expression was not related to patient age, sex, or histologic features. Conclusion: Epidermal growth factor receptor positivity was a significant and independent prognostic indicator for overall survival and recurrence-free survival for irradiated patients with astrocytic gliomas

  3. Stabilization of the bioactivity of tumor necrosis factor by its soluble receptors

    OpenAIRE

    1992-01-01

    The receptors for tumor necrosis factor (TNF) exist in cell-associated as well as soluble forms, both binding specifically to TNF. Since the soluble forms of TNF receptors (sTNF-Rs) can compete with the cell- associated TNF receptors for TNF, it was suggested that they function as inhibitors of TNF activity; at high concentrations, the sTNF-Rs indeed inhibit TNF effects. However, we report here that in the presence of low concentrations of the sTNF-Rs, effects of TNF whose induction depend on...

  4. Effects of glucagon-like peptide-1 (GLP-1) receptor agonists on cardiovascular risk factors

    DEFF Research Database (Denmark)

    Dalsgaard, Niels B; Vilsbøll, Tina; Knop, Filip K

    2018-01-01

    trials for diabetes therapies. To determine if different glucagon-like peptide-1 receptor agonists (GLP-1RAs) had varying effects on these CV risk factors, we reviewed 16 head-to-head trials directly comparing GLP-1RAs that included at least one of the five factors. Few trials reported statistical...

  5. Pharmacological targeting of the KIT growth factor receptor: a therapeutic consideration for mast cell disorders

    DEFF Research Database (Denmark)

    Jensen, Bettina Margrethe; Akin, C; Gilfillan, A M

    2008-01-01

    KIT is a member of the tyrosine kinase family of growth factor receptors which is expressed on a variety of haematopoietic cells including mast cells. Stem cell factor (SCF)-dependent activation of KIT is critical for mast cell homeostasis and function. However, when KIT is inappropriately activa...

  6. Integrin-mediated transactivation of P2X7R via hemichannel-dependent ATP release stimulates astrocyte migration.

    Science.gov (United States)

    Alvarez, Alvaro; Lagos-Cabré, Raúl; Kong, Milene; Cárdenas, Areli; Burgos-Bravo, Francesca; Schneider, Pascal; Quest, Andrew F G; Leyton, Lisette

    2016-09-01

    Our previous reports indicate that ligand-induced αVβ3 integrin and Syndecan-4 engagement increases focal adhesion formation and migration of astrocytes. Additionally, ligated integrins trigger ATP release through unknown mechanisms, activating P2X7 receptors (P2X7R), and the uptake of Ca(2+) to promote cell adhesion. However, whether the activation of P2X7R and ATP release are required for astrocyte migration and whether αVβ3 integrin and Syndecan-4 receptors communicate with P2X7R via ATP remains unknown. Here, cells were stimulated with Thy-1, a reported αVβ3 integrin and Syndecan-4 ligand. Results obtained indicate that ATP was released by Thy-1 upon integrin engagement and required the participation of phosphatidylinositol-3-kinase (PI3K), phospholipase-C gamma (PLCγ) and inositol trisphosphate (IP3) receptors (IP3R). IP3R activation leads to increased intracellular Ca(2+), hemichannel (Connexin-43 and Pannexin-1) opening, and ATP release. Moreover, silencing of the P2X7R or addition of hemichannel blockers precluded Thy-1-induced astrocyte migration. Finally, Thy-1 lacking the integrin-binding site did not stimulate ATP release, whereas Thy-1 mutated in the Syndecan-4-binding domain increased ATP release, albeit to a lesser extent and with delayed kinetics compared to wild-type Thy-1. Thus, hemichannels activated downstream of an αVβ3 integrin-PI3K-PLCγ-IP3R pathway are responsible for Thy-1-induced, hemichannel-mediated and Syndecan-4-modulated ATP release that transactivates P2X7Rs to induce Ca(2+) entry. These findings uncover a hitherto unrecognized role for hemichannels in the regulation of astrocyte migration via P2X7R transactivation induced by integrin-mediated ATP release. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Epidermal growth factor receptor immunohistochemistry: new opportunities in metastatic colorectal cancer.

    Science.gov (United States)

    Hutchinson, Ryan A; Adams, Richard A; McArt, Darragh G; Salto-Tellez, Manuel; Jasani, Bharat; Hamilton, Peter W

    2015-07-07

    The treatment of cancer is becoming more precise, targeting specific oncogenic drivers with targeted molecular therapies. The epidermal growth factor receptor has been found to be over-expressed in a multitude of solid tumours. Immunohistochemistry is widely used in the fields of diagnostic and personalised medicine to localise and visualise disease specific proteins. To date the clinical utility of epidermal growth factor receptor immunohistochemistry in determining monoclonal antibody efficacy has remained somewhat inconclusive. The lack of an agreed reproducible scoring criteria for epidermal growth factor receptor immunohistochemistry has, in various clinical trials yielded conflicting results as to the use of epidermal growth factor receptor immunohistochemistry assay as a companion diagnostic. This has resulted in this test being removed from the licence for the drug panitumumab and not performed in clinical practice for cetuximab. In this review we explore the reasons behind this with a particular emphasis on colorectal cancer, and to suggest a way of resolving the situation through improving the precision of epidermal growth factor receptor immunohistochemistry with quantitative image analysis of digitised images complemented with companion molecular morphological techniques such as in situ hybridisation and section based gene mutation analysis.

  8. The ligand specificities of the insulin receptor and the insulin-like growth factor I receptor reside in different regions of a common binding site

    Energy Technology Data Exchange (ETDEWEB)

    Kjeldsen, T.; Andersen, A.S.; Wiberg, F.C.; Rasmussen, J.S.; Schaeffer, L.; Balschmidt, P.; Moller, K.B.; Moller, N.P.H. (Novo Nordisk, Bagsvaerd (Denmark))

    1991-05-15

    To identify the region(s) of the insulin receptor and the insulin-like growth factor I (IGF-I) receptor responsible for ligand specificity (high-affinity binding), expression vectors encoding soluble chimeric insulin/IGF-I receptors were prepared. The chimeric receptors were expressed in mammalian cells and partially purified. Binding studies revealed that a construct comprising an IGF-I receptor in which the 68 N-terminal amino acids of the insulin receptor {alpha}-subunit had replaced the equivalent IGF-I receptor segment displayed a markedly increased affinity for insulin. In contrast, the corresponding IGF-I receptor sequence is not critical for high-affinity IGF-I binding. It is shown that part of the cysteine-rich domain determines IGF-I specificity. The authors have previously shown that exchanging exons 1, 2, and 3 of the insulin receptor with the corresponding IGF-I receptor sequence results in loss of high affinity for insulin and gain of high affinity for IGF-I. Consequently, it is suggested that the ligand specificities of the two receptors (i.e., the sequences that discriminate between insulin and IGF-I) reside in different regions of a binding site with common features present in both receptors.

  9. Ligand-independent recruitment of steroid receptor coactivators to estrogen receptor by cyclin D1

    NARCIS (Netherlands)

    Zwijsen, R.M.L.; Buckle, R.S.; Hijmans, E.M.; Loomans, C.J.M.; Bernards, R.A.

    1998-01-01

    The estrogen receptor (ER) is an important regulator of growth and differentiation of breast epithelium. Transactivation by ER depends on a leucine-rich motif, which constitutes a ligand-regulated binding site for steroid receptor coactivators (SRCs). Cyclin D1 is frequently amplified in breast

  10. Determination of vascular endothelial- and fibroblast- growth factor receptors in a mouse fibrosarcoma tumor model following photodynamic therapy

    International Nuclear Information System (INIS)

    Ziolkowski, P.; Osieka, B.J.; Symonwicz, K.; Chmielwski, P.; Latos-Grazynski, L.; Bronowicz, A.

    2004-01-01

    The role of angiogenic molecules, like vascular endothelial growth factor and fibroblast growth factor in tumor angio genesis was well confirmed. Photodynamic therapy action is, to very high degree, based on tumor vasculature damage. Therefore, it seemed to be important to evaluate growth factor receptors after photodynamic therapy. The extent of receptor expression was studied by immuno-histochemical method. In this study, vascular endothelial growth factor receptor and fibroblast growth factor receptor have been evaluated at different time points after photodynamic therapy of tumor- bearing BALB/c mice. Two sensitizer: hematoporphyrin derivative and 21, 23-dithia porphyrin were given intraperitoneally in doses: 1.25, 2.5 and 5.0 mg/kg followed by light irradiation at total doses: 50 and 100 J/sq.cm 24 hours later. The number of vascular endothelial growth factor and receptor and fibroblast growth factor in control samples did not exceed 40 per one vessel, whereas after photodynamic therapy, a significant decrease in the number of both receptors was observed. No differences between hematoporphyrin derivative and dithia porphyrin- photodynamic therapy in anti- receptor activities were observed (p<0.001 for vascular endothelial growth factor and p<0.002 for receptor and fibroblast growth factor ). The observed decrease in vascular endothelial growth factor and receptor and fibroblast growth factor amount confirms that after photodynamic therapy, some proteins are inactivated and such a decrease may influence photodynamic therapy effectiveness

  11. Fibroblast growth factor receptor 4 regulates tumor invasion by coupling fibroblast growth factor signaling to extracellular matrix degradation

    DEFF Research Database (Denmark)

    Sugiyama, Nami; Varjosalo, Markku; Meller, Pipsa

    2010-01-01

    Aberrant expression and polymorphism of fibroblast growth factor receptor 4 (FGFR4) has been linked to tumor progression and anticancer drug resistance. We describe here a novel mechanism of tumor progression by matrix degradation involving epithelial-to-mesenchymal transition in response...

  12. AML1/ETO trans-activates c-KIT expression through the long range interaction between promoter and intronic enhancer.

    Science.gov (United States)

    Tian, Ying; Wang, Genjie; Hu, Qingzhu; Xiao, Xichun; Chen, Shuxia

    2018-04-01

    The AML1/ETO onco-fusion protein is crucial for the genesis of t(8;21) acute myeloid leukemia (AML) and is well documented as a transcriptional repressor through dominant-negative effect. However, little is known about the transactivation mechanism of AML1/ETO. Through large cohort of patient's expression level data analysis and a series of experimental validation, we report here that AML1/ETO transactivates c-KIT expression through directly binding to and mediating the long-range interaction between the promoter and intronic enhancer regions of c-KIT. Gene expression analyses verify that c-KIT expression is significantly high in t(8;21) AML. Further ChIP-seq analysis and motif scanning identify two regulatory regions located in the promoter and intronic enhancer region of c-KIT, respectively. Both regions are enriched by co-factors of AML1/ETO, such as AML1, CEBPe, c-Jun, and c-Fos. Further luciferase reporter assays show that AML1/ETO trans-activates c-KIT promoter activity through directly recognizing the AML1 motif and the co-existence of co-factors. The induction of c-KIT promoter activity is reinforced with the existence of intronic enhancer region. Furthermore, ChIP-3C-qPCR assays verify that AML1/ETO mediates the formation of DNA-looping between the c-KIT promoter and intronic enhancer region through the long-range interaction. Collectively, our data uncover a novel transcriptional activity mechanism of AML1/ETO and enrich our knowledge of the onco-fusion protein mediated transcription regulation. © 2017 Wiley Periodicals, Inc.

  13. Internalization mechanisms of the epidermal growth factor receptor after activation with different ligands

    DEFF Research Database (Denmark)

    Henriksen, Lasse; Grandal, Michael Vibo; Knudsen, Stine Louise Jeppe

    2013-01-01

    after ligand activation focuses on the effect of epidermal growth factor (EGF) and transforming growth factor-α (TGF-α). For a long time it was believed that clathrin-mediated endocytosis was the major pathway for internalization of the receptor, but recent work suggests that different pathways exist....... Here we show that clathrin ablation completely inhibits internalization of EGF- and TGF-α-stimulated receptor, however the inhibition of receptor internalization in cells treated with heparin-binding EGF-like growth factor (HB-EGF) or betacellulin (BTC) was only partial. In contrast, clathrin knockdown...... fully inhibits EGFR degradation after all ligands tested. Furthermore, inhibition of dynamin function blocked EGFR internalization after stimulation with all ligands. Knocking out a number of clathrin-independent dynamin-dependent pathways of internalization had no effect on the ligand...

  14. Platelet-activating factor (PAF) receptor as a promising target for cancer cell repopulation after radiotherapy

    Science.gov (United States)

    da Silva-Jr, I A; Chammas, R; Lepique, A P; Jancar, S

    2017-01-01

    A major drawback of radiotherapy is the accelerated growth of the surviving tumor cells. Radiotherapy generates a variety of lipids that bind to the receptor for platelet-activating factor, expressed by cells in the tumor microenvironment. In the present study, using the TC-1 tumor cell line, we found that irradiation induced a twofold increase in receptor expression and generated agonists of receptor. Irradiated cells induced a 20-fold increase in live TC-1 proliferation in vitro. Furthermore, subcutaneous co-injection of irradiated TC-1 cells with TC-1 expressing luciferase (TC-1 fluc+) markedly increased TC-1 fluc+ proliferation in a receptor-dependent way. Moreover we used a human carcinoma cell line not expressing the PAF receptor (KBM) and the same cell transfected with the receptor gene (KBP). Following co-injection of live KBP cells with irradiated KBM in RAG mice, the tumor growth was significantly increased compared with tumor formed following co-injection of live KBM with irradiated KBM. This tumor cell repopulation correlated with increased infiltration of tumor-promoting macrophages (CD206+). We propose that receptor represents a possible target for improving the efficacy of radiotherapy through inhibition of tumor repopulation. PMID:28134937

  15. Identification of a second putative receptor of platelet activating factor on human polymorphonuclear leukocytes

    International Nuclear Information System (INIS)

    Hwang, S.B.

    1987-01-01

    Due to multiple molecular species of platelet activating factor (PAF) and the existence of high affinity binding sites in a variety of cells and tissues, possible existence of PAF receptor subtypes has been suggested. This report shows differences between specific PAF receptors on human leukocytes and platelets. Human PMN leukocyte membranes showed high affinity binding sites for PAF with an equilibrium dissociation constant (Kd) of 4.7 (+/- 1.4) x 10 -10 M. The maximal number (B/sub max/) of receptor sites was estimated to be 3.13 (+/- 1.4) x 10 -13 mol/mg protein. They compared the relative potencies of several PAF agonists and receptor antagonists between human platelet and human leukocyte membranes. One antagonist (Ono-6240) was found to be 8 times less potent at inhibiting the [ 3 H]PAF specific receptor binding to human leukocytes than to human platelets. Mg 2+ , Ca 2+ and K + ions potentiated the [ 3 H]PAF specific binding in both systems. Na + ions inhibited the [ 3 H]PAF specific binding to human platelets but showed no effects in human leukocytes. K + ions decreased the Mg 2+ -potentiated [ 3 H]PAF binding in human leukocytes but showed no effects in human platelets. These results suggest that the PAF specific receptors in human leukocytes are different structurally and possibly functionally from the receptors identified in human platelets

  16. Ric-8A, a Gα protein guanine nucleotide exchange factor potentiates taste receptor signaling

    Directory of Open Access Journals (Sweden)

    Claire J Fenech

    2009-10-01

    Full Text Available Taste receptors for sweet, bitter and umami tastants are G-protein coupled receptors (GPCRs. While much effort has been devoted to understanding G-protein-receptor interactions and identifying the components of the signalling cascade downstream of these receptors, at the level of the G-protein the modulation of receptor signal transduction remains relatively unexplored. In this regard a taste-specific regulator of G-protein signaling (RGS, RGS21, has recently been identified. To study whether guanine nucleotide exchange factors (GEFs are involved in the transduction of the signal downstream of the taste GPCRs we investigated the expression of Ric-8A and Ric-8B in mouse taste cells and their interaction with G-protein subunits found in taste buds. Mammalian Ric-8 proteins were initially identified as potent GEFs for a range of Gα subunits and Ric-8B has recently been shown to amplify olfactory signal transduction. We find that both Ric-8A and Ric-8B are expressed in a large portion of taste bud cells and that most of these cells contain IP3R-3 a marker for sweet, umami and bitter taste receptor cells. Ric-8A interacts with Gα-gustducin and Gαi2 through which it amplifies the signal transduction of hTas2R16, a receptor for bitter compounds. Overall, these findings are consistent with a role for Ric-8 in mammalian taste signal transduction.

  17. ErbB2 resembles an autoinhibited invertebrate epidermal growth factor receptor

    Energy Technology Data Exchange (ETDEWEB)

    Alvarado, Diego; Klein, Daryl E.; Lemmon, Mark A.; (UPENN-MED)

    2009-09-25

    The orphan receptor tyrosine kinase ErbB2 (also known as HER2 or Neu) transforms cells when overexpressed, and it is an important therapeutic target in human cancer. Structural studies have suggested that the oncogenic (and ligand-independent) signalling properties of ErbB2 result from the absence of a key intramolecular 'tether' in the extracellular region that autoinhibits other human ErbB receptors, including the epidermal growth factor (EGF) receptor. Although ErbB2 is unique among the four human ErbB receptors, here we show that it is the closest structural relative of the single EGF receptor family member in Drosophila melanogaster (dEGFR). Genetic and biochemical data show that dEGFR is tightly regulated by growth factor ligands, yet a crystal structure shows that it, too, lacks the intramolecular tether seen in human EGFR, ErbB3 and ErbB4. Instead, a distinct set of autoinhibitory interdomain interactions hold unliganded dEGFR in an inactive state. All of these interactions are maintained (and even extended) in ErbB2, arguing against the suggestion that ErbB2 lacks autoinhibition. We therefore suggest that normal and pathogenic ErbB2 signalling may be regulated by ligands in the same way as dEGFR. Our findings have important implications for ErbB2 regulation in human cancer, and for developing therapeutic approaches that target novel aspects of this orphan receptor.

  18. Thyroid hormone and retinoic acid nuclear receptors: specific ligand-activated transcription factors

    International Nuclear Information System (INIS)

    Brtko, J.

    1998-01-01

    Transcriptional regulation by both the thyroid hormone and the vitamin A-derived 'retinoid hormones' is a critical component in controlling many aspects of higher vertebrate development and metabolism. Their functions are mediated by nuclear receptors, which comprise a large super-family of ligand-inducible transcription factors. Both the thyroid hormone and the retinoids are involved in a complex arrangement of physiological and development responses in many tissues of higher vertebrates. The functions of 3,5,3'-triiodothyronine (T 3 ), the thyromimetically active metabolite of thyroxine as well as all-trans retinoic acid, the biologically active vitamin A metabolite are mediated by nuclear receptor proteins that are members of the steroid/thyroid/retinoid hormone receptor family. The functions of all members of the receptor super family are discussed. (authors)

  19. Human epidermal growth factor receptor 2, epidermal growth factor receptor, and c-MET overexpression and survival in biliary tract cancer: A meta-analysis.

    Science.gov (United States)

    Zhou, Wei; Jiang, Congqing; Zhan, Nan; Lv, Xiaoguang; Fan, Lifang; Ninu, Maskey

    2018-01-01

    Tyrosine kinase growth factor receptors (TKGFRs) play an important role in the progression of cancer. A variety of studies have investigated the clinicopathologic correlation of these receptors and their influences on patient survival in different types of cancer. As the members of TKGFRs, the biomarkers c-MET, epidermal growth factor receptor (EGFR), and human epidermal growth factor receptor 2 (HER-2) have been extensively investigated in biliary tract cancer (BTC). However, their prognostic value is still controversial. Our study aimed to evaluate the prognostic significance of the three markers in BTC patients based on the published studies. The correlation between high expression of these markers and clinical parameters or overall survival (OS) has been assumed in this paper. Including PubMed, EMBASE, China National Knowledge Infrastructure, and Springer, a comprehensive search for the related literature published in Chinese and English has been done. Finally, 31 studies were selected in our research. Surprisingly, the meta-analysis indicated that HER-2 high expression was not correlated with age, gender, primary tumor, tumor node metastasis (TNM) stage, lymph node status, and differentiation. We also found that EGFR high expression was not associated with the parameters, such as age, gender, TNM stage, differentiation, or lymph node status. c-MET high-expression was not associated with age, differentiation, gender, TNM stage, or lymph node status. In addition, our study showed that HER-2, EGFR, and c-MET high expression had an adverse influence on OS in BTC, the pooled hazard ratio for HER-2, EGFR, and c-MET was statistically significant. The present meta-analysis indicated that EGFR and HER-2 high expression have little impact on OS in patients with BTC while c-MET high expression influenced OS in patients with BTC to a large extent. However, c-MET, EGFR, and HER-2 expression did not show any correlation with those clinical parameters. c-MET may be a

  20. Metabotropic glutamate receptor 2 and corticotrophin-releasing factor receptor-1 gene expression is differently regulated by BDNF in rat primary cortical neurons

    DEFF Research Database (Denmark)

    Jørgensen, Christinna V; Klein, Anders B; El-Sayed, Mona

    2013-01-01

    Brain-derived neurotrophic factor (BDNF) is important for neuronal survival and plasticity. Incorporation of matured receptor proteins is an integral part of synapse formation. However, whether BDNF increases synthesis and integration of receptors in functional synapses directly is unclear. We...

  1. Insulin-like growth factor-II (IGF II) receptor from rat brain is of lower apparent molecular weight than the IGF II receptor from rat liver

    International Nuclear Information System (INIS)

    McElduff, A.; Poronnik, P.; Baxter, R.C.

    1987-01-01

    The binding subunits of the insulin and insulin-like growth factor-I (IGF I) receptors from rat brain are of lower molecular weight than the corresponding receptor in rat liver, possibly due to variations in sialic acid content. We have compared the IGF II receptor from rat brain and rat liver. The brain receptor is of smaller apparent mol wt (about 10 K) on sodium dodecyl sulfate polyacrylamide gel electrophoresis. This size difference is independent of ligand binding as it persists in iodinated and specifically immunoprecipitated receptors. From studies of wheat germ agglutinin binding and the effect of neuraminidase on receptor mobility, we conclude that this difference is not simply due to variations in sialic acid content. Treatment with endoglycosidase F results in reduction in the molecular size of both liver and brain receptors and after this treatment the aglycoreceptors are of similar size. We conclude that in rat brain tissue the IGF II receptor like the binding subunits of the insulin and IGF I receptors is of lower molecular size than the corresponding receptors in rat liver. This difference is due to differences in N-linked glycosylation

  2. Autoinhibition of the insulin-like growth factor I receptor by the juxtamembrane region.

    Science.gov (United States)

    Craddock, Barbara P; Cotter, Christopher; Miller, W Todd

    2007-07-10

    The juxtamembrane (JM) regions of several receptor tyrosine kinases are involved in autoinhibitory interactions that maintain the low basal activity of the receptors; mutations can give rise to constitutive kinase activity and signaling. In this report, we show that the JM region of the human insulin-like growth factor I receptor (IGF1R) plays a role in kinase regulation. We mutated JM residues that were conserved in this subfamily of receptor tyrosine kinases, and expressed and purified the cytoplasmic domains using the Sf9/baculovirus system. We show that a kinase-proximal mutation (Y957F) and (to a lesser extent) a mutation in the central part of the JM region (N947A) increase the autophosphorylation activity of the kinase. Steady-state kinetic measurements show the mutations cause an increase in V(max) for phosphorylation of peptide substrates. When the holoreceptors were expressed in fibroblasts derived from IGF1R-deficient mice, the Y957F mutation led to a large increase in basal and in IGF1-stimulated receptor autophosphorylation. Together, these data demonstrate that the JM region of IGF1R plays an important role in limiting the basal activity of the receptor.

  3. Association of nerve growth factor receptors with the triton X-100 cytoskeleton of PC12 cells

    International Nuclear Information System (INIS)

    Vale, R.D.; Ignatius, M.J.; Shooter, E.M.

    1985-01-01

    Triton X-100 solubilizes membranes of PC12 cells and leaves behind a nucleus and an array of cytoskeletal filaments. Nerve growth factor (NGF) receptors are associated with this Triton X-100-insoluble residue. Two classes of NGF receptors are found on PC12 cells which display rapid and slow dissociating kinetics. Although rapidly dissociating binding is predominant (greater than 75%) in intact cells, the majority of binding to the Triton X-100 cytoskeleton is slowly dissociating (greater than 75%). Rapidly dissociating NGF binding on intact cells can be converted to a slowly dissociating form by the plant lectin wheat germ agglutinin (WGA). This lectin also increases the number of receptors which associate with the Triton X-100 cytoskeleton by more than 10-fold. 125 I-NGF bound to receptors can be visualized by light microscopy autoradiography in Triton X-100-insoluble residues of cell bodies, as well as growth cones and neurites. The WGA-induced association with the cytoskeleton, however, is not specific for the NGF receptor. Concentrations of WGA which change the Triton X-100 solubility of membrane glycoproteins are similar to those required to alter the kinetic state of the NGF receptor. Both events may be related to the crossbridging of cell surface proteins induced by this multivalent lectin

  4. Sperm Impairment by Sperm Agglutinating Factor Isolated from Escherichia coli: Receptor Specific Interactions

    Directory of Open Access Journals (Sweden)

    Kiranjeet Kaur

    2013-01-01

    Full Text Available In an earlier work done in our laboratory, we have been able to isolate a sperm agglutinating strain of Escherichia coli from the semen sample of a male attending infertility clinic. Further, factor responsible for sperm agglutination (SAF was isolated and purified, and, using SAF as a tool, corresponding SAF binding receptor from human spermatozoa has been purified. Characterization of SAF and SAF binding receptor using MALDI-TOF showed homology to glutamate decarboxylase and MHC class I molecule, respectively. Coincubation of SAF with spermatozoa not only resulted in spermagglutination but could also compromise other sperm parameters, namely, Mg2+ dependent ATPase activity and apoptosis. Intravaginal administration of SAF could lead to infertility in Balb/c mice. SAF induced impairment of sperm parameters, and infertility was observed to be due to interaction of SAF with sperm surface receptor component as, when purified receptor was introduced, receptor completely inhibited all the detrimental effects induced by SAF. From these results, it could be concluded that interaction of SAF with spermatozoa is receptor mediated.

  5. The under-appreciated promiscuity of the epidermal growth factor receptor family.

    Directory of Open Access Journals (Sweden)

    Sean P Kennedy

    2016-08-01

    Full Text Available Each member of the epidermal growth factor receptor (EGFR family plays a key role in normal development, homeostasis and a variety of pathophysiological conditions, most notably in cancer. According to the prevailing dogma, these four receptor tyrosine kinases (RTKs; EGFR, ERBB2, ERBB3 and ERBB4 function exclusively through the formation of homodimers and heterodimers within the EGFR family. These combinatorial receptor interactions are known to generate increased interactome diversity and therefore influence signalling output, subcellular localization and function of the heterodimer. This molecular plasticity is also thought to play a role in the development of resistance towards targeted cancer therapies aimed at these known oncogenes. Interestingly, many studies now challenge this dogma and suggest that the potential for EGFR family receptors to interact with more distantly related RTKs is much greater than currently appreciated. Here we discuss how the promiscuity of these oncogenic receptors may lead to the formation of many unexpected receptor pairings and the significant implications for the efficiency of many targeted cancer therapies.

  6. Lack of the type III epidermal growth factor receptor mutation in colorectal cancer

    DEFF Research Database (Denmark)

    Spindler, Karen-Lise Garm; Olsen, Dorte A.; Nielsen, Jens Nederby

    2006-01-01

    BACKGROUND: Epidermal growth factor receptor (EGFR) analysis by traditional immunohistochemistry does not provide clinicians with a reliable tool for the selection of patients to EGFR-targeted treatment in colorectal cancer (CRC). Alternative methods and further understanding of the EGFR signalin...

  7. Imaging Diagnostic and Therapeutic Targets : Human Epidermal Growth Factor Receptor 2

    NARCIS (Netherlands)

    Gebhart, Geraldine; Flamen, Patrick; Vries, de Elisabeth G. E.; Jhaveri, Komal; Wimana, Zena

    2016-01-01

    Since the approval of trastuzumab, a humanized monoclonal antibody against the extracellular domain of human epidermal growth factor receptor 2 (HER2), 3 other HER2-targeting agents have gained regulatory approval: lapatinib, pertuzumab, and trastuzumab-emtansine. These agents have revolutionized

  8. The neural cell adhesion molecule binds to fibroblast growth factor receptor 2

    DEFF Research Database (Denmark)

    Christensen, Claus; Lauridsen, Jes B; Berezin, Vladimir

    2006-01-01

    The neural cell adhesion molecule (NCAM) can bind to and activate fibroblast growth factor receptor 1 (FGFR1). However, there are four major FGFR isoforms (FGFR1-FGFR4), and it is not known whether NCAM also interacts directly with the other three FGFR isoforms. In this study, we show by surface...

  9. Nectin-1 binds and signals through the fibroblast growth factor receptor

    DEFF Research Database (Denmark)

    Bojesen, Kirsten Borup; Clausen, Ole; Rohde, Kristian

    2012-01-01

    protein and induced neurite outgrowth in primary cultures of hippocampal and cerebellar granule neurons, an effect abolished by treatment with the fibroblast growth factor receptor (FGFR) inhibitor SU5402, or by transfection with a dominant-negative FGFR1 construct. We showed by surface plasmon resonance...

  10. Immunomodulator CD200 promotes neurotrophic activity by interacting with and activating the fibroblast growth factor receptor

    DEFF Research Database (Denmark)

    Pankratova, Stanislava; Björnsdóttir, Halla; Christensen, Claus

    2016-01-01

    in the suppression of microglia activation. We for the first time demonstrated that CD200 can interact with and transduce signaling through activation of the fibroblast growth factor receptor (FGFR), thereby inducing neuritogenesis and promoting neuronal survival in primary neurons. CD200-induced FGFR...

  11. Localization and functional roles of corticotropin-releasing factor receptor type 2 in the cerebellum

    NARCIS (Netherlands)

    Gounko, Natalia V.; Gramsbergen, Albert; van der Want, Johannes J. L.

    The corticotropin-releasing factor (CRF) type 2 receptor has three splice variants alpha, beta, and gamma. In the rodent brain only CRF-R2 alpha is present. In the cerebellum, CRF-R2 alpha has two different isoforms: a full-length form (fl) and truncated (tr). Both forms CRF-R2 have a unique

  12. Defects in articular cartilage metabolism and early arthritis in fibroblast growth factor receptor 3 deficient mice

    NARCIS (Netherlands)

    Valverde-Franco, G.; Binette, J.S.; Li, W.; Wang, H.; Chai, S.; Laflamme, F.; Tran-Khanh, N.; Quenneville, E.; Meijers, T.; Poole, A.R.; Mort, J.S.; Buschmann, M.D.; Henderson, J.E.

    2006-01-01

    Fibroblast growth factor (FGF) receptor 3 has been identified as a key regulator of endochondral bone development and of post-natal bone metabolism through its action on growth plate chondrocytes and osteoblasts, respectively. It has also been shown to promote chondrogenesis and cartilage production

  13. Soluble receptors for tumor necrosis factor as markers of disease activity in visceral leishmaniasis

    NARCIS (Netherlands)

    Zijlstra, E. E.; van der Poll, T.; Mevissen, M.

    1995-01-01

    Serum concentrations of soluble receptors for tumor necrosis factor (sTNFRs) were measured before and after antimony therapy in 25 Sudanese patients with active visceral leishmaniasis (VL). Both sTNFR types I and II were significantly elevated in patients with VL compared with healthy controls from

  14. Amplification of epidermal growth factor receptor gene in renal cell carcinoma

    DEFF Research Database (Denmark)

    El-Hariry, Iman; Powles, Thomas; Lau, Mike R

    2010-01-01

    Expression of epidermal growth factor receptor (EGFR) may be of prognostic value in renal cell cancer (RCC). Gene amplification of EGFR was investigated in a cohort of 315 patients with advanced RCC from a previously reported randomised study. Using fluorescent in situ hybridisation, only 2 patie...

  15. [From gene to disease; tumor necrosis factor receptor and a syndrome of familial periodic fever

    NARCIS (Netherlands)

    Simon, A.; Drenth, J.P.H.; Meer, J.W.M. van der

    2001-01-01

    Familial Hibernian fever (FHF) is a rare hereditary syndrome that causes periodic attacks of fever and inflammation. It is an autosomal dominantly inherited disorder. The gene involved in FHF encodes for a receptor for tumour necrosis factor (TNFR1). These mutations are thought to result in impaired

  16. Palliation of bone cancer pain by antagonists of platelet-activating factor receptors.

    Directory of Open Access Journals (Sweden)

    Katsuya Morita

    Full Text Available Bone cancer pain is the most severe among cancer pain and is often resistant to current analgesics. Thus, the development of novel analgesics effective at treating bone cancer pain are desired. Platelet-activating factor (PAF receptor antagonists were recently demonstrated to have effective pain relieving effects on neuropathic pain in several animal models. The present study examined the pain relieving effect of PAF receptor antagonists on bone cancer pain using the femur bone cancer (FBC model in mice. Animals were injected with osteolytic NCTC2472 cells into the tibia, and subsequently the effects of PAF receptor antagonists on pain behaviors were evaluated. Chemical structurally different type of antagonists, TCV-309, BN 50739 and WEB 2086 ameliorated the allodynia and improved pain behaviors such as guarding behavior and limb-use abnormalities in FBC model mice. The pain relieving effects of these antagonists were achieved with low doses and were long lasting. Blockade of spinal PAF receptors by intrathecal injection of TCV-309 and WEB 2086 or knockdown of the expression of spinal PAF receptor protein by intrathecal transfer of PAF receptor siRNA also produced a pain relieving effect. The amount of an inducible PAF synthesis enzyme, lysophosphatidylcholine acyltransferase 2 (LPCAT2 protein significantly increased in the spinal cord after transplantation of NCTC 2472 tumor cells into mouse tibia. The combination of morphine with PAF receptor antagonists develops marked enhancement of the analgesic effect against bone cancer pain without affecting morphine-induced constipation. Repeated administration of TCV-309 suppressed the appearance of pain behaviors and prolonged survival of FBC mice. The present results suggest that PAF receptor antagonists in combination with, or without, opioids may represent a new strategy for the treatment of persistent bone cancer pain and improve the quality of life of patients.

  17. [Methodology comparison and influence factors analysis of epidermal growth factor receptor mutation detection].

    Science.gov (United States)

    Qian, Xiaoyan; Wang, Shuai; Shen, Yinchen; Wu, Di; Zhang, Zhishang; Feng, Yun; Han, Xiaohong; Shi, Yuankai

    2015-01-13

    To compare two different methods in detecting epidermal growth factor receptor (EGFR) mutation status of non-small cell lung cancer (NSCLC) , and investing the influences of different conserving methods in DNA quantity, quality and detecting results of formalin-fixed paraffin embedded (FFPE) tissue samples. One hundred and fifty FFPE samples of advanced NSCLC were collected in Cancer Institute/Hospital, Chinese Academy of Medical Sciences from November 2010 to August 2011. Both scorpions amplification refractory mutation system (scorpions ARMS) and direct sequencing were used to detect EGFR mutation in exon 18, 19, 20 and 21, together analyzed the differences between these two methods. Samples with inconsistent results were collected clinical treatment and survival information for further analysis. Extracted DNA from 30 FFPE samples and conserved at -20 °C for 3 years, meanwhile the same 30 FFPE samples DNA were extracted after conserving for 3 years. The DNA quantity, quality and testing results of the two conserving methods were compared. The detection success rate of Scorpions ARMS (100%) was higher than direct sequencing (87.3%), with statistical significance ( χ(2) = 20.28, P < 0.05). The consistent rate of the two methods was 84.7%, and there were no significant differences between these two methods, with a high consistence (Kappa = 0.738, P < 0.001) . The clinical treatment and survival status of 5 patients with different testing results were analysed, which were consistent with ARMS testing results. The quality of DNA extracted from FFPE tissues which were conserved for 3 years according to the conventional way was worse and with more fragments compared with DNA conserved at -20 °C for 3 years, but the EGFR mutation detecting results of the DNA acquired from the two conserved ways were consistent. Combination of Scorpions ARMS and direct sequencing could make the detecting result more comprehensive and reliable. Clinical treatment and survival data may

  18. Histamine H1-receptor antagonists inhibit nuclear factor-kappaB and activator protein-1 activities via H1-receptor-dependent and -independent mechanisms.

    Science.gov (United States)

    Roumestan, C; Henriquet, C; Gougat, C; Michel, A; Bichon, F; Portet, K; Jaffuel, D; Mathieu, M

    2008-06-01

    Histamine H1-receptor antagonists are used to relieve the symptoms of an immediate allergic reaction. They have additional anti-inflammatory effects that could result from an inhibition of the transcription factors activator protein-1 (AP-1) and nuclear factor-kappa B (NF-kappaB). The implication of the H1-receptor in these effects is controversial. Diphenhydramine is a first-generation H1-receptor antagonist while mizolastine and desloratadine are second-generation compounds. Mizolastine is also an inhibitor of 5-lipoxygenase (5-LO), an enzyme that has been involved in NF-kappaB activation. We measured the ability of antihistamines to reverse histamine-induced smooth muscle contraction, an effect that involves the H1-receptor. We then investigated whether these drugs affect NF-kappaB and AP-1 activities in A549 lung epithelial cells, and whether this potential regulation involves H1-receptor and 5-LO. Muscle tone was measured on tracheal segments of guinea-pigs. The H1-receptor was overexpressed by transfection and detected by Western blotting and immunofluorescence microscopy. NF-kappaB and AP-1 activities were assessed by reporter gene assays in cells overexpressing or not overexpressing the H1-receptor. Production of regulated upon activation, normal T cell expressed andsecreted (RANTES), a chemokine whose expression is induced through NF-kappaB, was measured using an immunoassay. H1-receptor antagonists reversed histamine-induced contraction in a dose-dependent manner. Induction of AP-1 and NF-kappaB activities by histamine and the down-regulatory effect of antihistamines required overexpression of the H1-receptor. In contrast, when tumour necrosis factor-alpha and a phorbol ester were used to stimulate NF-kappaB and AP-1 activities, respectively, repression of these activities did not involve the H1-receptor. Indeed, repression was triggered only by a subset of H1-receptor antagonists and was not stronger after overexpression of the H1-receptor. Mizolastine

  19. Characterization of the regulatory domains of the human skn-1a/Epoc-1/Oct-11 POU transcription factor.

    Science.gov (United States)

    Hildesheim, J; Foster, R A; Chamberlin, M E; Vogel, J C

    1999-09-10

    The Skn-1a POU transcription factor is primarily expressed in keratinocytes of murine embryonic and adult epidermis. Although some POU factors expressed in a tissue-specific manner are important for normal differentiation, the biological function of Skn-1a remains unknown. Previous in vitro studies indicate that Skn-1a has the ability to transactivate markers of keratinocyte differentiation. In this study, we have characterized Skn-1a's transactivation domain(s) and engineered a dominant negative protein that lacked this transactivation domain. Deletional analysis of the human homologue of Skn-1a with three target promoters revealed the presence of two functional domains: a primary C-terminal transactivation domain and a combined N-terminal inhibitory domain and transactivation domain. Skn-1a lacking the C-terminal region completely lost transactivation ability, irrespective of the promoter tested, and was able to block transactivation by normal Skn-1a in competition assays. Compared with full-length, Skn-1a lacking the N-terminal region demonstrated either increased transactivation (bovine cytokeratin 6 promoter), comparable transactivation (human papillomavirus type 1a long control region), or loss of transactivation (human papillomavirus type 18 long control region). The identification of a primary C-terminal transactivation domain enabled us to generate a dominant negative Skn-1a factor, which will be useful in the quest for a better understanding of this keratinocyte-specific gene regulator.

  20. miR-29a suppresses MCF-7 cell growth by downregulating tumor necrosis factor receptor 1.

    Science.gov (United States)

    Zhao, Yiling; Yang, Fenghua; Li, Wenyuan; Xu, Chunyan; Li, Li; Chen, Lifei; Liu, Yancui; Sun, Ping

    2017-02-01

    Tumor necrosis factor receptor 1 is the main receptor mediating many tumor necrosis factor-alpha-induced cellular events. Some studies have shown that tumor necrosis factor receptor 1 promotes tumorigenesis by activating nuclear factor-kappa B signaling pathway, while other studies have confirmed that tumor necrosis factor receptor 1 plays an inhibitory role in tumors growth by inducing apoptosis in breast cancer. Therefore, the function of tumor necrosis factor receptor 1 in breast cancer requires clarification. In this study, we first found that tumor necrosis factor receptor 1 was significantly increased in human breast cancer tissues and cell lines, and knockdown of tumor necrosis factor receptor 1 by small interfering RNA inhibited cell proliferation by arresting the cell cycle and inducing apoptosis. In addition, miR-29a was predicted as a regulator of tumor necrosis factor receptor 1 by TargetScan and was shown to be inversely correlated with tumor necrosis factor receptor 1 expression in human breast cancer tissues and cell lines. Luciferase reporter assay further confirmed that miR-29a negatively regulated tumor necrosis factor receptor 1 expression by binding to the 3' untranslated region. In our functional study, miR-29a overexpression remarkably suppressed cell proliferation and colony formation, arrested the cell cycle, and induced apoptosis in MCF-7 cell. Furthermore, in combination with tumor necrosis factor receptor 1 transfection, miR-29a significantly reversed the oncogenic role caused by tumor necrosis factor receptor 1 in MCF-7 cell. In addition, we demonstrated that miR-29a suppressed MCF-7 cell growth by inactivating the nuclear factor-kappa B signaling pathway and by decreasing cyclinD1 and Bcl-2/Bax protein levels. Taken together, our results suggest that miR-29a is an important regulator of tumor necrosis factor receptor 1 expression in breast cancer and functions as a tumor suppressor by targeting tumor necrosis factor receptor 1 to

  1. Wnt signalling via the epidermal growth factor receptor: a role in breast cancer?

    International Nuclear Information System (INIS)

    Musgrove, Elizabeth A

    2004-01-01

    Recent data have suggested the epidermal-growth-factor receptor (EGFR) as a point of convergence for several different classes of receptor. Civenni and colleagues have now demonstrated crosstalk between Wnt signalling and the EGFR, showing that in breast epithelial cells Wnts activate downstream targets of the EGFR, including cyclin D1. Given the role of members of these pathways in the aetiology of breast cancer and as markers of outcome and potential therapeutic targets in breast cancer, this observation has a number of potential implications important for both the basic biology of breast cancer and the clinical management of the disease

  2. Mactosylceramide Prevents Glial Cell Overgrowth by Inhibiting Insulin and Fibroblast Growth Factor Receptor Signaling

    DEFF Research Database (Denmark)

    Gerdøe-Kristensen, Stine; Lund, Viktor K; Wandall, Hans H

    2017-01-01

    Receptor Tyrosine Kinase (RTK) signaling controls key aspects of cellular differentiation, proliferation, survival, metabolism, and migration. Deregulated RTK signaling also underlies many cancers. Glycosphingolipids (GSL) are essential elements of the plasma membrane. By affecting clustering...... hyperactivation is caused by absence of MacCer and not by GlcCer accumulation. We conclude that an early product in GSL biosynthesis, MacCer, prevents inappropriate activation of Insulin and Fibroblast Growth Factor Receptors in Drosophila glia. This article is protected by copyright. All rights reserved....

  3. Gastrin-Releasing Peptide Receptor Mediates Activation of the Epidermal Growth Factor Receptor in Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Sufi Mary Thomas

    2005-04-01

    Full Text Available Gastrin-releasing peptide receptor (GRPR and the epidermal growth factor receptor (EGFR are expressed in several cancers including non-small cell lung carcinoma (NSCLC. Here we demonstrate the activation of EGFR by the GRPR ligand, gastrin-releasing peptide (GRP, in NSCLC cells. GRP induced rapid activation of p44/42 MAPK in lung cancer cells through EGFR. GRP-mediated activation of MAPK in NSCLC cells was abrogated by pretreatment with the anti-EGFR-neutralizing antibody, C225. Pretreatment of NSCLC cells with neutralizing antibodies to the EGFR ligands, TGF-α or HB-EGF, also decreased GRP-mediated MAPK activation. On matrix metalloproteinase (MMP inhibition, GRP failed to activate MAPK in NSCLC cells. EGF and GRP both stimulated NSCLC proliferation, and inhibition of either EGFR or GRPR resulted in cell death. Combining a GRPR antagonist with the EGFR tyrosine kinase inhibitor, gefitinib, resulted in additive cytotoxic effects. Additive effects were seen at gefitinib concentrations from 1 to 18μM, encompassing the ID50 values of both gefitinib-sensitive and gefitinib-resistant NSCLC cell lines. Because a major effect of GRPR appears to be promoting the release of EGFR ligand, this study suggests that a greater inhibition of cell proliferation may occur by abrogating EGFR ligand release in consort with inhibition of EGFR.

  4. Mammographic features of calcifications in DCIS: correlation with oestrogen receptor and human epidermal growth factor receptor 2 status

    Energy Technology Data Exchange (ETDEWEB)

    Bae, Min Sun; Moon, Woo Kyung; Chang, Jung Min; Cho, Nariya [Seoul National University College of Medicine, Department of Radiology, Seoul (Korea, Republic of); Park, So Yeon; Won, Jae-Kyung; Jeon, Yoon-Kyung; Park, In Ae [Seoul National University College of Medicine, Department of Pathology, Seoul (Korea, Republic of); Moon, Hyeong-Gon; Han, Wonshik [Seoul National University College of Medicine, Department of Surgery, Seoul (Korea, Republic of)

    2013-08-15

    This study investigated the correlation of oestrogen receptor (ER) and human epidermal growth factor receptor 2 (HER2) status with the probability of malignancy (POM) of mammographic calcifications in ductal carcinoma in situ (DCIS). A total of 101 women (age range, 27-83 years) with pure DCIS that presented as mammographic calcifications were included. Three radiologists independently reviewed mammograms according to the BI-RADS lexicon and provided 100-point POM scores and a BI-RADS category. ER, HER2 and breast cancer subtypes were determined using immunohistochemistry (IHC) and fluorescence in situ hybridisation. Pairwise correlations between POM and IHC biomarker scores were calculated, and mammographic features were compared between breast cancer subtypes. HER2 level positively correlated with the POM score (P < 0.0001) and BI-RADS category (P < 0.0001), and ER level inversely correlated with the POM score (P < 0.013) and BI-RADS category (P < 0.010). Fine linear branching (P = 0.004) and segmental (P = 0.014) calcifications were significantly associated with HER2-positive cancers, and clustered calcifications were more frequently observed in ER-positive cancers (P = 0.014). HER2 status in DCIS correlated positively with the POM of mammographic calcifications, as determined by radiologists on the basis of the BI-RADS lexicon. (orig.)

  5. Multiple autophosphorylation sites of the epidermal growth factor receptor are essential for receptor kinase activity and internalization. Contrasting significance of tyrosine 992 in the native and truncated receptors

    DEFF Research Database (Denmark)

    Sorkin, A; Helin, K; Waters, C M

    1992-01-01

    for cells expressing kinase-negative receptor (A721). Moreover, tyrosine kinase activity of the Dc-123F receptor toward phospholipase C-gamma 1, compared to wild-type receptor, was reduced by 90%. Taken together, these results show that EGF receptor lacking five autophosphorylation sites functions similar...

  6. The diminished expression of proangiogenic growth factors and their receptors in gastric ulcers of cirrhotic patients.

    Science.gov (United States)

    Luo, Jiing-Chyuan; Peng, Yen-Ling; Hou, Ming-Chih; Huang, Kuang-Wei; Huang, Hui-Chun; Wang, Ying-Wen; Lin, Han-Chieh; Lee, Fa-Yauh; Lu, Ching-Liang

    2013-01-01

    The pathogenesis of the higher occurrence of peptic ulcer disease in cirrhotic patients is complex. Platelets can stimulate angiogenesis and promote gastric ulcer healing. We compared the expressions of proangiogenic growth factors and their receptors in the gastric ulcer margin between cirrhotic patients with thrombocytopenia and those of non-cirrhotic patients to elucidate possible mechanisms. Eligible cirrhotic patients (n = 55) and non-cirrhotic patients (n = 55) who had gastric ulcers were enrolled. Mucosa from the gastric ulcer margin and non-ulcer areas were sampled and the mRNA expressions of the proangiogenic growth factors (vascular endothelial growth factor [VEGF], platelet derived growth factor [PDGF], basic fibroblast growth factor [bFGF]) and their receptors (VEGFR1, VEGFR2, PDGFRA, PDGFRB, FGFR1, FGFR2) were measured and compared. Platelet count and the expressions of these growth factors and their receptors were correlated with each other. The two groups were comparable in terms of gender, ulcer size and infection rate of Helicobacter pylori. However, the cirrhotic group were younger in age, had a lower platelet count than those in the non-cirrhotic group (pexpressions of PDGFB, VEGFR2, FGFR1, and FGFR2 in gastric ulcer margin when compared with those of the non-cirrhotic patients (pexpressions of PDGFB and VEGFR2, FGFR1, and FGFR2 were well correlated with the degree of thrombocytopenia in these cirrhotic patients (ρ>0.5, pimplied that diminished activity of proangiogenic factors and their receptors may contribute to the pathogenesis of gastric ulcers in cirrhotic patients.

  7. A holistic approach combining factor analysis, positive matrix factorization, and chemical mass balance applied to receptor modeling.

    Science.gov (United States)

    Selvaraju, N; Pushpavanam, S; Anu, N

    2013-12-01

    Rapid urbanization and population growth resulted in severe deterioration of air quality in most of the major cities in India. Therefore, it is essential to ascertain the contribution of various sources of air pollution to enable us to determine effective control policies. The present work focuses on the holistic approach of combining factor analysis (FA), positive matrix factorization (PMF), and chemical mass balance (CMB) for receptor modeling in order to identify the sources and their contributions in air quality studies. Insight from the emission inventory was used to remove subjectivity in source identification. Each approach has its own limitations. Factor analysis can identify qualitatively a minimal set of important factors which can account for the variations in the measured data. This step uses information from emission inventory to qualitatively match source profiles with factor loadings. This signifies the identification of dominant sources through factors. PMF gives source profiles and source contributions from the entire receptor data matrix. The data from FA is applied for rank reduction in PMF. Whenever multiple solutions exist, emission inventory identifies source profiles uniquely, so that they have a physical relevance. CMB identifies the source contributions obtained from FA and PMF. The novel approach proposed here overcomes the limitations of the individual methods in a synergistic way. The adopted methodology is found valid for a synthetic data and also the data of field study.

  8. Growth Factor Receptors and Apoptosis Regulators: Signaling Pathways, Prognosis, Chemosensitivity and Treatment Outcomes of Breast Cancer

    Directory of Open Access Journals (Sweden)

    Siddik Sarkar

    2009-01-01

    Full Text Available Biomarkers of breast cancer are necessary for prognosis and prediction to chemotherapy. Prognostic biomarkers provide information regarding outcome irrespective of therapy, while predictive biomarkers provide information regarding response to therapy. Candidate prognostic biomarkers for breast cancers are growth factor receptors, steroid receptors, Ki-67, cyclins, urokinase plasminogen activator, p53, p21, pro- and anti-apoptotic factors, BRCA1 and BRCA2. But currently, the predictive markers are Estrogen and Progesterone receptors responding to endocrine therapy, and HER-2 responding to herceptin. But there are numerous breast cancer cases, where tamoxifen is ineffective even after estrogen receptor positivity. This lead to search of new prognostic and predictive markers and the number of potential markers is constantly increasing due to proteomics and genomics studies. However, most biomarkers individually have poor sensitivity or specificity, or other clinical value. It can be resolved by studying various biomarkers simultaneously, which will help in better prognosis and increasing sensitivity for chemotherapeutic agents. This review is focusing on growth factor receptors, apoptosis markers, signaling cascades, and their correlation with other associated biomarkers in breast cancers. As our knowledge regarding molecular biomarkers for breast cancer increases, prognostic indices will be developed that combine the predictive power of individual molecular biomarkers with specific clinical and pathologic factors. Rigorous comparison of these existing as well as emerging markers with current treatment selection is likely to see an escalation in an era of personalized medicines to ensure the breast cancer patients receive optimal treatment. This will also solve the treatment modalities and complications related to chemotherapeutic regimens.

  9. The role of endogenous epidermal growth factor receptor ligands in mediating corneal epithelial homeostasis.

    Science.gov (United States)

    Peterson, Joanne L; Phelps, Eric D; Doll, Mark A; Schaal, Shlomit; Ceresa, Brian P

    2014-05-01

    To provide a comprehensive study of the biological role and therapeutic potential of six endogenous epidermal growth factor receptor (EGFR) ligands in corneal epithelial homeostasis. Kinetic analysis and dose response curves were performed by using in vitro and in vivo wound-healing assays. Biochemical assays were used to determine receptor expression and activity. Human tears were collected and quantitatively analyzed by multianalyte profiling for endogenous EGFR ligands. Epidermal growth factor receptor ligands improved wound closure and activated EGFR, but betacellulin (BTC) was the most efficacious promoter of wound healing in vitro. In contrast, only epidermal growth factor (EGF) promoted wound healing in vivo. Human tears from 25 healthy individuals showed EGFR ligands at these average concentrations: EGF at 2053 ± 312.4 pg/mL, BTC at 207 ± 39.4 pg/mL, heparin-binding EGF at 44 ± 5.8 pg/mL, amphiregulin at 509 ± 28.8 pg/mL, transforming growth factor-α at 84 ± 19 pg/mL, and epiregulin at 52 ± 15 pg/mL. Under unwounded conditions, only EGF was present at concentrations near the ligand's Kd for the receptor, indicating it is the primary mediator of corneal epithelial homeostasis. Other ligands were present but at concentrations 11- to 7500-fold less their Kd, preventing significant ligand binding. Further, the high levels of EGF and its predicted binding preclude receptor occupancy by exogenous ligand and can explain the discrepancy between the in vitro and in vivo data. Therefore, therapeutic use of EGFR ligands may be unpredictable and impractical.

  10. B cell activating factor (BAFF) and BAFF receptors: fakes and facts.

    Science.gov (United States)

    Ferraccioli, G; Gremese, E

    2017-12-01

    Analysis of B cell activating factor (BAFF) receptors before and after B cell depletion therapy (BCDT) might offer a clue to the understanding of whether some B cell subsets may represent useful biomarkers of biological and clinical responses. Among the BAFF receptors in a cohort of rheumatoid arthritis (RA) patients, the AA have shown, by fluorescence activated cell sorter (FACS) analysis of median fluorescence intensity (MFI), that transmembrane activator and calcium-modulating cyclophilin ligand interactor (TACI) and B cell maturation antigen (BCMA) do not change, whereas the most important, BAFF receptor 3 (BR3), appears to be decreased before as well as after BCDT in all B cell subsets but not in plasmablasts, the most important subset, depleted by BCDT. © 2017 British Society for Immunology.

  11. Epidermal growth factor receptor and cancer: control of oncogenic signalling by endocytosis

    DEFF Research Database (Denmark)

    Grandal, Michael Vibo; Madshus, I.H.

    2008-01-01

    The epidermal growth factor receptor (EGFR) and other members of the EGFR/ErbB receptor family of receptor tyrosine kinases (RTKs) are important regulators of proliferation, angiogenesis, migration, tumorigenesis and metastasis. Overexpression, mutations, deletions and production of autocrine...... has been described, lysosomal degradation upon ligand-induced endocytosis seems to play the major role in EGFR down-regulation. Preclinical and clinical data have demonstrated that EGFR is a central player in cancer, especially in carcinomas, some brain tumours and in non-small cell lung cancer....... Such studies have further validated EGFR as an important molecular target in cancer treatment. This review focuses on mechanisms involved in ligand-induced EGFR activation and endocytic down-regulation. A better understanding of EGFR biology should allow development of more tumour-selective therapeutic...

  12. GLP-1/Exendin-4 induces β-cell proliferation via the epidermal growth factor receptor.

    Science.gov (United States)

    Fusco, Joseph; Xiao, Xiangwei; Prasadan, Krishna; Sheng, Qingfeng; Chen, Congde; Ming, Yung-Ching; Gittes, George

    2017-08-22

    Exendin-4 is a long acting glucagon-like peptide 1 (GLP-1) analogue that is an agonist for the GLP-1 receptor, a G-protein coupled receptor (GPCR). Exendin-4 is used to clinically improve glucose tolerance in diabetic patients due to its ability to enhance insulin secretion. In rodents, and possibly in humans, exendin-4 can stimulate β-cell proliferation. The exact mechanism of action to induce β-cell proliferation is not well understood. Here, using a β-cell specific epidermal growth factor receptor (EGFR) null mouse, we show that exendin-4 induced an increase in proliferation and β-cell mass through EGFR. Thus, our study sheds light on the role of EGFR signaling in the effects of exendin-4 on the control of blood glucose metabolism and β-cell mass.

  13. Cardio-oncology Related to Heart Failure: Epidermal Growth Factor Receptor Target-Based Therapy.

    Science.gov (United States)

    Kenigsberg, Benjamin; Jain, Varun; Barac, Ana

    2017-04-01

    Cancer therapy targeting the epidermal growth factor receptor (EGFR)/erythroblastic leukemia viral oncogene B (ErbB)/human EGFR receptor (HER) family of tyrosine kinases has been successfully used in treatment of several malignancies. The ErbB pathways play a role in the maintenance of cardiac homeostasis. This article summarizes current knowledge about EGFR/ErbB/HER receptor-targeted cancer therapeutics focusing on their cardiotoxicity profiles, molecular mechanisms, and implications in clinical cardio-oncology. The article discusses challenges in predicting, monitoring, and treating cardiac dysfunction and heart failure associated with ErbB-targeted cancer therapeutics and highlights opportunities for researchers and clinical investigators. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. FTZ-Factor1 and Fushi tarazu interact via conserved nuclear receptor and coactivator motifs

    Science.gov (United States)

    Schwartz, Carol J.E.; Sampson, Heidi M.; Hlousek, Daniela; Percival-Smith, Anthony; Copeland, John W.R.; Simmonds, Andrew J.; Krause, Henry M.

    2001-01-01

    To activate transcription, most nuclear receptor proteins require coactivators that bind to their ligand-binding domains (LBDs). The Drosophila FTZ-Factor1 (FTZ-F1) protein is a conserved member of the nuclear receptor superfamily, but was previously thought to lack an AF2 motif, a motif that is required for ligand and coactivator binding. Here we show that FTZ-F1 does have an AF2 motif and that it is required to bind a coactivator, the homeodomain-containing protein Fushi tarazu (FTZ). We also show that FTZ contains an AF2-interacting nuclear receptor box, the first to be found in a homeodomain protein. Both interaction motifs are shown to be necessary for physical interactions in vitro and for functional interactions in developing embryos. These unexpected findings have important implications for the conserved homologs of the two proteins. PMID:11157757

  15. Characterization of the epidermal growth factor receptor associated with cytoskeletons of A431 cells

    International Nuclear Information System (INIS)

    Roy, L.M.; Gittinger, C.K.; Landreth, G.E.

    1989-01-01

    Epidermal growth factor receptors (EGF-R) have been shown to be associated with the detergent-insoluble cytoskeleton of A431 cells, where they retained both a functional ligand-binding domain and tyrosine kinase activity. In the present study we have characterized the tyrosine kinase and ligand binding activities of this cytoskeletally associated EGF-R. The tyrosine kinase activity of the cytoskeletally associated EGF-R was stimulated by EGF treatment of intact cells as evidenced by increased autophosphorylation and phosphorylation of the exogenous substrate angiotensin II (AII). The kinetic behavior of the EGF-R associated with cytoskeletons of EGF-treated cells was similar to that of purified receptors. The stimulation of the receptor kinase activity required EGF treatment of intact cells prior to Triton extraction. If cytoskeletons were prepared from untreated cells and then incubated with EGF, there was no stimulation of the detergent-insoluble receptor kinase activity, indicating that the immobilized receptor was unable to undergo EGF-stimulated activation. Comparison of peptide maps from soluble and cytoskeletally associated EGF-R revealed qualitatively similar patterns; however, they are distinguished by a prominent 46 kD band in digests of the cytoskeletal EGF-R. Saturable binding of 125I-EGF to A431 cytoskeletons prepared from adherent and suspended cells demonstrated the presence of specific receptors on the cytoskeleton. High-affinity EGF-R were preferentially retained upon detergent extraction of adherent cells, whereas both low- and high-affinity receptors were solubilized from the cytoskeletons of suspended cells. Suspension of cells resulted in the solubilization of an additional 15% of the EGF-R to that solubilized in adherent cells, indicating that EGF-R can reversibly associate with the structural elements of the cell

  16. Transactivation Domain of Human c-Myc Is Essential to Alleviate Poly(Q)-Mediated Neurotoxicity in Drosophila Disease Models.

    Science.gov (United States)

    Raj, Kritika; Sarkar, Surajit

    2017-05-01

    Polyglutamine (poly(Q)) disorders, such as Huntington's disease (HD) and spinocerebellar ataxias, represent a group of neurological disorders which arise due to an atypically expanded poly(Q) tract in the coding region of the affected gene. Pathogenesis of these disorders inside the cells begins with the assembly of these mutant proteins in the form of insoluble inclusion bodies (IBs), which progressively sequester several vital cellular transcription factors and other essential proteins, and finally leads to neuronal dysfunction and apoptosis. We have shown earlier that targeted upregulation of Drosophila myc (dmyc) dominantly suppresses the poly(Q) toxicity in Drosophila. The present study examines the ability of the human c-myc proto-oncogene and also identifies the specific c-Myc isoform which drives the mitigation of poly(Q)-mediated neurotoxicity, so that it could be further substantiated as a potential drug target. We report for the first time that similar to dmyc, tissue-specific induced expression of human c-myc also suppresses poly(Q)-mediated neurotoxicity by an analogous mechanism. Among the three isoforms of c-Myc, the rescue potential was maximally manifested by the full-length c-Myc2 protein, followed by c-Myc1, but not by c-MycS which lacks the transactivation domain. Our study suggests that strategies focussing on the transactivation domain of c-Myc could be a very useful approach to design novel drug molecules against poly(Q) disorders.

  17. MB0 and MBI Are Independent and Distinct Transactivation Domains in MYC that Are Essential for Transformation.

    Science.gov (United States)

    Zhang, Qin; West-Osterfield, Kimberly; Spears, Erick; Li, Zhaoliang; Panaccione, Alexander; Hann, Stephen R

    2017-05-06

    MYC is a transcription factor that is essential for cellular proliferation and development. Deregulation or overexpression of MYC occurs in a variety of human cancers. Ectopic expression of MYC causes hyperproliferation and transformation of cells in culture and tumorigenesis in several transgenic mouse models. Deregulation of MYC can also induce apoptosis through activation of p53 and/or ARF tumor suppressors as a safeguard to prevent tumorigenesis. MYC binds to thousands of genomic sites and regulates hundreds of target genes in a context-dependent fashion to mediate these diverse biological roles. The N-terminal region of MYC contains several conserved domains or MYC Boxes (MB), which influence the different MYC transcriptional and biological activities to varying degrees. However, the specific domains that mediate the ability of MYC to activate transcription remain ill defined. In this report, we have identified a new conserved transactivation domain (TAD), MB0, which is essential for MYC transactivation and target gene induction. We demonstrate that MB0 and MBI represent two distinct and independent TADs within the N-terminal 62 amino acids of MYC. In addition, both MB0 and MBI are essential for MYC transformation of primary fibroblasts in cooperation with activated RAS, while MB0 is necessary for efficient MYC-induced p53-independent apoptosis.

  18. The Prostaglandin EP3 Receptor Is an Independent Negative Prognostic Factor for Cervical Cancer Patients.

    Science.gov (United States)

    Heidegger, Helene; Dietlmeier, Sebastian; Ye, Yao; Kuhn, Christina; Vattai, Aurelia; Aberl, Caroline; Jeschke, Udo; Mahner, Sven; Kost, Bernd

    2017-07-19

    We know that one of the main risk factors for cervical cancer is an infection with high-risk human papillomavirus (HR-HPV). Prostaglandins and their receptors are very important for the tumour growth and tumour-associated angiogenesis. Little is known about the expression of the Prostaglandin E receptor type 3 (EP3) or the Prostaglandin (PG)E₂-EP3 signalling in cervical cancer, so the aim of the study was to analyse the expression of the EP3 receptor in cervical cancer and find prognostic factors in relation to survival; EP3 immunohistological staining of 250 cervical cancer slides was performed and analysed with a semi-quantitative score. The statistical evaluation was performed with Statistical Package for the Social Sciences (SPSS) to evaluate the staining results and the survival analyses of the cervical cancer cases. A significant difference was observed in EP3 expression in Fédération Internationale de Gynécologie et d'Obstétrique (FIGO) stadium I versus FIGO stadium II-IV cases. High expression of EP3 (IRS ≥ 1.5) in cervical cancer patients was correlated with poor prognosis in overall survival rates. Survival in adenocarcinoma (AC) of the cervix was lower than in squamous cell carcinoma (SCC). Cox regression analysis shows that EP3 is an independent prognosticator. In this study we could show that the membrane-bound prostaglandin receptor EP3 is an independent prognosticator for cervical cancer patient survival. Targeting the EP3 receptor seems to be an interesting candidate for endocrine therapy. Therefore, more research is needed on the influence of the receptor system and its influence on cervical cancer growth.

  19. Oxidative stress effect on progesterone-induced blocking factor (PIBF) binding to PIBF-receptor in lymphocytes.

    Science.gov (United States)

    de la Haba, Carlos; Palacio, José R; Palkovics, Tamas; Szekeres-Barthó, Júlia; Morros, Antoni; Martínez, Paz

    2014-01-01

    Receptor-ligand binding is an essential interaction for biological function. Oxidative stress can modify receptors and/or membrane lipid dynamics, thus altering cell physiological functions. The aim of this study is to analyze how oxidative stress may alter receptor-ligand binding and lipid domain distribution in the case of progesterone-induced blocking factor/progesterone-induced blocking factor-receptor. For membrane fluidity regionalization analysis of MEC-1 lymphocytes, two-photon microscopy was used in individual living cells. Lymphocytes were also double stained with AlexaFluor647/progesterone-induced blocking factor and Laurdan to evaluate -induced blocking factor/progesterone-induced blocking factor-receptor distribution in the different membrane domains, under oxidative stress. A new procedure has been developed which quantitatively analyzes the regionalization of a membrane receptor among the lipid domains of different fluidity in the plasma membrane. We have been able to establish a new tool which detects and evaluates lipid raft clustering from two-photon microscopy images of individual living cells. We show that binding of progesterone-induced blocking factor to progesterone-induced blocking factor-receptor causes a rigidification of plasma membrane which is related to an increase of lipid raft clustering. However, this clustering is inhibited under oxidative stress conditions. In conclusion, oxidative stress decreases membrane fluidity, impairs receptor-ligand binding and reduces lipid raft clustering. © 2013.

  20. Brefeldin A-Inhibited Guanine Nucleotide-Exchange Factor 1 (BIG1 Governs the Recruitment of Tumor Necrosis Factor Receptor-Associated Factor 2 (TRAF2 to Tumor Necrosis Factor Receptor 1 (TNFR1 Signaling Complexes

    Directory of Open Access Journals (Sweden)

    Takuya Noguchi

    2016-11-01

    Full Text Available Tumor necrosis factor receptor-associated factor 2 (TRAF2 is a critical mediator of tumor necrosis factor-α (TNF-α signaling. However, the regulatory mechanisms of TRAF2 are not fully understood. Here we show evidence that TRAF2 requires brefeldin A-inhibited guanine nucleotide-exchange factor 1 (BIG1 to be recruited into TNF receptor 1 (TNFR1 signaling complexes. In BIG1 knockdown cells, TNF-α-induced c-Jun N-terminal kinase (JNK activation was attenuated and the sensitivity to TNF-α-induced apoptosis was increased. Since these trends correlated well with those of TRAF2 deficient cells as previously demonstrated, we tested whether BIG1 functions as an upstream regulator of TRAF2 in TNFR1 signaling. As expected, we found that knockdown of BIG1 suppressed TNF-α-dependent ubiquitination of TRAF2 that is required for JNK activation, and impaired the recruitment of TRAF2 to the TNFR1 signaling complex (complex I. Moreover, we found that the recruitment of TRAF2 to the death-inducing signaling complex termed complex II was also impaired in BIG1 knockdown cells. These results suggest that BIG1 is a key component of the machinery that drives TRAF2 to the signaling complexes formed after TNFR1 activation. Thus, our data demonstrate a novel and unexpected function of BIG1 that regulates TNFR1 signaling by targeting TRAF2.

  1. Insulin-like growth factor II: complexity of biosynthesis and receptor binding

    DEFF Research Database (Denmark)

    Gammeltoft, S; Christiansen, Jan; Nielsen, F C

    1991-01-01

    Insulin-like growth factor II (IGF-II) belongs to the insulin family of peptides and acts as a growth factor in many fetal tissues and tumors. The gene expression of IGF-II is initiated at three different promoters which gives rise to multiple transcripts. In a human rhabdomyosarcoma cell line......, Man-6-P induces cellular responses. We have studied rat brain neuronal precursor cells where Man-6-P acted as a mitogen suggesting that phosphomannosylated proteins may act as growth factors via the Man-6-P/IGF-II receptor. In conclusion, the gene expression and mechanism of action of IGF-II is very...

  2. Whole-genome cartography of p53 response elements ranked on transactivation potential.

    Science.gov (United States)

    Tebaldi, Toma; Zaccara, Sara; Alessandrini, Federica; Bisio, Alessandra; Ciribilli, Yari; Inga, Alberto

    2015-06-17

    Many recent studies using ChIP-seq approaches cross-referenced to trascriptome data and also to potentially unbiased in vitro DNA binding selection experiments are detailing with increasing precision the p53-directed gene regulatory network that, nevertheless, is still expanding. However, most experiments have been conducted in established cell lines subjected to specific p53-inducing stimuli, both factors potentially biasing the results. We developed p53retriever, a pattern search algorithm that maps p53 response elements (REs) and ranks them according to predicted transactivation potentials in five classes. Besides canonical, full site REs, we developed specific pattern searches for non-canonical half sites and 3/4 sites and show that they can mediate p53-dependent responsiveness of associated coding sequences. Using ENCODE data, we also mapped p53 REs in about 44,000 distant enhancers and identified a 16-fold enrichment for high activity REs within those sites in the comparison with genomic regions near transcriptional start sites (TSS). Predictions from our pattern search were cross-referenced to ChIP-seq, ChIP-exo, expression, and various literature data sources. Based on the mapping of predicted functional REs near TSS, we examined expression changes of thirteen genes as a function of different p53-inducing conditions, providing further evidence for PDE2A, GAS6, E2F7, APOBEC3H, KCTD1, TRIM32, DICER, HRAS, KITLG and TGFA p53-dependent regulation, while MAP2K3, DNAJA1 and potentially YAP1 were identified as new direct p53 target genes. We provide a comprehensive annotation of canonical and non-canonical p53 REs in the human genome, ranked on predicted transactivation potential. We also establish or corroborate direct p53 transcriptional control of thirteen genes. The entire list of identified and functionally classified p53 REs near all UCSC-annotated genes and within ENCODE mapped enhancer elements is provided. Our approach is distinct from, and complementary

  3. Noncanonical DNA motifs as transactivation targets by wild type and mutant p53.

    Directory of Open Access Journals (Sweden)

    Jennifer J Jordan

    2008-06-01

    Full Text Available Sequence-specific binding by the human p53 master regulator is critical to its tumor suppressor activity in response to environmental stresses. p53 binds as a tetramer to two decameric half-sites separated by 0-13 nucleotides (nt, originally defined by the consensus RRRCWWGYYY (n = 0-13 RRRCWWGYYY. To better understand the role of sequence, organization, and level of p53 on transactivation at target response elements (REs by wild type (WT and mutant p53, we deconstructed the functional p53 canonical consensus sequence using budding yeast and human cell systems. Contrary to early reports on binding in vitro, small increases in distance between decamer half-sites greatly reduces p53 transactivation, as demonstrated for the natural TIGER RE. This was confirmed with human cell extracts using a newly developed, semi-in vitro microsphere binding assay. These results contrast with the synergistic increase in transactivation from a pair of weak, full-site REs in the MDM2 promoter that are separated by an evolutionary conserved 17 bp spacer. Surprisingly, there can be substantial transactivation at noncanonical (1/2-(a single decamer and (3/4-sites, some of which were originally classified as biologically relevant canonical consensus sequences including PIDD and Apaf-1. p53 family members p63 and p73 yielded similar results. Efficient transactivation from noncanonical elements requires tetrameric p53, and the presence of the carboxy terminal, non-specific DNA binding domain enhanced transactivation from noncanonical sequences. Our findings demonstrate that RE sequence, organization, and level of p53 can strongly impact p53-mediated transactivation, thereby changing the view of what constitutes a functional p53 target. Importantly, inclusion of (1/2- and (3/4-site REs greatly expands the p53 master regulatory network.

  4. Fibroblast Growth Factor 10-Fibroblast Growth Factor Receptor 2b Mediated Signaling Is Not Required for Adult Glandular Stomach Homeostasis

    Science.gov (United States)

    Sala, Frederic G.; Ford, Henri R.; Bellusci, Saverio; Grikscheit, Tracy C.

    2012-01-01

    The signaling pathways that are essential for gastric organogenesis have been studied in some detail; however, those that regulate the maintenance of the gastric epithelium during adult homeostasis remain unclear. In this study, we investigated the role of Fibroblast growth factor 10 (FGF10) and its main receptor, Fibroblast growth factor receptor 2b (FGFR2b), in adult glandular stomach homeostasis. We first showed that mouse adult glandular stomach expressed Fgf10, its receptors, Fgfr1b and Fgfr2b, and most of the other FGFR2b ligands (Fgf1, Fgf7, Fgf22) except for Fgf3 and Fgf20. Fgf10 expression was mesenchymal whereas FGFR1 and FGFR2 expression were mostly epithelial. Studying double transgenic mice that allow inducible overexpression of Fgf10 in adult mice, we showed that Fgf10 overexpression in normal adult glandular stomach increased epithelial proliferation, drove mucous neck cell differentiation, and reduced parietal and chief cell differentiation. Although a similar phenotype can be associated with the development of metaplasia, we found that Fgf10 overexpression for a short duration does not cause metaplasia. Finally, investigating double transgenic mice that allow the expression of a soluble form of Fgfr2b, FGF10's main receptor, which acts as a dominant negative, we found no significant changes in gastric epithelial proliferation or differentiation in the mutants. Our work provides evidence, for the first time, that the FGF10-FGFR2b signaling pathway is not required for epithelial proliferation and differentiation during adult glandular stomach homeostasis. PMID:23133671

  5. Fibroblast growth factor 10-fibroblast growth factor receptor 2b mediated signaling is not required for adult glandular stomach homeostasis.

    Directory of Open Access Journals (Sweden)

    Allison L Speer

    Full Text Available The signaling pathways that are essential for gastric organogenesis have been studied in some detail; however, those that regulate the maintenance of the gastric epithelium during adult homeostasis remain unclear. In this study, we investigated the role of Fibroblast growth factor 10 (FGF10 and its main receptor, Fibroblast growth factor receptor 2b (FGFR2b, in adult glandular stomach homeostasis. We first showed that mouse adult glandular stomach expressed Fgf10, its receptors, Fgfr1b and Fgfr2b, and most of the other FGFR2b ligands (Fgf1, Fgf7, Fgf22 except for Fgf3 and Fgf20. Fgf10 expression was mesenchymal whereas FGFR1 and FGFR2 expression were mostly epithelial. Studying double transgenic mice that allow inducible overexpression of Fgf10 in adult mice, we showed that Fgf10 overexpression in normal adult glandular stomach increased epithelial proliferation, drove mucous neck cell differentiation, and reduced parietal and chief cell differentiation. Although a similar phenotype can be associated with the development of metaplasia, we found that Fgf10 overexpression for a short duration does not cause metaplasia. Finally, investigating double transgenic mice that allow the expression of a soluble form of Fgfr2b, FGF10's main receptor, which acts as a dominant negative, we found no significant changes in gastric epithelial proliferation or differentiation in the mutants. Our work provides evidence, for the first time, that the FGF10-FGFR2b signaling pathway is not required for epithelial proliferation and differentiation during adult glandular stomach homeostasis.

  6. Methylthioadenosine phosphorylase and activated insulin-like growth factor-1 receptor/insulin receptor: potential therapeutic targets in chordoma.

    Science.gov (United States)

    Sommer, Josh; Itani, Doha M; Homlar, Kelly C; Keedy, Vicki L; Halpern, Jennifer L; Holt, Ginger E; Schwartz, Herbert S; Coffin, Cheryl M; Kelley, Michael J; Cates, Justin M M

    2010-04-01

    Currently there is no effective chemotherapy for chordoma. Recent studies report co-expression of insulin-like growth factor-1 receptor (IGF1R) and its cognate ligand in chordoma, but it is unknown whether this receptor tyrosine kinase is activated in these tumours. Additionally, genetic studies have confirmed frequent deletions of chromosome 9p in chordomas, which encompasses the cyclin-dependent kinase inhibitor 2A (CDKN2A) locus. Another gene in this region, methylthioadenosine phosphorylase (MTAP), is an essential enzyme of the purine salvage pathway and has therapeutic relevance because MTAP-deficient cells are particularly sensitive to inhibitors of de novo purine synthesis. We investigated whether these pathways might be potential therapeutic targets for chordoma. Paraffin-embedded tissue samples from 30 chordomas were analysed by immunohistochemistry for expression of the phosphorylated isoforms of IGF1R or the insulin receptor (pIGF1R/pIR) and selected downstream signalling molecules, including BCL2-associated agonist of cell death protein (BAD). Expression of CDKN2A and MTAP proteins was also assessed. Skeletal chondrosarcomas, benign notochordal cell tumours, and fetal notochord were studied for comparison. Phosphorylated IGF1R/IR was detected in 41% of chordomas, together with activated downstream signalling molecules, and pIGF1R/pIR was absent in benign notochordal cell tumours and fetal notochord. Thirty-nine per cent of chordomas were negative for MTAP immunoreactivity. Patients with pIGF1R/pIR-positive tumours showed significantly decreased median disease-free survival in multivariate survival analysis (p = 0.036), whereas phosphorylation of BAD at serine-99 was found to be associated with a favourable prognosis (p = 0.002). Approximately 40% of chordomas demonstrate evidence of activation of the IGF1R/IR signalling pathway or loss of a key enzyme in the purine salvage pathway. Aberrant signalling cascades and disrupted metabolic pathways such as

  7. Distribution and number of epidermal growth factor receptors in skin is related to epithelial cell growth

    DEFF Research Database (Denmark)

    Green, M R; Basketter, D A; Couchman, J R

    1983-01-01

    receptors are detected on the epithelial cells overlying the basement membranes of the epidermis, sebaceous gland, and regions of the hair follicle all of which have proliferative capacity. In marked contrast, tissues which have started to differentiate and lost their growth potential, carry either......Epidermal growth factor (EGF), a low-molecular-weight polypeptide (G. Carpenter and S. Cohen, 1979, Annu. Rev. Biochem. 48, 193-216), stimulates the proliferation and keratinisation of cultured embryonic epidermis (S. Cohen, 1965, Dev. Biol. 12, 394-407) and promotes epidermal growth, thickening......, and keratinisation when injected into neonatal mice (S. Cohen and G.A. Elliott, 1963, J. Invest. Dermatol, 40, 1-5). We have determined the distribution of the available receptors for epidermal growth factor in rat skin using autoradiography following incubation of explants with 125I-labelled mouse EGF. EGF...

  8. BFV activates the NF-κB pathway through its transactivator (BTas) to enhance viral transcription

    International Nuclear Information System (INIS)

    Wang Jian; Tan Juan; Zhang Xihui; Guo Hongyan; Zhang Qicheng; Guo Tingting; Geng Yunqi; Qiao Wentao

    2010-01-01

    Multiple families of viruses have evolved sophisticated strategies to regulate nuclear factor-κB (NF-κB) signaling, which plays a pivotal role in diverse cellular events, including virus-host interactions. In this study, we report that bovine foamy virus (BFV) is able to activate the NF-κB pathway through the action of its transactivator, BTas. Both cellular IKKβ and IκBα also participate in this activation. In addition, we demonstrate that BTas induces the processing of p100, which implies that BTas can activate NF-κB through a noncanonical pathway as well. Co-immunoprecipitation analysis shows that BTas interacts with IKK catalytic subunits (IKKα and IKKβ), which may be responsible for regulation of IKK kinase activity and persistent NF-κB activation. Furthermore, our results indicate that the level of BTas-mediated LTR transcription correlates with the activity of cellular NF-κB. Together, this study suggests that BFV activates the NF-κB pathway through BTas to enhance viral transcription.

  9. Neurodegenerative and neuroprotective effects of tumor necrosis factor (TNF) in retinal ischemia : Opposite roles of TNF receptor 1 and TNF receptor 2

    NARCIS (Netherlands)

    Fontaine, Sharon; Mohand-Said, S; Hanoteau, N; Fuchs, L; Pfizenmaier, K; Eisel, U

    2002-01-01

    Tumor necrosis factor (TNF) is an important factor in various acute and chronic neurodegenerative disorders. In retinal ischemia, we show early, transient upregulation of TNF, TNF receptor 1 (TNF-R1), and TNF-R2 6 hr after reperfusion preceding neuronal cell loss. To assess the specific role of TNF

  10. Optimal Therapeutic Strategy for Non-small Cell Lung Cancer with Mutated Epidermal Growth Factor Receptor

    Directory of Open Access Journals (Sweden)

    Zhong SHI

    2015-02-01

    Full Text Available Although epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs have been widely used in non-small cell lung cancer (NSCLC patients, it is still controversial about how to combine EGFR-TKI with chemotherapy and other targeted drugs. We have made a summary on the current therapeutic models of EGFR-TKI combined with chemotherapy/bevacizumab in this review and aimed to find the optimal therapeutic strategy for NSCLC patients with EGFR mutation.

  11. NMR backbone assignments of the tyrosine kinase domain of human fibroblast growth factor receptor 1.

    Science.gov (United States)

    Vajpai, Navratna; Schott, Anne-Kathrin; Vogtherr, Martin; Breeze, Alexander L

    2014-04-01

    Members of the fibroblast growth factor receptor tyrosine kinase family (FGFR1-4) play an important role in many signalling cascades. Although tightly regulated, aberrant activity of these enzymes may lead to, or become features of, disease pathologies including cancer. FGFR isoforms have been the subject of drug discovery programmes, with a number of kinase-domain inhibitors in pre-clinical and clinical development. Here, we present the first (83% complete) backbone resonance assignments of apo-FGFR1 kinase.

  12. Insulin-Insulin-like Growth Factors Hybrids as Molecular Probes of Hormone:Receptor Binding Specificity

    Czech Academy of Sciences Publication Activity Database

    Křížková, Květoslava; Chrudinová, Martina; Povalová, Anna; Selicharová, Irena; Collinsová, Michaela; Vaněk, Václav; Brzozowski, A. M.; Jiráček, Jiří; Žáková, Lenka

    2016-01-01

    Roč. 55, č. 21 (2016), s. 2903-2913 ISSN 0006-2960 R&D Projects: GA ČR GA15-19018S Institutional support: RVO:61388963 Keywords : alanine scanning mutagenesis * high-affinity binding * type 1 IGF receptor Subject RIV: CE - Biochemistry Impact factor: 2.938, year: 2016 http://pubs.acs.org/doi/pdf/10.1021/acs.biochem.6b00140

  13. Identification of fibroblast growth factor receptor 3 (FGFR3 as a protein receptor for botulinum neurotoxin serotype A (BoNT/A.

    Directory of Open Access Journals (Sweden)

    Birgitte P S Jacky

    Full Text Available Botulinum neurotoxin serotype A (BoNT/A causes transient muscle paralysis by entering motor nerve terminals (MNTs where it cleaves the SNARE protein Synaptosomal-associated protein 25 (SNAP25206 to yield SNAP25197. Cleavage of SNAP25 results in blockage of synaptic vesicle fusion and inhibition of the release of acetylcholine. The specific uptake of BoNT/A into pre-synaptic nerve terminals is a tightly controlled multistep process, involving a combination of high and low affinity receptors. Interestingly, the C-terminal binding domain region of BoNT/A, HC/A, is homologous to fibroblast growth factors (FGFs, making it a possible ligand for Fibroblast Growth Factor Receptors (FGFRs. Here we present data supporting the identification of Fibroblast Growth Factor Receptor 3 (FGFR3 as a high affinity receptor for BoNT/A in neuronal cells. HC/A binds with high affinity to the two extra-cellular loops of FGFR3 and acts similar to an agonist ligand for FGFR3, resulting in phosphorylation of the receptor. Native ligands for FGFR3; FGF1, FGF2, and FGF9 compete for binding to FGFR3 and block BoNT/A cellular uptake. These findings show that FGFR3 plays a pivotal role in the specific uptake of BoNT/A across the cell membrane being part of a larger receptor complex involving ganglioside- and protein-protein interactions.

  14. Circulating tumour necrosis factor alpha & soluble TNF receptors in patients with Guillain-Barre syndrome.

    Science.gov (United States)

    Radhakrishnan, V V; Sumi, M G; Reuben, S; Mathai, A; Nair, M D

    2003-05-01

    Tumour necrosis factor-alpha (TNF-alpha) is regarded as one of the immune factors that can induce demyelination of peripheral nerves in patients with Guillian-Barre syndrome (GBS). This present study was undertaken to find out the role of TNF-alpha and soluble TNF receptors in the pathogenesis of GBS; and to study the effect of intravenous immunoglobulin (ivIg) therapy on the serum TNF-alpha and soluble TNF receptors in patients with GBS. Thirty six patients with GBS in progressive stages of motor weakness were included in this study. The serum TNF-alpha and soluble TNF receptors (TNF-RI, TNF-RII) were measured in the serum samples of these patients before and after ivIg therapy by a sandwich ELISA. Of the 36 patients with GBS, 26 (72.2%) showed elevated serum TNF-alpha levels prior to ivIg therapy. Following a complete course of ivIg therapy there was a progressive decrease in the serum TNF-alpha concentrations in these 26 patients. On the other hand, the soluble TNF receptors, particularly TNF-RII showed an increase in the serum of GBS patients following ivIg therapy. The results indicate that ivIg reduces the serum TNF-alpha concentrations in the GBS patients having elevated levels prior to ivIg therapy. Elevated serum levels of soluble TNF receptors following ivIg therapy may play a protective role by inhibiting the demyelinating effect of TNF-alpha in the peripheral nerves of patients with GBS.

  15. Prevalence of Epidermal Growth Factor Receptor Mutations in Patients with Non-Small Cell Lung Cancer in Turkish Population.

    Science.gov (United States)

    Güler Tezel, Gaye; Şener, Ebru; Aydın, Çisel; Önder, Sevgen

    2017-12-01

    Epidermal growth factor receptor mutation analysis in non-small cell lung cancer is important for selecting patients who will receive treatment with tyrosine kinase inhibitors. In this study, we aimed to investigate the prevalence of epidermal growth factor receptor mutations and mutation patterns in the Turkish population. We retrospectively reviewed molecular pathology reports of 959 cases with lung cancer analysed for epidermal growth factor receptor mutations. We analysed all four epidermal growth factor receptor exon mutations using a real-time polymerase chain reaction platform. In this study, the epidermal growth factor receptor mutation rate in the Turkish population was 16.7% (160 of 959). The epidermal growth factor receptor mutation frequency was significantly higher in women (37.1%, n=96) than in men (9.1%, n=64) (pmutation rate was higher in the adenocarcinoma histologic type (pmutations were older than those without mutations (p=0.003). The most frequent mutations were exon 19 deletions (48.8%, 78/160) and exon 21 L858R point mutations (38.1.1%, 61/160). We also detected compound mutation patterns in three cases (1.9%). The prevalence of epidermal growth factor receptor mutations in the Turkish population was slightly higher than that in the Caucasian population and lower than that in the East Asian population. The results of this study may provide guidance in personalized therapy of non-small cell lung cancer in the Turkish population.

  16. The Role of Vascular Endothelial Growth Factor Receptor-1 Signaling in the Recovery from Ischemia.

    Science.gov (United States)

    Amano, Hideki; Kato, Shintaro; Ito, Yoshiya; Eshima, Koji; Ogawa, Fumihiro; Takahashi, Ryo; Sekiguchi, Kazuki; Tamaki, Hideaki; Sakagami, Hiroyuki; Shibuya, Masabumi; Majima, Masataka

    2015-01-01

    Vascular endothelial growth factor (VEGF) is one of the most potent angiogenesis stimulators. VEGF binds to VEGF receptor 1 (VEGFR1), inducing angiogenesis through the receptor's tyrosine kinase domain (TK), but the mechanism is not well understood. We investigated the role of VEGFR1 tyrosine kinase signaling in angiogenesis using the ischemic hind limb model. Relative to control mice, blood flow recovery was significantly impaired in mice treated with VEGFA-neutralizing antibody. VEGFR1 tyrosine kinase knockout mice (TK-/-) had delayed blood flow recovery from ischemia and impaired angiogenesis, and this phenotype was unaffected by treatment with a VEGFR2 inhibitor. Compared to wild type mice (WT), TK-/- mice had no change in the plasma level of VEGF, but the plasma levels of stromal-derived cell factor 1 (SDF-1) and stem cell factor, as well as the bone marrow (BM) level of pro-matrix metalloproteinase-9 (pro-MMP-9), were significantly reduced. The recruitment of cells expressing VEGFR1 and C-X-C chemokine receptor type 4 (CXCR4) into peripheral blood and ischemic muscles was also suppressed. Furthermore, WT transplanted with TK-/- BM significantly impaired blood flow recovery more than WT transplanted with WT BM. These results suggest that VEGFR1-TK signaling facilitates angiogenesis by recruiting CXCR4+VEGFR1+ cells from BM.

  17. Thyroid hormone regulation of epidermal growth factor receptor levels in mouse mammary glands

    International Nuclear Information System (INIS)

    Vonderhaar, B.K.; Tang, E.; Lyster, R.R.; Nascimento, M.C.

    1986-01-01

    The specific binding of iodinated epidermal growth factor ([ 125 I]iodo-EGF) to membranes prepared from the mammary glands and spontaneous breast tumors of euthyroid and hypothyroid mice was measured in order to determine whether thyroid hormones regulate the EGF receptor levels in vivo. Membranes from hypothyroid mammary glands of mice at various developmental ages bound 50-65% less EGF than those of age-matched euthyroid controls. Treatment of hypothyroid mice with L-T4 before killing restored binding to the euthyroid control level. Spontaneous breast tumors arising in hypothyroid mice also bound 30-40% less EGF than tumors from euthyroid animals even after in vitro desaturation of the membranes of endogenous growth factors with 3 M MgCl2 treatment. The decrease in binding in hypothyroid membranes was due to a decrease in the number of binding sites, not to a change in affinity of the growth factor for its receptor, as determined by Scatchard analysis of the binding data. Both euthyroid and hypothyroid membranes bound EGF primarily to a single class of high affinity sites [dissociation constant (Kd) = 0.7-1.8 nM]. Euthyroid membranes bound 28.4 +/- (SE) 0.6 fmol/mg protein, whereas hypothyroid membranes bound 15.5 +/- 1.0 fmol/mg protein. These data indicate that EGF receptor levels in normal mammary glands and spontaneous breast tumors in mice are subject to regulation by thyroid status

  18. I-mfa domain proteins specifically interact with HTLV-1 Tax and repress its transactivating functions

    Energy Technology Data Exchange (ETDEWEB)

    Kusano, Shuichi, E-mail: skusano@m2.kufm.kagoshima-u.ac.jp [Division of Persistent and Oncogenic Viruses, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Yoshimitsu, Makoto; Hachiman, Miho [Division of Hematology and Immunology, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Ikeda, Masanori [Division of Persistent and Oncogenic Viruses, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan)

    2015-12-15

    The I-mfa domain proteins HIC (also known as MDFIC) and I-mfa (also known as MDFI) are candidate tumor suppressor genes that are involved in cellular and viral transcriptional regulation. Here, we show that HIC and I-mfa directly interact with human T-cell leukemia virus type-1 (HTLV-1) Tax protein in vitro. In addition, HIC and I-mfa repress Tax-dependent transactivation of an HTLV-1 long terminal repeat (LTR) reporter construct in COS-1, Jurkat and high-Tax-producing HTLV-1-infected T cells. HIC also interacts with Tax through its I-mfa domain in vivo and represses Tax-dependent transactivation of HTLV-1 LTR and NF-κB reporter constructs in an interaction-dependent manner. Furthermore, we show that HIC decreases the nuclear distribution and stimulates the proteasomal degradation of Tax. These data reveal that HIC specifically interacts with HTLV-1 Tax and negatively regulates Tax transactivational activity by altering its subcellular distribution and stability. - Highlights: • I-mfa domain proteins, HIC and I-mfa, specifically interact with HTLV-1 Tax. • HIC and I-mfa repress the Tax-dependent transactivation of HTLV-1 LTR. • HIC represses the Tax-dependent transactivation of NF-κΒ. • HIC decreases the nuclear distribution of Tax. • HIC stimulates the proteasomal degradation of Tax.

  19. Simultaneous Inhibition of Tumor Necrosis Factor Receptor 1 and Matrix Metalloproteinase 8 Completely Protects Against Acute Inflammation and Sepsis.

    Science.gov (United States)

    Steeland, Sophie; Van Ryckeghem, Sara; Vandewalle, Jolien; Ballegeer, Marlies; Van Wonterghem, Elien; Eggermont, Melanie; Decruyenaere, Johan; De Bus, Liesbet; Libert, Claude; Vandenbroucke, Roosmarijn E

    2018-01-01

    Sepsis causes very high mortality and morbidity rates and remains one of the biggest medical challenges. This study investigates whether plasma levels of both matrix metalloproteinase 8 and tumor necrosis factor receptor 1 are associated with sepsis severity and also investigates the therapeutic applicability of simultaneous inhibition of the two molecules in sepsis. Observational human pilot study-prospective controlled animal study. University hospital and research laboratory. Sepsis patients and C57BL/6 mice deficient for matrix metalloproteinase 8 and/or tumor necrosis factor receptor 1. Plasma and whole blood RNA were collected from 13 sepsis patients for 7 consecutive days and within 24 hours of admission to ICU. Matrix metalloproteinase 8 and tumor necrosis factor receptor 1 plasma and expression levels were determined in these patients. Mice deficient for both matrix metalloproteinase 8 and tumor necrosis factor receptor 1 were generated and subjected to endotoxemia and cecal ligation and puncture. Additionally, a bispecific Nanobody that simultaneously blocks matrix metalloproteinase 8 and tumor necrosis factor receptor 1 was created. Plasma levels of matrix metalloproteinase 8 and tumor necrosis factor receptor 1 were positively correlated with the Sequential Organ Failure Assessment score (r, 0.51 and 0.58) and interleukin 6 levels (r, 0.59 and 0.52) in 13 sepsis patients. Combined elimination of tumor necrosis factor receptor 1 and matrix metalloproteinase 8 in double knockout mice resulted in superior survival in endotoxemia and CLP compared with single knockouts and wild-type mice. Cotreatment with our bispecific Nanobody in CLP resulted in improved survival rates (28% vs 19%) compared with untreated mice. Inhibition of matrix metalloproteinase 8 and tumor necrosis factor receptor 1 might have therapeutic potential to treat sepsis and proof-of-principle was provided as therapeutics that inhibit both tumor necrosis factor receptor 1 and matrix

  20. The Neurotrophic Factor Receptor p75 in the Rat Dorsolateral Striatum Drives Excessive Alcohol Drinking

    Science.gov (United States)

    Darcq, Emmanuel; Morisot, Nadege; Phamluong, Khanhky; Warnault, Vincent; Jeanblanc, Jerome; Longo, Frank M.; Massa, Stephen M.

    2016-01-01

    Brain-derived neurotrophic factor (BDNF) signaling in the dorsolateral striatum (DLS) keeps alcohol intake in moderation. For example, activation of the BDNF receptor tropomyosin receptor kinase B (TrkB) in the DLS reduces intake in rats that consume moderate amounts of alcohol. Here, we tested whether long-term excessive consumption of alcohol produces neuroadaptations in BDNF signaling in the rat DLS. We found that BDNF was no longer able to gate alcohol self-administration after a history of repeated cycles of binge alcohol drinking and withdrawal. We then elucidated the possible neuroadaptations that could block the ability of BDNF to keep consumption of alcohol in moderation. We report that intermittent access to 20% alcohol in a two-bottle choice paradigm that models excessive alcohol drinking produces a mobilization of DLS p75 neurotrophin receptor (p75NTR), whose activities oppose those of the Trk receptors, including TrkB. These neuroadaptations were not observed in the DLS of rats exposed to continuous access to 10% alcohol or in rats consuming sucrose. Furthermore, short hairpin RNA (shRNA)-mediated knockdown of the p75NTR gene in the DLS, as well as intra-DLS infusion or systemic administration of the p75NTR modulator, LM11A-31, significantly reduced binge drinking of alcohol. Together, our results suggest that excessive alcohol consumption produces a change in BDNF signaling in the DLS, which is mediated by the recruitment of p75NTR. Our data also imply that modulators of p75NTR signaling could be developed as medications for alcohol abuse disorders. SIGNIFICANCE STATEMENT Neuroadaptations gate or drive excessive, compulsive alcohol drinking. We previously showed that brain-derived neurotrophic factor and its receptor, TrkB, in the dorsolateral striatum (DLS), are part of an endogenous system that keeps alcohol drinking in moderation. Here, we show that a history of excessive alcohol intake produces neuroadaptations in the DLS that preclude BDNF

  1. Involvement of platelet-derived growth factor ligands and receptors in tumorigenesis.

    Science.gov (United States)

    Heldin, C-H; Lennartsson, J; Westermark, B

    2018-01-01

    Platelet-derived growth factor (PDGF) isoforms and their receptors have important roles during embryogenesis, particularly in the development of various mesenchymal cell types in different organs. In the adult, PDGF stimulates wound healing and regulates tissue homeostasis. However, overactivity of PDGF signalling is associated with malignancies and other diseases characterized by excessive cell proliferation, such as fibrotic conditions and atherosclerosis. In certain tumours, genetic or epigenetic alterations of the genes for PDGF ligands and receptors drive tumour cell proliferation and survival. Examples include the rare skin tumour dermatofibrosarcoma protuberance, which is driven by autocrine PDGF stimulation due to translocation of a PDGF gene, and certain gastrointestinal stromal tumours and leukaemias, which are driven by constitute activation of PDGF receptors due to point mutations and formation of fusion proteins of the receptors, respectively. Moreover, PDGF stimulates cells in tumour stroma and promotes angiogenesis as well as the development of cancer-associated fibroblasts, both of which promote tumour progression. Inhibitors of PDGF signalling may thus be of clinical usefulness in the treatment of certain tumours. © 2017 The Association for the Publication of the Journal of Internal Medicine.

  2. Molecular Recognition of Corticotropin releasing Factor by Its G protein-coupled Receptor CRFR1

    Energy Technology Data Exchange (ETDEWEB)

    Pioszak, Augen A.; Parker, Naomi R.; Suino-Powell, Kelly; Xu, H. Eric (Van Andel)

    2009-01-15

    The bimolecular interaction between corticotropin-releasing factor (CRF), a neuropeptide, and its type 1 receptor (CRFR1), a class B G-protein-coupled receptor (GPCR), is crucial for activation of the hypothalamic-pituitary-adrenal axis in response to stress, and has been a target of intense drug design for the treatment of anxiety, depression, and related disorders. As a class B GPCR, CRFR1 contains an N-terminal extracellular domain (ECD) that provides the primary ligand binding determinants. Here we present three crystal structures of the human CRFR1 ECD, one in a ligand-free form and two in distinct CRF-bound states. The CRFR1 ECD adopts the alpha-beta-betaalpha fold observed for other class B GPCR ECDs, but the N-terminal alpha-helix is significantly shorter and does not contact CRF. CRF adopts a continuous alpha-helix that docks in a hydrophobic surface of the ECD that is distinct from the peptide-binding site of other class B GPCRs, thereby providing a basis for the specificity of ligand recognition between CRFR1 and other class B GPCRs. The binding of CRF is accompanied by clamp-like conformational changes of two loops of the receptor that anchor the CRF C terminus, including the C-terminal amide group. These structural studies provide a molecular framework for understanding peptide binding and specificity by the CRF receptors as well as a template for designing potent and selective CRFR1 antagonists for therapeutic applications.

  3. Dual role of the p75 tumor necrosis factor (TNF) receptor in TNF cytotoxicity

    DEFF Research Database (Denmark)

    Bigda, J; Beletsky, I; Brakebusch, C

    1994-01-01

    Whereas there is ample evidence for involvement of the p55 tumor necrosis factor (TNF) receptor (p55-R) in the cytocidal effect of TNF, the role of the p75 TNF receptor (p75-R) in this effect is a matter of debate. In this study, we probed the function of p75-R in cells sensitive...... to the cytotoxicity of TNF using a wide panel of antibodies (Abs) against the receptor's extracellular domain. Two distinct Ab effects were observed. The Abs triggered signaling for cytotoxicity. This effect: (a) was correlated with the extent of p75-R expression by the cells; (b) was dependent on receptor cross...... against p55-R. Moreover, it seemed to reverse induced desensitization to the cytocidal effect of anti p55-R Abs, suggesting that it involves mechanisms different from those of the signaling by the p55 TNF-R. In addition, the Abs affected the response to TNF in a way that does not involve the signaling...

  4. Functionality of intrinsic disorder in tumor necrosis factor-α and its receptors.

    Science.gov (United States)

    Uversky, Vladimir N; El-Baky, Nawal Abd; El-Fakharany, Esmail M; Sabry, Amira; Mattar, Ehab H; Uversky, Alexey V; Redwan, Elrashdy M

    2017-11-01

    Tumor necrosis factor-α (TNF-α) is a pleiotropic inflammatory cytokine that exerts potent cytotoxic effects on solid tumor cells, while not affecting their normal counterparts. It is also known that TNF-α exerts many of its biological functions via interaction with specific receptors. To understand the potential roles of intrinsic disorder in the functioning of this important cytokine, we explored the peculiarities of intrinsic disorder distribution in human TNF-α and its homologs from various species, ranging from zebrafish to chimpanzee. We also studied the peculiarities of intrinsic disorder distribution in human TNF-α receptors, TNFR1 and TNFR2. Analysis revealed that cytoplasmic domains of TNF-α and its receptors are expected to be highly disordered. Furthermore, although the sequence identities of analyzed TNF-α homologs range from 99.57% (between human and chimpanzee proteins) to 22.33% (between frog and fish proteins), their intrinsic disorder profiles are characterized by a remarkable similarity. These observations indicate that the peculiarities of distribution of the intrinsic disorder propensity within the amino acid sequences are evolutionary conserved, and therefore could be of functional importance for this family of proteins. We also show that disordered and flexible regions of human TNF-α and its TNFR1 and TNFR2 receptors are crucial for some of their biological activities. © 2017 Federation of European Biochemical Societies.

  5. Properties of virion transactivator proteins encoded by primate cytomegaloviruses

    Directory of Open Access Journals (Sweden)

    Barry Peter A

    2009-05-01

    Full Text Available Abstract Background Human cytomegalovirus (HCMV is a betaherpesvirus that causes severe disease in situations where the immune system is immature or compromised. HCMV immediate early (IE gene expression is stimulated by the virion phosphoprotein pp71, encoded by open reading frame (ORF UL82, and this transactivation activity is important for the efficient initiation of viral replication. It is currently recognized that pp71 acts to overcome cellular intrinsic defences that otherwise block viral IE gene expression, and that interactions of pp71 with the cell proteins Daxx and ATRX are important for this function. A further property of pp71 is the ability to enable prolonged gene expression from quiescent herpes simplex virus type 1 (HSV-1 genomes. Non-human primate cytomegaloviruses encode homologs of pp71, but there is currently no published information that addresses their effects on gene expression and modes of action. Results The UL82 homolog encoded by simian cytomegalovirus (SCMV, strain Colburn, was identified and cloned. This ORF, named S82, was cloned into an HSV-1 vector, as were those from baboon, rhesus monkey and chimpanzee cytomegaloviruses. The use of an HSV-1 vector enabled expression of the UL82 homologs in a range of cell types, and permitted investigation of their abilities to direct prolonged gene expression from quiescent genomes. The results show that all UL82 homologs activate gene expression, and that neither host cell type nor promoter target sequence has major effects on these activities. Surprisingly, the UL82 proteins specified by non-human primate cytomegaloviruses, unlike pp71, did not direct long term expression from quiescent HSV-1 genomes. In addition, significant differences were observed in the intranuclear localization of the UL82 homologs, and in their effects on Daxx. Strikingly, S82 mediated the release of Daxx from nuclear domain 10 substructures much more rapidly than pp71 or the other proteins tested. All

  6. Tissue factor is an angiogenic-specific receptor for factor VII-targeted immunotherapy and photodynamic therapy.

    Science.gov (United States)

    Hu, Zhiwei; Cheng, Jijun; Xu, Jie; Ruf, Wolfram; Lockwood, Charles J

    2017-02-01

    Identification of target molecules specific for angiogenic vascular endothelial cells (VEC), the inner layer of pathological neovasculature, is critical for discovery and development of neovascular-targeting therapy for angiogenesis-dependent human diseases, notably cancer, macular degeneration and endometriosis, in which vascular endothelial growth factor (VEGF) plays a central pathophysiological role. Using VEGF-stimulated vascular endothelial cells (VECs) isolated from microvessels, venous and arterial blood vessels as in vitro angiogenic models and unstimulated VECs as a quiescent VEC model, we examined the expression of tissue factor (TF), a membrane-bound receptor on the angiogenic VEC models compared with quiescent VEC controls. We found that TF is specifically expressed on angiogenic VECs in a time-dependent manner in microvessels, venous and arterial vessels. TF-targeted therapeutic agents, including factor VII (fVII)-IgG1 Fc and fVII-conjugated photosensitizer, can selectively bind angiogenic VECs, but not the quiescent VECs. Moreover, fVII-targeted photodynamic therapy can selectively and completely eradicate angiogenic VECs. We conclude that TF is an angiogenic-specific receptor and the target molecule for fVII-targeted therapeutics. This study supports clinical trials of TF-targeted therapeutics for the treatment of angiogenesis-dependent diseases such as cancer, macular degeneration and endometriosis.

  7. Macrophage colony-stimulating factor, CSF-1, and its proto-oncogene-encoded receptor

    International Nuclear Information System (INIS)

    Sherr, C.J.; Rettenmier, C.W.; Roussel, M.F.

    1988-01-01

    The macrophage colony-stimulating factor, CSF-1, or M-CSF, is one of a family of hematopoietic growth factors that stimulates the proliferation of monocytes, macrophages, and their committed bone marrow progenitors. Unlike pluripotent hemopoietins such as granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-3 (IL-3 or multi-CSF), which affect the growth of myeloid cells of several different hematopoietic lineages, CSF-1 acts only on cells of the mononuclear phagocyte series to stimulate their growth and enhance their survival. Retroviral transduction of the feline c-fms gene in the Susan McDonough and Hardy Zuckerman-5 (HZ-5) strains of feline sarcoma virus (FeSV) led to genetic alterations that endowed the recombined viral oncogene (v-fms) with the ability to transform cells in culture morphologically and to induce firbrosarcomas and hematopoietic neoplasms in susceptible animals. The v-fms oncogene product differs from the normal CSF-1 receptor in certain of its cardinal biochemical properties, most notably in exhibiting constitutively high basal levels of tyrosine kinase activity in the absence of its ligand. Comparative studies of the c-fms and v-fms genes coupled with analyses of engineered mutants and receptor chimeras have begun to pinpoint pertinent genetic alterations in the normal receptor gene that unmask its latent oncogenic potential. In addition, the availability of biologically active c-fms, v-fms, and CSF-1 cDNAs has allowed these genes to be mobilized and expressed in naive cells, thereby facilitating assays for receptor coupling with downstream components of the mitogenic pathway in diverse cell types

  8. Antibody-induced activation of the epidermal growth factor receptor tyrosine kinase requires the presence of detergent

    NARCIS (Netherlands)

    Spaargaren, M.; Defize, L. H.; de Laat, S. W.; Boonstra, J.

    1990-01-01

    Activation of the epidermal growth factor receptor (EGF-R) tyrosine kinase was investigated in membrane preparations as well as intact A431 cells, using anti-EGF-R antibodies directed against extra- and intracellular receptor domains. In vitro assay conditions were mimicked on whole cells by a mild

  9. Prognostic factors in advanced breast cancer: Race and receptor status are significant after development of metastasis.

    Science.gov (United States)

    Ren, Zhiyong; Li, Yufeng; Shen, Tiansheng; Hameed, Omar; Siegal, Gene P; Wei, Shi

    2016-01-01

    Prognostic factors are well established in early-stage breast cancer (BC), but less well-defined in advanced disease. We analyzed 323 BC patients who had distant relapse during follow-up from 1997 to 2010 to determine the significant clinicopathologic factors predicting survival outcomes. By univariate analysis, race, tumor grade, estrogen and progesterone receptors (ER/PR) and HER2 status were significantly associated with overall survival (OS) and post-metastasis survival (PMS). Applying a Cox regression model revealed that all these factors remained significant for PMS, while race, tumor grade and HER2 were independent factors for OS. Tumor grade was the only significant factor for metastasis-free survival by univariate and multivariate analyses. Our findings demonstrated that being Caucasian, hormonal receptor positive (HR+) and HER2 positive (HER2+) were all associated with a decreased hazard of death and that patients with HR+/HER2+ tumors had superior outcomes to those with HR+/HER2- disease. Further, PR status held a prognostic value over ER, thus reflecting the biologic mechanism of the importance of the functional ER pathway and the heterogeneity in the response to endocrine therapy. These observations indicate that the patients' genetic makeup and the intrinsic nature of the tumor principally govern BC progression and prognosticate the long-term outcomes in advanced disease. Copyright © 2015 Elsevier GmbH. All rights reserved.

  10. Activation of epidermal growth factor receptor mediates mucin production stimulated by p40, a Lactobacillus rhamnosus GG-derived protein.

    Science.gov (United States)

    Wang, Lihong; Cao, Hailong; Liu, Liping; Wang, Bangmao; Walker, W Allan; Acra, Sari A; Yan, Fang

    2014-07-18

    The mucus layer coating the gastrointestinal tract serves as the first line of intestinal defense against infection and injury. Probiotics promote mucin production by goblet cells in the intestine. p40, a Lactobacillus rhamnosus GG-derived soluble protein, has been shown to transactivate the EGF receptor (EGFR) in intestinal epithelial cells, which is required for inhibition of apoptosis and preservation of barrier function in the colon, thereby ameliorating intestinal injury and colitis. Because activation of EGFR has been shown to up-regulate mucin production in goblet cells, the purpose of this study was to investigate the effects and mechanisms of p40 regulation of mucin production. p40 activated EGFR and its downstream target, Akt, in a concentration-dependent manner in LS174T cells. p40 stimulated Muc2 gene expression and mucin production in LS174T cells, which were abolished by inhibition of EGFR kinase activity, down-regulation of EGFR expression by EGFR siRNA transfection, or suppression of Akt activation. Treatment with p40 increased mucin production in the colonic epithelium, thus thickening the mucus layer in the colon of wild type, but not of Egfr(wa5) mice, which have a dominant negative mutation in the EGFR kinase domain. Furthermore, inhibition of mucin-type O-linked glycosylation suppressed the effect of p40 on increasing mucin production and protecting intestinal epithelial cells from TNF-induced apoptosis in colon organ culture. Thus, these results suggest that p40-stimulated activation of EGFR mediates up-regulation of mucin production, which may contribute to the mechanisms by which p40 protects the intestinal epithelium from injury. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Expression, purification, and characterization of a diabody against the most important angiogenesis cell receptor: Vascular endothelial growth factor receptor 2

    Directory of Open Access Journals (Sweden)

    Mahdi Behdani

    2012-01-01

    Full Text Available Antibodies and their derivative fragments have long been used as tools in a variety of applications, in fundamental research work, biotechnology, diagnosis, and therapy. Camels produce single heavy-chain antibodies (VHH in addition to usual antibodies. These minimal-sized binders are very robust and bind the antigen with high affinity in a monomeric state. Vascular endothelial growth factor recepror-2 (VEGFR2 is an important tumor-associated receptor that blockade of its signaling can lead to the inhibition of neovascularization and tumor metastasis. Here, we describe the construction, expression, and purification VEGFR2-specific Diabody. Two variable fragments of a same camel anti-VEGFR2 antibody were linked together by the upper hinge segment of antibody to make a diabody. We showed the ability of diabody to recognition of VEGFR2 on the cell surface by FACS. Diabodies can be produced in the low-cost prokaryotic expression system, so they are suitable molecules for diagnostic and therapeutic issues.

  12. p53 transactivation and the impact of mutations, cofactors and small molecules using a simplified yeast-based screening system.

    Directory of Open Access Journals (Sweden)

    Virginia Andreotti

    Full Text Available The p53 tumor suppressor, which is altered in most cancers, is a sequence-specific transcription factor that is able to modulate the expression of many target genes and influence a variety of cellular pathways. Inactivation of the p53 pathway in cancer frequently occurs through the expression of mutant p53 protein. In tumors that retain wild type p53, the pathway can be altered by upstream modulators, particularly the p53 negative regulators MDM2 and MDM4.Given the many factors that might influence p53 function, including expression levels, mutations, cofactor proteins and small molecules, we expanded our previously described yeast-based system to provide the opportunity for efficient investigation of their individual and combined impacts in a miniaturized format. The system integrates i variable expression of p53 proteins under the finely tunable GAL1,10 promoter, ii single copy, chromosomally located p53-responsive and control luminescence reporters, iii enhanced chemical uptake using modified ABC-transporters, iv small-volume formats for treatment and dual-luciferase assays, and v opportunities to co-express p53 with other cofactor proteins. This robust system can distinguish different levels of expression of WT and mutant p53 as well as interactions with MDM2 or 53BP1.We found that the small molecules Nutlin and RITA could both relieve the MDM2-dependent inhibition of WT p53 transactivation function, while only RITA could impact p53/53BP1 functional interactions. PRIMA-1 was ineffective in modifying the transactivation capacity of WT p53 and missense p53 mutations. This dual-luciferase assay can, therefore, provide a high-throughput assessment tool for investigating a matrix of factors that can influence the p53 network, including the effectiveness of newly developed small molecules, on WT and tumor-associated p53 mutants as well as interacting proteins.

  13. Transforming growth factor betas and their signaling receptors in human hepatocellular carcinoma.

    Science.gov (United States)

    Abou-Shady, M; Baer, H U; Friess, H; Berberat, P; Zimmermann, A; Graber, H; Gold, L I; Korc, M; Büchler, M W

    1999-03-01

    Transforming growth factor betas (TGF-betas) are multifunctional polypeptides that have been suggested to influence tumor growth. They mediate their functions via specific cell surface receptors (type I ALK5 and type II TGF-beta receptors). The aim of this study was to analyze the roles of the three TGF-betas and their signaling receptors in human hepatocellular carcinoma (HCC). HCC tissue samples were obtained from 18 patients undergoing partial liver resection. Normal liver tissues from 7 females and 3 males served as controls. The tissues for histological analysis were fixed in Bouin's solution and paraffin embedded. For RNA analysis, freshly obtained tissue samples were snap frozen in liquid nitrogen and stored at -80 degrees C until used. Northern blot analysis was used in normal liver and HCC to examine the expression of TGF-beta1, -beta2, -beta3 and their receptors: type I ALK5 (TbetaR-I ALK5), type II (TbetaR-II), and type III (TbetaR-III). Immunohistochemistry was performed to localize the corresponding proteins. All three TGF-betas demonstrated a marked mRNA overexpression in HCC in comparison with normal controls, whereas the levels of all three TGF-beta receptors showed no significant changes. Intense TGF-beta1, TGF-beta2, and TGF-beta3 immunostaining was found in hepatocellular carcinoma cells and in the perineoplastic stroma with immunohistochemistry, whereas no or mild immunostaining was present in the normal liver. For TbetaR-I ALK5 and TbetaR-II, the immunostaining in both HCC and normal liver was mild to moderate, with a slightly higher intensity in the normal tissues. The upregulation of TGF-betas in HCC suggests an important role for these isoforms in hepatic carcinogenesis and tumor progression. Moreover, the localization of the immunoreactivity in both malignant hepatocytes and stromal cells suggests that TGF-betas act via autocrine and paracrine pathways in this neoplasm.

  14. Platelet activating factor receptor binding plays a critical role in jet fuel-induced immune suppression

    International Nuclear Information System (INIS)

    Ramos, Gerardo; Kazimi, Nasser; Nghiem, Dat X.; Walterscheid, Jeffrey P.; Ullrich, Stephen E.

    2004-01-01

    Applying military jet fuel (JP-8) or commercial jet fuel (Jet-A) to the skin of mice suppresses the immune response in a dose-dependant manner. The release of biological response modifiers, particularly prostaglandin E 2 (PGE 2 ), is a critical step in activating immune suppression. Previous studies have shown that injecting selective cyclooxygenase-2 inhibitors into jet fuel-treated mice blocks immune suppression. Because the inflammatory phospholipid mediator, platelet-activating factor (PAF), up-regulates cyclooxygenase-2 production and PGE 2 synthesis by keratinocytes, we tested the hypothesis that PAF-receptor binding plays a role in jet fuel-induced immune suppression. Treating keratinocyte cultures with PAF and/or jet fuel (JP-8 and Jet-A) stimulates PGE 2 secretion. Jet fuel-induced PGE 2 production was suppressed by treating the keratinocytes with specific PAF-receptor antagonists. Injecting mice with PAF, or treating the skin of the mice with JP-8, or Jet-A, induced immune suppression. Jet fuel-induced immune suppression was blocked when the jet fuel-treated mice were injected with PAF-receptor antagonists before treatment. Jet fuel treatment has been reported to activate oxidative stress and treating the mice with anti-oxidants (Vitamins C, or E or beta-hydroxy toluene), before jet fuel application, interfered with immune suppression. These findings confirm previous studies showing that PAF-receptor binding can modulate immune function. Furthermore, they suggest that PAF-receptor binding may be an early event in the induction of immune suppression by immunotoxic environmental agents that target the skin

  15. CD36 and Platelet-Activating Factor Receptor Promote House Dust Mite Allergy Development.

    Science.gov (United States)

    Patel, Preeyam S; Kearney, John F

    2017-08-01

    Over 89% of asthmatic children in underdeveloped countries demonstrate sensitivity to house dust mites (HDMs). The allergic response to HDMs is partially mediated by epithelial cell-derived cytokines that activate group 2 innate lymphoid cells, induce migration and activation of dendritic cells, and promote effector differentiation of HDM-specific TH2 cells. However, the contribution of innate receptor engagement on epithelial or dendritic cells by HDMs that ultimately mediates said innate and adaptive allergic responses is poorly understood. We and other investigators have demonstrated that HDMs express phosphorylcholine (PC) moieties. The major PC receptors involved in immune responses include CD36 and platelet-activating factor receptor (PAFR). Because CD36 and PAFR are expressed by epithelial cells and dendritic cells, and expression of these receptors is higher in human asthmatics, we determined whether engagement of CD36 or PAFR on epithelial or dendritic cells contributes to HDM allergy development. Testing bone marrow chimeric mice revealed that CD36 engagement on radioresistant cells and PAFR engagement on radioresistant and radiosensitive cells in the lung promote allergic responses to HDMs. Additionally, passive anti-PC IgM Abs administered intratracheally with HDMs decreased allergen uptake by epithelial cells and APCs in the lungs of C57BL/6 mice but not CD36 -/- or PAFR -/- mice. These results show that CD36 and PAFR are important mediators of HDM allergy development and that inhibiting HDM engagement with PC receptors in the lung protects against allergic airway disease. Copyright © 2017 by The American Association of Immunologists, Inc.

  16. Complex Relationship between Ligand Binding and Dimerization in the Epidermal Growth Factor Receptor

    Directory of Open Access Journals (Sweden)

    Nicholas J. Bessman

    2014-11-01

    Full Text Available The epidermal growth factor receptor (EGFR plays pivotal roles in development and is mutated or overexpressed in several cancers. Despite recent advances, the complex allosteric regulation of EGFR remains incompletely understood. Through efforts to understand why the negative cooperativity observed for intact EGFR is lost in studies of its isolated extracellular region (ECR, we uncovered unexpected relationships between ligand binding and receptor dimerization. The two processes appear to compete. Surprisingly, dimerization does not enhance ligand binding (although ligand binding promotes dimerization. We further show that simply forcing EGFR ECRs into preformed dimers without ligand yields ill-defined, heterogeneous structures. Finally, we demonstrate that extracellular EGFR-activating mutations in glioblastoma enhance ligand-binding affinity without directly promoting EGFR dimerization, suggesting that these oncogenic mutations alter the allosteric linkage between dimerization and ligand binding. Our findings have important implications for understanding how EGFR and its relatives are activated by specific ligands and pathological mutations.

  17. A Premature Termination of Human Epidermal Growth Factor Receptor Transcription in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Jihene Elloumi-Mseddi

    2014-01-01

    Full Text Available Our success in producing an active epidermal growth factor receptor (EGFR tyrosine kinase in Escherichia coli encouraged us to express the full-length receptor in the same host. Despite its large size, we were successful at producing the full-length EGFR protein fused to glutathione S-transferase (GST that was detected by Western blot analysis. Moreover, we obtained a majoritarian truncated GST-EGFR form detectable by gel electrophoresis and Western blot. This truncated protein was purified and confirmed by MALDI-TOF/TOF analysis to belong to the N-terminal extracellular region of the EGFR fused to GST. Northern blot analysis showed two transcripts suggesting the occurrence of a transcriptional arrest.

  18. Treatment related changes of the serum epidermal growth factor receptor in advanced colorectal cancer

    DEFF Research Database (Denmark)

    Spindler, K G; Aalund Olsen, Dorte; Brandslund, I

    2009-01-01

    e22096 Background: The epidermal growth factor receptor (EGFR) is an established target for therapy in colorectal cancer. The extracellular domain of the receptor is shed into circulation and detectable by ELISA. We investigated the changes in sEGFR levels during preoperative chemoradiation (CRT......) in rectal cancer patients and third-line treatment with cetuximab and irinotecan (CETIRI) in advanced disease, to elucidate the predictive or prognostic value in these settings. METHODS: We included 126 healthy controls and 118 patients with chemorefractory mCRC treated with cetuximab (initial 400/m(2) mg...... significant differences between the median pre-treatment sEGFR levels in controls, rectal cancer and mCRC (58 ng/ml(56-59 95% C-I), 53 ng/ml(51-55 95% C-I) and 51 ng/ml(49-53 95% C-I), respectively, ptreatment values during CETIRI (p

  19. Expression and purification of Nod factor receptors - Initial characterization of ligand binding

    DEFF Research Database (Denmark)

    Broghammer, Angelique

    . Lipochitooligosaccharides also serve as signals in the mutually beneficial interactions between arbuscular mycorrhiza (AM) and most land plants. In the model legume Lotus japonicus the Nod factor receptors, LjNFR1 and LjNFR5, two LysM receptor like kinases (LysM-RLK), are responsible for perceiving the rhizobial...... and LjNFR5 ectodomains were glycosylated; 3) LjNFR1 retained its in vitro kinase activity and 4) LjNFR1 and LjNFR5 were localized to the plasma membrane. In depth mass spectroscopy analysis of the N-glycan structure of LjNFR5 resulted in identification of two different glycan structures with identical...

  20. Detection and Quantification of Vascular Endothelial Growth Factor Receptor Tyrosine Kinases in Primary Human Endothelial Cells.

    Science.gov (United States)

    Fearnley, Gareth W; Wheatcroft, Stephen B; Ponnambalam, Sreenivasan

    2015-01-01

    Proteins differ widely in their pattern of expression depending on organism, tissue, and regulation in response to changing conditions. In the mammalian vasculature, the endothelium responds to vascular endothelial growth factors (VEGFs) via membrane-bound receptor tyrosine kinases (VEGFRs) to modulate many aspects of vascular physiology including vasculogenesis, angiogenesis, and blood pressure. Studies on VEGFR biology are thus dependent on detecting expression levels in different cell types and evaluating how changes in protein levels correlate with changing conditions including circulating VEGF levels. Here, we present a robust immunoblot-based protocol for detecting and quantifying VEGFRs in human endothelial cells. Using internal and external standards, we can rapidly evaluate receptor copy number and assess how this is altered in response to the cellular environment.

  1. Recent Advances of Colony-Stimulating Factor-1 Receptor (CSF-1R) Kinase and Its Inhibitors.

    Science.gov (United States)

    El-Gamal, Mohammed I; Al-Ameen, Shahad K; Al-Koumi, Dania M; Hamad, Mawadda G; Jalal, Nouran A; Oh, Chang-Hyun

    2018-01-17

    Colony stimulation factor-1 receptor (CSF-1R), which is also known as FMS kinase, plays an important role in initiating inflammatory, cancer, and bone disorders when it is overstimulated by its ligand, CSF-1. Innate immunity, as well as macrophage differentiation and survival, are regulated by the stimulation of the CSF-1R. Another ligand, interlukin-34 (IL-34), was recently reported to activate the CSF-1R receptor in a different manner. The relationship between CSF-1R and microglia has been reviewed. Both CSF-1 antibodies and small molecule CSF-1R kinase inhibitors have now been tested in animal models and in humans. In this Perspective, we discuss the role of CSF-1 and IL-34 in producing cancer, bone disorders, and inflammation. We also review the newly discovered and improved small molecule kinase inhibitors and monoclonal antibodies that have shown potent activity toward CSF-1R, reported from 2012 until 2017.

  2. [Clinical significance of tumor content of epidermal growth factor receptor in breast cancer].

    Science.gov (United States)

    Mulero, M; Fernández Raigoso, P; Vázquez, J; Lamelas, M L; Corte, D; Allende, M T; Rodríguez, J C; Vizoso, F

    2003-11-01

    To determine the content of epidermal growth factor receptor (EGFR) using a radioligand method in breast cancer and to analyze the relationship between the EGFR levels and the characteristics of patients and tumors. Prognostic significance was also analyzed. EGFR was measured by a single point radioligand assay in 265 invasive breast carcinomas tissues. In addition, estrogen and progesterone receptors (ER and PR) were measured by enzymatic immunoassays. We analyze the relationship of EGFR levels with the different clinico-pathologic parameters. EGFR levels in breast carcinomas varied widely (0.1 to 403) with a median at 4 fmol/mg prot. The significantly higher concentrations of EGFR were detected in patients under 60 years old (p = 0.042), undifferentiated tumors (p = 0.04), and carcinomas with negative ER and PR (p aggresive characteristics of the tumor.

  3. receptores

    Directory of Open Access Journals (Sweden)

    Salete Regina Daronco Benetti

    2006-01-01

    Full Text Available Se trata de un estudio etnográfico, que tuvo lo objetivo de interpretar el sistema de conocimiento y del significado atribuidos a la sangre referente a la transfusión sanguínea por los donadores y receptores de un banco de sangre. Para la colecta de las informaciones se observaron los participantes y la entrevista etnográfica se realizó el análisis de dominio, taxonómicos y temáticos. Los dominios culturales fueron: la sangre es vida: fuente de vida y alimento valioso; creencias religiosas: fuentes simbólicas de apoyos; donación sanguínea: un gesto colaborador que exige cuidarse, gratifica y trae felicidad; donación sanguínea: fuente simbólica de inseguridad; estar enfermo es una condición para realizar transfusión sanguínea; transfusión sanguínea: esperanza de vida; Creencias populares: transfusión sanguínea como riesgo para la salud; donadores de sangre: personas benditas; donar y recibir sangre: como significado de felicidad. Temática: “líquido precioso que origina, sostiene, modifica la vida, provoca miedo e inseguridad”.

  4. Receptors for insulin-like growth factors I and II: autoradiographic localization in rat brain and comparison to receptors for insulin

    International Nuclear Information System (INIS)

    Lesniak, M.A.; Hill, J.M.; Kiess, W.; Rojeski, M.; Pert, C.B.; Roth, J.

    1988-01-01

    Receptors for insulin-like growth factor I (IGF-I) in rat brain were visualized using autoradiography with [125I]IGF-I. The binding of the labeled peptide was competed for fully by high concentrations of unlabeled IGF-I. At intermediate concentrations of unlabeled peptide the binding of [125I]IGF-I was competed for by unlabeled IGF-I more effectively than by IGF-II or insulin, which is typical of receptors for IGF-I. Essentially every brain section shows specific binding of IGF-I, and the pattern of binding of IGF-I to its receptors correlated well with the cytoarchitectonic structures. In parallel studies we showed that [125I]IGF-II was bound to tissue sections of rat brain and that the binding was competed for by an excess of unlabeled IGF-II. However, intermediate concentrations of unlabeled peptides gave inconclusive results. To confirm that the binding of [125I]IGF-II was to IGF-II receptors, we showed that antibodies specific for the IGF-II receptor inhibited the binding of labeled IGF-II. Furthermore, the binding of the antibody to regions of the brain section, visualized by the application of [125I]protein-A, gave patterns indistinguishable from those obtained with [125I]IGF-II alone. Again, the binding was very widely distributed throughout the central nervous system, and the patterns of distribution corresponded well to the underlying neural structures. Densitometric analysis of the receptors enabled us to compare the distribution of IGF-I receptors with that of IGF-II receptors as well as retrospectively with that of insulin receptors

  5. The E3 ubiquitin ligase WWP2 facilitates RUNX2 protein transactivation in a mono-ubiquitination manner during osteogenic differentiation.

    Science.gov (United States)

    Zhu, Wei; He, Xinyu; Hua, Yue; Li, Qian; Wang, Jiyong; Gan, Xiaoqing

    2017-07-07

    Poly-ubiquitination-mediated RUNX2 degradation is an important cause of age- and inflammation-related bone loss. NEDD4 family E3 ubiquitin protein ligases are thought to be the major regulators of RUNX2 poly-ubiquitination. However, we observed a mono-ubiquitination of RUNX2 that was catalyzed by WWP2, a member of the NEDD4 family of E3 ubiquitin ligases. WWP2 has been reported to catalyze the mono-ubiquitination of Goosecoid in chondrocytes, facilitating craniofacial skeleton development. In this study, we found that osteogenic differentiation of mesenchymal stem cells promoted WWP2 expression and nuclear accumulation. Knockdown of Wwp2 in mesenchymal stem cells and osteoblasts led to significant deficiencies of osteogenesis, including decreased mineral deposition and down-regulation of osteogenic marker genes. Co-immunoprecipitation experiments showed the interaction of WWP2 with RUNX2 in vitro and in vivo Mono-ubiquitination by WWP2 leads to RUNX2 transactivation, as evidenced by the wild type of WWP2, but not its ubiquitin ligase-dead mutant, augmenting RUNX2-reponsive reporter activity. Moreover, deletion of WWP2-dependent mono-ubiquitination resulted in striking defects of RUNX2 osteoblastic activity. In addition, ectopic expression of the constitutively active type 1A bone morphogenetic protein receptor enhanced WWP2-dependent RUNX2 ubiquitination and transactivation, demonstrating a regulatory role of bone morphogenetic protein signaling in the WWP2-RUNX2 axis. Taken together, our results provide evidence that WWP2 serves as a positive regulator of osteogenesis by augmenting RUNX2 transactivation in a non-proteolytic mono-ubiquitination manner. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. C-terminal truncated forms of Met, the hepatocyte growth factor receptor.

    Science.gov (United States)

    Prat, M; Crepaldi, T; Gandino, L; Giordano, S; Longati, P; Comoglio, P

    1991-12-01

    The MET proto-oncogene encodes a transmembrane tyrosine kinase of 190 kDa (p190MET), which has recently been identified as the receptor for hepatocyte growth factor/scatter factor. p190MET is a heterodimer composed of two disulfide-linked chains of 50 kDa (p50 alpha) and 145 kDa (p145 beta). We have produced four different monoclonal antibodies that are specific for the extracellular domain of the Met receptor. These antibodies immunoprecipitate with p190MET two additional Met proteins of 140 and 130 kDa. The first protein (p140MET) is membrane bound and is composed of an alpha chain (p50 alpha) and an 85-kDa C-terminal truncated beta chain (p85 beta). The second protein (p130MET) is released in the culture supernatant and consists of an alpha chain (p50 alpha) and a 75-kDa C-terminal truncated beta chain (p75 beta). Both truncated forms lack the tyrosine kinase domain. p140MET and p130MET are consistently detected in vivo, together with p190MET, in different cell lines or their culture supernatants. p140MET is preferentially localized at the cell surface, where it is present in roughly half the amount of p190MET. The two C-terminal truncated forms of the Met receptor are also found in stable transfectants expressing the full-length MET cDNA, thus showing that they originate from posttranslational proteolysis. This process is regulated by protein kinase C activation. Together, these data suggest that the production of the C-terminal truncated Met forms may have a physiological role in modulating the Met receptor function.

  7. Advanced Research of Fibroblast Growth Factor Receptor 
in Non-small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Dan PU

    2013-11-01

    Full Text Available Lung cancer is severely threatening human health. In recent years, the treatment for lung adenocarcinoma has made a great progress, targeted therapy has been widely applied in clinic, and benefits amount of patients. However, in squamous cell lung cancer, the incidence of epidermal growth factor receptor (EGFR gene mutant and ALK fusion gene are low,and targeted therapy like Tarceva and crizotinib, can hardly work. Since the fibroblast growth factors (fibroblast growth factor, FGF pathway is considered to be related to tumor cell proliferation, metastasis and angiogenesis, more and more researches proved the amplification of fibroblast growth factor receptor (FGFR in squamous cell lung cancer. Experiments in vivo and in vitro found that blocking FGF pathway could reduce the proliferation of tumor cells and inhibit metastasis. The FGF pathway might be a new target for treatment of squamous cell lung cancer. This article reviews the effect of FGFR in tumorigenesis,as well as the prospect as a therapeutic target in non-small cell lung cancer.

  8. Prognostic value of insulin- like growth factor-I receptor expression in renal cell carcinoma

    Directory of Open Access Journals (Sweden)

    Sichani Mehrdad

    2010-01-01

    Full Text Available The Insulin-Like growth factor-I receptor (IGF-IR, a tyrosine-kinas receptor over expressed in many tumor cell lines and in some human tumors, plays a critical role in trans-formation, tumorigenicity and metastasis. The aim of the present study is to investigate the role of IGF-IR expression as a prognostic factor in RCC. This study was conducted in a historical cohort of 82 patients who had RCC treated with radical nephrectomy from 1994 to 2005. Specimens were reevaluated with regard to histological subtype, nuclear grade, stage and IGF-IR expression. The IGF-IR stain was semi- quantitatively evaluated using the Allred score system. Kaplan-Meier analysis demonstrated a significant positive correlation between Fuhrman nuclear grade and IGF-IR Allred score (P< 0.0001. Survival in patients with score IGF-I ≤ 4 was 90.21 month and in patients with score IGF-1R> 4 was 33.39 month (P Value < 0.0001. Cox regression analysis in-dicated that expression of IGF-IR is a prognostic factor in patients with RCC (P Value < 0.0001, odds Ratio = 2.38. In conclusion, a statistically significant correlation was demonstrated between IGF-IR expression and Fuhrman nuclear grading and survival in patients with RCC. In stage-by-stage and grade-by-grade analysis; however, it seems that we cannot consider IGF-IR as an inde-pendent prognostic factor.

  9. Prognostic value of insulin- like growth factor-I receptor expression in renal cell carcinoma.

    Science.gov (United States)

    Sichani, Mehrdad Mohammadi; Yazdi, Fateme Sarreshtedar; Moghaddam, Noushin Afshar; Chehrei, Ali; Kabiri, Mahmud; Naeimi, Amin; Taheri, Diana

    2010-01-01

    The insulin-like growth factor-I receptor (IGF-IR), a tyrosine kinase receptor over expressed in many tumor cell lines and in some human tumors, plays a critical role in transformation, tumorigenicity and metastasis. The aim of the present study is to investigate the role of IGF-IR expression as a prognostic factor in RCC. This study was conducted in a historical cohort of 82 patients who had RCC treated with radical nephrectomy from 1994 to 2005. Specimens were reevaluated with regard to histological subtype, nuclear grade, stage and IGF-IR expression. The IGF-IR stain was semi-quantitatively evaluated using the Allred score system. Kaplan-Meier analysis demonstrated a significant positive correlation between Fuhrman nuclear grade and IGF-IR Allred score (P 4 was 33.39 month (P Value < 0.0001). Cox regression analysis indicated that expression of IGF-IR is a prognostic factor in patients with RCC (P Value < 0.0001, odds Ratio = 2.38). In conclusion, a statistically significant correlation was demonstrated between IGF-IR expression and Fuhrman nuclear grading and survival in patients with RCC. In stage-by-stage and grade-by-grade analysis; however, it seems that we cannot consider IGF-IR as an independent prognostic factor.

  10. Contribution to the investigation of the p53 in vivo and in vitro trans-activation activity

    International Nuclear Information System (INIS)

    Meiller, A.

    2004-03-01

    Among the body's defence mechanisms, the programmed cellular death or apoptosis is an important safeguard way which allows the body to get rid of the injured cells before they acquire steady genetic modifications leading to an anarchistic multiplication. As p53 tumor suppressor gene plays a predominant role within this process, this research report first presents the p53 protein, its structure, its activities as a transcription factor, its modifications and the implications on its functional activities, its biological activities, and describes the p53 intracellular rate regulation and the use of this protein in radiology, particularly in 'in vivo' investigations on irradiated mice. It also presents the p53 family. Then, the author reports experimental investigations on possible other genes which could be trans-activated by p53. A gene is identified as a new target gene. She also demonstrates a new p53 activation path induced by another member of the p53 family, the p73 alpha protein

  11. Stimulation of LDL receptor activity in Hep-G2 cells by a serum factor(s)

    International Nuclear Information System (INIS)

    Ellsworth, J.L.; Brown, C.; Cooper, A.D.

    1988-01-01

    The regulation of low-density lipoprotein (LDL) receptor activity in the human hepatoma cell line Hep-G2 by serum components was examined. Incubation of dense monolayers of Hep-G2 cells with fresh medium containing 10% fetal calf serum (FM) produced a time-dependent increase in LDL receptor activity. Uptake and degradation of 125I-LDL was stimulated two- to four-fold, as compared with that of Hep-G2 cells cultured in the same media in which they had been grown to confluence (CM); the maximal 125I-LDL uptake plus degradation increased from 0.2 microgram/mg cell protein/4 h to 0.8 microgram/mg cell protein/4 h. In addition, a two-fold increase in cell surface binding of 125I-LDL to Hep-G2 cells was observed when binding was measured at 4 degrees C. There was no change in the apparent Kd. The stimulation of LDL receptor activity was suppressed in a concentration-dependent manner by the addition of cholesterol, as LDL, to the cell medium. In contrast to the stimulation of LDL receptor activity, FM did not affect the uptake or degradation of 125I-asialoorosomucoid. Addition of FM increased the protein content per dish, and DNA synthesis was stimulated approximately five-fold, as measured by [3H]thymidine incorporation into DNA; however, the cell number did not change. Cellular cholesterol biosynthesis was also stimulated by FM; [14C]acetate incorporation into unesterified and esterified cholesterol was increased approximately five-fold. Incubation of Hep-G2 cells with high-density lipoproteins (200 micrograms protein/ml) or albumin (8.0 mg/ml) in the absence of the serum factor did not significantly increase the total processed 125I-LDL. Stimulation of LDL receptor activity was dependent on a heat-stable, nondialyzable serum component that eluted in the inclusion volume of a Sephadex G-75 column

  12. BS69 : A novel adenovirus E1A-associated protein that inhibits E1A transactivation

    NARCIS (Netherlands)

    Hateboer, G.; Gennissen, A.M.C.; Ramos, Y.F.M.; Kerkhoven, R.; Sonntag-Buck, V.; Stunnenberg, H.G.; Bernards, R.A.

    1995-01-01

    The adenovirus ElA gene products are nuclear phosphoproteins that can transactivate the other adenovirus early genes as well as several cellular genes, and can transform primary rodent cells in culture. Transformation and transactivation by ElA proteins is most likely to be mediated through

  13. Novel targeted approaches to treating biliary tract cancer: the dual epidermal growth factor receptor and ErbB-2 tyrosine kinase inhibitor NVP-AEE788 is more efficient than the epidermal growth factor receptor inhibitors gefitinib and erlotinib.

    Science.gov (United States)

    Wiedmann, Marcus; Feisthammel, Jürgen; Blüthner, Thilo; Tannapfel, Andrea; Kamenz, Thomas; Kluge, Annett; Mössner, Joachim; Caca, Karel

    2006-08-01

    Aberrant activation of the epidermal growth factor receptor is frequently observed in neoplasia, notably in tumors of epithelial origin. Attempts to treat such tumors with epidermal growth factor receptor antagonists resulted in remarkable success in recent studies. Little is known, however, about the efficacy of this therapy in biliary tract cancer. Protein expression of epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 was assessed in seven human biliary tract cancer cell lines by immunoblotting. In addition, histological sections from 19 patients with extrahepatic cholangiocarcinoma were analyzed for epidermal growth factor receptor, ErbB-2 and vascular endothelial growth factor receptor-2 expression by immunohistochemistry. Moreover, we sequenced the cDNA products representing the entire epidermal growth factor receptor coding region of the seven cell lines, and searched for genomic epidermal growth factor receptor amplifications and polysomy by fluorescence in-situ hybridization. Cell growth inhibition by gefitinib erlotinib and NVP-AEE788 was studied in vitro by automated cell counting. In addition, the anti-tumoral effect of erlotinib and NVP-AEE788 was studied in a chimeric mouse model. The anti-tumoral drug mechanism in this model was assessed by MIB-1 antibody staining, terminal deoxynucleotidyl transfer-mediated dUTP nick end-labelling assay, von Willebrand factor staining, and immunoblotting for p-p42/44 (p-Erk1/2, p-MAPK) and p-AKT. Immunoblotting revealed expression of epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 in all biliary tract cancer cell lines. EGFR was detectable in six of 19 (32%) extrahepatic human cholangiocarcinoma tissue samples, ErbB-2 in 16 of 19 (84%), and vascular endothelial growth factor receptor-2 in nine of 19 (47%). Neither epidermal growth factor receptor mutations nor amplifications or polysomy were found in the seven biliary tract cancer

  14. Mammary tumors that become independent of the type I insulin-like growth factor receptor express elevated levels of platelet-derived growth factor receptors

    Directory of Open Access Journals (Sweden)

    Campbell Craig I

    2011-11-01

    Full Text Available Abstract Background Targeted therapies are becoming an essential part of breast cancer treatment and agents targeting the type I insulin-like growth factor receptor (IGF-IR are currently being investigated in clinical trials. One of the limitations of targeted therapies is the development of resistant variants and these variants typically present with unique gene expression patterns and characteristics compared to the original tumor. Results MTB-IGFIR transgenic mice, with inducible overexpression of the IGF-IR were used to model mammary tumors that develop resistance to IGF-IR targeting agents. IGF-IR independent mammary tumors, previously shown to possess characteristics associated with EMT, were found to express elevated levels of PDGFRα and PDGFRβ. Furthermore, these receptors were shown to be inversely expressed with the IGF-IR in this model. Using cell lines derived from IGF-IR-independent mammary tumors (from MTB-IGFIR mice, it was demonstrated that PDGFRα and to a lesser extent PDGFRβ was important for cell migration and invasion as RNAi knockdown of PDGFRα alone or PDGFRα and PDGFRβ in combination, significantly decreased tumor cell migration in Boyden chamber assays and suppressed cell migration in scratch wound assays. Somewhat surprisingly, concomitant knockdown of PDGFRα and PDGFRβ resulted in a modest increase in cell proliferation and a decrease in apoptosis. Conclusion During IGF-IR independence, PDGFRs are upregulated and function to enhance tumor cell motility. These results demonstrate a novel interaction between the IGF-IR and PDGFRs and highlight an important, therapeutically relevant pathway, for tumor cell migration and invasion.

  15. The viral transactivator HBx protein exhibits a high potential for regulation via phosphorylation through an evolutionarily conserved mechanism

    Directory of Open Access Journals (Sweden)

    Hernández Sergio

    2012-10-01

    Full Text Available Abstract Background Hepatitis B virus (HBV encodes an oncogenic factor, HBx, which is a multifunctional protein that can induce dysfunctional regulation of signaling pathways, transcription, and cell cycle progression, among other processes, through interactions with target host factors. The subcellular localization of HBx is both cytoplasmic and nuclear. This dynamic distribution of HBx could be essential to the multiple roles of the protein at different stages during HBV infection. Transactivational functions of HBx may be exerted both in the nucleus, via interaction with host DNA-binding proteins, and in the cytoplasm, via signaling pathways. Although there have been many studies describing different pathways altered by HBx, and its innumerable binding partners, the molecular mechanism that regulates its different roles has been difficult to elucidate. Methods In the current study, we took a bioinformatics approach to investigate whether the viral protein HBx might be regulated via phosphorylation by an evolutionarily conserved mechanism. Results We found that the phylogenetically conserved residues Ser25 and Ser41 (both within the negative regulatory domain, and Thr81 (in the transactivation domain are predicted to be phosphorylated. By molecular 3D modeling of HBx, we further show these residues are all predicted to be exposed on the surface of the protein, making them easily accesible to these types of modifications. Furthermore, we have also identified Yin Yang sites that might have the potential to be phosphorylated and O-β-GlcNAc interplay at the same residues. Conclusions Thus, we propose that the different roles of HBx displayed in different subcellular locations might be regulated by an evolutionarily conserved mechanism of posttranslational modification, via phosphorylation.

  16. Prognostic role of fibroblast growth factor receptor 2 in human solid tumors: A systematic review and meta-analysis.

    Science.gov (United States)

    Liu, Gang; Xiong, Disheng; Xiao, Rui; Huang, Zhengjie

    2017-06-01

    In the past decades, the oncogenic role of fibroblast growth factor receptor 2 has been demonstrated in a number of cancer types. However, studies have reported contradictory findings concerning the correlation between fibroblast growth factor receptor 2 expression and prognosis in solid tumors. To address this discrepancy, we performed a meta-analysis with 18 published studies (2975 patients) retrieved from PubMed, EMBASE, and Web of science. Data were extracted and computed into odds ratios. The results showed that fibroblast growth factor receptor 2 overexpression was significantly associated with decreased 3-year overall survival (odds ratio = 1.93, 95% confidence interval: 1.30-2.85, p = 0.001) and 5-year overall survival (odds ratio = 1.62, 95% confidence interval: 1.07-2.44, p = 0.02) in patients with solid tumors. Subgroup analysis revealed that high fibroblast growth factor receptor 2 expression was also associated with poor prognosis of gastric cancer, hepatocellular carcinoma, and esophageal cancer, but not correlated with pancreatic cancer. In conclusion, fibroblast growth factor receptor 2 overexpression is correlated with decreased survival in most solid tumors, suggesting that the expression status of fibroblast growth factor receptor 2 is a valuable prognostic biomarker and a novel therapeutic target in human solid tumors.

  17. Self-renewal of human embryonic stem cells requires insulin-like growth factor-1 receptor and ERBB2 receptor signaling

    Science.gov (United States)

    Wang, Linlin; Schulz, Thomas C.; Sherrer, Eric S.; Dauphin, Derek S.; Shin, Soojung; Nelson, Angelique M.; Ware, Carol B.; Zhan, Mei; Song, Chao-Zhong; Chen, Xiaoji; Brimble, Sandii N.; McLean, Amanda; Galeano, Maria J.; Uhl, Elizabeth W.; D'Amour, Kevin A.; Chesnut, Jonathan D.; Rao, Mahendra S.

    2007-01-01

    Despite progress in developing defined conditions for human embryonic stem cell (hESC) cultures, little is known about the cell-surface receptors that are activated under conditions supportive of hESC self-renewal. A simultaneous interrogation of 42 receptor tyrosine kinases (RTKs) in hESCs following stimulation with mouse embryonic fibroblast (MEF) conditioned medium (CM) revealed rapid and prominent tyrosine phosphorylation of insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R); less prominent tyrosine phosphorylation of epidermal growth factor receptor (EGFR) family members, including ERBB2 and ERBB3; and trace phosphorylation of fibroblast growth factor receptors. Intense IGF1R and IR phosphorylation occurred in the absence of MEF conditioning (NCM) and was attributable to high concentrations of insulin in the proprietary KnockOut Serum Replacer (KSR). Inhibition of IGF1R using a blocking antibody or lentivirus-delivered shRNA reduced hESC self-renewal and promoted differentiation, while disruption of ERBB2 signaling with the selective inhibitor AG825 severely inhibited hESC proliferation and promoted apoptosis. A simple defined medium containing an IGF1 analog, heregulin-1β (a ligand for ERBB2/ERBB3), fibroblast growth factor-2 (FGF2), and activin A supported long-term growth of multiple hESC lines. These studies identify previously unappreciated RTKs that support hESC proliferation and self-renewal, and provide a rationally designed medium for the growth and maintenance of pluripotent hESCs. PMID:17761519

  18. The far and distal enhancers in the CYP3A4 gene co-ordinate the proximal promoter in responding similarly to the pregnane X receptor but differentially to hepatocyte nuclear factor-4alpha.

    Science.gov (United States)

    Liu, Fu-Jun; Song, Xiulong; Yang, Dongfang; Deng, Ruitang; Yan, Bingfang

    2008-01-01

    CYP3A4 (cytochrome P450 3A4) is involved in the metabolism of more than 50% of drugs and other xenobiotics. The expression of CYP3A4 is induced by many structurally dissimilar compounds. The PXR (pregnane X receptor) is recognized as a key regulator for the induction, and the PXR-directed transactivation of the CYP3A4 gene is achieved through a co-ordinated mechanism of the distal module with the proximal promoter. Recently, a far module was found to support constitutive expression of CYP3A4. The far module, like the distal module, is structurally clustered by a PXR response element (F-ER6) and elements recognized by HNF-4alpha (hepatocyte nuclear receptor-4alpha). We hypothesized that the far module supports PXR transactivation of the CYP3A4 gene. Consistent with the hypothesis, fusion of the far module to the proximal promoter of CYP3A4 markedly increased rifampicin-induced reporter activity. The increase was synergistically enhanced when both the far and distal modules were fused to the proximal promoter. The increase, however, was significantly reduced when the F-ER6 was disrupted. Chromatin immunoprecipitation detected the presence of PXR in the far module. Interestingly, HNF-4alpha increased the activity of the distal-proximal fused promoter, but decreased the activity of the far-proximal fused promoter. Given the fact that induction of CYP3A4 represents an important detoxification mechanism, the functional redundancy and synergistic interaction in supporting PXR transactivation suggest that the far and distal modules ensure the induction of CYP3A4 during chemical insults. The difference in responding to HNF-4alpha suggests that the magnitude of the induction is under control through various transcriptional networks.

  19. Effects of glucagon-like peptide-1 receptor agonists on cardiovascular risk factors

    DEFF Research Database (Denmark)

    Dalsgaard, Niels B; Vilsbøll, Tina; Knop, Filip K

    2018-01-01

    trials for diabetes therapies. To determine if different glucagon-like peptide-1 receptor agonists (GLP-1RAs) had varying effects on these CV risk factors, we reviewed 16 head-to-head trials directly comparing GLP-1RAs that included at least one of the five factors. Few trials reported statistical...... differences between GLP-1RAs in terms of systolic blood pressure (SBP), body weight and total cholesterol. Liraglutide increased heart rate vs its comparators in three separate trials. All GLP-1RAs reduced glycated haemoglobin (HbA1c), but exenatide twice daily and lixisenatide had statistically smaller...... effects compared with other GLP-1RAs. These descriptive data indicate that individual GLP-1RAs affect CV risk factors differently, potentially because of their individual pharmacokinetics and/or size. Short-acting GLP-1RAs appeared to result in smaller changes in SBP and total cholesterol compared...

  20. Modulation of Tumor-Associated Macrophages (TAM) Phenotype by Platelet-Activating Factor (PAF) Receptor.

    Science.gov (United States)

    da Silva Junior, Ildefonso Alves; Stone, Simone Cardozo; Rossetti, Renata Marques; Jancar, Sonia; Lepique, Ana Paula

    2017-01-01

    Platelet-activating factor (PAF) plays an important role in the pathogenesis of several types of tumors. The biological effects of PAF are mediated by the PAF receptor (PAFR), which can be expressed by tumor cells and host cells that infiltrate the tumor microenvironment. In the present study, we investigated the role of PAFR expressed by leukocytes that infiltrate two types of tumors, one that expresses PAFR (TC-1 carcinoma) and another that does not express the receptor (B16F10 melanoma) implanted in mice that express the receptor or not (PAFR KO). It was found that both tumors grew significantly less in PAFR KO than in wild-type (WT) mice. Analysis of the leukocyte infiltration shown in PAFR KO increased the frequency of neutrophils (Gr1 + ) and of CD8 + lymphocytes in B16F10 tumors and of CD4 + lymphocytes in TC-1 tumors. PAFR KO also had a higher frequency of M1-like (CD11c + ) and lower M2-like (CD206 + ) macrophages infiltrated in both tumors. This was confirmed in macrophages isolated from the tumors that showed higher iNOS, lower arginase activity, and lower IL10 expression in PAFR KO tumors than WT mice. These data suggest that in the tumor microenvironment, endogenous PAF-like activity molecules bind PAFR in macrophages which acquire an M2-like profile and this promotes tumor growth.

  1. Platelet-Activating Factor Receptor Ligands Protect Tumor Cells from Radiation-Induced Cell Death

    Directory of Open Access Journals (Sweden)

    Ildefonso Alves da Silva-Junior

    2018-02-01

    Full Text Available Irradiation generates oxidized phospholipids that activate platelet-activating factor receptor (PAFR associated with pro-tumorigenic effects. Here, we investigated the involvement of PAFR in tumor cell survival after irradiation. Cervical cancer samples presented higher levels of PAF-receptor gene (PTAFR when compared with normal cervical tissue. In cervical cancer patients submitted to radiotherapy (RT, the expression of PTAFR was significantly increased. Cervical cancer-derived cell lines (C33, SiHa, and HeLa and squamous carcinoma cell lines (SCC90 and SCC78 express higher levels of PAFR mRNA and protein than immortalized keratinocytes. Gamma radiation increased PAFR expression and induced PAFR ligands and prostaglandin E2 (PGE2 in these tumor cells. The blocking of PAFR with the antagonist CV3938 before irradiation inhibited PGE2 and increased tumor cells death. Similarly, human carcinoma cells transfected with PAFR (KBP were more resistant to radiation compared to those lacking the receptor (KBM. PGE2 production by irradiated KBP cells was also inhibited by CV3988. These results show that irradiation of carcinoma cells generates PAFR ligands that protect tumor cells from death and suggests that the combination of RT with a PAFR antagonist could be a promising strategy for cancer treatment.

  2. Orexin-corticotropin-releasing factor receptor heteromers in the ventral tegmental area as targets for cocaine.

    Science.gov (United States)

    Navarro, Gemma; Quiroz, César; Moreno-Delgado, David; Sierakowiak, Adam; McDowell, Kimberly; Moreno, Estefanía; Rea, William; Cai, Ning-Sheng; Aguinaga, David; Howell, Lesley A; Hausch, Felix; Cortés, Antonio; Mallol, Josefa; Casadó, Vicent; Lluís, Carme; Canela, Enric I; Ferré, Sergi; McCormick, Peter J

    2015-04-29

    Release of the neuropeptides corticotropin-releasing factor (CRF) and orexin-A in the ventral tegmental area (VTA) play an important role in stress-induced cocaine-seeking behavior. We provide evidence for pharmacologically significant interactions between CRF and orexin-A that depend on oligomerization of CRF1 receptor (CRF1R) and orexin OX1 receptors (OX1R). CRF1R-OX1R heteromers are the conduits of a negative crosstalk between orexin-A and CRF as demonstrated in transfected cells and rat VTA, in which they significantly modulate dendritic dopamine release. The cocaine target σ1 receptor (σ1R) also associates with the CRF1R-OX1R heteromer. Cocaine binding to the σ1R-CRF1R-OX1R complex promotes a long-term disruption of the orexin-A-CRF negative crosstalk. Through this mechanism, cocaine sensitizes VTA cells to the excitatory effects of both CRF and orexin-A, thus providing a mechanism by which stress induces cocaine seeking. Copyright © 2015 the authors 0270-6474/15/356639-15$15.00/0.

  3. Orexin–Corticotropin-Releasing Factor Receptor Heteromers in the Ventral Tegmental Area as Targets for Cocaine

    Science.gov (United States)

    Navarro, Gemma; Quiroz, César; Moreno-Delgado, David; Sierakowiak, Adam; McDowell, Kimberly; Moreno, Estefanía; Rea, William; Cai, Ning-Sheng; Aguinaga, David; Howell, Lesley A.; Hausch, Felix; Cortés, Antonio; Mallol, Josefa; Casadó, Vicent; Lluís, Carme; Canela, Enric I.

    2015-01-01

    Release of the neuropeptides corticotropin-releasing factor (CRF) and orexin-A in the ventral tegmental area (VTA) play an important role in stress-induced cocaine-seeking behavior. We provide evidence for pharmacologically significant interactions between CRF and orexin-A that depend on oligomerization of CRF1 receptor (CRF1R) and orexin OX1 receptors (OX1R). CRF1R–OX1R heteromers are the conduits of a negative crosstalk between orexin-A and CRF as demonstrated in transfected cells and rat VTA, in which they significantly modulate dendritic dopamine release. The cocaine target σ1 receptor (σ1R) also associates with the CRF1R–OX1R heteromer. Cocaine binding to the σ1R–CRF1R–OX1R complex promotes a long-term disruption of the orexin-A–CRF negative crosstalk. Through this mechanism, cocaine sensitizes VTA cells to the excitatory effects of both CRF and orexin-A, thus providing a mechanism by which stress induces cocaine seeking. PMID:25926444

  4. Function of insulin-like growth factor 1 receptor in cancer resistance to chemotherapy.

    Science.gov (United States)

    Yuan, Jingsheng; Yin, Zhijie; Tao, Kaixiong; Wang, Guobing; Gao, Jinbo

    2018-01-01

    Drug resistance is a primary cause of chemotherapeutic failure; however, how this resistance develops is complex. A comprehensive understanding of chemotherapeutic resistance mechanisms may aid in identifying more effective drugs and improve the survival rates of patients with cancer. Insulin-like growth factor 1 receptor (IGF1R), a member of the insulin receptor family, has been extensively assessed for biological activity, and its putative contribution to tumor cell development and progression. Furthermore, researchers have attended to drugs that target IGF1R since IGF1R functions as a membrane receptor. However, how IGF1R participates in chemotherapeutic resistance remains unclear. Therefore, the present study described the IGF1R gene and its associated signaling pathways, and offered details of IGF1R-induced tumor chemoresistance associated with promoting cell proliferation, inhibition of apoptosis, regulation of ATP-binding cassette transporter proteins and interactions with the extracellular matrix. The present study offered additional explanations for tumor chemotherapy resistance and provided a theoretical basis of IGF1R and its downstream pathways for future possible chemotherapy treatment options.

  5. Thrombospondin-1 modulates vascular endothelial growth factor activity at the receptor level.

    Science.gov (United States)

    Zhang, Xuefeng; Kazerounian, Shideh; Duquette, Mark; Perruzzi, Carole; Nagy, Janice A; Dvorak, Harold F; Parangi, Sareh; Lawler, Jack

    2009-10-01

    Vascular endothelial growth factor (VEGF) is a well-established stimulator of vascular permeability and angiogenesis, whereas thrombospondin-1 (TSP-1) is a potent angiogenic inhibitor. In this study, we have found that the TSP-1 receptors CD36 and beta1 integrin associate with the VEGF receptor 2 (VEGFR2). The coclustering of receptors that regulate angiogenesis may provide the endothelial cell with a platform for integration of positive and negative signals in the plane of the membrane. Thus, this complex may represent a molecular switch that regulates angiogenesis and determines endothelial cell behavior. In this context, physiological levels of TSP-1 appear to support VEGFR2 function on both the cellular and tissue level, because phosphorylation of VEGFR2 and vascular permeability in response to VEGF are decreased in TSP-1-null mice and isolated endothelial cells. A therapeutic agent based on the antiangiogenic domain of TSP-1, designated 3TSR (for three TSP-1 type 1 repeats), has significant antiangiogenic and antitumor efficacy. Systemic treatment of wild-type mice with 3TSR significantly decreased VEGF-induced permeability. Consistent with this result, VEGF-stimulated phosphorylation of VEGFR2 was also significantly decreased in lung extracts from 3TSR-treated mice. Moreover, 3TSR significantly decreased VEGF-stimulated VEGFR2 phosphorylation in human dermal microvascular endothelial cells in culture. Taken together, the results indicate that TSP-1 and 3TSR modulate the function of VEGFR2.

  6. Phosphorylation of the transcription factor Sp4 is reduced by NMDA receptor signaling.

    Science.gov (United States)

    Saia, Gregory; Lalonde, Jasmin; Sun, Xinxin; Ramos, Belén; Gill, Grace

    2014-05-01

    The regulation of transcription factor function in response to neuronal activity is important for development and function of the nervous system. The transcription factor Sp4 regulates the developmental patterning of dendrites, contributes to complex processes including learning and memory, and has been linked to psychiatric disorders such as schizophrenia and bipolar disorder. Despite its many roles in the nervous system, the molecular mechanisms regulating Sp4 activity are poorly understood. Here, we report a site of phosphorylation on Sp4 at serine 770 that is decreased in response to membrane depolarization. Inhibition of the voltage-dependent NMDA receptor increased Sp4 phosphorylation. Conversely, stimulation with NMDA reduced the levels of Sp4 phosphorylation, and this was dependent on the protein phosphatase 1/2A. A phosphomimetic substitution at S770 impaired the Sp4-dependent maturation of cerebellar granule neuron primary dendrites, whereas a non-phosphorylatable Sp4 mutant behaved like wild type. These data reveal that transcription factor Sp4 is regulated by NMDA receptor-dependent activation of a protein phosphatase 1/2A signaling pathway. Our findings also suggest that the regulated control of Sp4 activity is an important mechanism governing the developmental patterning of dendrites. © 2014 International Society for Neurochemistry.

  7. Divergent effects of insulin-like growth factor-1 receptor expression on prognosis of estrogen receptor positive versus triple negative invasive ductal breast carcinoma

    NARCIS (Netherlands)

    Hartog, Hermien; Horlings, Hugo M; van der Vegt, Bert; Kreike, Bas; Ajouaou, Abderrahim; van de Vijver, Marc J; Boezen, Hendrika; de Bock, Geertruida H; van der Graaf, Wilhelmina; Wesseling, Jelle

    2011-01-01

    The insulin-like growth factor type 1 receptor (IGF1R) is involved in progression of breast cancer and resistance to systemic treatment. Targeting IGF1R signaling may, therefore, be beneficial in systemic treatment. We report the effect of IGF1R expression on prognosis in invasive ductal breast

  8. Localization of the gene for the ciliary neutrotrophic factor receptor (CNTFR) to human chromosome 9

    Energy Technology Data Exchange (ETDEWEB)

    Donaldson, D.H.; Jones, C.; Patterson, D. (Eleanor Roosevelt Institute, Denver, CO (United States) Univ. of Colorado Health Science Center, Denver, CO (United States)); Britt, D.E.; Jackson, C.L. (Brown Univ., Providence, RI (United States))

    1993-09-01

    Ciliary neurotrophic factor (CNTF) has recently been found to be important for the survival of motor neurons and has shown activity in animal models of amyotrophic lateral sclerosis (ALS). CNTF therefore holds promise as a treatment for ALS, and it and its receptor (CNTFR) are candidates for a gene involved in familial ALS. The CNTFR gene was mapped to chromosome 9 by PCR on a panel of human/CHO somatic cell hybrids and localized to 9p13 by PCR on a panel of radiation hybrids. 18 ref., 1 fig., 2 tabs.

  9. Nanostructured materials detect epidermal growth factor receptor, neuron specific enolase and carcinoembryonic antigen

    Science.gov (United States)

    Stefan-van Staden, Raluca-Ioana; Comnea-Stancu, Ionela Raluca; Surdu-Bob, Carmen Cristina; Badulescu, Marius

    2015-09-01

    New nanostructured materials based on thin films of Cu and Ni deposited on textile material (veil), as well as gold nanostructured microspheres were used for the design of new stochastic sensors. The stochastic sensors were able to detect simultaneously a panel of biomarkers comprising epidermal growth factor receptor, neuron specific enolase, and carcinoembryonic antigen from whole blood samples with high reliabilities - recovery tests higher than 97.00%, with a RSD (%) lower than 0.1%. The stochastic sensors had shown high sensitivities and low determination levels for the detection of the proposed panel of biomarkers making early detection of lung cancer possible by fast screening of whole blood.

  10. The N363S polymorphism of the glucocorticoid receptor and metabolic syndrome factors in men

    DEFF Research Database (Denmark)

    Buemann, Benjamin; Black, Eva; Holst, Claus

    2005-01-01

    OBJECTIVE: To test the associations between the N363S polymorphism of the glucocorticoid receptor gene (NR3C1) and factors related to the metabolic syndrome in middle-aged men with and without juvenile-onset obesity. RESEARCH METHODS AND PROCEDURES: This study included two groups of middle-aged men...... with the obese men (n = 299; age, 50 +/- 7 years). The subjects were genotyped for the N363S polymorphism by polymerase chain reaction-restriction fragment length polymorphism. Body composition was measured by DXA. Glucose metabolism was evaluated by an oral glucose tolerance test, and the Matsudas index...

  11. Regulation of tumour necrosis factor (TNF) induced apoptosis by soluble TNF receptors in Helicobacter pylori infection

    OpenAIRE

    Shibata, J; Goto, H; Arisawa, T; Niwa, Y; Hayakawa, T; Nakayama, A; Mori, N

    1999-01-01

    BACKGROUND—Tumour necrosis factor (TNF) is a predominant cytokine produced in the gastric mucosa of patients with Helicobacter pylori infection. TNF induces apoptosis in a variety of cells. The soluble TNF receptors (sTNF-Rs) can be divided into sTNF-RI and sTNF-RII, both of which inhibit TNF activity. However, their precise mechanisms remain unclear.
AIM—To investigate the role of sTNF-Rs in H pylori infection.
METHODS—In 40 patients, production of TNF and sTNF-Rs in gastric mucosa was measu...

  12. Expression of granulocyte colony-stimulating factor and its receptor in childhood neuroblastoma

    OpenAIRE

    Xin WU; Da-wei HE; Yong-bo ZHANG; Wen-fei HE; Ze-dong BIAN; Qin-jun YI; Guang-hui WEI

    2012-01-01

    Objective  To study the expression of granulocyte colony-stimulating factor (G-CSF) and its receptor (G-CSFR) in neuroblastoma of children. Methods  Twenty-five specimens of neuroblastoma were collected in our department during 2009.1–2011.6. G-CSF and G-CSFR were determined by immunohistochemistry. The correlation between expressions of G-CSF and G-CSFR and age, gender and clinical stage were analyzed. Results  The expression of G-CSF and G-CSFR in neuroblastoma specimens was 68%, 72% respec...

  13. Role of epidermal growth factor receptor activation in regulating mucin synthesis

    Directory of Open Access Journals (Sweden)

    Nadel Jay A

    2001-02-01

    Full Text Available Abstract Healthy individuals have few goblet cells in their airways, but in patients with hypersecretory diseases goblet-cell upregulation results in mucus hypersecretion, airway plugging, and death. Multiple stimuli produce hypersecretion via epidermal growth factor receptor (EGFR expression and activation, causing goblet-cell metaplasia from Clara cells by a process of cell differentiation. These cells are also believed to be the cells of origin of non-small-cell lung cancer, but this occurs via cell multiplication. The mechanisms that determine which pathway is chosen are critical but largely unknown. Although no effective therapy exists for hypersecretion at present, the EGFR cascade suggests methods for effective therapeutic intervention.

  14. Strong association of epidermal growth factor receptor status with breast cancer FDG uptake

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Joohee; Moon, Seung Hwan; Hyun, Seung Hyup; Cho, Young Seok; Choi, Joon Young; Kim, Byung-Tae; Lee, Kyung-Han [Sungkyunkwan University School of Medicine, Department of Nuclear Medicine, Samsung Medical Center, Seoul (Korea, Republic of); Lee, Eun Jeong [Seoul Medical Center, Department of Nuclear Medicine, Seoul (Korea, Republic of); Kim, Seokhwi [Sungkyunkwan University School of Medicine, Department of Pathology, Samsung Medical Center, Seoul (Korea, Republic of)

    2017-08-15

    Imaging tumor FDG uptake could complement breast cancer biomarkers of risk and treatment response. Although breast cancer FDG uptake is reputedly influenced by major biomarker states, the role of epidermal growth factor receptor (EGFR) expression remains largely unexplored. This is a retrospective study that included 499 patients with primary breast cancer at initial presentation. Tumor FDG uptake was measured on pretreatment PET/CT as maximum standardized uptake value (SUVmax), and biomarkers were assessed by immunohistochemistry of tumor tissue. Regression analysis was performed for predictors of high tumor FDG uptake (SUVmax ≥ 8.6). SUVmax was higher in ER- (36.5%; 11.2 ± 6.0 vs. 8.3 ± 5.3), PR- (42.3%; 10.9 ± 6.0 vs. 8.2 ± 5.2), and triple-negative tumors (19.8%; 12.0 ± 6.9 vs. 8.7 ± 5.2; all p < 0.0001). EGFR expression (28.5%) was more frequent in ER-, PR-, triple-negative, cytokeratin 5/6 (CK5/6) + and mutant P53 (mP53) + tumors (all p < 0.0001). EGFR+ was associated with higher SUVmax among all tumors (11.9 ± 6.0 vs. 8.3 ± 5.3), ER- tumors (p < 0.0001), PR- and + tumors (p < 0.0001 and 0.027), hormone receptor- and + tumors (p < 0.0001 and 0.004), human epidermal growth factor receptor 2 (HER2)- and + tumors (p < 0.0001 and 0.006), non-triple negative tumors (p < 0.0001), CK5/6- and + tumors (p = 0.021 and <0.0001), and mP53- and + tumors (p < 0.0001 and 0.008). Tumors had high FDG uptake in 73.2% of EGFR+ and 40.6% of EGFR- tumors. On regression analysis, significant multivariate predictors of high tumor FDG uptake were large size, EGFR+ and CK5/6+ for the entire subjects, and EGFR+ and CK5/6+ for ER- and hormone receptor negative subgroups. High FDG uptake was able to sub-stratify EGFR+ tumors that were more likely to be ER- and CK5/6+, and EGFR- tumors more likely to be mP53 +. Primary breast tumor FDG uptake is strongly influenced by EGFR status beyond that by other major biomarkers including hormone receptor and HER2 status, and EGFR

  15. Parabens and Human Epidermal Growth Factor Receptor Ligand Cross-Talk in Breast Cancer Cells.

    Science.gov (United States)

    Pan, Shawn; Yuan, Chaoshen; Tagmount, Abderrahmane; Rudel, Ruthann A; Ackerman, Janet M; Yaswen, Paul; Vulpe, Chris D; Leitman, Dale C

    2016-05-01

    Xenoestrogens are synthetic compounds that mimic endogenous estrogens by binding to and activating estrogen receptors. Exposure to estrogens and to some xenoestrogens has been associated with cell proliferation and an increased risk of breast cancer. Despite evidence of estrogenicity, parabens are among the most widely used xenoestrogens in cosmetics and personal-care products and are generally considered safe. However, previous cell-based studies with parabens do not take into account the signaling cross-talk between estrogen receptor α (ERα) and the human epidermal growth factor receptor (HER) family. We investigated the hypothesis that the potency of parabens can be increased with HER ligands, such as heregulin (HRG). The effects of HER ligands on paraben activation of c-Myc expression and cell proliferation were determined by real-time polymerase chain reaction, Western blots, flow cytometry, and chromatin immunoprecipitation assays in ERα- and HER2-positive human BT-474 breast cancer cells. Butylparaben (BP) and HRG produced a synergistic increase in c-Myc mRNA and protein levels in BT-474 cells. Estrogen receptor antagonists blocked the synergistic increase in c-Myc protein levels. The combination of BP and HRG also stimulated proliferation of BT-474 cells compared with the effects of BP alone. HRG decreased the dose required for BP-mediated stimulation of c-Myc mRNA expression and cell proliferation. HRG caused the phosphorylation of serine 167 in ERα. BP and HRG produced a synergistic increase in ERα recruitment to the c-Myc gene. Our results show that HER ligands enhanced the potency of BP to stimulate oncogene expression and breast cancer cell proliferation in vitro via ERα, suggesting that parabens might be active at exposure levels not previously considered toxicologically relevant from studies testing their effects in isolation. Pan S, Yuan C, Tagmount A, Rudel RA, Ackerman JM, Yaswen P, Vulpe CD, Leitman DC. 2016. Parabens and human epidermal

  16. Enhancement by histamine of vascular endothelial growth factor production in granulation tissue via H2 receptors

    Science.gov (United States)

    Ghosh, Ajoy Kumar; Hirasawa, Noriyasu; Ohuchi, Kazuo

    2001-01-01

    Roles of histamine in the production of vascular endothelial growth factor (VEGF) in the carrageenin-induced granulation tissue in rats were analysed in vitro and in vivo.Incubation of the minced granulation tissue in the presence of histamine (1 and 10 μM) increased the content of VEGF protein in the conditioned medium in a time- and concentration-dependent manner. The levels of VEGF mRNA in the minced granulation tissue were also increased by histamine in a concentration-dependent manner.The increase in the content of VEGF protein in the conditioned medium by histamine (10 μM) was suppressed by the H2 receptor antagonist cimetidine (IC50 0.37 μM), but not by the H1 receptor antagonist pyrilamine maleate, the H3 receptor antagonist thioperamide or the cyclo-oxygenase inhibitor indomethacin.The histamine-induced increase in the content of VEGF protein in the conditioned medium was inhibited by the cyclic AMP antagonist Rp-cAMP (IC50 6.8 μM), and the protein kinase A inhibitor H-89 (IC50 12.5 μM), but not by the protein kinase C inhibitors Ro 31-8425 and calphostin C or the tyrosine kinase inhibitor genistein.Simultaneous injection of cimetidine (400 μg) and indomethacin (100 μg) into the air pouch of rats additively reduced the carrageenin-induced increase in VEGF protein levels and angiogenesis in the granulation tissue as assessed by using carmine dye.These findings indicate that histamine has an activity to induce VEGF production in the granulation tissue via the H2 receptor-cyclic AMP-protein kinase A pathway and augments angiogenesis in the granulation tissue. PMID:11724747

  17. The cytokines cardiotrophin-like cytokine/cytokine-like factor-1 (CLC/CLF) and ciliary neurotrophic factor (CNTF) differ in their receptor specificities.

    Science.gov (United States)

    Tormo, Aurélie Jeanne; Letellier, Marie-Claude; Lissilaa, Rami; Batraville, Laurie-Anne; Sharma, Mukut; Ferlin, Walter; Elson, Greg; Crabé, Sandrine; Gauchat, Jean-François

    2012-12-01

    Ciliary neurotrophic factor (CNTF) and cardiotrophin-like cytokine (CLC) are two cytokines with neurotrophic and immunomodulatory activities. CNTF is a cytoplasmic factor believed to be released upon cellular damage, while CLC requires interaction with a soluble cytokine receptor, cytokine-like factor 1 (CLF), to be efficiently secreted. Both cytokines activate a receptor complex comprising the cytokine binding CNTF receptor α (CNTFRα) and two signaling chains namely, leukemia inhibitory factor receptor β (LIFRβ) and gp130. Human CNTF can recruit and activate an alternative receptor in which CNTFRα is substituted by IL-6Rα. As both CNTF and CLC have immune-regulatory activities in mice, we compared their ability to recruit mouse receptors comprising both gp130 and LIFRβ signaling chains and either IL-6Rα or IL-11Rα which, unlike CNTFRα, are expressed by immune cells. Our results indicate that 1) mouse CNTF, like its human homologue, can activate cells expressing gp130/LIFRβ with either CNTFRα or IL-6Rα and, 2) CLC/CLF is more restricted in its specificity in that it activates only the tripartite CNTFR. Several gp130 signaling cytokines influence T helper cell differentiation. We therefore investigated the effect of CNTF on CD4 T cell cytokine production. We observed that CNTF increased the number of IFN-γ producing CD4 T cells. As IFN-γ is considered a mediator of the therapeutic effect of IFN-β in multiple sclerosis, induction of IFN-γ by CNTF may contribute to the beneficial immunomodulatory effect of CNTF in mouse multiple sclerosis models. Together, our results indicate that CNTF activates the same tripartite receptors in mouse and human cells and further validate rodent models for pre-clinical investigation of CNTF and CNTF derivatives. Furthermore, CNTF and CLC/CLF differ in their receptor specificities. The receptor α chain involved in the immunomodulatory effects of CLC/CLF remains to be identified. Crown Copyright © 2012. Published by

  18. Trans-activation of an artificial dTam3 transposable element in transgenic tobacco plants

    NARCIS (Netherlands)

    Haring, Michel A.; Teeuwen-de Vroomen, Marianne J.; Nijkamp, H. John J.; Hille, Jacques

    1991-01-01

    In Antirrhinum majus only autonomous Tam3 transposons have been characterized. We investigated whether an artificial dTam3 element, with a deletion in the presumptive transposase coding region, can be trans-activated in tobacco by an activator Tam3 element, which was immobilized by the deletion of

  19. Fibroblast growth factor 21, fibroblast growth factor receptor 1, and β-Klotho expression in bovine growth hormone transgenic and growth hormone receptor knockout mice.

    Science.gov (United States)

    Brooks, Nicole E; Hjortebjerg, Rikke; Henry, Brooke E; List, Edward O; Kopchick, John J; Berryman, Darlene E

    Although growth hormone (GH) and fibroblast growth factor 21 (FGF21) have a reported relationship, FGF21 and its receptor, fibroblast growth factor receptor 1 (FGFR1) and cofactor β-Klotho (KLB), have not been analyzed in chronic states of altered GH action. The objective of this study was to quantify circulating FGF21 and tissue specific expression of Fgf21, Fgfr1, and Klb in mice with modified GH action. Based on previous studies, we hypothesized that bovine GH transgenic (bGH) mice will be FGF21 resistant and GH receptor knockout (GHR-/-) mice will have normal FGF21 action. Seven-month-old male bGH mice (n=9) and wild type (WT) controls (n=10), and GHR-/- mice (n=8) and WT controls (n=8) were used for all measurements. Body composition was determined before dissection, and tissue weights were measured at the time of dissection. Serum FGF21 levels were evaluated by ELISA. Expression of Fgf21, Fgfr1, and Klb mRNA in white adipose tissue (AT), brown AT, and liver were evaluated by reverse transcription quantitative PCR. As expected, bGH mice had increased body weight (p=3.70E -8 ) but decreased percent fat mass (p=4.87E -4 ). Likewise, GHR-/- mice had decreased body weight (p=1.78E -10 ) but increased percent fat mass (p=1.52E -9 ), due to increased size of the subcutaneous AT depot when normalized to body weight (p=1.60E -10 ). Serum FGF21 levels were significantly elevated in bGH mice (p=0.041) and unchanged in GHR-/- mice (p=0.88). Expression of Fgf21, Fgfr1, and Klb mRNA in white AT and liver were downregulated or unchanged in both bGH and GHR-/- mice. The only exception was Fgf21 expression in brown AT of GHR-/-, which trended toward increased expression (p=0.075). In accordance with our hypothesis, we provide evidence that circulating FGF21 is increased in bGH animals, but remains unchanged in GHR-/- mice. Downregulation or no change in Fgf21, Fgfr1, and Klb expression are seen in white AT, brown AT, and liver of bGH and GHR-/- mice when compared to their

  20. Adenosine inhibits neutrophil vascular endothelial growth factor release and transendothelial migration via A2B receptor activation.

    LENUS (Irish Health Repository)

    Wakai, A

    2012-02-03

    The effects of adenosine on neutrophil (polymorphonuclear neutrophils; PMN)-directed changes in vascular permeability are poorly characterized. This study investigated whether adenosine modulates activated PMN vascular endothelial growth factor (vascular permeability factor; VEGF) release and transendothelial migration. PMN activated with tumour necrosis factor-alpha (TNF-alpha, 10 ng\\/mL) were incubated with adenosine and its receptor-specific analogues. Culture supernatants were assayed for VEGF. PMN transendothelial migration across human umbilical vein endothelial cell (HUVEC) monolayers was assessed in vitro. Adhesion molecule receptor expression was assessed flow cytometrically. Adenosine and some of its receptor-specific analogues dose-dependently inhibited activated PMN VEGF release. The rank order of potency was consistent with the affinity profile of human A2B receptors. The inhibitory effect of adenosine was reversed by 3,7-dimethyl-1-propargylxanthine, an A2 receptor antagonist. Adenosine (100 microM) or the A2B receptor agonist 5\\'-N-ethylcarboxamidoadenosine (NECA, 100 microM) significantly reduced PMN transendothelial migration. However, expression of activated PMN beta2 integrins and HUVEC ICAM-1 were not significantly altered by adenosine or NECA. Adenosine attenuates human PMN VEGF release and transendothelial migration via the A2B receptor. This provides a novel target for the modulation of PMN-directed vascular hyperpermeability in conditions such as the capillary leak syndrome.

  1. DNA alkylating agents alleviate silencing of class II transactivator gene expression in L1210 lymphoma cells.

    Science.gov (United States)

    Murphy, Shawn P; Holtz, Renae; Lewandowski, Nicole; Tomasi, Thomas B; Fuji, Hiroshi

    2002-09-15

    MHC class II (Ia) Ag expression is inversely correlated with tumorigenicity and directly correlated with immunogenicity in clones of the mouse L1210 lymphoma (1 ). Understanding the mechanisms by which class II Ag expression is regulated in L1210 lymphoma may facilitate the development of immunotherapeutic approaches for the treatment of some types of lymphoma and leukemia. This study demonstrates that the variation in MHC class II Ag expression among clones of L1210 lymphoma is due to differences in the expression of the class II transactivator (CIITA). Analysis of stable hybrids suggests that CIITA expression is repressed by a dominant mechanism in class II-negative L1210 clones. DNA-alkylating agents such as ethyl methanesulfonate and the chemotherapeutic drug melphalan activate CIITA and class II expression in class II negative L1210 cells, and this effect appears to be restricted to transformed cell lines derived from the early stages of B cell ontogeny. Transient transfection assays demonstrated that the CIITA type III promoter is active in class II(-) L1210 cells, despite the fact that the endogenous gene is not expressed, which suggests that these cells have all of the transacting factors necessary for CIITA transcription. An inverse correlation between methylation of the CIITA transcriptional regulatory region and CIITA expression was observed among L1210 clones. Furthermore, 5-azacytidine treatment activated CIITA expression in class II-negative L1210 cells. Collectively, our results suggest that 1) CIITA gene expression is repressed in class II(-) L1210 cells by methylation of the CIITA upstream regulatory region, and 2) treatment with DNA-alkylating agents overcomes methylation-based silencing of the CIITA gene in L1210 cells.

  2. FOXM1 regulates glycolysis in hepatocellular carcinoma by transactivating glucose transporter 1 expression.

    Science.gov (United States)

    Shang, Runze; Pu, Meng; Li, Yu; Wang, Desheng

    2017-04-01

    The Forkhead box M1 (FOXM1) transcription factor plays crucial roles in the initiation and progression of various malignancies, including hepatocellular carcinoma (HCC). However, the mechanism by which FOXM1 regulates cancer metabolism remains unclear. In the present study, overexpression and RNA interference (RNAi) approaches were used to investigate the role of FOXM1 in the regulation of glycolysis in vitro. Luciferase reporter assays were used to explore the transcriptional regulation of the glucose transporter 1 (GLUT1) promoter by FOXM1. Then, immunohistochemical staining was used to examine the expression of FOXM1 and GLUT1 in 100 paired HCC and adjacent non-cancerous liver tissues. Chi-square test and logistic regression analysis were performed to evaluate the association between FOXM1 and GLUT1 expression with clinicopathological characteristics. Our data showed that FOXM1 promoted glycolysis in the HCC cells. FOXM1 knockdown significantly reduced the expression of GLUT1 among key glycolysis-related molecules in the different HCC cell lines. Glucose uptake and lactate production assay showed that FOXM1 positively regulated glycolysis based on GLUT1 expression. Moreover, FOXM1 overexpression increased and knockdown decreased GLUT1 expression. Luciferase reporter assays showed that the -206 to -199 bp region of the GLUT1 promoter is important for FOXM1 to enhance GLUT1 promoter activity. The results of the IHC analysis showed that the protein expression of FOXM1 and GLUT1 was closely related to the tumor histological grade and TNM stage. In addition, GLUT1 expression was also related to microvascular invasion. In conclusion, overexpression of FOXM1 and GLUT1 may play critical roles in HCC. FOXM1 promotes HCC glycolysis by transactivating GLUT1 expression.

  3. Role of PU.1 in MHC class II expression through transcriptional regulation of class II transactivator pI in dendritic cells.

    Science.gov (United States)

    Kitamura, Nao; Yokoyama, Hokuto; Yashiro, Takuya; Nakano, Nobuhiro; Nishiyama, Makoto; Kanada, Shunsuke; Fukai, Tatsuo; Hara, Mutsuko; Ikeda, Shigaku; Ogawa, Hideoki; Okumura, Ko; Nishiyama, Chiharu

    2012-03-01

    PU.1 is a hematopoietic cell-specific transcription factor belonging to the Ets family. We hypothesized that PU.1 is involved in MHC class II expression in dendritic cells (DCs). The role of PU.1 in MHC class II expression in DCs was analyzed. Transcriptional regulation of the DC-specific pI promoter of the class II transactivator (CIITA) gene and subsequent MHC class II expression was investigated by using PU.1 small interfering RNA (siRNA) and reporter, chromatin immunoprecipitation, and electrophoretic mobility shift assays. PU.1 siRNA introduction suppressed MHC class II expression, allogeneic and syngeneic T-cell activation activities of bone marrow-derived DCs (BMDCs) with reduction of CIITA mRNA driven by the DC-specific promoter pI, and MHC class II mRNA. The chromatin immunoprecipitation assay showed constitutive binding of PU.1 to the pI region in BMDCs, whereas acetylation of histone H3 on pI was suppressed by LPS stimulation in parallel with shutdown of CIITA transcription. PU.1 transactivated the pI promoter through cis-elements at -47/-44 and -30/-27 in a reporter assay and to which PU.1 directly bound in an electrophoretic mobility shift assay. Acetylation of histones H3 and H4 on pI was reduced in PU.1 siRNA-introduced BMDCs. Knockdown of interferon regulatory factor 4 or 8, which is a heterodimer partner of PU.1, by siRNA did not affect pI-driven CIITA transcription or MHC class II expression. PU.1 basally transactivates the CIITA pI promoter in DCs by functioning as a monomeric transcription factor and by affecting histone modification, resulting in the subsequent expression and function of MHC class II. Copyright © 2011 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

  4. Nutrition, anthropometry, gastrointestinal dysfunction, and circulating levels of tumour necrosis factor alpha receptor I and interleukin-1 receptor antagonist in children during stem cell transplantation

    DEFF Research Database (Denmark)

    Andreassen, B. U.; Pærregaard, Anders; Michaelsen, Kim F.

    2008-01-01

    To evaluate anthropometry, nutrition and gastrointestinal dysfunction, and to characterize the relation between these parameters and the inflammatory activity evaluated by plasma levels of soluble tumour necrosis factor alpha receptor I (sTNFRI) and interleukin-1 receptor antagonist (IL-1Ra) levels...... during stem cell transplantation (SCT) in children. Clinical assessments and blood sampling were performed on days -3, 0, +7, +15 and +31 in eight children undergoing SCT. Energy intake, anthropometry, gastrointestinal dysfunction (WHO toxicity score) and sTNFRI and IL-1Ra were evaluated. The energy...

  5. A peptide derived from the CD loop-D helix region of ciliary neurotrophic factor (CNTF) induces neuronal differentiation and survival by binding to the leukemia inhibitory factor (LIF) receptor and common cytokine receptor chain gp130

    DEFF Research Database (Denmark)

    Rathje, Mette; Pankratova, Stanislava; Nielsen, Janne

    2011-01-01

    Ciliary neurotrophic factor (CNTF) induces neuronal differentiation and promotes the survival of various neuronal cell types by binding to a receptor complex formed by CNTF receptor a (CNTFRa), gp130, and the leukemia inhibitory factor (LIF) receptor (LIFR). The CD loop-D helix region of CNTF has...... that these receptors are involved in the effects of cintrofin. The C-terminal part of the peptide, corresponding to the D helix region of CNTF, was shown to be essential for the neuritogenic action of the peptide. CNTF and LIF induced neurite outgrowth in CGNs plated on laminin-coated slides. On uncoated slides, CNTF...... similar to those induced by CNTF and may be a valuable survival agent with possible therapeutic potential....

  6. Significance of Phosphorylated Epidermal Growth Factor Receptor and Its Signal Transducers in Human Soft Tissue Sarcoma

    Directory of Open Access Journals (Sweden)

    Jia-Lin Yang

    2017-05-01

    Full Text Available Previous studies have shown that total epidermal growth factor receptor (EGFR protein is highly expressed in soft tissue sarcoma (STS. We aimed to investigate the significance of phosphorylated-EGFR (pEGFR and its activated-downstream signal transducers in STS tissue samples. A tissue microarray comprising 87 STS samples was assessed for total EGFR, pEGFR and its phosphorylated signal transducers and expression was correlated with clinicopathlogical parameters including patient outcome. Although the expression of total EGFR was significantly associated with adverse STS histologic grade (p = 0.004 and clinical stage (p = 0.012 similar to pEGFR, phosphorylated protein kinase B (pAkt and phosphorylated extracellular signal regulated kinase (pERK, it is not a prognostic factor for survival. By contrast, the expression of pEGFR is an independent factor for cancer specific survival, while pERK is an independent prognostic factor for both overall and cancer specific survival in STS (p < 0.05, Cox proportional hazard model and log-rank test in addition to the recognised factors of tumour grade and clinical stage. pERK and pEGFR are new independent prognostic factors for overall and/or cancer specific survival in STS. The expression of EGFR/pEGFR, and their associated downstream signal transducers, was associated with STS progression, suggesting that EGFR downstream signalling pathways may jointly support STS cell survival.

  7. B Cell Activating Factor (BAFF) and BAFF Receptor Expression in Autoimmune and Nonautoimmune Thyroid Diseases.

    Science.gov (United States)

    Campi, Irene; Tosi, Delfina; Rossi, Stefania; Vannucchi, Guia; Covelli, Danila; Colombo, Federico; Trombetta, Elena; Porretti, Laura; Vicentini, Leonardo; Cantoni, Gianmaria; Currò, Nicola; Beck-Peccoz, Paolo; Bulfamante, Gaetano; Salvi, Mario

    2015-09-01

    The B cell activating factor (BAFF) is a member of the tumor necrosis factor family, which controls the survival/proliferation of B cells and is involved in the pathogenesis of a number of autoimmune diseases. The objective of the present study was to investigate the expression of BAFF and BAFF receptor (BAFF-R) in the thyroid tissue of patients affected with autoimmune thyroid disorders (AITD) or multinodular goiter (MNG) compared with those with normal thyroids. Immunohistochemistry was performed using a panel of antibodies against BAFF, BAFF-R, CD3, CD4, CD8, CD20, CD34, CD79a, CD1a, CD68, and CD163 on the thyroid sections of 27 patients affected with Graves' disease (GD), 23 with Hashimoto's thyroiditis (HT), 16 with nontoxic nodular goiter (NTG), and 15 with toxic nodular goiter (TG), submitted to total thyroidectomy between 2000 and 2011. The overall BAFF-R expression in thyrocytes was weak and not different in AITD and MNG. Conversely, a stronger BAFF expression was observed in MNG compared with AITD. BAFF and BAFF-R expression in the infiltrating lymphocytes was higher in AITD compared with MNG. Interestingly, in lymphocytes of follicular-like structures observed in HT, BAFF and BAFF-R were localized in the germinal center or in the mantle, respectively. This study shows that BAFF and BAFF-R are expressed in the thyrocytes derived from patients with either AITD or MNG, in addition to the expected expression of BAFF and its receptor in the infiltrating immune cells of GD and HT. These findings suggest a possible involvement of BAFF and its receptors in the pathophysiology of AITD.

  8. Expression of osteoprotegerin, receptor activator of nuclear factor kappa-B ligand, tumor necrosis factor-related apoptosis-inducing ligand, stromal cell-derived factor-1 and their receptors in epithelial metastatic breast cancer cell lines

    Directory of Open Access Journals (Sweden)

    Labovsky Vivian

    2012-06-01

    Full Text Available Abstract Background While breast cancer (BC is the major cause of death among women worldwide, there is no guarantee of better patient survival because many of these patients develop primarily metastases, despite efforts to detect it in its early stages. Bone metastasis is a common complication that occurs in 65-80 % of patients with disseminated disease, but the molecular basis underlying dormancy, dissemination and establishment of metastasis is not understood. Our objective has been to evaluate simultaneously osteoprotegerin (OPG, receptor activator of nuclear factor kappa B ligand (RANKL, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL, stromal cell-derived factor-1 (SDF-1, and their receptors (R in 2 human BC cell lines, MDA-MB-231 and MCF-7. Methods OPG, RANKL, TRAIL and SDF-1 expression and release, in addition to the expression of their receptors has been investigated using immunofluorescence, immunocytochemistry and ELISA analyses. Results MCF-7 cells released higher levels of OPG in conditioned media (CM than MDA-MB-231 cells; 100 % of both types of cell expressed OPG, RANKL, TRAIL and SDF-1. Moreover, 100 % in both lines expressed membrane RANKL and RANK, whereas only 50 % expressed CXCR4. Furthermore, 100 % expressed TRAIL-R1 and R4, 30-50 % TRAIL-R2, and 40-55 % TRAIL-R3. Conclusions MCF-7 and MDA-MB-231 cells not only released OPG, but expressed RANKL, TRAIL and SDF-1. The majority of the cells also expressed RANK, CXCR4 and TRAIL-R. Since these ligands and their receptors are implicated in the regulation of proliferation, survival, migration and future bone metastasis during breast tumor progression, assessment of these molecules in tumor biopsies of BC patients could be useful in identifying patients with more aggressive tumors that are also at risk of bone metastasis, which may thus improve the available options for therapeutic intervention.

  9. The nucleotide-binding domain of NLRC5 is critical for nuclear import and transactivation activity

    Energy Technology Data Exchange (ETDEWEB)

    Meissner, Torsten B. [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA 02215 (United States); Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02215 (United States); Li, Amy; Liu, Yuen-Joyce [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA 02215 (United States); Gagnon, Etienne [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA 02215 (United States); Institut de Recherche en Immunologie et Cancerologie, Departement de Microbiologie et Immunologie, Universite de Montreal, Montreal, Canada H3T1J4 (Canada); Kobayashi, Koichi S., E-mail: Koichi_Kobayashi@dfci.harvard.edu [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA 02215 (United States); Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02215 (United States)

    2012-02-24

    Highlights: Black-Right-Pointing-Pointer NLRC5 requires an intact NLS for its function as MHC class I transactivator. Black-Right-Pointing-Pointer Nuclear presence of NLRC5 is required for MHC class I induction. Black-Right-Pointing-Pointer Nucleotide-binding controls nuclear import and transactivation activity of NLRC5. -- Abstract: Major histocompatibility complex (MHC) class I and class II are crucial for the function of the human adaptive immune system. A member of the NLR (nucleotide-binding domain, leucine-rich repeat) protein family, NLRC5, has recently been identified as a transcriptional regulator of MHC class I and related genes. While a 'master regulator' of MHC class II genes, CIITA, has long been known, NLRC5 specifically associates with and transactivates the proximal promoters of MHC class I genes. In this study, we analyzed the molecular requirements of NLRC5 nuclear import and transactivation activity. We show that NLRC5-mediated MHC class I gene induction requires an intact nuclear localization signal and nuclear distribution of NLRC5. In addition, we find that the nucleotide-binding domain (NBD) of NLRC5 is critical not only for nuclear translocation but also for the transactivation of MHC class I genes. Changing the cellular localization of NLRC5 is likely to immediately impact MHC class I expression as well as MHC class I-mediated antigen presentation. NLRC5 may thus provide a promising target for the modulation of MHC class I antigen presentation, especially in the setting of transplant medicine.

  10. Recurrent exposure to nicotine differentiates human bronchial epithelial cells via epidermal growth factor receptor activation

    International Nuclear Information System (INIS)

    Martinez-Garcia, Eva; Irigoyen, Marta; Anso, Elena; Martinez-Irujo, Juan Jose; Rouzaut, Ana

    2008-01-01

    Cigarette smoking is the major preventable cause of lung cancer in developed countries. Nicotine (3-(1-methyl-2-pyrrolidinyl)-pyridine) is one of the major alkaloids present in tobacco. Besides its addictive properties, its effects have been described in panoply of cell types. In fact, recent studies have shown that nicotine behaves as a tumor promoter in transformed epithelial cells. This research focuses on the effects of acute repetitive nicotine exposure on normal human bronchial epithelial cells (NHBE cells). Here we show that treatment of NHBE cells with recurrent doses of nicotine up to 500 μM triggered cell differentiation towards a neuronal-like phenotype: cells emitted filopodia and expressed neuronal markers such as neuronal cell adhesion molecule, neurofilament-M and the transcription factors neuronal N and Pax-3. We also demonstrate that nicotine treatment induced NF-kB translocation to the nucleus, phosphorylation of the epidermal growth factor receptor (EGFR), and accumulation of heparin binding-EGF in the extracellular medium. Moreover, addition of AG1478, an inhibitor of EGFR tyrosine phosphorylation, or cetuximab, a monoclonal antibody that precludes ligand binding to the same receptor, prevented cell differentiation by nicotine. Lastly, we show that differentiated cells increased their adhesion to the extracellular matrix and their protease activity. Given that several lung pathologies are strongly related to tobacco consumption, these results may help to better understand the damaging consequences of nicotine exposure

  11. Fibroblast growth factor receptor-2 mutation analysis in human prostate cancer.

    Science.gov (United States)

    Mehta, P; Robson, C N; Neal, D E; Leung, H Y

    2000-10-01

    To assess whether mutations in the hot-spots of the fibroblast growth factor (FGF) receptor-2 gene (FGFR2, exons encoding the IIIa, IIIb, IIIc and transmembrane domain, TMD) are associated with the development of prostate cancer, as the IIIb variant is the specific receptor for FGF7/KGF, an androgen-inducible paracrine factor regulating prostatic growth. Materials and methods Single-strand conformational polymorphism-polymerase chain reaction (SSCP-PCR) and cycle-sequencing analysis were used to screen FGFR2 mutations in 30 patients with prostate cancer; corresponding blood samples were analysed from 11 of the patients. The human prostate cell lines, LNCaP, PC3, DU145, PNT1A and PNT1B were also examined. In addition, 10 foci of invasive cancer from three patients who underwent radical prostatectomy were also assessed. Positive controls containing FGFR2 mutations (Crouzon disease and Pfieffer syndrome) were confirmed by SSCP-PCR and sequencing. Analysis of all prostate tumour samples and prostate-derived cell lines revealed no polymorphisms or mutations in the IIIa, IIIb, IIIc and TMD regions of FGFR2. FGFR2 mutations in the-FGF binding domain and the TMD are not frequent events in human prostate cancer.

  12. Therapies based on inhibitors of the epidermal growth factor receptor: enclosing the future

    International Nuclear Information System (INIS)

    Diaz, Arlhee; Lage, Agustin

    2007-01-01

    The Epidermal Growth Factor Receptor (EGFR) is considered an important target for rational drug design due to its key role in numerous tumors. Potential contribution of EGFR-related signaling pathways to promote tumorigenic processes, including cell proliferation, angiogenesis, and resistance to apoptosis has been well established. Two classes of anti-EGFR agents in late-stage clinical testing include monoclonal antibodies against extracellular EGFR domain (Cetuximab, Nimotuzumab) and small molecules tyrosine kinase inhibitors, which inhibit the receptor enzyme activity (Gefitinib, Erlotinib). A considerable body of evidence has emerged since its introduction in the treatment of cancer patients. However, important questions such as reliable surrogate markers to predict response to the treatment, or optimal sequence and combination of these agents with conventional therapies remain to be addressed. Identify and validate predictive factors to select patients likely to respond to EGFR inhibitors, such as mutations that confer resistance versus those associated with sensitivity is required. A better understanding of molecular mechanisms associated with antitumor activity will useful to predict the interaction of these agents with other therapies in order to avoid antagonisms or overlapping effects resulting in no adding effects. Finally, the benefits derived from EGFR inhibitors as first-line therapy in selected populations, and the optimal doses and ways to delivery to the tumor site resulting in optimal target modulation should be established by the ongoing investigation. (Author)

  13. Physiological epidermal growth factor concentrations activate high affinity receptors to elicit calcium oscillations.

    Directory of Open Access Journals (Sweden)

    Béatrice Marquèze-Pouey

    Full Text Available Signaling mediated by the epidermal growth factor (EGF is crucial in tissue development, homeostasis and tumorigenesis. EGF is mitogenic at picomolar concentrations and is known to bind its receptor on high affinity binding sites depending of the oligomerization state of the receptor (monomer or dimer. In spite of these observations, the cellular response induced by EGF has been mainly characterized for nanomolar concentrations of the growth factor, and a clear definition of the cellular response to circulating (picomolar concentrations is still lacking. We investigated Ca2+ signaling, an early event in EGF responses, in response to picomolar doses in COS-7 cells where the monomer/dimer equilibrium is unaltered by the synthesis of exogenous EGFR. Using the fluo5F Ca2+ indicator, we found that picomolar concentrations of EGF induced in 50% of the cells a robust oscillatory Ca2+ signal quantitatively similar to the Ca2+ signal induced by nanomolar concentrations. However, responses to nanomolar and picomolar concentrations differed in their underlying mechanisms as the picomolar EGF response involved essentially plasma membrane Ca2+ channels that are not activated by internal Ca2+ store depletion, while the nanomolar EGF response involved internal Ca2+ release. Moreover, while the picomolar EGF response was modulated by charybdotoxin-sensitive K+ channels, the nanomolar response was insensitive to the blockade of these ion channels.

  14. Auxin signal transcription factor regulates expression of the brassinosteroid receptor gene in rice.

    Science.gov (United States)

    Sakamoto, Tomoaki; Morinaka, Yoichi; Inukai, Yoshiaki; Kitano, Hidemi; Fujioka, Shozo

    2013-02-01

    The phytohormones auxins and brassinosteroids are both essential regulators of physiological and developmental processes, and it has been suggested that they act inter-dependently and synergistically. In rice (Oryza sativa), auxin co-application improves the brassinosteroid response in the rice lamina inclination bioassay. Here, we showed that auxins stimulate brassinosteroid perception by regulating the level of brassinosteroid receptor. Auxin treatment increased expression of the rice brassinosteroid receptor gene OsBRI1. The promoter of OsBRI1 contains an auxin-response element (AuxRE) that is targeted by auxin-response factor (ARF) transcription factors. An AuxRE mutation abolished the induction of OsBRI1 expression by auxins, and OsBRI1 expression was down-regulated in an arf mutant. The AuxRE motif in the OsBRI1 promoter, and thus the transient up-regulation of OsBRI1 expression caused by treatment with indole-3-acetic acid, is essential for the indole-3-acetic acid-induced increase in sensitivity to brassinosteroids. These findings demonstrate that some ARFs control the degree of brassinosteroid perception required for normal growth and development in rice. Although multi-level interactions between auxins and brassinosteroids have previously been reported, our findings suggest a mechanism by which auxins control cellular sensitivity to brassinosteroids, and further support the notion that interactions between auxins and brassinosteroids are extensive and complex. © 2012 The Authors The Plant Journal © 2012 Blackwell Publishing Ltd.

  15. Familial clustering of recurrent pericarditis may disclose tumour necrosis factor receptor-associated periodic syndrome.

    Science.gov (United States)

    Cantarini, Luca; Lucherini, Orso Maria; Baldari, Cosima Tatiana; Laghi Pasini, Franco; Galeazzi, Mauro

    2010-01-01

    Although several causes of recurrent pericarditis have been identified, the etiology remains obscure in most cases. The tumour necrosis factor receptor-1 associated periodic syndrome (TRAPS) is the most common autosomal dominant autoinflammatory disorder and is caused by mutations in the TNFRSF1A gene encoding the 55-kD receptor for tumour necrosis factor-(TNF)-alpha. Serosal membrane inflammation is a common feature of TRAPS, usually in the form of polyserositis. In addition, patients affected with recurrent pericarditis as the only clinical manifestation of TRAPS have been recently described. Our aim was to investigate the possible involvement of mutations in the TNFRSF1A gene in a cohort of patients affected with idiopathic recurrent pericarditis. Twenty consecutive patients diagnosed with idiopathic recurrent pericarditis were enrolled. Each patient underwent detailed examinations in order to rule out underlying diseases such as infections, connective tissue disorders and malignancies, and mutations of the TNFRSF1A gene were searched for by amplifying, using polymerase chain reaction (PCR), genomic DNA, and direct sequencing. TNFRSF1A mutations were found in 2 of the 20 patients. They were siblings, and they both carried a heterozygous low-penetrance R92Q mutation in the TNFRSF1A gene. Familial clustering has been recently reported in up to 10% of patients with recurrent pericarditis, thus suggesting in some cases a possible genetic predisposition. Our study suggests that familial clustering may represent a clue for investigating mutations in the TNFRSF1A gene in these patients and eventually disclose TRAPS.

  16. Genomic organization of the mouse fibroblast growth factor receptor 3 (Fgfr3) gene

    Energy Technology Data Exchange (ETDEWEB)

    Perez-Castro, A.V.; Wilson, J.; Altherr, M.R. [Los Alamos National Lab., NM (United States)

    1995-11-20

    The fibroblast growth factor receptor 3 (Fgfr3) protein is a tyrosine kinase receptor involved in the signal transduction of various fibroblast growth factors. Recent studies suggest its important role in normal development. In humans, mutation in Fgfr3 is responsible for growth disorders such as achondroplasia, hypoachondroplasia, and thanatophoric dysplasia. Here, we report the complete genomic organization of the mouse Fgfr3 gene. The murine gene spans approximately 15 kb and consists of 19 exons and 18 introns. One major and one minor transcription initiation site were identified. Position +1 is located 614 nucleotides upstream from the ATG initiation codon. The translation initiation and termination sites are located in exons 2 and 19, respectively. Five Sp1 sites, two AP2 sites, one Zeste site, and one Krox 24 site were observed in the 5{prime}-flanking region. The Fgfr3 promoter appears to be contained within a CpG island and, as is common in genes having multiple Sp1-binding sites, lacks a TATA box. 35 refs., 3 figs., 1 tab.

  17. Nutritional factors, physical activity, and breast cancer by hormonal receptor status.

    Science.gov (United States)

    Rosato, Valentina; Bertuccio, Paola; Bosetti, Cristina; Negri, Eva; Edefonti, Valeria; Ferraroni, Monica; Decarli, Adriano; Talamini, Renato; Dal Maso, Luigino; Falcini, Fabio; Montella, Maurizio; Franceschi, Silvia; La Vecchia, Carlo

    2013-10-01

    To investigate the association between food and nutrient intake, occupational and leisure-time physical activity, and body mass index and breast cancer risk by estrogen receptor (ER) and progesterone receptor (PR) status. We analyzed data from a hospital-based case-control study conducted between 1991 and 1994 in Italy, including 1075 women with incident breast cancer and 1477 controls. The associations with breast cancer risk were similar according to ER status for all risk factors considered. In particular, significant reduced risk of ER- and ER+ breast cancers were observed for raw vegetables (multivariate odds ratio, OR, for high vs low consumption: 0.6 and 0.7, respectively) and for polyunsaturated fats (OR: 0.6 and 0.7, respectively). No significant heterogeneous risk estimates were observed for combinations of ER and PR status. Our study does not suggest major differences risk for various dietary and lifestyle factors according to ER and PR breast cancer subtypes. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. The Epidermal Growth Factor Receptor Is a Regulator of Epidermal Complement Component Expression and Complement Activation

    DEFF Research Database (Denmark)

    Abu-Humaidan, Anas H A; Ananthoju, Nageshwar; Mohanty, Tirthankar

    2014-01-01

    The complement system is activated in response to tissue injury. During wound healing, complement activation seems beneficial in acute wounds but may be detrimental in chronic wounds. We found that the epidermal expression of many complement components was only increased to a minor extent in skin...... wounds in vivo and in cultured keratinocytes after exposure to supernatant from stimulated mononuclear cells. In contrast, the epidermal expression of complement components was downregulated in ex vivo injured skin lacking the stimulation from infiltrating inflammatory cells but with intact injury......-induced epidermal growth factor receptor (EGFR)-mediated growth factor response. In cultured primary keratinocytes, stimulation with the potent EGFR ligand, TGF-α, yielded a significant downregulation of complement component expression. Indeed, EGFR inhibition significantly enhanced the induction of complement...

  19. Mechanical stretch augments insulin-induced vascular smooth muscle cell proliferation by insulin-like growth factor-1 receptor

    International Nuclear Information System (INIS)

    Liu, Gang; Hitomi, Hirofumi; Hosomi, Naohisa; Lei, Bai; Nakano, Daisuke; Deguchi, Kazushi; Mori, Hirohito; Masaki, Tsutomu; Ma, Hong; Griendling, Kathy K.; Nishiyama, Akira

    2011-01-01

    Insulin resistance and hypertension have been implicated in the pathogenesis of cardiovascular disease; however, little is known about the roles of insulin and mechanical force in vascular smooth muscle cell (VSMC) remodeling. We investigated the contribution of mechanical stretch to insulin-induced VSMC proliferation. Thymidine incorporation was stimulated by insulin in stretched VSMCs, but not in un-stretched VSMCs. Insulin increased 2-deoxy-glucose incorporation in both stretched and un-stretched VSMCs. Mechanical stretch augmented insulin-induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation. Inhibitors of epidermal growth factor (EGF) receptor tyrosine kinase and Src attenuated insulin-induced ERK and Akt phosphorylation, as well as thymidine incorporation, whereas 2-deoxy-glucose incorporation was not affected by these inhibitors. Moreover, stretch augmented insulin-like growth factor (IGF)-1 receptor expression, although it did not alter the expression of insulin receptor and insulin receptor substrate-1. Insulin-induced ERK and Akt activation, and thymidine incorporation were inhibited by siRNA for the IGF-1 receptor. Mechanical stretch augments insulin-induced VSMC proliferation via upregulation of IGF-1 receptor, and downstream Src/EGF receptor-mediated ERK and Akt activation. Similar to in vitro experiment, IGF-1 receptor expression was also augmented in hypertensive rats. These results provide a basis for clarifying the molecular mechanisms of vascular remodeling in hypertensive patients with hyperinsulinemia. -- Highlights: → Mechanical stretch augments insulin-induced VSMC proliferation via IGF-1 receptor. → Src/EGFR-mediated ERK and Akt phosphorylation are augmented in stretched VSMCs. → Similar to in vitro experiment, IGF-1 receptor is increased in hypertensive rats. → Results provide possible mechanisms of vascular remodeling in hypertension with DM.

  20. Mechanical stretch augments insulin-induced vascular smooth muscle cell proliferation by insulin-like growth factor-1 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Gang [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Department of Anesthesiology, First Affiliated Hospital of China Medical University, Shenyang (China); Hitomi, Hirofumi, E-mail: hitomi@kms.ac.jp [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Hosomi, Naohisa [Department of Cardiorenal and Cerebrovascular Medicine, Faculty of Medicine, Kagawa University, Kagawa (Japan); Lei, Bai; Nakano, Daisuke [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Deguchi, Kazushi; Mori, Hirohito; Masaki, Tsutomu [Department of Gastroenterology and Neurology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Ma, Hong [Department of Anesthesiology, First Affiliated Hospital of China Medical University, Shenyang (China); Griendling, Kathy K. [Department of Medicine, Division of Cardiology, Emory University School of Medicine, Atlanta, GA (United States); Nishiyama, Akira [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan)

    2011-10-15

    Insulin resistance and hypertension have been implicated in the pathogenesis of cardiovascular disease; however, little is known about the roles of insulin and mechanical force in vascular smooth muscle cell (VSMC) remodeling. We investigated the contribution of mechanical stretch to insulin-induced VSMC proliferation. Thymidine incorporation was stimulated by insulin in stretched VSMCs, but not in un-stretched VSMCs. Insulin increased 2-deoxy-glucose incorporation in both stretched and un-stretched VSMCs. Mechanical stretch augmented insulin-induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation. Inhibitors of epidermal growth factor (EGF) receptor tyrosine kinase and Src attenuated insulin-induced ERK and Akt phosphorylation, as well as thymidine incorporation, whereas 2-deoxy-glucose incorporation was not affected by these inhibitors. Moreover, stretch augmented insulin-like growth factor (IGF)-1 receptor expression, although it did not alter the expression of insulin receptor and insulin receptor substrate-1. Insulin-induced ERK and Akt activation, and thymidine incorporation were inhibited by siRNA for the IGF-1 receptor. Mechanical stretch augments insulin-induced VSMC proliferation via upregulation of IGF-1 receptor, and downstream Src/EGF receptor-mediated ERK and Akt activation. Similar to in vitro experiment, IGF-1 receptor expression was also augmented in hypertensive rats. These results provide a basis for clarifying the molecular mechanisms of vascular remodeling in hypertensive patients with hyperinsulinemia. -- Highlights: {yields} Mechanical stretch augments insulin-induced VSMC proliferation via IGF-1 receptor. {yields} Src/EGFR-mediated ERK and Akt phosphorylation are augmented in stretched VSMCs. {yields} Similar to in vitro experiment, IGF-1 receptor is increased in hypertensive rats. {yields} Results provide possible mechanisms of vascular remodeling in hypertension with DM.

  1. Expression of hepatocyte growth factor/scatter factor and its receptor, MET, suggests roles in human embryonic organogenesis.

    Science.gov (United States)

    Kolatsi-Joannou, M; Moore, R; Winyard, P J; Woolf, A S

    1997-05-01

    Hepatocyte growth factor/scatter factor (HGF/SF) is secreted by mesenchymal cells and elicits proliferation, motility, differentiation, and morphogenesis of epithelia and other cells. These effects are mediated by binding to MET, a receptor tyrosine kinase. Genetically engineered mice lacking HGF/SF die in utero due to a failure of placental and hepatocyte differentiation, but little information exists regarding the expression of this signaling system in human development. Using reverse transcriptase-polymerase chain reaction, Western blots, and immunohistochemistry, we report that HGF/SF and MET are expressed during critical early periods of human organogenesis from 6 to 13 wk of gestation. Organs that expressed both genes included liver, metanephric kidney, intestine, and lung, each of which develop by inductive interactions between mesenchyme and epithelia. Of all organs studied, the placenta contained the highest levels of HGF/SF protein, and MET was detected in trophoblastic cells of chorionic villi as early as the 5th wk of gestation. Finally, examination of a human multicystic dysplastic kidney demonstrated that malformed, hyperproliferative tubules expressed MET, whereas HGF/SF protein was immunolocalized to the same epithelia and also to the surrounding undifferentiated cells. Hence HGF/SF might be an important growth factor in normal human embryogenesis and may additionally play a role in human organ malformations.

  2. Vascular endothelial growth factor A and vascular endothelial growth factor receptor 2 expression in non-small cell lung cancer patients: relation to prognosis

    DEFF Research Database (Denmark)

    Bonnesen, Barbara; Pappot, Helle; Holmstav, Julie

    2009-01-01

    elements in neoplastic cells and their microenvironment have recently been and are continuously developed including drugs inhibiting the angiogenic system. Angiogenic factor vascular endothelial growth factor (VEGF) and its receptor vascular endothelial growth factor receptor 2 (VEGFR2) seem to play key......BACKGROUND: The majority of patients with non-small cell lung cancer (NSCLC) are diagnosed with advanced inoperable disease. While treatment with conventional chemotherapy has improved during the last decade the 5 years survival is still modest. Novel drugs, which selectively target aberrant...

  3. A novel interaction between adrenergic receptors and the alpha-subunit of eukaryotic initiation factor 2B.

    Science.gov (United States)

    Klein, U; Ramirez, M T; Kobilka, B K; von Zastrow, M

    1997-08-01

    The alpha-subunit of eukaryotic initiation factor 2B (eIF-2B), a guanine nucleotide exchange protein that functions in regulation of translation, was observed to associate with the carboxyl-terminal cytoplasmic domains of the alpha2A- and alpha2B-adrenergic receptors in a yeast two-hybrid screen of a cDNA library prepared from 293 cells. This protein association was confirmed in vitro by affinity chromatography and was shown to be specific for a subset of G protein-coupled receptors, including the alpha2A-, alpha2B-, alpha2C-, and beta2-adrenergic receptors, but not the vasopressin (V2) receptor. Association of these proteins in vivo was confirmed by specific co-immunoprecipitation of eIF-2Balpha with full-length beta2-adrenergic receptors expressed in transfected 293 cells and by fluorescence microscopy showing co-localization of these proteins in intact cells. Remarkably, eIF-2Balpha co-localized with receptors exclusively in regions of the plasma membrane that are in contact with the extracellular medium, but failed to associate with membranes making cell-cell contacts. Overexpression of eIF-2Balpha in 293 cells caused a small (approximately 15%) but significant enhancement of beta2-adrenergic receptor-mediated activation of adenylyl cyclase, without affecting forskolin or V2 receptor-mediated activation. These observations suggest a new role for a previously identified guanine nucleotide exchange protein in membrane biology and cell signaling.

  4. Search for a platelet-activating factor receptor in the Trypanosoma cruzi proteome: a potential target for Chagas disease chemotherapy

    Directory of Open Access Journals (Sweden)

    Daniel Fábio Kawano

    2011-12-01

    Full Text Available Chagas disease (CD causes the highest burden of parasitic diseases in the Western Hemisphere and is therefore a priority for drug research and development. Platelet-activating factor (PAF causes the CD parasite Trypanosoma cruzi to differentiate, which suggests that the parasite may express PAF receptors. Here, we explored the T. cruzi proteome for PAF receptor-like proteins. From a total of 23,000 protein sequences, we identified 29 hypothetical proteins that are predicted to have seven transmembrane domains (TMDs, which is the main characteristic of the G protein-coupled receptors (GPCRs, including the PAF receptor. The TMDs of these sequences were independently aligned with domains from 25 animal PAF receptors and the sequences were analysed for conserved residues. The conservation score mean values for the TMDs of the hypothetical proteins ranged from 31.7-44.1%, which suggests that if the putative T. cruzi PAF receptor is among the sequences identified, the TMDs are not highly conserved. These results suggest that T. cruzi contains several GPCR-like proteins and that one of these GPCRs may be a PAF receptor. Future studies may further validate the PAF receptor as a target for CD chemotherapy.

  5. Selective Endothelin-B Receptor Stimulation Increases Vascular Endothelial Growth Factor in the Rat Brain during Postnatal Development.

    Science.gov (United States)

    Leonard, M G; Prazad, P; Puppala, B; Gulati, A

    2015-11-01

    Endothelin, vascular endothelial growth factor and nerve growth factor play important roles in development of the central nervous system. ET(B) receptors have been shown to promote neurovascular remodeling in the adult ischemic brain through an increase in VEGF and NGF. It is possible that ET(B) receptors may be involved in postnatal development of the brain through VEGF and NGF. In the present study, the brains of male rat pups on postnatal days 1, 7, 14 and 28 were analyzed for expression of ET(B) receptors, VEGF and NGF. In order to determine the effect of ET(B) receptor stimulation, a separate group of pups were administered saline or ET(B) receptor agonist, IRL-1620, on day 21, and their brains were analyzed on day 28. The intensity of ET(B) receptor and VEGF staining in the vasculature as well as the number of blood vessels of normal pups increased with age and was significantly higher on postnatal day 14 compared to day 1 and day 7. In contrast, both ET(B) and NGF staining intensity in the cortex and subventricular zones decreased (Pbrain (Pdevelopment of the CNS and selective stimulation of ET(B) receptors enhances VEGF but not NGF in the postnatal rat brain. © Georg Thieme Verlag KG Stuttgart · New York.

  6. The Hinge Region as a Key Regulatory Element of Androgen Receptor Dimerization, DNA Binding and Transactivation

    Science.gov (United States)

    2006-05-01

    Chambon (Institut de Génétique et de Biologie Moléculaire et Cellulaire , Illkirch, France). The pCMV-hGal vector was obtained from Stratagene ( La Jolla...oral communication ). I.E. Selectivity of the androgen responses (milestone I.B) A selective ARE was defined as such when the hormone responses...the ∆11 motif indicates a possible communication between these different functions. Also the AR is a phosphoprotein with a function of Serine 650

  7. Transcription factor krüppel-like factor 9 (klf9) as potential predictor of dysfunctional estrogen receptor-a signaling in the uterus

    Science.gov (United States)

    Kruppel-like factors (KLFs) are zinc finger-containing transcription factors that are implicated in diverse physiological processes in the reproductive tract. We previously showed that KLF9, a 33kDa protein family member, influenced estrogen receptor-alpha (ER alpha) expression and activity in mouse...

  8. Flow cytometric detection of growth factor receptors in autografts and analysis of growth factor concentrations in autologous stem cell transplantation: possible significance for platelet recovery

    DEFF Research Database (Denmark)

    Schiødt, I; Jensen, Charlotte Harken; Kjaersgaard, E

    2000-01-01

    In order to improve prediction of hematopoietic recovery, we conducted a pilot study, analyzing the significance of growth factor receptor expression in autografts as well as endogenous growth factor levels in blood before, during and after stem cell transplantation. Three early acting (stem cell...

  9. Functional variation in the arginine vasopressin 2 receptor as a modifier of human plasma von Willebrand factor levels

    DEFF Research Database (Denmark)

    Nossent, Anne Yaël; Robben, J H; Deen, P M T

    2010-01-01

    SUMMARY OBJECTIVES: Stimulation of arginine vasopressin 2 receptor (V2R) with arginine vasopressin (AVP) results in a rise in von Willebrand factor (VWF) and factor VIII plasma levels. We hypothesized that gain-of-function variations in the V2R gene (AVPR2) would lead to higher plasma levels of V...

  10. Effect of corticotropin-releasing factor receptor antagonist on psychologically suppressed masculine sexual behavior in rats.

    Science.gov (United States)

    Miwa, Yoshiji; Nagase, Keiko; Oyama, Nobuyuki; Akino, Hironobu; Yokoyama, Osamu

    2011-03-01

    Corticotropin-releasing factor (CRF) coordinates various responses of the body to stress, and CRF receptors are important targets of treatment for stress-related disorders. To investigate the effect of a nonselective CRF receptor antagonist, astressin, on suppression of masculine sexual behavior by psychological stress in rats. First, we investigated the influence of psychological stress, induced 2 hours per day for three consecutive days, on sexual behavior. Then, rats were divided into 4 groups: a control group, an astressin administration group (A), a psychological stress loading group (PS), and a psychological stress loading and astressin administration group (PS + A). The rats were exposed to sham or psychological stress for three consecutive days. After the last stress loading, the rats were injected with vehicle or astressin, and their sexual behavior was observed. We also measured serum levels of adrenocorticotropic hormone (ACTH). The effects of astressin on sexual behavior and serum levels of ACTH in rats affected by psychological stress were determined. Sexual behavior was reduced after psychological stress loading. The PS rats had significantly longer mount, intromission, and ejaculation latencies and lower ejaculation frequency than did the control, A, and PS + A rats. The intromission latency and ejaculation frequency in the PS + A rats did not achieve the level observed in the controls. There was no significant difference in these parameters between the control and A rats. Serum ACTH levels were significantly lower in PS + A rats than in PS rats. Psychologically suppressed masculine sexual behavior could be partially recovered with astressin administration in rats. These data provide a rationale for the further study of CRF receptor antagonists as novel agents for treating psychological sexual disorders. © 2010 International Society for Sexual Medicine.

  11. The epidermal growth factor receptor as a target for gastrointestinal cancer therapy.

    Science.gov (United States)

    Tedesco, Karen L; Lockhart, A Craig; Berlin, Jordan D

    2004-10-01

    The epidermal growth factor receptor (EGFR) is a member of the family of transmembrane protein kinase receptors known as the erbB or HER receptor family. When activated, EGFR phosphorylates and activates other intracellular proteins that affect cell signaling pathways, cellular proliferation, control of apoptosis and angiogenesis. EGFR signaling is best thought of as a network of activating and inactivating proteins with EGFR as the entry point into the network. EGFR overexpression occurs in most GI malignancies and while data are not entirely consistent, EGFR overexpression often confers a poor prognosis in those GI malignancies that have been studied. It often correlates with poorly differentiated histology, more advanced stage and other known poor prognostic markers. The EGFR is a tempting target because of its presence and overexpression on so many tumor types. However, downstream of the EGFR are several proteins that may be activated without EGFR thus allowing blockade to be overcome. Therefore, while blocking the activity of the EGFR protein appears to be a promising anticancer strategy, a simplistic strategy of blocking only EGFR is likely to only impact a minority of patients. It is time for the laboratory and clinical researchers to work closely together to develop this treatment strategy, moving back and forth from clinical to laboratory to best understand how to block this network effectively enough to produce a broader antitumor effect. While multiple methods of targeting the EGFR pathway are under development, including the inhibition of downstream proteins, only two modalities have entered clinical trials in GI malignancies: small molecule inhibitors of the intracellular kinase domain of EGFR and antibodies designed to block the extracellular ligand-binding domain of EGFR. EGFR inhibitors are still experimental in every GI malignancy with the notable exception of cetuximab that is approved for second or third-line therapy of metastatic colorectal

  12. P2X7R suppression promotes glioma growth through epidermal growth factor receptor signal pathway.

    Science.gov (United States)

    Fang, Jingqin; Chen, Xiao; Zhang, Letian; Chen, Jinhua; Liang, Yi; Li, Xue; Xiang, Jianbo; Wang, Lili; Guo, Guangkuo; Zhang, Bo; Zhang, Weiguo

    2013-06-01

    P2X7 receptor (P2X7R) has been shown to mediate an anticancer effect via apoptosis in different types of cancer. However, whether P2X7R exerts a promoting or suppressive effect on brain glioma is still a controversial issue and its underlying mechanism remains unknown. We showed here that P2X7R suppression exerted a pro-growth effect on glioma through directly promoting cells proliferation and pro-angiogenesis, which was associated with epidermal growth factor receptor (EGFR) signaling. The P2X7R was markedly downregulated by cells exposure to the P2X7R antagonist, brilliant blue G (BBG), moreover, the cells proliferation was enhanced in a dose-dependent manner and the expression of EGFR or p-EGFR protein was significantly upregulated. By constructing C6 cells with reduced expression of P2X7R using shRNA, we also demonstrated strong upregulation in cells proliferation and EGFR/p-EGFR expression. However, this effect of BBG was reversed in the presence of gefitinib or suramin. Magnetic resonance imaging and computed tomography perfusion showed that the BBG or P2X7R shRNA promoted the tumor growth by about 40% and 50%, respectively, and significantly increased angiogenesis. Nissl and Ki-67 staining also confirmed that BBG or P2X7R shRNA notably increased the tumor growth. More importantly, either BBG or P2X7R shRNA could markedly upregulated the expression of EGFR, p-EGFR, HIF-1α and VEGF in glioma cells. In conclusion, P2X7R suppression exerts a promoting effect on glioma growth, which is likely to be related to upregulated EGFR, HIF-1α and VEGF expression. These findings provide important clues to the molecular basis of anticancer effect of targeting purinergic receptors. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. NF-κB Protects NKT Cells from Tumor Necrosis Factor Receptor 1-induced Death.

    Science.gov (United States)

    Kumar, Amrendra; Gordy, Laura E; Bezbradica, Jelena S; Stanic, Aleksandar K; Hill, Timothy M; Boothby, Mark R; Van Kaer, Luc; Joyce, Sebastian

    2017-11-15

    Semi-invariant natural killer T (NKT) cells are innate-like lymphocytes with immunoregulatory properties. NKT cell survival during development requires signal processing by activated RelA/NF-κB. Nonetheless, the upstream signal(s) integrated by NF-κB in developing NKT cells remains incompletely defined. We show that the introgression of Bcl-x L -coding Bcl2l1 transgene into NF-κB signalling-deficient IκBΔN transgenic mouse rescues NKT cell development and differentiation in this mouse model. We reasoned that NF-κB activation was protecting developing NKT cells from death signals emanating either from high affinity agonist recognition by the T cell receptor (TCR) or from a death receptor, such as tumor necrosis factor receptor 1 (TNFR1) or Fas. Surprisingly, the single and combined deficiency in PKC-θ or CARMA-1-the two signal transducers at the NKT TCR proximal signalling node-only partially recapitulated the NKT cell deficiency observed in IκBΔN tg mouse. Accordingly, introgression of the Bcl2l1 transgene into PKC-θ null mouse failed to rescue NKT cell development. Instead, TNFR1-deficiency, but not the Fas-deficiency, rescued NKT cell development in IκBΔN tg mice. Consistent with this finding, treatment of thymocytes with an antagonist of the inhibitor of κB kinase -which blocks downstream NF-κB activation- sensitized NKT cells to TNF-α-induced cell death in vitro. Hence, we conclude that signal integration by NF-κB protects developing NKT cells from death signals emanating from TNFR1, but not from the NKT TCR or Fas.

  14. Targeting the epidermal growth factor receptor in radiotherapy: radiobiological mechanisms, preclinical and clinical results

    International Nuclear Information System (INIS)

    Baumann, Michael; Krause, Mechthild

    2004-01-01

    Background and purpose: Inhibition of the epidermal growth factor receptor (EGFR) is a fastly developing field in preclinical and clinical cancer research. This review presents the current status of knowledge and discusses radiobiological mechanisms which may underly the efficacy of EGFR inhibitors combined with irradiation. Materials and methods: Preclinical and clinical results on combined targeting of the EGFR and irradiation from the literature and from this laboratory are reviewed. Focus is given to the radiobiological rationale of this approach and to endpoints of experimental radiotherapy. Results: Overexpression of the EGFR is associated with decreased local tumour control after radiotherapy, especially when the overall treatment time is long. Inhibition of the EGFR either alone or in combination with irradiation decreases the growth rate of tumours expressing this receptor. Preclinical data provide proof-of-principle that local tumour control may be improved by combining irradiation with C225 mAb. In a randomised phase III clinical trial, simultaneous irradiation and treatment with the EGFR antibody Cetuximab (Erbitux[reg]; C225) in head and neck cancer patients resulted in significantly improved locoregional tumour control and survival compared to curative irradiation alone. Acute skin reactions increased in the experimental arm. The underlying mechanisms of enhanced radiation effects of combined EGFR inhibition with irradiation and of the partly conflicting results in different studies are poorly understood. There is increasing evidence, that important intertumoral heterogeneity in the response to EGFR inhibition alone and combined with irradiation exists, which appears to be at least partly dependent on specific mutations of the receptor as well as of molecules that are involved in the intracellular signal transduction pathway. Conclusions and outlook: Further investigations at all levels of the translational research chain exploring the mechanisms of

  15. A possible new target in lung-cancer cells: The orphan receptor, bombesin receptor subtype-3.

    Science.gov (United States)

    Moreno, Paola; Mantey, Samuel A; Lee, Suk H; Ramos-Álvarez, Irene; Moody, Terry W; Jensen, Robert T

    2018-03-01

    Human bombesin receptors, GRPR and NMBR, are two of the most frequently overexpressed G-protein-coupled-receptors by lung-cancers. Recently, GRPR/NMBR are receiving considerable attention because they act as growth factor receptors often in an autocrine manner in different lung-cancers, affect tumor angiogenesis, their inhibition increases the cytotoxic potency of tyrosine-kinase inhibitors reducing lung-cancer cellular resistance/survival and their overexpression can be used for sensitive tumor localization as well as to target cytotoxic agents to the cancer. The orphan BRS-3-receptor, because of homology is classified as a bombesin receptor but has received little attention, despite the fact that it is also reported in a number of studies in lung-cancer cells and has growth effects in these cells. To address its potential importance, in this study, we examined the frequency/relative quantitative expression of human BRS-3 compared to GRPR/NMBR and the effects of its activation on cell-signaling/growth in 13 different human lung-cancer cell-lines. Our results showed that BRS-3 receptor is expressed in 92% of the cell-lines and that it is functional in these cells, because its activation stimulates phospholipase-C with breakdown of phosphoinositides and changes in cytosolic calcium, stimulates ERK/MAPK and stimulates cell growth by EGFR transactivation in some, but not all, the lung-cancer cell-lines. These results suggest that human BRS-3, similar to GRPR/NMBR, is frequently ectopically-expressed by lung-cancer cells in which, it is functional, affecting cell signaling/growth. These results suggest that similar to GRPR/NMBR, BRS-3 should receive increased attention as possible approach for the development of novel treatments and/or diagnosis in lung-cancer. Published by Elsevier Inc.

  16. Pioneer Factors FOXA1 and FOXA2 Assist Selective Glucocorticoid Receptor Signaling in Human Endometrial Cells.

    Science.gov (United States)

    Whirledge, Shannon; Kisanga, Edwina P; Taylor, Robert N; Cidlowski, John A

    2017-11-01

    Successful pregnancy relies on dynamic control of cell signaling to achieve uterine receptivity and the necessary biological changes required for endometrial decidualization, embryo implantation, and fetal development. Glucocorticoids are master regulators of intracellular signaling and can directly regulate embryo implantation and endometrial remodeling during murine pregnancy. In immortalized human uterine cells, we have shown that glucocorticoids and estradiol (E2) coregulate thousands of genes. Recently, glucocorticoids and E2 were shown to coregulate the expression of Left-right determination factor 1 (LEFTY1), previously implicated in the regulation of decidualization. To elucidate the molecular mechanism by which glucocorticoids and E2 regulate the expression of LEFTY1, immortalized and primary human endometrial cells were evaluated for gene expression and receptor recruitment to regulatory regions of the LEFTY1 gene. Glucocorticoid administration induced expression of LEFTY1 messenger RNA and protein and recruitment of the glucocorticoid receptor (GR) and activated polymerase 2 to the promoter of LEFTY1. Glucocorticoid-mediated recruitment of GR was dependent on pioneer factors FOXA1 and FOXA2. E2 was found to antagonize glucocorticoid-mediated induction of LEFTY1 by reducing recruitment of GR, FOXA1, FOXA2, and activated polymerase 2 to the LEFTY1 promoter. Gene expression analysis identified several genes whose glucocorticoid-dependent induction required FOXA1 and FOXA2 in endometrial cells. These results suggest a molecular mechanism by which E2 antagonizes GR-dependent induction of specific genes by preventing the recruitment of the pioneer factors FOXA1 and FOXA2 in a physiologically relevant model. Copyright © 2017 Endocrine Society.

  17. Selective tumor necrosis factor receptor I blockade is antiinflammatory and reveals immunoregulatory role of tumor necrosis factor receptor II in collagen-induced arthritis.

    Science.gov (United States)

    McCann, Fiona E; Perocheau, Dany P; Ruspi, Gerhard; Blazek, Katrina; Davies, Marie L; Feldmann, Marc; Dean, Jonathan L E; Stoop, A Allart; Williams, Richard O

    2014-10-01

    Tumor necrosis factor (TNF) signals via 2 receptors, TNFR type I (TNFRI) and TNFRII, with distinct cellular distribution and signaling functions. In rheumatoid arthritis (RA), the net effect of TNFR signaling favors inflammatory responses while inhibiting the activity of regulatory T cells. TNFRII signaling has been shown to promote Treg cell function. To assess the relative contributions of TNFRI and TNFRII signaling to inflammatory and regulatory responses in vivo, we compared the effect of TNF blockade, hence TNFRI/II, versus TNFRI alone in collagen-induced arthritis (CIA) as a model of RA. Mice with established arthritis were treated for 10 days with anti-mouse TNFRI domain antibody (dAb; DMS5540), an isotype control dAb (DMS5538), or murine TNFRII genetically fused with mouse IgG1 Fc domain (mTNFRII-Fc) beginning on the day of arthritis onset, and disease progression was monitored. Systemic cytokine concentrations and numbers of T cell subsets in lymph nodes and spleens were measured, and intrinsic Treg cell function was determined by ex vivo suppression assays. Progression of CIA was suppressed similarly by TNFRI (DMS5540) and TNFRI/II (mTNFRII-Fc) blockade. However, blockade of TNFRI/II led to increased effector T cell activity, which was not observed after selective TNFRI blockade, suggesting an immunoregulatory role of TNFRII. In support of this, TNFRI blockade, but not TNFRI/II blockade, expanded and activated Treg cells. Furthermore, a dramatic increase in expression of the Treg cell signature genes FoxP3 and TNFRII was observed in joints undergoing remission, which supports the notion that these molecules have a physiologic role in the resolution of inflammation. We propose that a therapeutic strategy that targets TNFRI while sparing TNFRII has the potential to both inhibit inflammation and promote Treg cell activity, which might be superior to TNF blockade. Copyright © 2014 by the American College of Rheumatology.

  18. Amphiregulin triggered epidermal growth factor receptor activation confers in vivo crizotinib-resistance of EML4-ALK lung cancer and circumvention by epidermal growth factor receptor inhibitors.

    Science.gov (United States)

    Taniguchi, Hirokazu; Takeuchi, Shinji; Fukuda, Koji; Nakagawa, Takayuki; Arai, Sachiko; Nanjo, Shigeki; Yamada, Tadaaki; Yamaguchi, Hiroyuki; Mukae, Hiroshi; Yano, Seiji

    2017-01-01

    Crizotinib, a first-generation anaplastic lymphoma kinase (ALK) tyrosine-kinase inhibitor, is known to be effective against echinoderm microtubule-associated protein-like 4 (EML4)-ALK-positive non-small cell lung cancers. Nonetheless, the tumors subsequently become resistant to crizotinib and recur in almost every case. The mechanism of the acquired resistance needs to be deciphered. In this study, we established crizotinib-resistant cells (A925LPE3-CR) via long-term administration of crizotinib to a mouse model of pleural carcinomatous effusions; this model involved implantation of the A925LPE3 cell line, which harbors the EML4-ALK gene rearrangement. The resistant cells did not have the secondary ALK mutations frequently occurring in crizotinib-resistant cells, and these cells were cross-resistant to alectinib and ceritinib as well. In cell clone #2, which is one of the clones of A925LPE3-CR, crizotinib sensitivity was restored via the inhibition of epidermal growth factor receptor (EGFR) by means of an EGFR tyrosine-kinase inhibitor (erlotinib) or an anti-EGFR antibody (cetuximab) in vitro and in the murine xenograft model. Cell clone #2 did not have an EGFR mutation, but the expression of amphiregulin (AREG), one of EGFR ligands, was significantly increased. A knockdown of AREG with small interfering RNAs restored the sensitivity to crizotinib. These data suggest that overexpression of EGFR ligands such as AREG can cause resistance to crizotinib, and that inhibition of EGFR signaling may be a promising strategy to overcome crizotinib resistance in EML4-ALK lung cancer. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  19. Vascular endothelial growth factor receptor-1 contributes to resistance to anti-epidermal growth factor receptor drugs in human cancer cells.

    Science.gov (United States)

    Bianco, Roberto; Rosa, Roberta; Damiano, Vincenzo; Daniele, Gennaro; Gelardi, Teresa; Garofalo, Sonia; Tarallo, Valeria; De Falco, Sandro; Melisi, Davide; Benelli, Roberto; Albini, Adriana; Ryan, Anderson; Ciardiello, Fortunato; Tortora, Giampaolo

    2008-08-15

    The resistance to selective EGFR inhibitors involves the activation of alternative signaling pathways, and Akt activation and VEGF induction have been described in EGFR inhibitor-resistant tumors. Combined inhibition of EGFR and other signaling proteins has become a successful therapeutic approach, stimulating the search for further determinants of resistance as basis for novel therapeutic strategies. We established human cancer cell lines with various degrees of EGFR expression and sensitivity to EGFR inhibitors and analyzed signal transducers under the control of EGFR-dependent and EGFR-independent pathways. Multitargeted inhibitor vandetanib (ZD6474) inhibited the growth and the phosphorylation of Akt and its effector p70S6 kinase in both wild-type and EGFR inhibitor-resistant human colon, prostate, and breast cancer cells. We found that the resistant cell lines exhibit, as common feature, VEGFR-1/Flt-1 overexpression, increased secretion of VEGF and placental growth factor, and augmented migration capabilities and that vandetanib is able to antagonize them. Accordingly, a new kinase assay revealed that in addition to VEGF receptor (VEGFR)-2, RET, and EGFR, vandetanib efficiently inhibits also VEGFR-1. The contribution of VEGFR-1 to the resistant phenotype was further supported by the demonstration that VEGFR-1 silencing in resistant cells restored sensitivity to anti-EGFR drugs and impaired migration capabilities, whereas exogenous VEGFR-1 overexpression in wild-type cells conferred resistance to these agents. This study shows that VEGFR-1 contributes to anti-EGFR drug resistance in different human cancer cells. Moreover, vandetanib inhibits VEGFR-1 activation, cell proliferation, and migration, suggesting its potential utility in patients resistant to EGFR inhibitors.

  20. Association of Polymorphisms in Connective Tissue Growth Factor and Epidermal Growth Factor Receptor Genes With Human Longevity.

    Science.gov (United States)

    Donlon, Timothy A; Morris, Brian J; He, Qimei; Chen, Randi; Masaki, Kamal H; Allsopp, Richard C; Willcox, D Craig; Tranah, Gregory J; Parimi, Neeta; Evans, Daniel S; Flachsbart, Friederike; Nebel, Almut; Kim, Duk-Hwan; Park, Joobae; Willcox, Bradley J

    2017-08-01

    Growth pathways play key roles in longevity. The present study tested single-nucleotide polymorphisms (SNPs) in the connective tissue growth factor gene (CTGF) and the epidermal growth factor receptor gene (EGFR) for association with longevity. Comparison of allele and genotype frequencies of 12 CTGF SNPs and 41 EGFR SNPs between 440 American men of Japanese ancestry aged ≥95 years and 374 men of average life span revealed association with longevity at the p cases, consistent with heterozygote advantage in living to extreme old age. No associations of the most significant SNPs were observed in whites or Koreans. In conclusion, the present findings indicate that genetic variation in CTGF and EGFR may contribute to the attainment of extreme old age in Japanese. More research is needed to confirm that genetic variation in CTGF and EGFR contributes to the attainment of extreme old age across human populations. © The Author 2016. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  1. Binding characteristics of brain-derived neurotrophic factor to its receptors on neurons from the chick embryo

    International Nuclear Information System (INIS)

    Rodriguez-Tebar, A.; Barde, Y.A.

    1988-01-01

    Brain-derived neurotrophic factor (BDNF), a protein known to support the survival of embryonic sensory neurons and retinal ganglion cells, was derivatized with 125I-Bolton-Hunter reagent and obtained in a biologically active, radioactive form (125I-BDNF). Using dorsal root ganglion neurons from chick embryos at 9 d of development, the basic physicochemical parameters of the binding of 125I-BDNF with its receptors were established. Two different classes of receptors were found, with dissociation constants of 1.7 x 10(-11) M (high-affinity receptors) and 1.3 x 10(-9) M (low-affinity receptors). Unlabeled BDNF competed with 125I-BDNF for binding to the high-affinity receptors with an inhibition constant essentially identical to the dissociation constant of the labeled protein: 1.2 x 10(-11) M. The association and dissociation rates from both types of receptors were also determined, and the dissociation constants calculated from these kinetic experiments were found to correspond to the results obtained from steady-state binding. The number of high-affinity receptors (a few hundred per cell soma) was 15 times lower than that of low-affinity receptors. No high-affinity receptors were found on sympathetic neurons, known not to respond to BDNF, although specific binding of 125I-BDNF to these cells was detected at a high concentration of the radioligand. These results are discussed and compared with those obtained with nerve growth factor on the same neuronal populations

  2. Suppression of estrogen receptor transcriptional activity by connective tissue growth factor.

    Directory of Open Access Journals (Sweden)

    Long Cheng

    Full Text Available Secreted growth factors have been shown to stimulate the transcriptional activity of estrogen receptors (ER that are responsible for many biological processes. However, whether these growth factors physically interact with ER remains unclear. Here, we show for the first time that connective tissue growth factor (CTGF physically and functionally associates with ER. CTGF interacted with ER both in vitro and in vivo. CTGF interacted with ER DNA-binding domain. ER interaction region in CTGF was mapped to the thrombospondin type I repeat, a cell attachment motif. Overexpression of CTGF inhibited ER transcriptional activity as well as the expression of estrogen-responsive genes, including pS2 and cathepsin D. Reduction of endogenous CTGF with CTGF small interfering RNA enhanced ER transcriptional activity. The interaction between CTGF and ER is required for the repression of estrogen-responsive transcription by CTGF. Moreover, CTGF reduced ER protein expression, whereas the CTGF mutant that did not repress ER transcriptional activity also did not alter ER protein levels. The results suggested the transcriptional regulation of estrogen signaling through interaction between CTGF and ER, and thus may provide a novel mechanism by which cross-talk between secreted growth factor and ER signaling pathways occurs.

  3. Pluripotency factors and Polycomb Group proteins repress aryl hydrocarbon receptor expression in murine embryonic stem cells

    Directory of Open Access Journals (Sweden)

    Chia-I Ko

    2014-01-01

    Full Text Available The aryl hydrocarbon receptor (AHR is a transcription factor and environmental sensor that regulates expression of genes involved in drug-metabolism and cell cycle regulation. Chromatin immunoprecipitation analyses, Ahr ablation in mice and studies with orthologous genes in invertebrates suggest that AHR may also play a significant role in embryonic development. To address this hypothesis, we studied the regulation of Ahr expression in mouse embryonic stem cells and their differentiated progeny. In ES cells, interactions between OCT3/4, NANOG, SOX2 and Polycomb Group proteins at the Ahr promoter repress AHR expression, which can also be repressed by ectopic expression of reprogramming factors in hepatoma cells. In ES cells, unproductive RNA polymerase II binds at the Ahr transcription start site and drives the synthesis of short abortive transcripts. Activation of Ahr expression during differentiation follows from reversal of repressive marks in Ahr promoter chromatin, release of pluripotency factors and PcG proteins, binding of Sp factors, establishment of histone marks of open chromatin, and engagement of active RNAPII to drive full-length RNA transcript elongation. Our results suggest that reversible Ahr repression in ES cells holds the gene poised for expression and allows for a quick switch to activation during embryonic development.

  4. Emerging role of epidermal growth factor receptor inhibition in therapy for advanced malignancy: focus on NSCLC

    International Nuclear Information System (INIS)

    Langer, Corey J.

    2004-01-01

    Combination chemotherapy regimens have emerged as the standard approach in advanced non-small-cell lung cancer. Meta-analyses have demonstrated a 2-month increase in median survival after platinum-based therapy vs. best supportive care, and an absolute 10% improvement in the 1-year survival rate. Just as importantly, cytotoxic therapy has produced benefits in symptom control and quality of life. Newer agents, including the taxanes, vinorelbine, gemcitabine, and irinotecan, have expanded our therapeutic options in the treatment of advanced non-small-cell lung cancer. Despite their contributions, we have reached a therapeutic plateau, with response rates seldom exceeding 30-40% in cooperative group studies and 1-year survival rates stable between 30% and 40%. It is doubtful that substituting one agent for another in various combinations will lead to any further improvement in these rates. The thrust of current research has focused on targeted therapy, and epidermal growth factor receptor inhibition is one of the most promising clinical strategies. Epidermal growth factor receptor inhibitors currently under investigation include the small molecules gefitinib (Iressa, ZD1839) and erlotinib (Tarceva, OSI-774), as well as monoclonal antibodies such as cetuximab (IMC-225, Erbitux). Agents that have only begun to undergo clinical evaluation include CI-1033, an irreversible pan-erbB tyrosine kinase inhibitor, and PKI166 and GW572016, both examples of dual kinase inhibitors (inhibiting epidermal growth factor receptor and Her2). Preclinical models have demonstrated synergy for all these agents in combination with either chemotherapy or radiotherapy, leading to great enthusiasm regarding their ultimate contribution to lung cancer therapy. However, serious clinical challenges persist. These include the identification of the optimal dose(s); the proper integration of these agents into popular, established cytotoxic regimens; and the selection of the optimal setting(s) in which

  5. DNA homologous recombination factor SFR1 physically and functionally interacts with estrogen receptor alpha.

    Directory of Open Access Journals (Sweden)

    Yuxin Feng

    Full Text Available Estrogen receptor alpha (ERα, a ligand-dependent transcription factor, mediates the expression of its target genes by interacting with corepressors and coactivators. Since the first cloning of SRC1, more than 280 nuclear receptor cofactors have been identified, which orchestrate target gene transcription. Aberrant activity of ER or its accessory proteins results in a number of diseases including breast cancer. Here we identified SFR1, a protein involved in DNA homologous recombination, as a novel binding partner of ERα. Initially isolated in a yeast two-hybrid screen, the interaction of SFR1 and ERα was confirmed in vivo by immunoprecipitation and mammalian one-hybrid assays. SFR1 co-localized with ERα in the nucleus, potentiated ER's ligand-dependent and ligand-independent transcriptional activity, and occupied the ER binding sites of its target gene promoters. Knockdown of SFR1 diminished ER's transcriptional activity. Manipulating SFR1 expression by knockdown and overexpression revealed a role for SFR1 in ER-dependent and -independent cancer cell proliferation. SFR1 differs from SRC1 by the lack of an intrinsic activation function. Taken together, we propose that SFR1 is a novel transcriptional modulator for ERα and a potential target in breast cancer therapy.

  6. Insulin and insulin-like growth factor receptors in the brain: physiological and pathological aspects.

    Science.gov (United States)

    Werner, Haim; LeRoith, Derek

    2014-12-01

    The involvement of insulin, the insulin-like growth factors (IGF1, IGF2) and their receptors in central nervous system development and function has been the focus of scientific interest for more than 30 years. The insulin-like peptides, both locally-produced proteins as well as those transported from the circulation into the brain via the blood-brain barrier, are involved in a myriad of biological activities. These actions include, among others, neuronal survival, neurogenes, angiogenesis, excitatory and inhibitory neurotransmission, regulation of food intake, and cognition. In recent years, a linkage between brain insulin/IGF1 and certain neuropathologies has been identified. Epidemiological studies have demonstrated a correlation between diabetes (mainly type 2) and Alzheimer׳s disease. In addition, an aberrant decline in IGF1 values was suggested to play a role in the development of Alzheimer׳s disease. The present review focuses on the expression and function of insulin, IGFs and their receptors in the brain in physiological and pathological conditions. Copyright © 2014 Elsevier B.V. and ECNP. All rights reserved.

  7. Pertuzumab: Unprecedented benefit in human epidermal growth factor receptor 2-positive breast cancer

    Directory of Open Access Journals (Sweden)

    Amit Rauthan

    2015-01-01

    Full Text Available Human epidermal growth factor receptor 2 (HER2-positive breast cancer is a more aggressive subtype of breast cancer and targeting the HER2 receptor has proven effective in improving the prognosis of these patients. Pertuzumab, a recombinant humanized monoclonal antibody and the first in a class of HER2 dimerization inhibitors approved for treating HER2+ breast cancer. It blocks ligand-dependent heterodimerization and ligand-independent homodimerization of HER2 with other HER members. When used in combination with trastuzumab and taxane, pertuzumab complements the action of trastuzumab and results in a comprehensive blockade of HER2 signaling pathway. This review article traces the development of pertuzumab from concept to its current use in HER2+ breast cancer treatment. A search of Medical Literature Published since 2007 was performed in PubMed using the keywords "pertuzumab," "HER2+ breast cancer," "HER2 targeted therapy," "metastatic breast cancer," and in search engines for ongoing trials with pertuzumab and incidence of cancer and breast cancer in India. A total of 35 publications and abstracts from the American Society of Clinical Oncology were selected for this review. Pertuzumab is approved in combination with trastuzumab and docetaxel for the treatment of patients with HER2+ metastatic BC, who have not received prior anti-HER2 therapy or chemotherapy for metastatic disease. The dual HER2 blockade of pertuzumab and trastuzumab is now accepted worldwide as a standard of care by various guidelines.

  8. Alterations in epidermal growth factor receptors 1 and 2 in esophageal squamous cell carcinomas.

    Science.gov (United States)

    Gonzaga, Isabela Martins; Soares-Lima, Sheila Coelho; de Santos, Paulo Thiago Souza; Blanco, Tania Cristina Moita; de Reis, Bruno Souza Bianchi; Quintella, Danielle Carvalho; de Oliveira, Ivanir Martins; de Faria, Paulo Antonio Silvestre; Kruel, Cleber Dario Pinto; Andreollo, Nelson Adami; de Simão, Tatiana Almeida; Pinto, Luis Felipe Ribeiro

    2012-12-04

    Esophageal squamous cell carcinoma (ESCC) shows a 5-year survival rate below 10%, demonstrating the urgency in improving its treatment. Alterations in epidermal growth factor receptors are closely related to malignancy transformation in a number of tumors and recent successful targeted therapies have been directed to these molecules. Therefore, in this study, we analyzed the expression of EGFR and HER2 and evaluated EGFR mutation profile as well as the presence of mutations in hotspots of KRAS and BRAF in ESCC patients. We performed RT-qPCR, immunohistochemistry and Fluorescent in situ hybridization to determine EGFR and HER2 expression in ESCC patients, and direct sequencing and PCR-RFLP for mutations and polymorphism analysis. Our results showed an increased EGFR mRNA expression in tumors compared to surrounding tissue (p T) in 2.1% of the patients, and at codon 787 (G>A) in 79.2% of the cases. This last polymorphism was also evaluated in 304 healthy controls, which presented a similar frequency (73.7%) in comparison with ESCC patients. The absence of mutations of EGFR, KRAS and BRAF as well as the overexpression of EGFR and HER2 in less than 10% of the patients suggest that this signaling pathway is altered in only a small proportion of patients with ESCC. HER receptors target therapies may have the potential to be effective in only a minor fraction of patients with ESCC.

  9. Insulin-like growth factor-1 receptor expression in oral squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Devipriyaa B. Sundaram

    2011-12-01

    Full Text Available Objectives: The Insulin-like growth factor-I receptor (IGF-1R plays critical roles in cancer development, proliferation, motility and survival. IGF-1R over expression is frequently found in various tumours and is often associated with an aggressive phenotype. Hence, the aim of the present study was to examine the expression of IGF-1R in normal oral mucosa, fibroepithelial polyps, dysplastic oral mucosa and well-differentiated squamous cell carcinomas.Materials and methods: A 3-layered streptavidin peroxidase immunohistochemical method was used to detect the expression of IGF-1R in normal oral mucosa, fibroepithelial polyps, dysplastic oral mucosa and well-differentiated squamous cell carcinomas.Results: All squamous cell carcinomas (15 out of 15 patients showed intense immunoreactivity for IGF-1R. Moderate immunoreactivity was seen in dysplastic oral lesions (12 out of 12 lesions with positive staining in the prickle cell layer. The staining distribution in the benign lesions (14 out of 14 lesions was weaker and similar to that seen in normal oral mucosa (10 out of 10 samples when compared to squamous cell carcinomas and dysplastic lesions.Conclusions: Our results demonstrate increased IGF-1 receptor expression in oral squamous cell carcinomas which suggests that IGF-1 may have an important role in the development of oral cancer. J Clin Exp Invest 2011; 2 (4: 354-361

  10. Mechanism for activation of mutated epidermal growth factor receptors in lung cancer.

    Science.gov (United States)

    Red Brewer, Monica; Yun, Cai-Hong; Lai, Darson; Lemmon, Mark A; Eck, Michael J; Pao, William

    2013-09-17

    The initiation of epidermal growth factor receptor (EGFR) kinase activity proceeds via an asymmetric dimerization mechanism in which a "donor" tyrosine kinase domain (TKD) contacts an "acceptor" TKD, leading to its activation. In the context of a ligand-induced dimer, identical wild-type EGFR TKDs are thought to assume the donor or acceptor roles in a random manner. Here, we present biochemical reconstitution data demonstrating that activated EGFR mutants found in lung cancer preferentially assume the acceptor role when coexpressed with WT EGFR. Mutated EGFRs show enhanced association with WT EGFR, leading to hyperphosphorylation of the WT counterpart. Mutated EGFRs also hyperphosphorylate the related erythroblastic leukemia viral oncogene (ErbB) family member, ErbB-2, in a similar manner. This directional "superacceptor activity" is particularly pronounced in the drug-resistant L834R/T766M mutant. A 4-Å crystal structure of this mutant in the active conformation reveals an asymmetric dimer interface that is essentially the same as that in WT EGFR. Asymmetric dimer formation induces an allosteric conformational change in the acceptor subunit. Thus, superacceptor activity likely arises simply from a lower energetic cost associated with this conformational change in the mutant EGFR compared with WT, rather than from any structural alteration that impairs the donor role of the mutant. Collectively, these findings define a previously unrecognized mode of mutant-specific intermolecular regulation for ErbB receptors, knowledge of which could potentially be exploited for therapeutic benefit.

  11. The Influence of Adnectin Binding on the Extracellular Domain of Epidermal Growth Factor Receptor

    Science.gov (United States)

    Iacob, Roxana E.; Chen, Guodong; Ahn, Joomi; Houel, Stephane; Wei, Hui; Mo, Jingjie; Tao, Li; Cohen, Daniel; Xie, Dianlin; Lin, Zheng; Morin, Paul E.; Doyle, Michael L.; Tymiak, Adrienne A.; Engen, John R.

    2014-12-01

    The precise and unambiguous elucidation and characterization of interactions between a high affinity recognition entity and its cognate protein provides important insights for the design and development of drugs with optimized properties and efficacy. In oncology, one important target protein has been shown to be the epidermal growth factor receptor (EGFR) through the development of therapeutic anticancer antibodies that are selective inhibitors of EGFR activity. More recently, smaller protein derived from the 10th type III domain of human fibronectin termed an adnectin has also been shown to inhibit EGFR in clinical studies. The mechanism of EGFR inhibition by either an adnectin or an antibody results from specific binding of the high affinity protein to the extracellular portion of EGFR (exEGFR) in a manner that prevents phosphorylation of the intracellular kinase domain of the receptor and thereby blocks intracellular signaling. Here, the structural changes induced upon binding were studied by probing the solution conformations of full length exEGFR alone and bound to a cognate adnectin through hydrogen/deuterium exchange mass spectrometry (HDX MS). The effects of binding in solution were identified and compared with the structure of a bound complex determined by X-ray crystallography.

  12. Regulation of granulocyte colony-stimulating factor receptor-mediated granulocytic differentiation by C-mannosylation.

    Science.gov (United States)

    Otani, Kei; Niwa, Yuki; Suzuki, Takehiro; Sato, Natsumi; Sasazawa, Yukiko; Dohmae, Naoshi; Simizu, Siro

    2018-04-06

    Granulocyte colony-stimulating factor (G-CSF) receptor (G-CSFR) is a type I cytokine receptor which is involved in hematopoietic cell maturation. G-CSFR has three putative C-mannosylation sites at W253, W318, and W446; however, it is not elucidated whether G-CSFR is C-mannosylated or not. In this study, we first demonstrated that G-CSFR was C-mannosylated at only W318. We also revealed that C-mannosylation of G-CSFR affects G-CSF-dependent downstream signaling through changing ligand binding capability but not cell surface localization. Moreover, C-mannosylation of G-CSFR was functional and regulated granulocytic differentiation in myeloid 32D cells. In conclusion, we found that G-CSFR is C-mannosylated at W318 and that this C-mannosylation has role(s) for myeloid cell differentiation through regulating downstream signaling. Copyright © 2018 Elsevier Inc. All rights reserved.

  13. Heterodimerization of wild-type and mutant fibroblast growth factor receptors in cell-derived vesicles

    Science.gov (United States)

    Hristova, Kalina; Del Piccolo, Nuala; Sarabipour, Sarvenaz

    The activity of receptor tyrosine kinases (RTKs) is controlled through their lateral dimerization in the plasma membrane. RTKs are believed to form both homodimers and heterodimers, and the different dimers are believed to play unique roles in cell signaling. However, RTK heterodimers remain poorly characterized, as compared to homodimers, due to limitations in current experimental methods. Here, we develop a Förster Resonance Energy Transfer (FRET)-based methodology to assess the thermodynamics of hetero-interactions in the plasma membrane. To demonstrate the utility of the methodology, we use it to study the hetero-interactions between three Fibroblast Growth Factor Receptors - FGFR1, FGFR2, and FGFR3 - in the absence of ligand. Our results show that all possible FGFR heterodimers form, suggesting that the biological roles of FGFR heterodimers may be as significant as the homodimer roles. We further investigate the effect of two pathogenic point mutations in FGFR3 (A391E and G380R) on heterodimerization. We show that each of these mutations stabilize most of the heterodimers, with the largest effects observed for FGFR3 wild-type/mutant heterodimers. We thus demonstrate that the methodology presented here can yield new knowledge about RTK interactions and can further our understanding of signal transduction across the plasma membrane..

  14. Homogeneous expansion of human T-regulatory cells via tumor necrosis factor receptor 2.

    Science.gov (United States)

    Okubo, Yoshiaki; Mera, Toshiyuki; Wang, Limei; Faustman, Denise L

    2013-11-06

    T-regulatory cells (T(regs)) are a rare lymphocyte subtype that shows promise for treating infectious disease, allergy, graft-versus-host disease, autoimmunity, and asthma. Clinical applications of T(regs) have not been fully realized because standard methods of expansion ex vivo produce heterogeneous progeny consisting of mixed populations of CD4 + T cells. Heterogeneous progeny are risky for human clinical trials and face significant regulatory hurdles. With the goal of producing homogeneous T(regs), we developed a novel expansion protocol targeting tumor necrosis factor receptors (TNFR) on T(regs). In in vitro studies, a TNFR2 agonist was found superior to standard methods in proliferating human T(regs) into a phenotypically homogeneous population consisting of 14 cell surface markers. The TNFR2 agonist-expanded T(regs) also were functionally superior in suppressing a key T(reg) target cell, cytotoxic T-lymphocytes. Targeting the TNFR2 receptor during ex vivo expansion is a new means for producing homogeneous and potent human T(regs) for clinical opportunities.

  15. Fcgamma receptor IIIA polymorphism as a risk-factor for coronary artery disease.

    Science.gov (United States)

    Gavasso, Sonia; Nygård, Ottar; Pedersen, Eva Ringdal; Aarseth, Jan H; Bleie, Oyvind; Myhr, Kjell-Morten; Vedeler, Christian A

    2005-06-01

    Inflammation is important in the pathogenesis of atherosclerosis. Polymorphisms of Fc receptors for IgG (FcgammaR) are associated with modifying effects of several infectious and autoimmune diseases. We have assessed the relationship between polymorphisms in three different FcgammaR genes and coronary artery disease (CAD). We genotyped for the FcgammaRIIA-R/H131, the FcgammaRIIIB-Na1/Na2, and the FcgammaRIIIA-F/V158 polymorphisms in 882 patients undergoing diagnostic coronary angiography. Significant CAD was defined as >/=50% lumen diameter stenosis in at least one coronary artery. In the analysis, no association was found between the FcgammaRIIA and FcgammaRIIIB genotypes and CAD, whereas the FcgammaRIIIA genotype was strongly related. Compared to those being heterozygous, or homozygous for the F allele, patients homozygous for the V allele had significantly reduced risk: OR, 0.53; (CI, 0.32-0.90). Additional adjustment for classical risk factors and sedimentation rate did not affect the results. The V/V genotype was also inversely related to the extent of CAD defined as no CAD, single, double or triple vessel disease (P trend=0.002). Our data provide evidence for an association between FcgammaRIIIA allelic variants and coronary atherosclerosis. Genetic variation in this IgG-receptor may influence the clearance of antibodies by monocyte-derived macrophages involved in the pathogenesis of CAD.

  16. Characterization of the receptors for mycobacterial cord factor in Guinea pig.

    Directory of Open Access Journals (Sweden)

    Kenji Toyonaga

    Full Text Available Guinea pig is a widely used animal for research and development of tuberculosis vaccines, since its pathological disease process is similar to that present in humans. We have previously reported that two C-type lectin receptors, Mincle (macrophage inducible C-type lectin, also called Clec4e and MCL (macrophage C-type lectin, also called Clec4d, recognize the mycobacterial cord factor, trehalose-6,6'-dimycolate (TDM. Here, we characterized the function of the guinea pig homologue of Mincle (gpMincle and MCL (gpMCL. gpMincle directly bound to TDM and transduced an activating signal through ITAM-bearing adaptor molecule, FcRγ. Whereas, gpMCL lacked C-terminus and failed to bind to TDM. mRNA expression of gpMincle was detected in the spleen, lymph nodes and peritoneal macrophages and it was strongly up-regulated upon stimulation of zymosan and TDM. The surface expression of gpMincle was detected on activated macrophages by a newly established monoclonal antibody that also possesses a blocking activity. This antibody potently suppressed TNF production in BCG-infected macrophages. Collectively, gpMincle is the TDM receptor in the guinea pig and TDM-Mincle axis is involved in host immune responses against mycobacteria.

  17. β-adrenergic receptor-dependent alterations in murine cardiac transcript expression are differentially regulated by gefitinib in vivo.

    Directory of Open Access Journals (Sweden)

    Jennifer A Talarico

    Full Text Available β-adrenergic receptor (βAR-mediated transactivation of epidermal growth factor receptor (EGFR has been shown to promote cardioprotection in a mouse model of heart failure and we recently showed that this mechanism leads to enhanced cell survival in part via regulation of apoptotic transcript expression in isolated primary rat neonatal cardiomyocytes. Thus, we hypothesized that this process could regulate cardiac transcript expression in vivo. To comprehensively assess cardiac transcript alterations in response to acute βAR-dependent EGFR transactivation, we performed whole transcriptome analysis of hearts from C57BL/6 mice given i.p. injections of the βAR agonist isoproterenol in the presence or absence of the EGFR antagonist gefitinib for 1 hour. Total cardiac RNA from each treatment group underwent transcriptome analysis, revealing a substantial number of transcripts regulated by each treatment. Gefitinib alone significantly altered the expression of 405 transcripts, while isoproterenol either alone or in conjunction with gefitinib significantly altered 493 and 698 distinct transcripts, respectively. Further statistical analysis was performed, confirming 473 transcripts whose regulation by isoproterenol were significantly altered by gefitinib (isoproterenol-induced up/downregulation antagonized/promoted by gefinitib, including several known to be involved in the regulation of numerous processes including cell death and survival. Thus, βAR-dependent regulation of cardiac transcript expression in vivo can be modulated by the EGFR antagonist gefitinib.

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

    LENUS (Irish Health Repository)

    Gleeson, Eimear M

    2013-07-19

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

  19. The DNA replication licensing factor miniature chromosome maintenance 7 is essential for RNA splicing of epidermal growth factor receptor, c-Met, and platelet-derived growth factor receptor.

    Science.gov (United States)

    Chen, Zhang-Hui; Yu, Yan P; Michalopoulos, George; Nelson, Joel; Luo, Jian-Hua

    2015-01-16

    Miniature chromosome maintenance 7 (MCM7) is an essential component of DNA replication licensing complex. Recent studies indicate that MCM7 is amplified and overexpressed in a variety of human malignancies. In this report, we show that MCM7 binds SF3B3. The binding motif is located in the N terminus (amino acids 221-248) of MCM7. Knockdown of MCM7 or SF3B3 significantly increased unspliced RNA of epidermal growth factor receptor, platelet-derived growth factor receptor, and c-Met. A dramatic drop of reporter gene expression of the oxytocin exon 1-intron-exon 2-EGFP construct was also identified in SF3B3 and MCM7 knockdown PC3 and DU145 cells. The MCM7 or SF3B3 depleted cell extract failed to splice reporter RNA in in vitro RNA splicing analyses. Knockdown of SF3B3 and MCM7 leads to an increase of cell death of both PC3 and DU145 cells. Such cell death induction is partially rescued by expressing spliced c-Met. To our knowledge, this is the first report suggesting that MCM7 is a critical RNA splicing factor, thus giving significant new insight into the oncogenic activity of this protein. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Single-domain antibodies that compete with the natural ligand fibroblast growth factor block the internalization of the fibroblast growth factor receptor 1

    International Nuclear Information System (INIS)

    Highlights: → Recombinant antibodies for FGFR1 were isolated from a llama naive library in VHH format. → These antibodies compete with the natural ligand FGF-2 for the same epitope on FGFR1. → The antibody competition inhibits the FGF-2-dependent internalization of FGFR1. -- Abstract: Single-domain antibodies in VHH format specific for fibroblast growth factor receptor 1 (FGFR1) were isolated from a phage-display llama naive library. In particular, phage elution in the presence of the natural receptor ligand fibroblast growth factor (FGF) allowed for the identification of recombinant antibodies that compete with FGF for the same region on the receptor surface. These antibodies posses a relatively low affinity for FGFR1 and were never identified when unspecific elution conditions favoring highly affine binders were applied to panning procedures. Two populations of competitive antibodies were identified that labeled specifically the receptor-expressing cells in immunofluorescence and recognize distinct epitopes. Antibodies from both populations effectively prevented FGF-dependent internalization and nuclear accumulation of the receptor in cultured cells. This achievement indicates that these antibodies have a capacity to modulate the receptor physiology and, therefore, constitute powerful reagents for basic research and a potential lead for therapeutic applications.

  1. Single-domain antibodies that compete with the natural ligand fibroblast growth factor block the internalization of the fibroblast growth factor receptor 1

    Energy Technology Data Exchange (ETDEWEB)

    Veggiani, Gianluca; Ossolengo, Giuseppe; Aliprandi, Marisa; Cavallaro, Ugo [IFOM-IEO Campus, Via Adamello 16, 20139 Milano (Italy); Marco, Ario de, E-mail: ario.demarco@ung.si [IFOM-IEO Campus, Via Adamello 16, 20139 Milano (Italy); Dept. Environmental Sciences, University of Nova Gorica (UNG), Vipavska 13, P.O. Box 301-SI-5000, Rozna Dolina, Nova Gorica (Slovenia)

    2011-05-20

    Highlights: {yields} Recombinant antibodies for FGFR1 were isolated from a llama naive library in VHH format. {yields} These antibodies compete with the natural ligand FGF-2 for the same epitope on FGFR1. {yields} The antibody competition inhibits the FGF-2-dependent internalization of FGFR1. -- Abstract: Single-domain antibodies in VHH format specific for fibroblast growth factor receptor 1 (FGFR1) were isolated from a phage-display llama naive library. In particular, phage elution in the presence of the natural receptor ligand fibroblast growth factor (FGF) allowed for the identification of recombinant antibodies that compete with FGF for the same region on the receptor surface. These antibodies posses a relatively low affinity for FGFR1 and were never identified when unspecific elution conditions favoring highly affine binders were applied to panning procedures. Two populations of competitive antibodies were identified that labeled specifically the receptor-expressing cells in immunofluorescence and recognize distinct epitopes. Antibodies from both populations effectively prevented FGF-dependent internalization and nuclear accumulation of the receptor in cultured cells. This achievement indicates that these antibodies have a capacity to modulate the receptor physiology and, therefore, constitute powerful reagents for basic research and a potential lead for therapeutic applications.

  2. Transcriptional mechanisms that control expression of the macrophage colony-stimulating factor receptor locus.

    Science.gov (United States)

    Rojo, Rocio; Pridans, Clare; Langlais, David; Hume, David A

    2017-08-15

    The proliferation, differentiation, and survival of cells of the macrophage lineage depends upon signals from the macrophage colony-stimulating factor (CSF) receptor (CSF1R). CSF1R is expressed by embryonic macrophages and induced early in adult hematopoiesis, upon commitment of multipotent progenitors to the myeloid lineage. Transcriptional activation of CSF1R requires interaction between members of the E26 transformation-specific family of transcription factors (Ets) (notably PU.1), C/EBP, RUNX, AP-1/ATF, interferon regulatory factor (IRF), STAT, KLF, REL, FUS/TLS (fused in sarcoma/ranslocated in liposarcoma) families, and conserved regulatory elements within the mouse and human CSF1R locus. One element, the Fms-intronic regulatory element (FIRE), within intron 2, is conserved functionally across all the amniotes. Lineage commitment in multipotent progenitors also requires down-regulation of specific transcription factors such as MYB, FLI1, basic leucine zipper transcriptional factor ATF-like (BATF3), GATA-1, and PAX5 that contribute to differentiation of alternative lineages and repress CSF1R transcription. Many of these transcription factors regulate each other, interact at the protein level, and are themselves downstream targets of CSF1R signaling. Control of CSF1R transcription involves feed-forward and feedback signaling in which CSF1R is both a target and a participant; and dysregulation of CSF1R expression and/or function is associated with numerous pathological conditions. In this review, we describe the regulatory network behind CSF1R expression during differentiation and development of cells of the mononuclear phagocyte system. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  3. Differential regulation of the transcriptional activity of the glucocorticoid receptor through site-specific phosphorylation

    Directory of Open Access Journals (Sweden)

    Raj Kumar

    2008-08-01

    Full Text Available Raj Kumar1, William J Calhoun21Division of Gastroenterology; 2Division of Allergy, Pulmonary, Immunology, Critical Care, and Sleep (APICS, Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX, USAAbstract: Post-translational modifications such as phosphorylation are known to play an important role in the gene regulation by the transcription factors including the nuclear hormone receptor superfamily of which the glucocorticoid receptor (GR is a member. Protein phosphorylation often switches cellular activity from one state to another. Like many other transcription factors, the GR is a phosphoprotein, and phosphorylation plays an important role in the regulation of GR activity. Cell signaling pathways that regulate phosphorylation of the GR and its associated proteins are important determinants of GR function under various physiological conditions. While the role of many phosphorylation sites in the GR is still not fully understood, the role of others is clearer. Several aspects of transcription factor function, including DNA binding affinity, interaction of transactivation domains with the transcription initiation complex, and shuttling between the cytoplasmic compartments, have all been linked to site-specific phosphorylation. All major phosphorylation sites in the human GR are located in the N-terminal domain including the major transactivation domain, AF1. Available literature clearly indicates that many of these potential phosphorylation sites are substrates for multiple kinases, suggesting the potential for a very complex regulatory network. Phosphorylated GR interacts favorably with critical coregulatory proteins and subsequently enhances transcriptional activity. In addition, the activities and specificities of coregulators may be subject to similar regulation by phosphorylation. Regulation of the GR activity due to phosphorylation appears to be site-specific and dependent upon specific cell signaling cascade

  4. The N363S polymorphism of the glucocorticoid receptor and metabolic syndrome factors in men

    DEFF Research Database (Denmark)

    Buemann, Benjamin; Black, Eva; Holst, Claus

    2005-01-01

    OBJECTIVE: To test the associations between the N363S polymorphism of the glucocorticoid receptor gene (NR3C1) and factors related to the metabolic syndrome in middle-aged men with and without juvenile-onset obesity. RESEARCH METHODS AND PROCEDURES: This study included two groups of middle-aged men...... was calculated as a proxy for insulin sensitivity. Serum triglycerides and total and high-density lipoprotein-cholesterol were measured in the fasting state. RESULTS: Among the men with juvenile-onset obesity, carriers (n = 17) of the 363S allele had a lower whole body fat percentage, after accounting...... for differences in BMI and higher Matsudas index, compared with the noncarriers. The difference in Matsudas index lost statistical significance after the difference in body fat was accounted for. In the randomly sampled men, these variables did not relate to genotype. No relationship between carriers...

  5. Mutation of the KIT (mast/stem cell growth factor receptor) protooncogene in human piebaldism

    Energy Technology Data Exchange (ETDEWEB)

    Giebel, L.B.; Spritz, R.A. (Univ .of Wisconsin, Madison (United States))

    1991-10-01

    Piebaldism is an autosomal dominant genetic disorder characterized by congenital patches of skin and hair from which melanocytes are completely absent. A similar disorder of mouse, dominant white spotting (W), results from mutations of the c-Kit protooncogene, which encodes the receptor for mast/stem cell growth factor. The authors identified a KIT gene mutation in a proband with classic autosomal dominant piebaldism. This mutation results in a Gly {yields} Arg substitution at codon 664, within the tyrosine kinase domain. This substitution was not seen in any normal individuals and was completely linked to the piebald phenotype in the proband's family. Piebaldism in this family thus appears to be the human homologue to dominant white spotting (W) of the mouse.

  6. Expression of epithelial growth factor receptor in oral epithelial dysplastic lesions.

    Science.gov (United States)

    Rajeswari, Manickam Rajakili Chockalingam; Saraswathi, Thillai Rajasekaran

    2012-05-01

    The aim of the study was to assess the expression of epithelial growth factor receptor (EGFR) in normal oral mucosa and varying grades of oral epithelial dysplasia (OED) and to correlate these findings, with clinicopathologic features and findings on routine hematoxylin and eosin-stained (H and E) sections. Twenty-nine formalin-fixed, paraffin-embedded blocks of various grades of OED and 10 normal mucosa were stained with routine H and E and immunostained with EGFR by avidin-biotin method. The results showed a significant increase in the staining reactions in varying grades of dysplasia as compared with normal mucosa. EGFR can be considered as an early marker of a cell proliferation and maturation as well as early marker of epithelial dysplasia and onset of cancer in oral dysplasia. However, further studies with a larger sample size and continuous followup is suggested to determine its role and significance precisely.

  7. Lack of the type III epidermal growth factor receptor mutation in colorectal cancer

    DEFF Research Database (Denmark)

    Spindler, Karen-Lise Garm; Olsen, Dorte A.; Nielsen, Jens Nederby

    2006-01-01

    BACKGROUND: Epidermal growth factor receptor (EGFR) analysis by traditional immunohistochemistry does not provide clinicians with a reliable tool for the selection of patients to EGFR-targeted treatment in colorectal cancer (CRC). Alternative methods and further understanding of the EGFR signaling...... therapy. The aim of the present study was to investigate the presence of EGFRvIII in CRC by PCR and protein analysis. MATERIALS AND METHODS: The study included 79 colorectal cancer patients for PCR analysis and 50 patients for protein analysis by Western blots, in two different laboratories. RESULTS......: No type III mutations were detected in our material. CONCLUSION: The EGFRvIII mutations are rare in colorectal adenocarcinomas and overall probability does not appear to contribute to the malignant phenotype of this disease....

  8. A tale of the epidermal growth factor receptor: The quest for structural resolution on cells.

    Science.gov (United States)

    Tynan, Christopher J; Lo Schiavo, Valentina; Zanetti-Domingues, Laura; Needham, Sarah R; Roberts, Selene K; Hirsch, Michael; Rolfe, Daniel J; Korovesis, Dimitrios; Clarke, David T; Martin-Fernandez, Marisa L

    2016-02-15

    The challenge of determining the architecture and geometry of oligomers of the epidermal growth factor receptor (EGFR) on the cell surface has been approached using a variety of biochemical and biophysical methods. This review is intended to provide a narrative of how key concepts in the field of EGFR research have evolved over the years, from the origins of the prevalent EGFR signalling dimer hypothesis through to the development and implementation of methods that are now challenging the conventional view. The synergy between X-ray crystallography and cellular fluorescence microscopy has become particularly important, precisely because the results from these two methods diverged and highlighted the complexity of the challenge. We illustrate how developments in super-resolution microscopy are now bridging this gap. Exciting times lie ahead where knowledge of the nature of the complexes can assist with the development of a new generation of anti-cancer drugs. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Epidermal growth factor receptor is a preferred target for treating amyloid-β-induced memory loss.

    Science.gov (United States)

    Wang, Lei; Chiang, Hsueh-Cheng; Wu, Wenjuan; Liang, Bin; Xie, Zuolei; Yao, Xinsheng; Ma, Weiwei; Du, Shuwen; Zhong, Yi

    2012-10-09

    Current understanding of amyloid-β (Aβ) metabolism and toxicity provides an extensive list of potential targets for developing drugs for treating Alzheimer's disease. We took two independent approaches, including synaptic-plasticity-based analysis and behavioral screening of synthetic compounds, for identifying single compounds that are capable of rescuing the Aβ-induced memory loss in both transgenic fruit fly and transgenic mouse models. Two clinically available drugs and three synthetic compounds not only showed positive effects in behavioral tests but also antagonized the Aβ oligomers-induced activation of the epidermal growth factor receptor (EGFR). Such surprising converging outcomes from two parallel approaches lead us to conclude that EGFR is a preferred target for treating Aβ-induced memory loss.

  10. Transforming growth factor β receptor 1 is a new candidate prognostic biomarker after acute myocardial infarction

    Directory of Open Access Journals (Sweden)

    Devaux Yvan

    2011-12-01

    Full Text Available Abstract Background Prediction of left ventricular (LV remodeling after acute myocardial infarction (MI is clinically important and would benefit from the discovery of new biomarkers. Methods Blood samples were obtained upon admission in patients with acute ST-elevation MI who underwent primary percutaneous coronary intervention. Messenger RNA was extracted from whole blood cells. LV function was evaluated by echocardiography at 4-months. Results In a test cohort of 32 MI patients, integrated analysis of microarrays with a network of protein-protein interactions identified subgroups of genes which predicted LV dysfunction (ejection fraction ≤ 40% with areas under the receiver operating characteristic curve (AUC above 0.80. Candidate genes included transforming growth factor beta receptor 1 (TGFBR1. In a validation cohort of 115 MI patients, TGBFR1 was up-regulated in patients with LV dysfunction (P Conclusions We identified TGFBR1 as a new candidate prognostic biomarker after acute MI.

  11. Anticipating and managing the cutaneous side effects of epidermal growth factor receptor inhibitors.

    Science.gov (United States)

    Sinclair, Rod

    2014-03-01

    Epidermal growth factor receptor inhibitor (EGFRi) therapy for tumor suppression produces significant cutaneous toxicity that may necessitate dose reduction or treatment cessation. This manuscript aims to describe the skin toxicity associated with these agents and equip oncologists with the tools to best manage these patients. A literature review of skin toxicity associated with EGFRi was conducted by a clinical dermatologist experienced in the management of these patients and a management strategy developed for EGFRi skin toxicity The sequence of development of EGFRi cutaneous toxicity is predictable. Many skin side effects can be ameliorated by prophylactic therapy. Management of established skin toxicity is complex and needs to be continued throughout the period of EGFRi drug treatment. Early referral to a dermatologist experienced in the management of these patients is recommended. © 2014 Wiley Publishing Asia Pty Ltd.

  12. 99m Tc-anti-epidermal growth factor receptor nanobody for tumor imaging.

    Science.gov (United States)

    Piramoon, Majid; Hosseinimehr, Seyed Jalal; Omidfar, Kobra; Noaparast, Zohreh; Abedi, Seyed Mohammad

    2017-04-01

    Nanobodies are important biomolecules for tumor targeting. In this study, we synthesized and labeled anti-epidermal growth factor receptor (EGFR) nanobody OA-cb6 with 99m Tc(CO) 3 + and evaluated its characteristics for targeting the EGFR in the A431 human epidermal carcinoma cell line. Nanobody radiolabeling was achieved with high yield and radiochemical purity, and the radioconjugate was stable. Biodistribution results in nude mice exhibited a favorable tumor-to-muscle ratio at 4-hr postinjection, and tumor location was visualized at 4 hr after injection of radiolabeled nanobody. Our result showed that the OA-cb6- 99m Tc-tricarbonyl radiolabeled nanobody is a promising radiolabeled biomolecule for tumor imaging in cancers with high EGFR overexpression. © 2016 John Wiley & Sons A/S.

  13. Vascular Endothelial Growth Factor Receptor 3 Controls Neural Stem Cell Activation in Mice and Humans

    Directory of Open Access Journals (Sweden)

    Jinah Han

    2015-02-01

    Full Text Available Neural stem cells (NSCs continuously produce new neurons within the adult mammalian hippocampus. NSCs are typically quiescent but activated to self-renew or differentiate into neural progenitor cells. The molecular mechanisms of NSC activation remain poorly understood. Here, we show that adult hippocampal NSCs express vascular endothelial growth factor receptor (VEGFR 3 and its ligand VEGF-C, which activates quiescent NSCs to enter the cell cycle and generate progenitor cells. Hippocampal NSC activation and neurogenesis are impaired by conditional deletion of Vegfr3 in NSCs. Functionally, this is associated with compromised NSC activation in response to VEGF-C and physical activity. In NSCs derived from human embryonic stem cells (hESCs, VEGF-C/VEGFR3 mediates intracellular activation of AKT and ERK pathways that control cell fate and proliferation. These findings identify VEGF-C/VEGFR3 signaling as a specific regulator of NSC activation and neurogenesis in mammals.

  14. Effect of angiotensin II type 1 receptor blocker and angiotensin converting enzyme inhibitor on the intraocular growth factors and their receptors in streptozotocin-induced diabetic rats

    Directory of Open Access Journals (Sweden)

    Ik Soo Byon

    2017-06-01

    Full Text Available AIM: To investigate the effect of angiotensin II type 1 receptor blocker (ARB and angiotensin converting enzyme inhibitor (ACEI on intraocular growth factors and their receptors in streptozotocin-induced diabetic rats. METHODS: Forty Sprague-Dawley rats were divided into 4 groups: control, diabetes mellitus (DM, candesartan-treated DM, and enalapril-treated DM (each group, n=10. After the induction of DM by streptozotocin, candesartan [ARB, 5 mg/(kg·d] and enalapril [ACEI, 10 mg/(kg·d] were administered to rats orally for 4wk. Vascular endothelial growth factor (VEGF and angiotensin II (Ang II concentrations in the vitreous were measured using enzyme-linked immunosorbent assays, and VEGF receptor 2 and angiotensin II type 1 receptor (AT1R levels were assessed at week 4 by Western blotting. RESULTS: Vitreous Ang II levels were significantly higher in the DM group and candesartan-treated DM group than in the control (P=0.04 and 0.005, respectively. Vitreous AT1R increased significantly in DM compared to the other three groups (P<0.007. Candesartan-treated DM rats showed higher vitreal AT1R concentration than the enalapril-treated DM group and control (P<0.001 and P=0.005, respectively. No difference in vitreous Ang II and AT1R concentration was found between the enalapril-treated DM group and control. VEGF and its receptor were below the minimum detection limit in all 4 groups. CONCLUSION: Increased Ang II and AT1R in the hyperglycemic state indicate activated the intraocular renin-angiotensin system, which is inhibited more effectively by systemic ACEI than systemic ARB.

  15. Mechanisms of Inhibition of the Epidermal Growth Factor Receptor: Implications for Novel Anti-Cancer Therapies

    National Research Council Canada - National Science Library

    Klein, Daryl E

    2005-01-01

    .... No secreted or extracellular ErbB receptor inhibitors have been reported in mammals. However, two natural inhibitors of the highly homologous Drosophila EGF receptor are found in Drosophila melanogaster...

  16. Epidermal Growth Factor Receptor activation promotes ADA3 acetylation through the AKT-p300 pathway.

    Science.gov (United States)

    Srivastava, Shashank; Mohibi, Shakur; Mirza, Sameer; Band, Hamid; Band, Vimla

    2017-08-18

    The ADA3 (Alteration/Deficiency in Activation 3) protein is an essential adaptor component of several Lysine Acetyltransferase (KAT) complexes involved in chromatin modifications. Previously, we and others have demonstrated a crucial role of ADA3 in cell cycle progression and in maintenance of genomic stability. Recently, we have shown that acetylation of ADA3 is key to its role in cell cycle progression. Here, we demonstrate that AKT activation downstream of Epidermal Growth Factor Receptor (EGFR) family proteins stimulation leads to phosphorylation of p300, which in turn promotes the acetylation of ADA3. Inhibition of upstream receptor tyrosine kinases (RTKs), HER1 (EGFR)/HER2 by lapatinib and the accompanying reduction of phospho-AKT levels led to a decrease in p300 phosphorylation and ADA3 protein levels. The p300/PCAF inhibitor garcinol also destabilized the ADA3 protein in a proteasome-dependent manner and an ADA3 mutant with K→R mutations exhibited a marked increase in half-life, consistent with opposite role of acetylation and ubiquitination of ADA3 on shared lysine residues. ADA3 knockdown led to cell cycle inhibitory effects, as well as apoptosis similar to those induced by lapatinib treatment of HER2+ breast cancer cells, as seen by accumulation of CDK inhibitor p27, reduction in mitotic marker pH3(S10), and a decrease in the S-phase marker PCNA, as well as the appearance of cleaved PARP. Taken together our results reveal a novel RTK-AKT-p300-ADA3 signaling pathway involved in growth factor-induced cell cycle progression.

  17. Retinal hypoxia induces vascular endothelial growth factor through induction of estrogen-related receptor γ

    Energy Technology Data Exchange (ETDEWEB)

    Do, Ji Yeon; Choi, Young Keun [Leading-edge Research Center for Drug Discovery and Development for Diabetes and Metabolic Disease, Kyungpook National University School of Medicine, Kyungpook National University, Daegu (Korea, Republic of); Kook, Hyun [Department of Pharmacology, Chonnam National University Medical School, Gwangju (Korea, Republic of); Suk, Kyoungho [Department of Pharmacology, Brain Science & Engineering Institute, Kyungpook National University School of Medicine, Daegu (Korea, Republic of); Lee, In-Kyu [Leading-edge Research Center for Drug Discovery and Development for Diabetes and Metabolic Disease, Kyungpook National University School of Medicine, Kyungpook National University, Daegu (Korea, Republic of); Division of Endocrinology and Metabolism, Department of Internal Medicine, Research Institute of Aging and Metabolism, Kyungpook National University School of Medicine, Daegu (Korea, Republic of); Park, Dong Ho, E-mail: sarasate2222@gmail.com [Department of Ophthalmology, Kyungpook National University School of Medicine, Daegu (Korea, Republic of)

    2015-05-01

    Ischemic retinopathies causing overexpression of pro-angiogenic factors, including vascular endothelial growth factor (VEGF), are the most common cause of blindness. Thus, understanding the pathophysiology of targetable pathways that regulate retinal VEGF is of great interest. A conserved binding site for estrogen-related receptor γ (ERRγ) has been identified in the promoter of the Vegfa gene. ERRγ is a constitutively active orphan nuclear receptor and its expression is increased by hypoxic stimuli in metabolically active tissues. This study evaluated the role of ERRγ in the ischemic retina and the anti-VEGF potential of GSK5182, a selective inverse agonist of ERRγ. In an oxygen-induced retinopathy (OIR) mouse model, immunohistochemistry showed significantly increased ERRγ expression in the ganglion cell layer at postnatal day (P) 17. In a ganglion cell line (RGC-5), mRNA and protein levels of ERRγ were increased by desferrioxamine treatment and hypoxic conditions (1% O{sub 2}). Transient transfection of RGC-5 cells revealed that ERRγ regulated Vegfa expression and this was inhibited by GSK5182. Intravitreal injection of GSK5182 into the OIR model at P14 inhibited retinal Vegfa mRNA expression at P17. GSK5182 suppresses hypoxia-induced VEGF expression via ERRγ; therefore, ERRγ could be a treatment target for ischemic retinopathies. - Highlights: • OIR mice exhibited increased ERRγ expression in the ganglion cell layer. • Hypoxia-induced ERRγ expression was observed in retinal ganglion cells. • ERRγ overexpression increased VEGFA expression in retinal ganglion cells. • An ERRγ inverse agonist suppressed VEGFA expression in retinal ganglion cells. • Intravitreal injection of an ERRγ inverse agonist suppressed VEGFA in OIR mice.

  18. Stage specific requirement of platelet-derived growth factor receptor-α in embryonic development.

    Science.gov (United States)

    Qian, Chen; Wong, Carol Wing Yan; Wu, Zhongluan; He, Qiuming; Xia, Huimin; Tam, Paul Kwong Hang; Wong, Kenneth Kak Yuen; Lui, Vincent Chi Hang

    2017-01-01

    Platelet-derived growth factor receptor alpha (PDGFRα) is a cell-surface receptor tyrosine kinase for platelet-derived growth factors. Correct timing and level of Pdgfra expression is crucial for embryo development, and deletion of Pdgfra caused developmental defects of multiple endoderm and mesoderm derived structures, resulting in a complex phenotypes including orofacial cleft, spina bifida, rib deformities, and omphalocele in mice. However, it is not clear if deletion of Pdgfra at different embryonic stages differentially affects these structures. To address the temporal requirement of Pdgfra in embryonic development. We have deleted the Pdgfra in Pdgfra-expressing tissues at different embryonic stages in mice, examined and quantified the developmental anomalies. Current study showed that (i) conditional deletion of Pdgfra at different embryonic days (between E7.5 and E10.5) resulted in orofacial cleft, spina bifida, rib cage deformities, and omphalocele, and (ii) the day of Pdgfra deletion influenced the combinations, incidence and severities of these anomalies. Deletion of Pdgfra caused apoptosis of Pdgfra-expressing tissues, and developmental defects of their derivatives. Orofacial cleft, spina bifida and omphalocele are among the commonest skeletal and abdominal wall defects of newborns, but their genetic etiologies are largely unknown. The remarkable resemblance of our conditional Pdgfra knockout embryos to theses human congenital anomalies, suggesting that dysregulated PDGFRA expression could cause these anomalies in human. Future work should aim at defining (a) the regulatory elements for the expression of the human PDGFRA during embryonic development, and (b) if mutations / sequence variations of these regulatory elements cause these anomalies.

  19. Effects of radiation on the epidermal growth factor receptor pathway in the heart.

    Science.gov (United States)

    Sridharan, Vijayalakshmi; Sharma, Sunil K; Moros, Eduardo G; Corry, Peter M; Tripathi, Preeti; Lieblong, Benjamin J; Guha, Chandan; Hauer-Jensen, Martin; Boerma, Marjan

    2013-07-01

    Radiation-induced heart disease (RIHD) is a serious side-effect of thoracic radiotherapy. The epidermal growth factor receptor (EGFR) pathway is essential for the function and survival of cardiomyocytes. Hence, agents that target the EGFR pathway are cardiotoxic. Tocotrienols protect from radiation injury, but may also enhance the therapeutic effects of EGFR pathway inhibitors in cancer treatment. This study investigated the effects of local irradiation on the EGFR pathway in the heart and tests whether tocotrienols may modify radiation-induced changes in this pathway. Male Sprague-Dawley rats received image-guided localized heart irradiation with 21 Gy. Twenty four hours before irradiation, rats received a single dose of tocotrienol-enriched formulation or vehicle by oral gavage. At time points from 2 h to 9 months after irradiation, left ventricular expression of EGFR pathway mediators was studied. Irradiation caused a decrease in the expression of epidermal growth factor (EGF) and neuregulin-1 (Nrg-1) mRNA from 6 h up to 10 weeks, followed by an upregulation of these ligands and the receptor erythroblastic leukemia viral oncogene homolog (ErbB)4 at 6 months. In addition, the upregulation of Nrg-1 was statistically significant up to 9 months after irradiation. A long-term upregulation of ErbB2 protein did not coincide with changes in transcription or post-translational interaction with the chaperone heat shock protein 90 (HSP90). Pretreatment with tocotrienols prevented radiation-induced changes at 2 weeks. Local heart irradiation causes long-term changes in the EGFR pathway. Studies have to address how radiation may interact with cardiotoxic effects of EGFR inhibitors.

  20. Quantitative super-resolution single molecule microscopy dataset of YFP-tagged growth factor receptors.

    Science.gov (United States)

    Lukeš, Tomáš; Pospíšil, Jakub; Fliegel, Karel; Lasser, Theo; Hagen, Guy M

    2018-01-19

    Super-resolution single molecule localization microscopy (SMLM) is a method for achieving resolution beyond the classical limit in optical microscopes (approx. 200 nm laterally). Yellow fluorescent protein (YFP) has been used for super-resolution single molecule localization microscopy, but less frequently than other fluorescent probes. Working with YFP in SMLM is a challenge because a lower number of photons are emitted per molecule compared to organic dyes which are more commonly used. Publically available experimental data can facilitate development of new data analysis algorithms. Four complete, freely available single molecule super-resolution microscopy datasets on YFP-tagged growth factor receptors expressed in a human cell line are presented including both raw and analyzed data. We report methods for sample preparation, for data acquisition, and for data analysis, as well as examples of the acquired images. We also analyzed the SMLM data sets using a different method: super-resolution optical fluctuation imaging (SOFI). The two modes of analysis offer complementary information about the sample. A fifth single molecule super-resolution microscopy dataset acquired with the dye Alexa 532 is included for comparison purposes. This dataset has potential for extensive reuse. Complete raw data from SMLM experiments has typically not been published. The YFP data exhibits low signal to noise ratios, making data analysis a challenge. These data sets will be useful to investigators developing their own algorithms for SMLM, SOFI, and related methods. The data will also be useful for researchers investigating growth factor receptors such as ErbB3. © The Author(s) 2018. Published by Oxford University Press.

  1. Gene silencing for epidermal growth factor receptor variant III induces cell-specific cytotoxicity.

    Science.gov (United States)

    Yamoutpour, Farnaz; Bodempudi, Vidya; Park, Shay E; Pan, Weihong; Mauzy, Mary Jean; Kratzke, Robert A; Dudek, Arkadiusz; Potter, David A; Woo, Richard A; O'Rourke, Donald M; Tindall, Donald J; Farassati, Faris

    2008-11-01

    Epidermal growth factor receptor variant III (EGFRvIII) is a constitutively active mutant form of EGFR that is expressed in 40% to 50% of gliomas and several other malignancies. Here, we describe the therapeutic effects of silencing EGFRvIII on glioma cell lines in vitro and in vivo. A small interfering RNA molecule against EGFRvIII was introduced into EGFRvIII-expressing glioma cells (U87Delta) by electroporation resulting in complete inhibition of expression of EGFRvIII as early as 48 h post-treatment. During EGFRvIII silencing, a decrease in the proliferation and invasiveness of U87Delta cells was accompanied by an increase in apoptosis (P < 0.05). Notably, EGFRvIII silencing inhibited the signal transduction machinery downstream of EGFRvIII as evidenced by decreases in the activated levels of Ras and extracellular signal-regulated kinase. A lentivirus capable of expressing anti-EGFRvIII short hairpin RNA was also able to achieve progressive silencing of EGFRvIII in U87Delta cells in addition to inhibiting cell proliferation, invasiveness, and colony formation in a significant manner (P < 0.05). Silencing EGFRvIII in U87Delta cultures with this virus reduced the expression of factors involved in epithelial-mesenchymal transition including N-cadherin, beta-catenin, Snail, Slug, and paxillin but not E-cadherin. The anti-EGFRvIII lentivirus also affected the cell cycle progression of U87Delta cells with a decrease in G(1) and increase in S and G(2) fractions. In an in vivo model, tumor growth was completely inhibited in severe combined immunodeficient mice (n = 10) injected s.c. with U87Delta cells treated with the anti-EGFRvIII lentivirus (P = 0.005). We conclude that gene specific silencing of EGFRvIII is a promising strategy for treating cancers that contain this mutated receptor.

  2. Genomic organization and chromosomal localization of the human and mouse genes encoding the {alpha} receptor component for ciliary neurotrophic factor

    Energy Technology Data Exchange (ETDEWEB)

    Valenzuela, D.M.; Rojas, E.; McClain, J. [Regeneron Pharmaceuticals, Inc., Tarrytown, NY (United States)] [and others

    1995-01-01

    Ciliary neurotrophic factor (CNTF) has recently been found to share receptor components with, and to be structurally related to, a family of broadly acting cytokines, including interleukin-6, leukemia inhibitory factor, and oncostatin M. However, the CNTF receptor complex also includes a CNTF-specific component known as CNTF receptor {alpha} (CNTFR{alpha}). Here we describe the molecular cloning of the human and mouse genes encoding CNTFR. We report that the human and mouse genes have an identical intron-exon structure that correlates well with the domain structure of CNTFR{alpha}. That is, the signal peptide and the immunoglobulin-like domain are each encoded by single exons, the cytokine receptor-like domain is distributed among 4 exons, and the C-terminal glycosyl phosphatidylinositol recognition domain in encoded by the final coding exon. The position of the introns within the cytokine receptor-like domain corresponds to those found in other members of the cytokine receptor superfamily. Confirming a recent study using radiation hybrids, we have also mapped the human CNTFR gene to chromosome band 9p13 and the mouse gene to a syntenic region of chromosome 4. 24 refs., 4 figs.

  3. Differential expression of the insulin-like growth factor receptor among early breast cancer subtypes.

    Directory of Open Access Journals (Sweden)

    Giannis Mountzios

    Full Text Available INTRODUCTION: We sought to determine the level of protein expression of the critical components of the insulin-like growth factor receptor (IGFR pathway and to evaluate their prognostic significance across the different early breast cancer subtypes. PATIENTS AND METHODS: Archival tumor tissue from 1,021 women with early, node positive breast cancer, who were prospectively evaluated within two randomized clinical trials, was used to construct tissue microarrays that were stained for hormone receptors (HR, Ki67, HER2, epidermal growth factor receptor (EGFR and cytokeratins 5/6, to classify tumors into five immunophenotypical subgroups. Immunohistochemical (IHC expression of IGF1R-alpha and beta subunits, IGF2R and IGF-binding protein 2 (IGFBP2 was assessed using the immunoreactive score (IRS. Repeated internal cross-validation was performed to examine the statistical validity of the cut off points for all biomarkers. RESULTS: After a median follow-up time of 105.4 months, overall 370 women (36.2% had relapsed and 270 (26.4% had died. Tumors expressing IGF1R-alpha above the median IRS were significantly more frequently HR positive (luminal A+B+HER2, as compared to HER2-enriched and triple negative ones (p<0.001 for both comparisons. IGF2R was overexpressed significantly more frequently in HR negative tumors (p = 0.001 and had an inverse correlation with all other biomarkers. Patients with luminal A and B tumors with high IGF1R-alpha and negative EGFR expression (N = 190 had significantly higher 4-year survival rates, as compared to the rest (log-rank p = 0.046, as did patients with luminal A and B tumors with high IGF1R-alpha and low IGF2R expression, as compared to the rest (N = 91, (log-rank p = 0.035. After adjustment for significant variables, patients in the latter group had a relative 45% reduction in the risk of death, as compared to the rest (p = 0.035. CONCLUSION: Aberrant expression of components of the IGF1R

  4. Estrogen receptor positive breast cancers and their association with environmental factors

    Directory of Open Access Journals (Sweden)

    Mannel Sylvio

    2011-05-01

    Full Text Available Abstract Background Epidemiological studies to assess risk factors for breast cancer often do not differentiate between different types of breast cancers. We applied a general linear model to determine whether data from the Surveillance, Epidemiology, and End Results Program on annual county level age-adjusted incidence rates of breast cancer with and without estrogen receptors (ER+ and ER- were associated with environmental pollutants. Results Our final model explained approximately 38% of the variation in the rate of ER+ breast cancer. In contrast, we were only able to explain 14% of the variation in the rate of ER- breast cancer with the same set of environmental variables. Only ER+ breast cancers were positively associated with the EPA's estimated risk of cancer based on toxic air emissions and the proportion of agricultural land in a county. Meteorological variables, including short wave radiation, temperature, precipitation, and water vapor pressure, were also significantly associated with the rate of ER+ breast cancer, after controlling for age, race, premature mortality from heart disease, and unemployment rate. Conclusions Our findings were consistent with what we expected, given the fact that many of the commonly used pesticides and air pollutants included in the EPA cancer risk score are classified as endocrine disruptors and ER+ breast cancers respond more strongly to estrogen than ER- breast cancers. The findings of this study suggest that ER+ and ER- breast cancers have different risk factors, which should be taken into consideration in future studies that seek to understand environmental risk factors for breast cancer.

  5. Frequency of Epidermal Growth Factor Receptor Mutation in Smokers with Lung Cancer Without Pulmonary Emphysema.

    Science.gov (United States)

    Takeda, Kenichi; Yamasaki, Akira; Igishi, Tadashi; Kawasaki, Yuji; Ito-Nishii, Shizuka; Izumi, Hiroki; Sakamoto, Tomohiro; Touge, Hirokazu; Kodani, Masahiro; Makino, Haruhiko; Yanai, Masaaki; Tanaka, Natsumi; Matsumoto, Shingo; Araki, Kunio; Nakamura, Hiroshige; Shimizu, Eiji

    2017-02-01

    Chronic obstructive pulmonary disease is a smoking-related disease, and is categorized into the emphysema and airway dominant phenotypes. We examined the relationship between emphysematous changes and epidermal growth factor receptor (EGFR) mutation status in patients with lung adenocarcinoma. The medical records for 250 patients with lung adenocarcinoma were retrospectively reviewed. All patients were categorized into the emphysema or non-emphysema group. Wild-type EGFR was detected in 136 (54%) and mutant EGFR in 48 (19%). Emphysematous changes were observed in 87 (36%) patients. EGFR mutation was highly frequent in the non-emphysema group (p=0.0014). Multivariate logistic regression analysis showed that emphysema was an independent risk factor for reduced frequency of EGFR mutation (Odds Ratio=3.47, p=0.005). Our data showed a relationship between emphysematous changes and EGFR mutation status. There might be mutually exclusive genetic risk factors for carcinogenesis and development of emphysematous changes. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  6. Differences in human skin between the epidermal growth factor receptor distribution detected by EGF binding and monoclonal antibody recognition

    DEFF Research Database (Denmark)

    Green, M R; Couchman, J R

    1985-01-01

    Two methods have been used to examine epidermal growth factor (EGF) receptor distribution in human scalp and foreskin. The first employed [125I]EGF viable explants and autoradiography to determine the EGF binding pattern while the second used a monoclonal antibody to the human EGF receptor to map...... the distribution on frozen skin sections of an extracellular epitope on the EGF receptor. The [125I]EGF binding experiments showed accessible, unoccupied EGF receptors to be present on the epidermal basal cells (with reduced binding to spinous cells), the basal cells of the hair shaft and sebaceous gland......, the eccrine sweat glands, capillary system, and the hair follicle outer root sheath, generally similar in pattern to that previously reported for full-thickness rat skin and human epidermis. The same areas also bound EGF-R1 but in addition the monoclonal antibody recognized a cone of melanin containing...

  7. Structure and function of the Juxta membrane domain of the human epidermal growth factor receptor by NMR spectroscopy

    International Nuclear Information System (INIS)

    Choowongkomon, Kiattawee; Carlin, Cathleen; Sonnichsen, Frank D.

    2005-10-01

    The epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase family involved in the regulation of cellular proliferation and differentiation. Its juxta membrane domain (JX), the region located between the transmembrane and kinase domains, plays important roles in receptor trafficking since both basolateral sorting in polarized epithelial cells and lysosomal sorting signals are identified in this region. In order to understand the regulation of these signals, we characterized the structural properties of recombinant JX domain in dodecyl phosphocholine detergent (DPC) by nuclear magnetic resonance (NMR) spectroscopy. In DPC micelles, structures derived from NMR data showed three amphipathic, helical segments. Two equivalent average structural models on the surface of micelles were obtained that differ only in the relative orientation between the first and second helices. Our data suggests that the activity of sorting signals may be regulated by their membrane association and restricted accessibility in the intact receptor

  8. Epidermal growth factor treatment of A431 cells alters the binding capacity and electrophoretic mobility of the cytoskeletally associated epidermal growth factor receptor

    International Nuclear Information System (INIS)

    Roy, L.M.; Gittinger, C.K.; Landreth, G.E.

    1991-01-01

    Epidermal growth factor receptor interacts with structural elements of A431 cells and remains associated with the cytoskeleton following extraction with nonionic detergents. Extraction of cells with 0.15% Triton X-100 resulted in detection of only approximately 40% of the EGF binding sites on the cytoskeleton. If the cells were exposed to EGF prior to extraction, approximately twofold higher levels of low-affinity EGF binding sites were detected. The difference in number of EGF binding sites was not a consequence of differences in numbers of EGF receptors associated with the cytoskeleton; equal amounts of 35S-labeled receptor were immunoprecipitated from the cytoskeletons of both control and EGF-treated cells. The effect of EGF pretreatment on binding activity was coincident with a change in the mobility of the receptor from a doublet of Mr approximately 160-180 kDa to a single sharp band at 180 kDa. The alteration in receptor mobility was not a simple consequence of receptor phosphorylation in that the alteration was not reversed by alkaline phosphatase treatment, nor was the shift produced by treatment of the cells with phorbol ester. The two EGF receptor species demonstrated differential susceptibility to V8 proteinase digestion. The EGF-induced 180 kDa species was preferentially digested by the proteinase relative to the 160 kDa species, indicating that EGF binding results in a conformational change in the receptor. The EGF-mediated preservation of binding activity and altered conformation may be related to receptor oligomerization

  9. An improved reprogrammable mouse model harbouring the reverse tetracycline-controlled transcriptional transactivator 3

    Directory of Open Access Journals (Sweden)

    S. Alaei

    2016-07-01

    Full Text Available Reprogrammable mouse models engineered to conditionally express Oct-4, Klf-4, Sox-2 and c-Myc (OKSM have been instrumental in dissecting molecular events underpinning the generation of induced pluripotent stem cells. However, until now these models have been reported in the context of the m2 reverse tetracycline-controlled transactivator, which results in low reprogramming efficiency and consequently limits the number of reprogramming intermediates that can be isolated for downstream profiling. Here, we describe an improved OKSM mouse model in the context of the reverse tetracycline-controlled transactivator 3 with enhanced reprogramming efficiency (>9-fold and increased numbers of reprogramming intermediate cells albeit with similar kinetics, which we believe will facilitate mechanistic studies of the reprogramming process.

  10. Epidermal growth factor receptor amplification does not have prognostic significance in patients with glioblastoma multiforme

    International Nuclear Information System (INIS)

    Quan, Aimee L.; Barnett, Gene H.; Lee, S.-Y.; Vogelbaum, Michael A.; Toms, Steven A.; Staugaitis, Susan M.; Prayson, Richard A.; Peereboom, David M.; Stevens, Glen H.J.; Cohen, Bruce H.; Suh, John H.

    2005-01-01

    Purpose: There have been conflicting reports in the literature regarding the prognostic significance of epidermal growth factor receptor (EGFR) amplification in patients with glioblastoma multiforme (GBM). The purpose of this study is to determine the prognostic significance of EGFR amplification in patients with GBM treated at Cleveland Clinic Foundation. Methods and Materials: A retrospective review of GBM patients treated with surgery at Cleveland Clinic Foundation was performed. Amplification of EGFR was evaluated with fluorescence in situ hybridization in a total of 107 patients diagnosed between December 1995 and May 2003. In addition to EGFR status, various prognostic factors were evaluated to determine the factors that influenced survival and radiographic response rate. The median follow-up was 9 months. Results: The overall median survival was 9.8 months, with a 1-year survival of 40%. Of the 107 patients in whom EGFR status was evaluated, 36 (33.6%) were found to have EGFR amplification. On multivariate analysis, median survival was found to be significantly improved for patients with age <60 (12.6 months vs. 8 months, p = 0.0061), patients with Karnofsky Performance Status ≥70 (12.1 months vs. 4.4 months, p < 0.0001), patients who had undergone subtotal resection or gross total resection (11.1 months vs. 4.1 months, p = 0.002), and patients who received a radiation dose ≥60 Gy compared with no radiation (12.7 months vs. 3 months, p < 0.0001). There was no association of EGFR amplification with survival. When stratified by age (<60 vs. ≥60), EGFR status still did not reach statistical significance in predicting for survival. For the 81 patients who had radiographic follow-up, the 1-year overall local control was 14%. On univariate analysis, only treatment with radiation (<60 Gy vs. ≥60 Gy vs. no radiation, p = 0.03) was found to predict for improved local control. Treatment with radiation did not remain statistically significant on multivariate

  11. Adenosine Receptor Activation in the “Trigger” Limb of Remote Pre-Conditioning Mediates Human Endothelial Conditioning and Release of Circulating Cardioprotective Factor(s

    Directory of Open Access Journals (Sweden)

    Hussain Contractor, MBChB, DPhil

    2016-10-01

    Full Text Available Remote ischemic pre-conditioning (rIPC has emerged as a potential mechanism to reduce ischemia-reperfusion injury. Clinical data, however, have been mixed, and its physiological basis remains unclear, although it appears to involve release of circulating factor(s and/or neural pathways. Here, the authors demonstrate that adenosine receptor activation is an important step in initiating human pre-conditioning; that pre-conditioning liberates circulating cardioprotective factor(s; and that exogenous adenosine infusion is able to recapitulate release of this factor. However, blockade of adenosine receptors in ischemic tissue does not block the protection afforded by pre-conditioning. These data have important implications for defining the physiology of human pre-conditioning and its translation to future clinical trials.

  12. Identification of Essential Genetic Baculoviral Elements for Recombinant Protein Expression by Transactivation in Sf21 Insect Cells.

    Directory of Open Access Journals (Sweden)

    Maren Bleckmann

    Full Text Available The Baculovirus Expression Vector System (BEVS is widely used to produce high amounts of recombinant proteins. Nevertheless, generating recombinant baculovirus in high quality is rather time-consuming and labor-intensive. Alternatively, virus-free expression in insect cells did not achieve similar expression levels for most proteins so far. The transactivation method is a promising approach for protein expression in Sf21 cells. It combines advantages of BEVS and plasmid-based expression by activating strong virus-dependent promoters on a transfected plasmid by baculoviral coinfection. Here, we identified expression elements required for transactivation. Therefore, we designed several vectors comprising different viral promoters or promoter combinations and tested them for eGFP expression using the automated BioLector microcultivation system. Remarkably, only the combination of the very late promoter p10 together with the homologous region 5 (hr5 could boost expression during transactivation. Other elements, like p10 alone or the late viral promoter polH, did not respond to transactivation. A new combination of hr5 and p10 with the strongest immediate early OpMNPV viral promoter OpIE2 improved the yield of eGFP by ~25% in comparison to the previous applied hr5-IE1-p10 expression cassette. Furthermore, we observed a strong influence of the transcription termination sequence and vector backbone on the level of expression. Finally, the expression levels for transactivation, BEVS and solely plasmid-based expression were compared for the marker protein eGFP, underlining the potential of transactivation for fast recombinant protein expression in Sf21 cells. In conclusion, essential elements for transactivation could be identified. The optimal elements were applied to generate an improved vector applicable in virus-free plasmid-based expression, transactivation and BEVS.

  13. Identification of Essential Genetic Baculoviral Elements for Recombinant Protein Expression by Transactivation in Sf21 Insect Cells

    Science.gov (United States)

    Chen, Fang-Fang; Yen, Zen-Zen; Lindemann, Nils; Meyer, Steffen; Spehr, Johannes; van den Heuvel, Joop

    2016-01-01

    The Baculovirus Expression Vector System (BEVS) is widely used to produce high amounts of recombinant proteins. Nevertheless, generating recombinant baculovirus in high quality is rather time-consuming and labor-intensive. Alternatively, virus-free expression in insect cells did not achieve similar expression levels for most proteins so far. The transactivation method is a promising approach for protein expression in Sf21 cells. It combines advantages of BEVS and plasmid-based expression by activating strong virus-dependent promoters on a transfected plasmid by baculoviral coinfection. Here, we identified expression elements required for transactivation. Therefore, we designed several vectors comprising different viral promoters or promoter combinations and tested them for eGFP expression using the automated BioLector microcultivation system. Remarkably, only the combination of the very late promoter p10 together with the homologous region 5 (hr5) could boost expression during transactivation. Other elements, like p10 alone or the late viral promoter polH, did not respond to transactivation. A new combination of hr5 and p10 with the strongest immediate early OpMNPV viral promoter OpIE2 improved the yield of eGFP by ~25% in comparison to the previous applied hr5-IE1-p10 expression cassette. Furthermore, we observed a strong influence of the transcription termination sequence and vector backbone on the level of expression. Finally, the expression levels for transactivation, BEVS and solely plasmid-based expression were compared for the marker protein eGFP, underlining the potential of transactivation for fast recombinant protein expression in Sf21 cells. In conclusion, essential elements for transactivation could be identified. The optimal elements were applied to generate an improved vector applicable in virus-free plasmid-based expression, transactivation and BEVS. PMID:26934632

  14. Analysis of the epidermal growth factor receptor specific transcriptome: effect of receptor expression level and an activating mutation

    DEFF Research Database (Denmark)

    Pedersen, Mikkel W; Pedersen, Nina; Damstrup, Lars

    2005-01-01

    moderately expressed or overexpressed at an in-itself transforming level. These changes were compared to those induced by the naturally occurring constitutively active variant EGFRvIII. This study provides novel insight on the activities and mechanisms of EGFRvIII and EGFR mediated transformation, as genes...... by interferons. Expression of this module was absent in the EGFRvIII-expressing cell line and the parental cell line. Treatment with the specific EGFR inhibitor AG1478 indicated that the regulations were primary, receptor-mediated events. Furthermore, activation of this module correlated with activation of STAT1...

  15. Smoking-associated lung cancer prevention by blockade of the beta-adrenergic receptor-mediated insulin-like growth factor receptor activation.

    Science.gov (United States)

    Min, Hye-Young; Boo, Hye-Jin; Lee, Ho Jin; Jang, Hyun-Ji; Yun, Hye Jeong; Hwang, Su Jung; Smith, John Kendal; Lee, Hyo-Jong; Lee, Ho-Young

    2016-10-25

    Activation of receptor tyrosine kinases (RTKs) is associated with carcinogenesis, but its contribution to smoking-associated lung carcinogenesis is poorly understood. Here we show that a tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced insulin-like growth factor 1 receptor (IGF-1R) activation via β-adrenergic receptor (β-AR) is crucial for smoking-associated lung carcinogenesis. Treatment with NNK stimulated the IGF-1R signaling pathway in a time- and dose-dependent manner, which was suppressed by pharmacological or genomic blockade of β-AR and the downstream signaling including a Gβγ subunit of β-AR and phospholipase C (PLC). Consistently, β-AR agonists led to increased IGF-1R phosphorylation. The increase in IGF2 transcription via β-AR, signal transducer and activator of transcription 3 (STAT3), and nuclear factor-kappa B (NF-κB) was associated with NNK-induced IGF-1R activation. Finally, treatment with β-AR antagonists suppressed the acquisition of transformed phenotypes in lung epithelial cells and lung tumor formation in mice. These results suggest that blocking β-AR-mediated IGF-1R activation can be an effective strategy for lung cancer prevention in smokers.

  16. Matrix metalloproteinase-3, vitamin D receptor gene polymorphisms, and occupational risk factors in lumbar disc degeneration.

    Science.gov (United States)

    Zawilla, N H; Darweesh, H; Mansour, N; Helal, S; Taha, F M; Awadallah, M; El Shazly, R

    2014-06-01

    Lumbar disc degeneration (LDD) is a process that begins early in life, contributing to the development of low back pain. LDD is a consequence of a variety of factors, and its etiology remains poorly understood. Objectives to investigate occupational and genetic risk factors inducing lumbar disc degeneration, and to evaluate the possible association of genetic polymorphisms of matrix metalloproteinase 3 (MMP-3) and vitamin D receptor (VDR) with the severity of LDD in an Egyptian population. A case control study involving 84 LDD and 60 controls was carried out. Five types of work related factors were investigated by questionnaire, complete neurological examination for all subjects and MRI for the cases. Polymerase chain reaction and restriction fragment length polymorphism methods were applied to detect polymorphisms in MMP-3 Promoter (-1,171 6A/5A) (rs 731236) and VDR-Apa (rs 35068180). We found that family history, back injury, smoking, high level of sitting, bending/twisting, physical workload, lifting, whole body vibration, mutant allele 5A of MMP-3 and mutant allele T of VDR were significantly associated with LDD (OR = 2.9, 3.1, 2.1, 11.1, 15.9, 11.7, 8.2, 12.6, 2.5 and 3.1 respectively, p < 0.05). Cases that carry allele 5A and/or allele T were associated with LDD severity. LDD is closely associated in occurrence and severity with occupational, environmental risk factors and susceptibility genes namely MMP-3, and VDR (ApaI). This study throws light on the importance of screening for early detection of susceptible individuals and disease prevention.

  17. Critical role of tumor necrosis factor receptor 1 in the pathogenesis of pulmonary emphysema in mice

    Directory of Open Access Journals (Sweden)

    Fujita M

    2016-07-01

    Full Text Available Masaki Fujita,1 Hiroshi Ouchi,2 Satoshi Ikegame,2 Eiji Harada,2 Takemasa Matsumoto,1 Junji Uchino,1 Yoichi Nakanishi,2 Kentaro Watanabe1 1Department of Respiratory Medicine, Faculty of Medicine, Fukuoka University, 2Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan Abstract: COPD is a major cause of chronic morbidity and mortality throughout the world. Although tumor necrosis factor-α (TNF-α has a critical role in the development of COPD, the role of different TNF receptors (TNFRs in pulmonary emphysema has not been resolved. We aimed to clarify the role of TNFRs in the development of pulmonary emphysema. TNF-α transgenic mice, a murine model of COPD in which the mice spontaneously develop emphysema with a large increase in lung volume and pulmonary hypertension, were crossed with either TNFR1-deficient mice or TNFR2-deficient mice. After 6 months, the gross appearance of the lung, lung histology, and pulmonary and cardiac physiology were determined. In addition, the relationship between apoptosis and emphysema was investigated. Pulmonary emphysema-like changes disappeared with deletion of TNFR1. However, slight improvements were attained with deletion of TNFR2. Apoptotic cells in the interstitium of the lung were observed in TNF-α transgenic mice. The apoptotic signals through TNFR1 appear critical for the pathogenesis of pulmonary emphysema. In contrast, the inflammatory process has a less important role for the development of emphysema. Keywords: TNF-α, receptor, emphysema, apoptosis

  18. Effects of different ligands on epidermal growth factor receptor (EGFR) nuclear translocation

    Energy Technology Data Exchange (ETDEWEB)

    Faria, Jerusa A.Q.A.; Andrade, Carolina de; Goes, Alfredo M. [Department of Biochemistry and Immunology, Universidade Federal de Minas Gerais, Av. Antonio Carlos, 6627, Belo Horizonte, MG, 31270-901 (Brazil); Rodrigues, Michele A. [Department of Biochemistry and Immunology, Universidade Federal de Minas Gerais, Av. Antonio Carlos, 6627, Belo Horizonte, MG, 31270-901 (Brazil); Department of General Pathology, Universidade Federal de Minas Gerais, Av. Antonio Carlos, 6627, Belo Horizonte, MG, 31270-901 (Brazil); Gomes, Dawidson A., E-mail: dawidson@ufmg.br [Department of Biochemistry and Immunology, Universidade Federal de Minas Gerais, Av. Antonio Carlos, 6627, Belo Horizonte, MG, 31270-901 (Brazil)

    2016-09-09

    The epidermal growth factor receptor (EGFR) is activated through binding to specific ligands and generates signals for proliferation, differentiation, migration, and cell survival. Recent data show the role of nuclear EGFR in tumors. Although many EGFR ligands are upregulated in cancers, little is known about their effects on EGFR nuclear translocation. We have compared the effects of six EGFR ligands (EGF, HB-EGF, TGF-α, β-Cellulin, amphiregulin, and epiregulin) on nuclear translocation of EGFR, receptor phosphorylation, migration, and proliferation. Cell fractionation and confocal immunofluorescence detected EGFR in the nucleus after EGF, HB-EGF, TGF-α and β-Cellulin stimulation in a dose-dependent manner. In contrast, amphiregulin and epiregulin did not generate nuclear translocation of EGFR. EGF, HB-EGF, TGF-α and β-Cellulin showed correlations between a higher rate of wound closure and increased phosphorylation of residues in the carboxy-terminus of EGFR, compared to amphiregulin and epiregulin. The data indicate that EGFR is translocated to the nucleus after stimulation with EGF, HB-EGF, TGF-α and β-Cellulin, and that these ligands are related to increased phosphorylation of EGFR tyrosine residues, inducing migration of SkHep-1 cells. - Highlights: • EGF, HB-EGF, TGF-α, β-Cellulin are involved in the EGFR nuclear translocation. • Amphiregulin and epiregulin did not promote nuclear translocation of EGFR. • EGF, HB-EGF, TGF-α and β-Cellulin have a role in SkHep-1 cells migration. • EGFR ligands associated with better prognosis don't stimulate EGFR translocation.

  19. Platelet-activating factor (PAF) receptor binding activity of the roots of Enicosanthellum pulchrum.

    Science.gov (United States)

    Nordin, Noraziah; Jalil, Juriyati; Jantan, Ibrahim; Murad, Shahnaz

    2012-03-01

    Enicosanthellum pulchrum (King) Heusden (Annonaceae) is a coniferous tree that is confined to mountain forests. The chemical constituents of this species have been studied previously; however, its biological activity has never been investigated before and is reported here for the first time. The extracts, fractions and compounds from the roots of E. pulchrum were investigated for their inhibitory effects on platelet-activating factor (PAF) receptor binding to rabbit platelets using (3)H-PAF as a ligand. The PAF receptor binding inhibitory effect using rabbit platelets was determined in vitro by measuring the difference between total amount of bound (3)H-PAF in the presence and the absence of excess unlabelled PAF. The compounds were isolated by bioassay-guided fractionation and their structures were elucidated by spectroscopic techniques. Among the extracts tested, the ethyl acetate extract was the most active with 85.6% inhibition, while hexane and methanol extracts showed 40.2 and 42.5% inhibition, respectively. Fractionation of the ethyl acetate extract using vacuum liquid chromatography (VLC) yielded six fractions AEA(I--VI). Chromatography fraction AEA(VI) yielded a new compound, 1-(2',3',4'-trimethoxyphenyl)hexan-1-ol, while fraction AEA(III) afforded three compounds, namely liriodenine, cleistopholine and dehydroanonaine. 1-(2',3',4'-Trimethoxyphenyl)hexan-1-ol, cleistopholine and dehydroanonaine showed relatively strong inhibition with IC(50) values of 26.6, 50.2 and 45.4 µM, respectively. The results suggest that these compounds could be responsible for the PAF antagonistic activity of the ethyl acetate extract of this plant.

  20. Epidermal growth factor receptor mutation in combination with expression of MIG6 alters gefitinib sensitivity

    Directory of Open Access Journals (Sweden)

    Tanaka Hiroshi

    2011-02-01

    Full Text Available Abstract Background Epidermal growth factor receptor (EGFR signaling plays an important role in the regulation of cell proliferation, survival, metastasis, and invasion in various tumors. Earlier studies showed that the EGFR is frequently overexpressed in non-small-cell lung cancer (NSCLC and EGFR mutations at specific amino acid residues in the kinase domain induce altered responsiveness to gefitinib, a small molecule EGFR tyrosine kinase inhibitor. However, the mechanism underlying the drug response modulated by EGFR mutation is still largely unknown. To elucidate drug response in EGFR signal transduction pathway in which complex dynamics of multiple molecules involved, a systematic approach is necessary. In this paper, we performed experimental and computational analyses to clarify the underlying mechanism of EGFR signaling and cell-specific gefitinib responsiveness in three H1299-derived NSCLC cell lines; H1299 wild type (H1299WT, H1299 with an overexpressed wild type EGFR (H1299EGFR-WT, and H1299 with an overexpressed mutant EGFR L858R (H1299L858R; gefitinib sensitive mutant. Results We predicted and experimentally verified that Mig6, which is a known negative regulator of EGFR and specifically expressed in H1299L858R cells, synergized with gefitinib to suppress cellular growth. Computational analyses indicated that this inhibitory effect is amplified at the phosphorylation/dephosphorylation steps of MEK and ERK. Conclusions Thus, we showed that L858R receptor mutation in combination with expression of its negative regulator, Mig6, alters signaling outcomes and results in variable drug sensitivity.

  1. Ciliary neurotrophic factor mediates dopamine D2 receptor-induced CNS neurogenesis in adult mice.

    Science.gov (United States)

    Yang, Peng; Arnold, Sheila A; Habas, Agata; Hetman, Michal; Hagg, Theo

    2008-02-27

    Neurogenesis continues in the adult forebrain subventricular zone (SVZ) and the dentate gyrus of the hippocampal formation. Degeneration of dopaminergic projections in Parkinson's disease and animals reduces, whereas ciliary neurotrophic factor (CNTF) promotes, neurogenesis. We tested whether the dopaminergic system promotes neurogenesis through CNTF. Astrocytes of the SVZ and dentate gyrus expressed CNTF and were close to dopaminergic terminals. Dopaminergic denervation in adult mice reduced CNTF mRNA by approximately 60%, whereas systemic treatment with the D2 agonist quinpirole increased CNTF mRNA in the SVZ and hippocampal formation, and in cultured astrocytes by 1.5-5 fold. The effect of quinpirole in vitro was blocked by the D2 antagonist eticlopride and did not cause astroglial proliferation or hypertrophy. Systemic quinpirole injections increased proliferation in wild-type mice by approximately 25-75% but not in CNTF-/- littermates or in the SVZ of mice infused with CNTF antibodies. Quinpirole increased the number of neuroblasts in wild-type but not in CNTF-/- littermates. Neurogenesis was reduced by approximately 20% in CNTF-/- mice, confirming the endogenous role of CNTF. Nigrostriatal denervation did not affect SVZ proliferation in CNTF-/- mice, suggesting that the dopaminergic innervation normally regulates neurogenesis through CNTF. Quinpirole acted on postsynaptic receptors as it reversed the reduced proliferation seen after dopaminergic denervation in wild-type mice. Thus, CNTF mediates dopaminergic innervation- and D2 receptor-induced neurogenesis in the adult forebrain. Because CNTF is predominantly expressed in the nervous system, this mechanism and the ability to pharmacologically modulate it have implications for Parkinson's disease and cell-replacement therapies for other disorders.

  2. Vascular endothelial growth factor receptor-3 directly interacts with phosphatidylinositol 3-kinase to regulate lymphangiogenesis.

    Directory of Open Access Journals (Sweden)

    Sanja Coso

    Full Text Available BACKGROUND: Dysfunctional lymphatic vessel formation has been implicated in a number of pathological conditions including cancer metastasis, lymphedema, and impaired wound healing. The vascular endothelial growth factor (VEGF family is a major regulator of lymphatic endothelial cell (LEC function and lymphangiogenesis. Indeed, dissemination of malignant cells into the regional lymph nodes, a common occurrence in many cancers, is stimulated by VEGF family members. This effect is generally considered to be mediated via VEGFR-2 and VEGFR-3. However, the role of specific receptors and their downstream signaling pathways is not well understood. METHODS AND RESULTS: Here we delineate the VEGF-C/VEGF receptor (VEGFR-3 signaling pathway in LECs and show that VEGF-C induces activation of PI3K/Akt and MEK/Erk. Furthermore, activation of PI3K/Akt by VEGF-C/VEGFR-3 resulted in phosphorylation of P70S6K, eNOS, PLCγ1, and Erk1/2. Importantly, a direct interaction between PI3K and VEGFR-3 in LECs was demonstrated both in vitro and in clinical cancer specimens. This interaction was strongly associated with the presence of lymph node metastases in primary small cell carcinoma of the lung in clinical specimens. Blocking PI3K activity abolished VEGF-C-stimulated LEC tube formation and migration. CONCLUSIONS: Our findings demonstrate that specific VEGFR-3 signaling pathways are activated in LECs by VEGF-C. The importance of PI3K in VEGF-C/VEGFR-3-mediated lymphangiogenesis provides a potential therapeutic target for the inhibition of lymphatic metastasis.

  3. Epidermal growth factor receptor (EGFR) involvement in successful growth hormone (GH) signaling in GH transduction defect.

    Science.gov (United States)

    Kostopoulou, Eirini; Rojas-Gil, Andrea Paola; Karvela, Alexia; Spiliotis, Bessie E

    2017-02-01

    Growth hormone (GH) transduction defect (GHTD) is a growth disorder with impaired signal transducer and activator of transcription 3 (STAT3) phosphorylation mediated by overexpression of cytokine-inducible SH2-containing protein (CIS), which causes increased growth hormone receptor (GHR) degradation. This study investigated the role of epidermal growth factor (EGF) in the restoration of normal GH signaling in GHTD. Protein expression, cellular localization and physical contact of proteins of the GH and EGF signaling pathways were studied by Western immunoblotting, immunofluorescence and co-immunoprecipitation, respectively. These were performed in fibroblasts of one GHTD patient (P) and one control child (C) at the basal state and after induction with human GH (hGH) 200 μg/L (GH200), either with or without silencing of CIS mRNA, and after induction with hGH 1000 μg/L (GH1000) or 50 ng/mL EGF. The membrane availability of the EGF receptor (EGFR) and the activated EGFR (pEGFR) was increased in P only after simultaneous GH200 and silencing of CIS mRNA or with GH1000, whereas this occurred in C after GH200 alone. After EGF induction, the membrane localization of GHR, STAT3 and that of EGFR were increased in P more than in C. In conclusion, in GHTD, the EGFR seems to participate in successful GH signaling, but induction of GHTD fibroblasts with a higher dose of hGH is needed. The EGF/EGFR pathway, in contrast to the GH/GHR pathway, seems to function normally in P and is more primed compared to C. The involvement of the EGFR in successful GH signaling may explain the catch-up growth seen in the Ps when exogenous hGH is administered.

  4. Osteoblast-specific transcription factor Osterix increases vitamin D receptor gene expression in osteoblasts.

    Directory of Open Access Journals (Sweden)

    Chi Zhang

    Full Text Available Osterix (Osx is an osteoblast-specific transcription factor required for osteoblast differentiation from mesenchymal stem cells. In Osx knock-out mice, no bone formation occurs. The vitamin D receptor (VDR is a member of the nuclear hormone receptor superfamily that regulates target gene transcription to ensure appropriate control of calcium homeostasis and bone development. Here, we provide several lines of evidence that show that the VDR gene is a target for transcriptional regulation by Osx in osteoblasts. For example, calvaria obtained from Osx-null embryos displayed dramatic reductions in VDR expression compared to wild-type calvaria. Stable overexpression of Osx stimulated VDR expression in C2C12 mesenchymal cells. Inhibition of Osx expression by siRNA led to downregulation of VDR. In contrast, Osx levels remained unchanged in osteoblasts in VDR-null mice. Mechanistic approaches using transient transfection assays showed that Osx directly activated a 1 kb fragment of the VDR promoter in a dose-dependent manner. To define the region of the VDR promoter that was responsive to Osx, a series of VDR promoter deletion mutants were examined and the minimal Osx-responsive region was refined to the proximal 120 bp of the VDR promoter. Additional point mutants were used to identify two GC-rich regions that were responsible for VDR promoter activation by Osx. Chromatin immunoprecipitation assays demonstrated that endogenous Osx was associated with the native VDR promoter in primary osteoblasts in vivo. Cumulatively, these data strongly support a direct regulatory role for Osx in VDR gene expression. They further provide new insight into potential mechanisms and pathways that Osx controls in osteoblasts and during the process of osteoblastic cell differentiation.

  5. Insulin like growth factor 2 regulation of aryl hydrocarbon receptor in MCF-7 breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Tomblin, Justin K.; Salisbury, Travis B., E-mail: salisburyt@marshall.edu

    2014-01-17

    Highlights: •IGF-2 stimulates concurrent increases in AHR and CCND1 expression. •IGF-2 promotes the binding of AHR to the endogenous cyclin D1 promoter. •AHR knockdown inhibits IGF-2 stimulated increases in CCND1 mRNA and protein. •AHR knockdown inhibits IGF-2 stimulated increases in MCF-7 proliferation. -- Abstract: Insulin like growth factor (IGF)-1 and IGF-2 stimulate normal growth, development and breast cancer cell proliferation. Cyclin D1 (CCND1) promotes cell cycle by inhibiting retinoblastoma protein (RB1). The aryl hydrocarbon receptor (AHR) is a major xenobiotic receptor that also regulates cell cycle. The purpose of this study was to investigate whether IGF-2 promotes MCF-7 breast cancer proliferation by inducing AHR. Western blot and quantitative real time PCR (Q-PCR) analysis revealed that IGF-2 induced an approximately 2-fold increase (P < .001) in the expression of AHR and CCND1. Chromatin immunoprecipitation (ChIP), followed by Q-PCR indicated that IGF-2 promoted (P < .001) a 7-fold increase in AHR binding on the CCND1 promoter. AHR knockdown significantly (P < .001) inhibited IGF-2 stimulated increases in CCND1 mRNA and protein. AHR knockdown cells were less (P < .001) responsive to the proliferative effects of IGF-2 than control cells. Collectively, our findings have revealed a new regulatory mechanism by which IGF-2 induction of AHR promotes the expression of CCND1 and the proliferation of MCF-7 cells. This previously uncharacterized pathway could be important for the proliferation of IGF responsive cancer cells that also express AHR.

  6. Molecular basis for the Kallmann syndrome-linked fibroblast growth factor receptor mutation

    Energy Technology Data Exchange (ETDEWEB)

    Thurman, Ryan D.; Kathir, Karuppanan Muthusamy; Rajalingam, Dakshinamurthy [Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, AR 72701 (United States); Kumar, Thallapuranam K. Suresh, E-mail: sthalla@uark.edu [Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, AR 72701 (United States)

    2012-08-31

    Highlights: Black-Right-Pointing-Pointer The structural basis of the Kallmann syndrome is elucidated. Black-Right-Pointing-Pointer Kallmann syndrome mutation (A168S) induces a subtle conformational change(s). Black-Right-Pointing-Pointer Structural interactions mediated by beta-sheet G are most perturbed. Black-Right-Pointing-Pointer Ligand (FGF)-receptor interaction(s) is completely abolished by Kallmann mutation. Black-Right-Pointing-Pointer Kallmann mutation directly affects the FGF signaling process. -- Abstract: Kallmann syndrome (KS) is a developmental disease that expresses in patients as hypogonadotropic hypogonadism and anosmia. KS is commonly associated with mutations in the extracellular D2 domain of the fibroblast growth factor receptor (FGFR). In this study, for the first time, the molecular basis for the FGFR associated KS mutation (A168S) is elucidated using a variety of biophysical experiments, including multidimensional NMR spectroscopy. Secondary and tertiary structural analysis using far UV circular dichroism, fluorescence and limited trypsin digestion assays suggest that the KS mutation induces subtle tertiary structure change in the D2 domain of FGFR. Results of isothermal titration calorimetry experiments show the KS mutation causes a 10-fold decrease in heparin binding affinity and also a complete loss in ligand (FGF-1) binding. {sup 1}H-{sup 15}N chemical perturbation data suggest that complete loss in the ligand (FGF) binding affinity is triggered by a subtle conformational change that disrupts crucial structural interactions in both the heparin and the FGF binding sites in the D2 domain of FGFR. The novel findings reported in this study are expected to provide valuable clues toward a complete understanding of the other genetic diseases linked to mutations in the FGFR.

  7. Effects of different ligands on epidermal growth factor receptor (EGFR) nuclear translocation

    International Nuclear Information System (INIS)

    Faria, Jerusa A.Q.A.; Andrade, Carolina de; Goes, Alfredo M.; Rodrigues, Michele A.; Gomes, Dawidson A.

    2016-01-01

    The epidermal growth factor receptor (EGFR) is activated through binding to specific ligands and generates signals for proliferation, differentiation, migration, and cell survival. Recent data show the role of nuclear EGFR in tumors. Although many EGFR ligands are upregulated in cancers, little is known about their effects on EGFR nuclear translocation. We have compared the effects of six EGFR ligands (EGF, HB-EGF, TGF-α, β-Cellulin, amphiregulin, and epiregulin) on nuclear translocation of EGFR, receptor phosphorylation, migration, and proliferation. Cell fractionation and confocal immunofluorescence detected EGFR in the nucleus after EGF, HB-EGF, TGF-α and β-Cellulin stimulation in a dose-dependent manner. In contrast, amphiregulin and epiregulin did not generate nuclear translocation of EGFR. EGF, HB-EGF, TGF-α and β-Cellulin showed correlations between a higher rate of wound closure and increased phosphorylation of residues in the carboxy-terminus of EGFR, compared to amphiregulin and epiregulin. The data indicate that EGFR is translocated to the nucleus after stimulation with EGF, HB-EGF, TGF-α and β-Cellulin, and that these ligands are related to increased phosphorylation of EGFR tyrosine residues, inducing migration of SkHep-1 cells. - Highlights: • EGF, HB-EGF, TGF-α, β-Cellulin are involved in the EGFR nuclear translocation. • Amphiregulin and epiregulin did not promote nuclear translocation of EGFR. • EGF, HB-EGF, TGF-α and β-Cellulin have a role in SkHep-1 cells migration. • EGFR ligands associated with better prognosis don't stimulate EGFR translocation.

  8. The role of epidermal growth factor receptor in chordoma pathogenesis: a potential therapeutic target.

    Science.gov (United States)

    Shalaby, Asem; Presneau, Nadège; Ye, Hongtao; Halai, Dina; Berisha, Fitim; Idowu, Bernadine; Leithner, Andreas; Liegl, Bernadette; Briggs, Timothy R W; Bacsi, Krisztian; Kindblom, Lars-Gunnar; Athanasou, Nicholas; Amary, Maria Fernanda; Hogendoorn, Pancras C W; Tirabosco, Roberto; Flanagan, Adrienne M

    2011-02-01

    Chordoma, the molecular hallmark of which is T (brachyury), is a rare malignant bone tumour with a high risk of local recurrence and a tumour from which metastatic disease is a common late event. Currently, there is no effective drug therapy for treating chordomas, although there is evidence that some patients respond to the empirical use of epidermal growth factor receptor (EGFR) antagonists. The aim of this study was to determine the role of EGFR in the pathogenesis of chordoma. Paraffin-embedded material from 173 chordomas from 160 patients [sacro-coccygeal (n = 94), skull-based (n = 50), and mobile spine (n = 16)] was analysed by immunohistochemistry and revealed total EGFR expression in 69% of cases analysed. Of 147 informative chordomas analysed by FISH, 38% revealed high-level EGFR polysomy, 4% high-level polysomy with focal amplification, 18% low-level polysomy, and 39% disomy. Phospho-receptor tyrosine kinase array membranes showed EGFR activation in the chordoma cell line U-CH1 and all of the three chordomas analysed. Direct sequencing of EGFR (exons 18-21), KRAS, NRAS, HRAS (exons 2, 3), and BRAF (exons 11, 15) using DNA from 62 chordomas failed to reveal mutations. PTEN expression was absent by immunohistochemistry in 19 of 147 (13%) analysed chordomas, only one of which revealed high-level polysomy of EGFR. The EGFR inhibitor tyrphostin (AG 1478) markedly inhibited proliferation of the chordoma cell line U-CH1 in vitro and diminished EGFR phosphorylation in a dose-dependant manner, a finding supported by inhibition of phosphorylated Erk1/2. p-Akt was suppressed to a much lesser degree in these experiments. There was no reduction of T as assessed by western blotting. These data implicate aberrant EGFR signalling in the pathogenesis of chordoma. This study provides a strategy for patient stratification for treatment with EGFR antagonists. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  9. IQGAP1 Protein Binds Human Epidermal Growth Factor Receptor 2 (HER2) and Modulates Trastuzumab Resistance*

    Science.gov (United States)

    White, Colin D.; Li, Zhigang; Dillon, Deborah A.; Sacks, David B.

    2011-01-01

    Human epidermal growth factor receptor 2 (HER2) is overexpressed in 20–25% of breast cancers. Increased HER2 expression is an adverse prognostic factor and correlates with decreased patient survival. HER2-positive (HER2(+)) breast cancer is treated with trastuzumab. Unfortunately, some patients are intrinsically refractory to therapy, and many who do respond initially become resistant within 1 year. Understanding the molecular mechanisms underlying HER2 signaling and trastuzumab resistance is essential to reduce breast cancer mortality. IQGAP1 is a ubiquitously expressed scaffold protein that contains multiple protein interaction domains. By regulating its binding partners IQGAP1 integrates signaling pathways, several of which contribute to breast tumorigenesis. We show here that IQGAP1 is overexpressed in HER2(+) breast cancer tissue and binds directly to HER2. Knockdown of IQGAP1 decreases HER2 expression, phosphorylation, signaling, and HER2-stimulated cell proliferation, effects that are all reversed by reconstituting cells with IQGAP1. Reducing IQGAP1 up-regulates p27, and blocking this increase attenuates the growth inhibitory effects of IQGAP1 knockdown. Importantly, IQGAP1 is overexpressed in trastuzumab-resistant breast epithelial cells, and reducing IQGAP1 both augments the inhibitory effects of trastuzumab and restores trastuzumab sensitivity to trastuzumab-resistant SkBR3 cells. These data suggest that inhibiting IQGAP1 function may represent a rational strategy for treating HER2(+) breast carcinoma. PMID:21724847

  10. Fibroblast growth factor receptor 4 polymorphism is associated with liver cirrhosis in hepatocarcinoma.

    Directory of Open Access Journals (Sweden)

    Ming-Jen Sheu

    Full Text Available Fibroblast growth factor receptor 4 (FGFR4 polymorphisms are positively correlated with tumor progression in numerous malignant tumors. However, the association between FGFR4 genetic variants and the risk of hepatocellular carcinoma (HCC has not yet been determined. In this study, we investigated the potential associations of FGFR4 single nucleotide polymorphisms (SNPs with HCC susceptibility and its clinicopathological characteristics.Four SNPs in FGFR4 (rs1966265, rs351855, rs2011077, and rs7708357 were analyzed among 884 participants, including 595 controls and 289 patients with HCC. The samples were further analyzed to clarify the associations between these gene polymorphisms and the risk of HCC, and the impact of these SNPs on the susceptibility and clinicopathological characteristics of HCC. After adjusting for other covariants, HCC patients who carrying at least one A genotype (GA and AA at rs351855 were observed to have a higher risk of liver cirrhosis compared with those carrying the wild-type genotype (GG (OR: 2.113, 95% CI: 1.188-3.831. Moreover, the patients with at least one A genotype were particularly showed a high level of alpha-fetoprotein (AFP.Our findings suggest that genetic polymorphism in FGFR4 rs351855 may be associated with the risk of HCC coupled with liver cirrhosis and may markedly increase the AFP level in Taiwanese patients with HCC. In addition, this is the first study that evaluated the risk factors associated with FGFR4 polymorphism variants in Taiwanese patients with HCC.

  11. Involvement of Corticotropin-Releasing Factor and Receptors in Immune Cells in Irritable Bowel Syndrome.

    Science.gov (United States)

    Chatoo, Mahanand; Li, Yi; Ma, Zhiqiang; Coote, John; Du, Jizeng; Chen, Xuequn

    2018-01-01

    Irritable bowel syndrome (IBS) is a common functional gastrointestinal disorder defined by ROME IV criteria as pain in the lower abdominal region, which is associated with altered bowel habit or defecation. The underlying mechanism of IBS is not completely understood. IBS seems to be a product of interactions between various factors with genetics, dietary/intestinal microbiota, low-grade inflammation, and stress playing a key role in the pathogenesis of this disease. The crosstalk between the immune system and stress in IBS mechanism is increasingly recognized. Corticotropin-releasing factor (CRF), a major mediator in the stress response, is involved in altered function in GI, including inflammatory processes, colonic transit time, contractile activity, defecation pattern, pain threshold, mucosal secretory function, and barrier functions. This mini review focuses on the recently establish local GI-CRF system, its involvement in modulating the immune response in IBS, and summarizes current IBS animal models and mapping of CRF, CRFR1, and CRFR2 expression in colon tissues. CRF and receptors might be a key molecule involving the immune and movement function via brain-gut axis in IBS.

  12. Involvement of Corticotropin-Releasing Factor and Receptors in Immune Cells in Irritable Bowel Syndrome

    Directory of Open Access Journals (Sweden)

    Mahanand Chatoo

    2018-02-01

    Full Text Available Irritable bowel syndrome (IBS is a common functional gastrointestinal disorder defined by ROME IV criteria as pain in the lower abdominal region, which is associated with altered bowel habit or defecation. The underlying mechanism of IBS is not completely understood. IBS seems to be a product of interactions between various factors with genetics, dietary/intestinal microbiota, low-grade inflammation, and stress playing a key role in the pathogenesis of this disease. The crosstalk between the immune system and stress in IBS mechanism is increasingly recognized. Corticotropin-releasing factor (CRF, a major mediator in the stress response, is involved in altered function in GI, including inflammatory processes, colonic transit time, contractile activity, defecation pattern, pain threshold, mucosal secretory function, and barrier functions. This mini review focuses on the recently establish local GI-CRF system, its involvement in modulating the immune response in IBS, and summarizes current IBS animal models and mapping of CRF, CRFR1, and CRFR2 expression in colon tissues. CRF and receptors might be a key molecule involving the immune and movement function via brain–gut axis in IBS.

  13. Human epidermal growth factor receptor2 expression in unresectable gastric cancers: Relationship with CT characteristics

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jeong Sub [Dept. of Radiology, Jeju National University Hospital, Jeju (Korea, Republic of); Kim, Se Hyung; Im, Seock Ah; Kim, Min A; Han, Joon Koo [Seoul National University Hospital, Seoul (Korea, Republic of)

    2017-09-15

    To retrospectively analyze the qualitative CT features that correlate with human epidermal growth factor receptor 2 (HER2)-expression in pathologically-proven gastric cancers. A total of 181 patients with pathologically-proven unresectable gastric cancers with HER2-expression (HER2-positive [n = 32] and negative [n = 149]) were included. CT features of primary gastric and metastatic tumors were reviewed. The prevalence of each CT finding was compared in both groups. Thereafter, binary logistic regression determined the most significant differential CT features. Clinical outcomes were compared using Kaplan-Meier method. HER2-postive cancers showed lower clinical T stage (21.9% vs. 8.1%; p = 0.015), hyperattenuation on portal phase (62.5% vs. 30.9%; p = 0.003), and was more frequently metastasized to the liver (62.5% vs. 32.2%; p = 0.001), than HER2-negative cancers. On binary regression analysis, hyperattenuation of the tumor (odds ratio [OR], 4.68; p < 0.001) and hepatic metastasis (OR, 4.43; p = 0.001) were significant independent factors that predict HER2-positive cancers. Median survival of HER2-positive cancers (13.7 months) was significantly longer than HER2-negative cancers (9.6 months) (p = 0.035). HER2-positive gastric cancers show less-advanced T stage, hyperattenuation on the portal phase, and frequently metastasize to the liver, as compared to HER2-negative cancers.

  14. Citrus pectin: characterization and inhibitory effect on fibroblast growth factor-receptor interaction.

    Science.gov (United States)

    Liu, Y; Ahmad, H; Luo, Y; Gardiner, D T; Gunasekera, R S; McKeehan, W L; Patil, B S

    2001-06-01

    This study was undertaken to characterize the pectin from four citrus species and to determine their in vitro inhibitory activities on the binding of fibroblast growth factor (FGF) to the FGF receptor (FGFR). Pectin from various parts of lemon, grapefruit, tangerine, and orange were isolated and characterized. Tangerine had the highest pectin content among the four citrus species. Segment membrane contained as much as or more pectin than flavedo/albedo. Anhydrogalacturonic content was highest in pectin from segment membrane of tangerine and flavedo/albedo of grapefruit. Lemon pectin contained the highest methoxyl content (MC), and grapefruit contained the largest proportion of lower molecular weight (lemon was the most potent inhibitor. The inhibition activity was significantly correlated with sugar content, MC, and size of pectin. Kinetic studies revealed a competitive nature of pectin inhibition with the heparin, a crucial component of the FGF signal transduction process. The observation that the heparin-dependent biological activity of FGF signal transduction is antagonized by citrus pectin should be further investigated for the use of these pectins as anti-growth factor agents for potential health benefits.

  15. Optical Molecular Imaging of Epidermal Growth Factor Receptor Expression to Improve Detection of Oral Neoplasia

    Directory of Open Access Journals (Sweden)

    Nitin Nitin

    2009-06-01

    Full Text Available Background: The development of noninvasive molecular imaging approaches has the potential to improve management of cancer. Methods: In this study, we demonstrate the potential of noninvasive topical delivery of an epidermal growth factor-Alexa 647 (EGF-Alexa 647 conjugate to image changes in epidermal growth factor receptor expression associated with oral neoplasia. We report a series of preclinical analyses to evaluate the optical contrast achieved after topical delivery of EGF-Alexa 647 in a variety of model systems, including cells, three-dimensional tissue cultures, and intact human tissue specimens using wide-field and high-resolution fluorescence imaging. Data were collected from 17 different oral cancer patients: eight pairs of normal and abnormal biopsies and nine resected tumors were examined. Results: The EGF-dye conjugate can be uniformly delivered throughout the oral epithelium with a penetration depth exceeding 500 µm and incubation time of less than 30 minutes. After EGF-Alexa 647 incubation, the presence of oral neoplasia is associated with a 1.5- to 6.9-fold increase in fluorescence contrast compared with grossly normal mucosa from the same patient with both wide-field and high-resolution fluorescence imaging. Conclusions: Results illustrate the potential of EGF-targeted fluorescent agents for in vivo molecular imaging, a technique that may aid in the diagnosis and characterization of oral neoplasia and allow real-time detection of tumor margins.

  16. Eps15 is recruited to the plasma membrane upon epidermal growth factor receptor activation and localizes to components of the endocytic pathway during receptor internalization

    DEFF Research Database (Denmark)

    Torrisi, M R; Lotti, L V; Belleudi, F

    1999-01-01

    role for eps15 in receptor-mediated endocytosis. In this study we show that, upon activation of the EGFR kinase, eps15 undergoes dramatic relocalization consisting of 1) initial relocalization to the plasma membrane and 2) subsequent colocalization with the EGFR in various intracellular compartments......Eps15 is a substrate for the tyrosine kinase of the epidermal growth factor receptor (EGFR) and is characterized by the presence of a novel protein:protein interaction domain, the EH domain. Eps15 also stably binds the clathrin adaptor protein complex AP-2. Previous work demonstrated an essential......, suggesting sustained phosphorylation in endocytic compartments. Our results are consistent with a model in which eps15 undergoes cycles of association:dissociation with membranes and suggest multiple roles for this protein in the endocytic pathway....

  17. Kinetic analysis of platelet-derived growth factor receptor/phosphoinositide 3-kinase/Akt signaling in fibroblasts.

    Science.gov (United States)

    Park, Chang Shin; Schneider, Ian C; Haugh, Jason M

    2003-09-26

    Isoforms of the serine-threonine kinase Akt coordinate multiple cell survival pathways in response to stimuli such as platelet-derived growth factor (PDGF). Activation of Akt is a multistep process, which relies on the production of 3'-phosphorylated phosphoinositide (PI) lipids by PI 3-kinases. To quantitatively assess the kinetics of PDGF receptor/PI 3-kinase/Akt signaling in fibroblasts, a systematic study of this pathway was performed, and a mechanistic mathematical model that describes its operation was formulated. We find that PDGF receptor phosphorylation exhibits positive cooperativity with respect to PDGF concentration, and its kinetics are quantitatively consistent with a mechanism in which receptor dimerization is initially mediated by the association of two 1:1 PDGF/PDGF receptor complexes. Receptor phosphorylation is transient at high concentrations of PDGF, consistent with the loss of activated receptors upon endocytosis. By comparison, Akt activation responds to lower PDGF concentrations and exhibits more sustained kinetics. Further analysis and modeling suggest that the pathway is saturated at the level of PI 3-kinase activation, and that the p110alpha catalytic subunit of PI 3-kinase contributes most to PDGF-stimulated 3'-PI production. Thus, at high concentrations of PDGF the kinetics of 3'-PI production are limited by the turnover rate of these lipids, while the Akt response is additionally influenced by the rate of Akt deactivation.

  18. Radiotherapy and receptor of epidermal growth factor; Radiotherapie et recepteur de l'Epidermal Growth Factor

    Energy Technology Data Exchange (ETDEWEB)

    Deberne, M. [Institut Gustave-Roussy, 94 - Villejuif (France)

    2009-10-15

    The expression level of the receptor of the epidermal growth factor is in correlation with the tumor cells radiosensitivity. An overexpression of the E.G.F.R. is often present in the bronchi cancer, epidermoid carcinomas of the O.R.L. sphere, esophagus, uterine cervix, and anal duct but also in the rectum cancers and glioblastomas. At the clinical level, the E.G.F.R. expression is in correlation with an unfavourable prognosis after radiotherapy in numerous tumoral localizations. In the rectum cancers it is an independent prognosis factor found in multifactorial analysis: increase of the rate of nodes and local recurrence when the E.G.F.R. is over expressed. In the uterine cervix cancers, the survival is is negatively affected in multifactorial analysis by the E.G.F.R. membranes expression level. At the therapy level, the development of anti E.G.F.R. targeted therapies (tyrosine kinase inhibitors and monoclonal antibodies) opens a new therapy field at radio-sensitivity potentiality. The irradiation makes an activation of the E.G.F.R. way that would be partially responsible of the post irradiation tumoral repopulation. This activation leads the phosphorylation of the PI3 kinase ways and M.A.P. kinase ones, then the Akt protein one that acts an apoptotic modulator part. It has been shown that blocking the E.G.F.R. way acts on three levels: accumulation of ells in phase G1, reduction of the cell repair and increasing of apoptosis. he inhibition of post irradiation action of the E.G.F.R. signal way is a factor explaining the ionizing radiation - anti E.G.F.R. synergy. The preclinical data suggest that the E.G.F.R. blocking by the monoclonal antibodies is more important than the use of tyrosine kinase inhibitors. A first positive randomized study with the cetuximab, published in 2006 in the epidermoid carcinomas of the O.R.L. sphere lead to its authorization on the market with the radiotherapy for this localization. The use of cetuximab in other indication with or in

  19. Tumour necrosis factor-alpha (TNF), lymphotoxin and TNF receptor levels in serum from patients with Wegener's granulomatosis

    DEFF Research Database (Denmark)

    Jónasdóttir, O; Petersen, J; Bendtzen, K

    2001-01-01

    Wegener's granulomatosis (WG) is a systemic inflammatory disease with vasculitis as the key feature. Abnormal expression of tumour necrosis factor alpha (TNFalpha) is considered of prime pathogenic importance in several inflammatory diseases. The effects of TNFa are mediated by TNF receptors (TNF...

  20. Neurite outgrowth induced by a synthetic peptide ligand of neural cell adhesion molecule requires fibroblast growth factor receptor activation

    DEFF Research Database (Denmark)

    Rønn, L C; Doherty, P; Holm, A

    2000-01-01

    The neural cell adhesion molecule NCAM is involved in axonal outgrowth and target recognition in the developing nervous system. In vitro, NCAM-NCAM binding has been shown to induce neurite outgrowth, presumably through an activation of fibroblast growth factor receptors (FGFRs). We have recently...

  1. Complement factor 5a receptor chimeras reveal the importance of lipid-facing residues in transport competence

    DEFF Research Database (Denmark)

    Klco, Jeffery M; Sen, Saurabh; Hansen, Jakob L

    2009-01-01

    oligomerization studies demonstrated energy transfer between the wild-type complement factor 5a receptor and the lipid-facing chimeras, suggesting that the lipid-facing residues within a single TM segment are not essential for oligomerization. These studies highlight the importance of the lipid-facing residues...

  2. Elucidation of the mechanism of the regulatory function of the Ig1 module of the fibroblast growth factor receptor 1

    DEFF Research Database (Denmark)

    Kiselyov, Vladislav; Kochoyan, Artur; Poulsen, Flemming

    2006-01-01

    The extracellular part of the fibroblast growth factor (FGF) receptor (FGFR) consists of up to three Ig modules (Ig1-Ig3), in which the Ig2 and Ig3 modules determine affinity and specificity for FGF and heparin. The FGFR isoforms lacking the Ig1 module have higher affinity for FGF and heparin than...

  3. Primary cilia and coordination of receptor tyrosine kinase (RTK) and transforming growth factor β (TGF-β) signaling

    DEFF Research Database (Denmark)

    Christensen, Søren Tvorup; Morthorst, Stine Kjær; Mogensen, Johanne Bay

    2017-01-01

    are at the root of a pleiotropic group of diseases and syndromic disorders called ciliopathies. In this review, we present an overview of primary cilia-mediated regulation of receptor tyrosine kinase (RTK) and transforming growth factor β (TGF-β) signaling. Further, we discuss how defects in the coordination...

  4. Molecular cloning and functional characterization of the human platelet-derived growth factor alpha receptor gene promoter

    NARCIS (Netherlands)

    Afink, G. B.; Nistér, M.; Stassen, B. H.; Joosten, P. H.; Rademakers, P. J.; Bongcam-Rudloff, E.; van Zoelen, E. J.; Mosselman, S.

    1995-01-01

    Expression of the platelet-derived growth factor alpha receptor (PDGF alpha R) is strictly regulated during mammalian development and tumorigenesis. The molecular mechanisms involved in the specific regulation of PDGF alpha R expression are unknown, but transcriptional regulation of the PDGF alpha R

  5. Frontolimbic serotonin 2A receptor binding in healthy subjects is associated with personality risk factors for affective disorder

    DEFF Research Database (Denmark)

    Frokjaer, Vibe G.; Mortensen, Erik L.; Nielsen, Finn Årup

    2008-01-01

    Background: Serotonergic dysfunction has been associated with affective disorders. High trait neuroticism, as measured on personality inventories, is a risk factor for major depression. In this study we investigated whether neuroticism is associated with serotonin 2A receptor binding in brain reg...

  6. Magnesium wasting associated with epidermal-growth-factor receptor-targeting antibodies in colorectal cancer: a prospective study.

    NARCIS (Netherlands)

    Tejpar, S.; Piessevaux, H.; Claes, K.; Piront, P.; Hoenderop, J.G.J.; Verslype, C.; Cutsem, E. van

    2007-01-01

    BACKGROUND: Preliminary evidence suggests that magnesium wasting occurs in patients who are treated with epidermal-growth-factor receptor (EGFR)-targeting antibodies for colorectal cancer. The mechanism of this side-effect is unknown, and if all or a subset of patients are affected is also unclear.

  7. Granulocyte colony-stimulating factor receptors on granulocytes are down-regulated after endotoxin administration to healthy humans

    NARCIS (Netherlands)

    Dekkers, P. E.; Juffermans, N. P.; Hove, T. t; de Jonge, E.; van Deventer, S. J.; van der Poll, T.

    2000-01-01

    Granulocyte colony-stimulating factor (G-CSF) is considered an important mediator of host defense against infection, and recombinant G-CSF is administered to patients with various infections. G-CSF binds to a specific receptor that is expressed on granulocytes and monocytes. To obtain insight about

  8. Intraoperative fluorescence delineation of head and neck cancer with a fluorescent Anti-epidermal growth factor receptor nanobody

    NARCIS (Netherlands)

    Van Driel, P.B.A.A.; Van Der Vorst, J.R.; Verbeek, F.P.R.; Oliveira, S.|info:eu-repo/dai/nl/304841455; Snoeks, T.J.A.; Keereweer, S.; Chan, B.; Boonstra, M.C.; Frangioni, J.V.; Van Bergen En Henegouwen, P.M.P.|info:eu-repo/dai/nl/071919481; Vahrmeijer, A.L.; Lowik, C.W.G.M.

    2014-01-01

    Intraoperative near-infrared (NIR) fluorescence imaging is a technology with high potential to provide the surgeon with real-time visualization of tumors during surgery. Our study explores the feasibility for clinical translation of an epidermal growth factor receptor (EGFR)-targeting nanobody for

  9. Limited human epidermal growth factor receptor 2 discordance in metastatic breast cancer patients treated with trastuzumab, a population based study

    NARCIS (Netherlands)

    van Rooijen, J.M.; de Munck, L.; de Graaf, J.C.; Siesling, Sabine; de Vries, Erik G.; Boers, J.E.

    2014-01-01

    Background Accurate assessment of the human epidermal growth factor receptor 2 (HER2) in breast cancer is essential for proper treatment decisions. HER2 positivity confirmation rates in breast cancer trials by central testing pathology laboratories were reported to be approximately 85%. The aim of

  10. Limited human epidermal growth factor receptor 2 discordance in metastatic breast cancer patients treated with trastuzumab, a population based study

    NARCIS (Netherlands)

    van Rooijen, J. M.; de Munck, L.; de Graaf, J. C.; Siesling, S.; de Vries, E. G.; Boers, J. E.

    Background: Accurate assessment of the human epidermal growth factor receptor 2 (HER2) in breast cancer is essential for proper treatment decisions. HER2 positivity confirmation rates in breast cancer trials by central testing pathology laboratories were reported to be approximately 85%. The aim of

  11. Cloning of Human Tumor Necrosis Factor (TNF) Receptor cDNA and Expression of Recombinant Soluble TNF-Binding Protein

    Science.gov (United States)

    Gray, Patrick W.; Barrett, Kathy; Chantry, David; Turner, Martin; Feldmann, Marc

    1990-10-01

    The cDNA for one of the receptors for human tumor necrosis factor (TNF) has been isolated. This cDNA encodes a protein of 455 amino acids that is divided into an extracellular domain of 171 residues and a cytoplasmic domain of 221 residues. The extracellular domain has been engineered for expression in mammalian cells, and this recombinant derivative binds TNFα with high affinity and inhibits its cytotoxic activity in vitro. The TNF receptor exhibits similarity with a family of cell surface proteins that includes the nerve growth factor receptor, the human B-cell surface antigen CD40, and the rat T-cell surface antigen OX40. The TNF receptor contains four cysteine-rich subdomains in the extra-cellular portion. Mammalian cells transfected with the entire TNF receptor cDNA bind radiolabeled TNFα with an affinity of 2.5 x 10-9 M. This binding can be competitively inhibited with unlabeled TNFα or lymphotoxin (TNFβ).

  12. Decreased numbers of chemotactic factor receptors in chronic neutropenia with defective chemotaxis: spontaneous recovery from the neutrophil abnormalities during early childhood

    International Nuclear Information System (INIS)

    Yasui, K.; Yamazaki, M.; Miyagawa, Y.; Komiyama, A.; Akabane, T.

    1987-01-01

    Childhood chronic neutropenia with decreased numbers of chemotactic factor receptors as well as defective chemotaxis was first demonstrated in an 8-month-old girl. Chemotactic factor receptors on neutrophils were assayed using tritiated N-formyl-methionyl-leucyl-phenylalanine ( 3 H-FMLP). The patient's neutrophils had decreased numbers of the receptors: numbers of the receptors were 20,000 (less than 3 SD) as compared with those of control cells of 52,000 +/- 6000 (mean +/- SD) (n = 10). The neutropenia disappeared spontaneously by 28 months of age parallel with the improvement of chemotaxis and increase in numbers of chemotactic factor receptors. These results demonstrate a transient decrease of neutrophil chemotactic factor receptors as one of the pathophysiological bases of a transient defect of neutrophil chemotaxis in this disorder

  13. Experimental diabetes increases insulin-like growth factor I and II receptor concentration and gene expression in kidney

    International Nuclear Information System (INIS)

    Werner, H.; Shen-Orr, Z.; Stannard, B.; Burguera, B.; Roberts, C.T. Jr.; LeRoith, D.

    1990-01-01

    Insulinlike growth factor I (IGF-I) is a mitogenic hormone with important regulatory roles in growth and development. One of the target organs for IGF-I action is the kidney, which synthesizes abundant IGF-I receptors and IGF-I itself. To study the involvement of IGF-I and the IGF-I receptor in the development of nephropathy, one of the major complications of diabetes mellitus, we measured the expression of these genes in the kidney and in other tissues of the streptozocin-induced diabetic rat. The binding of 125I-labeled IGF-I to crude membranes was measured in the same tissues. We observed a 2.5-fold increase in the steady-state level of IGF-I-receptor mRNA in the diabetic kidney, which was accompanied by a 2.3-fold increase in IGF-I binding. In addition to this increase in IGF-I binding to the IGF-I receptor, there was also binding to a lower-molecular-weight material that may represent an IGF-binding protein. No change was detected in the level of IGF-I-peptide mRNA. Similarly, IGF-II-receptor mRNA levels and IGF-II binding were significantly increased in the diabetic kidney. IGF-I- and IGF-II-receptor mRNA levels and IGF-I and IGF-II binding returned to control values after insulin treatment. Because the IGF-I receptor is able to transduce mitogenic signals on activation of its tyrosine kinase domain, we hypothesize that, among other factors, high levels of receptor in the diabetic kidney may also be involved in the development of diabetic nephropathy. Increased IGF-II-receptor expression in the diabetic kidney may be important for the intracellular transport and packaging of lysosomal enzymes, although a role for this receptor in signal transduction cannot be excluded. Finally, the possible role of IGF-binding proteins requires further study

  14. Reproductive factors and risk of hormone receptor positive and negative breast cancer: a cohort study

    International Nuclear Information System (INIS)

    Ritte, Rebecca; Grote, Verena; Boeing, Heiner; Aleksandrova, Krasimira; Trichopoulou, Antonia; Lagiou, Pagona; Trichopoulos, Dimitrios; Palli, Domenico; Berrino, Franco; Mattiello, Amalia; Tumino, Rosario; Tikk, Kaja; Sacerdote, Carlotta; Quirós, José Ramón; Buckland, Genevieve; Molina-Montes, Esther; Chirlaque, María-Dolores; Ardanaz, Eva; Amiano, Pilar; Bueno-de-Mesquita, H Bas; Gils, Carla H van; Peeters, Petra HM; Lukanova, Annekatrin; Wareham, Nick; Khaw, Kay-Tee; Key, Timothy J; Travis, Ruth C; Weiderpass, Elisabete; Dumeaux, Vanessa; Lund, Eliv; Sund, Malin; Andersson, Anne; Romieu, Isabelle; Tjønneland, Anne; Rinaldi, Sabina; Vineis, Paulo; Merritt, Melissa A; Riboli, Elio; Kaaks, Rudolf; Olsen, Anja; Overvad, Kim; Dossus, Laure; Fournier, Agnès; Clavel-Chapelon, Françoise

    2013-01-01

    The association of reproductive factors with hormone receptor (HR)-negative breast tumors remains uncertain. Within the EPIC cohort, Cox proportional hazards models were used to describe the relationships of reproductive factors (menarcheal age, time between menarche and first pregnancy, parity, number of children, age at first and last pregnancies, time since last full-term childbirth, breastfeeding, age at menopause, ever having an abortion and use of oral contraceptives [OC]) with risk of ER-PR- (n = 998) and ER+PR+ (n = 3,567) breast tumors. A later first full-term childbirth was associated with increased risk of ER+PR+ tumors but not with risk of ER-PR- tumors (≥35 vs. ≤19 years HR: 1.47 [95% CI 1.15-1.88] p trend < 0.001 for ER+PR+ tumors; ≥35 vs. ≤19 years HR: 0.93 [95% CI 0.53-1.65] p trend = 0.96 for ER-PR- tumors; P het = 0.03). The risk associations of menarcheal age, and time period between menarche and first full-term childbirth with ER-PR-tumors were in the similar direction with risk of ER+PR+ tumors (p het = 0.50), although weaker in magnitude and statistically only borderline significant. Other parity related factors such as ever a full-term birth, number of births, age- and time since last birth were associated only with ER+PR+ malignancies, however no statistical heterogeneity between breast cancer subtypes was observed. Breastfeeding and OC use were generally not associated with breast cancer subtype risk. Our study provides possible evidence that age at menarche, and time between menarche and first full-term childbirth may be associated with the etiology of both HR-negative and HR-positive malignancies, although the associations with HR-negative breast cancer were only borderline significant

  15. Type I Insulin-like Growth Factor Receptor Induces Pulmonary Tumorigenesis

    Directory of Open Access Journals (Sweden)

    Nicolle M. Linnerth

    2009-07-01

    Full Text Available Despite the type I insulin-like growth factor receptor (IGF-IR being highly expressed in more than 80% of human lung tumors, a transgenic model of IGF-IR overexpression in the lung has not been created. We produced two novel transgenic mouse models in which IGF-IR is overexpressed in either lung type II alveolar cells (surfactant protein C [SPC]-IGFIR or Clara cells (CCSP-IGFIR in a doxycycline-inducible manner. Overexpression of IGF-IR in either cell type caused multifocal adenomatous alveolar hyperplasia with papillary and solid adenomas. These tumors expressed thyroid transcription factor 1 and Kruppel-like factor 5 in most tumor cells. Similar to our previous work with lung tumors that developed in the mouse mammary tumor virus-IGF-II transgenic mice, the lung tumors that develop in the SPC-IGFIR and CCSP-IGFIR transgenic mice expressed high levels of the cyclic adenosine monophosphate response element binding protein that was localized primarily to the nucleus. Although elevated IGF-IR expression can initiate lung tumor development, tumors can become independent of IGF-IR signaling as IGF-IR down-regulation in established tumors produced tumor regression in some, but not all, of the tumors. These findings implicate IGF-IR as an important initiator of lung tumorigenesis and suggest that the SPC-IGFIR and CCSP-IGFIR transgenic mice can be used to further our understanding of human lung cancer and the role IGF-IR plays in this disease.

  16. Ac-mediated trans-activation of the Ds element in rice (Oryza sativa L.) cells as revealed by GUS assay.

    Science.gov (United States)

    Solis, R; Takumi, S; Mori, N; Nakamura, C

    1999-01-01

    A method using particle bombardment and beta-glucuronidase (GUS) assay was applied to rice callus for detecting the excision of the maize Ds element trans-activated by the Ac transposase source. Excision of Ds biolistically introduced into rice callus resulted in the restoration of the interrupted gus gene expression, allowing visual detection of trans-activation two days after bombardment. Only the transgenic callus lines expressing the Ac transposase gene and the wild-type callus co-transformed with Ac and Ds revealed GUS activity. Frequency of excision, estimated based on the relative GUS activity, ranged from 0.3% to 2.2%. Callus lines showing different levels of Ac transcripts revealed varying excision frequencies. At the later stages of callus growth after selection for the Ac/Ds transformed lines, excision events were detected by GUS assay and confirmed by PCR and sequence analyses of the excision sites in individual colonies. GUS activity was also demonstrated in the primary regenerants from the Ac/Ds-transformed callus colonies. The method described in this study may be used as an approach for rapid detection of excision events and assessment of various factors limiting Ac/Ds activity in rice cells.

  17. Human Epidermal Growth Factor Receptor 2 Overexpression in Micropapillary and Other Variants of Urothelial Carcinoma.

    Science.gov (United States)

    Behzatoğlu, Kemal; Yörükoğlu, Kutsal; Demir, Hale; Bal, Nebil

    2016-06-21

    Human epidermal growth factor receptor 2 (HER2) protein overexpression or gene amplification has been shown in urothelial bladder cancer. This could be helpful when using targeted anti-HER2 therapy on these tumors. To evaluate HER2 immunohistochemical expression in conventional urothelial carcinoma (UC), in situ UC, and UC variants primarily in micropapillary urothelial carcinoma (MPUC). The study evaluated 60 MPUC cases; 25 invasive, 20 low-grade noninvasive, and 10 high-grade noninvasive UC cases; 8 in situ UC cases; and 69 UC variant cases. The immunohistochemistry staining was scored according to recommendations of the American Society of Clinical Oncology/College of American Pathologists 2013 HER2 test guideline established for breast cancer and only 3+ staining was considered HER2 overexpression. HER2 overexpression was determined by 3+ staining. 34 of 60 MPUC cases (56%) showed HER2 overexpression (3+ staining). We observed 3+ staining HER2 overexpression in nine of 25 conventional invasive UC cases (36%), four of eight in situ UC cases (50%), and three of six lipid cell variant cases (50%). 3+ staining HER2 overexpression was not seen in eight glandular, six small cell, and five sarcomatoid variant cases. HER2 overexpression was negative in the 20 low-grade noninvasive UC cases but positive in two of the 10 high-grade noninvasive UC cases (20%). We observed HER2 overexpression most commonly in MPUC cases. We also found HER2 overexpression in conventional invasive and in situ UC cases. Pure in situ UC and conventional invasive UC, especially MPUC, could be candidate tumors for treatment with anti-HER2 antibody (trastuzumab therapy). Targeted therapy has a limited place in treatment of bladder cancer. In this study, human epidermal growth factor receptor 2 (HER2) overexpression in bladder carcinomas was evaluated in a large number of cases. Anti-HER2 therapy could be used in bladder cancers, as in breast and gastric cancers. Copyright © 2016 European

  18. Epidermal growth factor receptor (EGFR) mutations in non-small cell lung cancer (NSCLC) of Yunnan in southwestern China

    OpenAIRE

    Zhou, Yongchun; Yang, Yanlong; Yang, Chenggang; Chen, Yunlan; Yang, Changshao; Du, Yaxi; Zhao, Guangqiang; Ye, Lianhua; Huang, Yunchao

    2017-01-01

    To investigate the Epidermal Growth Factor Receptor (EGFR) mutation status in non-small cell lung cancer (NSCLC) in Yunnan province in southwestern China, we detected EGFR mutation by Amplification Refractory Mutation System (ARMS) polymerase chain reaction (PCR) using DNA samples from 447 pathologically confirmed NSCLC specimens (175 tissue, 256 plasma and 16 cytologic samples). The relationship between EGFR mutations and demographic and clinical factors were further explored. Subgroup analy...

  19. Significance of epidermal growth factor receptor signaling for acquisition of meiotic and developmental competence in mammalian oocytes

    Czech Academy of Sciences Publication Activity Database

    Procházka, Radek; Blaha, Milan; Němcová, Lucie

    2017-01-01

    Roč. 97, č. 4 (2017), s. 537-549 ISSN 0006-3363 R&D Projects: GA MZe(CZ) QJ1510138; GA MŠk EF15_003/0000460 Institutional support: RVO:67985904 Keywords : amphiregulin * cumulus cells * epidermal growth factor receptor Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Reproductive biology (medical aspects to be 3) Impact factor: 3.432, year: 2016

  20. Enhancement of insulin-like growth factor 2 receptors in glioblastoma

    International Nuclear Information System (INIS)

    Sara, V.; Prisell, Per; Sjoegren, Barbro; Enberg, Goesta

    1986-01-01

    The somatomedins (IGF-1/IGF-2) are a family of growth-promoting hormones which have been identified in the human central nervous system where their specific receptors are distributed. The present study identified somatomedin receptors in glioblastoma and compared them with those found in normal brain. A significant enhancement in the binding of 125 1-IGF-2 but not 125 1-IGF-1 to glioblastoma membranes was found. A fourfold increase in IGF-2 receptor concentration was observed. These findings indicate enhanced expression of the IGF-2 receptor in glioblastoma. (author)