WorldWideScience

Sample records for cd137 ligand signaling

  1. Deciphering CD137 (4-1BB) signaling in T-cell costimulation for translation into successful cancer immunotherapy.

    Science.gov (United States)

    Sanchez-Paulete, Alfonso R; Labiano, Sara; Rodriguez-Ruiz, Maria E; Azpilikueta, Arantza; Etxeberria, Iñaki; Bolaños, Elixabet; Lang, Valérie; Rodriguez, Manuel; Aznar, M Angela; Jure-Kunkel, Maria; Melero, Ignacio

    2016-03-01

    CD137 (4-1BB, TNF-receptor superfamily 9) is a surface glycoprotein of the TNFR family which can be induced on a variety of leukocyte subsets. On T and NK cells, CD137 is expressed following activation and, if ligated by its natural ligand (CD137L), conveys polyubiquitination-mediated signals via TNF receptor associated factor 2 that inhibit apoptosis, while enhancing proliferation and effector functions. CD137 thus behaves as a bona fide inducible costimulatory molecule. These functional properties of CD137 can be exploited in cancer immunotherapy by systemic administration of agonist monoclonal antibodies, which increase anticancer CTLs and enhance NK-cell-mediated antibody-dependent cell-mediated cytotoxicity. Reportedly, anti-CD137 mAb and adoptive T-cell therapy strongly synergize, since (i) CD137 expression can be used to select the T cells endowed with the best activities against the tumor, (ii) costimulation of the lymphocyte cultures to be used in adoptive T-cell therapy can be done with CD137 agonist antibodies or CD137L, and (iii) synergistic effects upon coadministration of T cells and antibodies are readily observed in mouse models. Furthermore, the signaling cytoplasmic tail of CD137 is a key component of anti-CD19 chimeric antigen receptors that are used to redirect T cells against leukemia and lymphoma in the clinic. Ongoing phase II clinical trials with agonist antibodies and the presence of CD137 sequence in these successful chimeric antigen receptors highlight the importance of CD137 in oncoimmunology. PMID:26773716

  2. Deciphering CD137 (4-1BB) signaling in T-cell costimulation for translation into successful cancer immunotherapy.

    Science.gov (United States)

    Sanchez-Paulete, Alfonso R; Labiano, Sara; Rodriguez-Ruiz, Maria E; Azpilikueta, Arantza; Etxeberria, Iñaki; Bolaños, Elixabet; Lang, Valérie; Rodriguez, Manuel; Aznar, M Angela; Jure-Kunkel, Maria; Melero, Ignacio

    2016-03-01

    CD137 (4-1BB, TNF-receptor superfamily 9) is a surface glycoprotein of the TNFR family which can be induced on a variety of leukocyte subsets. On T and NK cells, CD137 is expressed following activation and, if ligated by its natural ligand (CD137L), conveys polyubiquitination-mediated signals via TNF receptor associated factor 2 that inhibit apoptosis, while enhancing proliferation and effector functions. CD137 thus behaves as a bona fide inducible costimulatory molecule. These functional properties of CD137 can be exploited in cancer immunotherapy by systemic administration of agonist monoclonal antibodies, which increase anticancer CTLs and enhance NK-cell-mediated antibody-dependent cell-mediated cytotoxicity. Reportedly, anti-CD137 mAb and adoptive T-cell therapy strongly synergize, since (i) CD137 expression can be used to select the T cells endowed with the best activities against the tumor, (ii) costimulation of the lymphocyte cultures to be used in adoptive T-cell therapy can be done with CD137 agonist antibodies or CD137L, and (iii) synergistic effects upon coadministration of T cells and antibodies are readily observed in mouse models. Furthermore, the signaling cytoplasmic tail of CD137 is a key component of anti-CD19 chimeric antigen receptors that are used to redirect T cells against leukemia and lymphoma in the clinic. Ongoing phase II clinical trials with agonist antibodies and the presence of CD137 sequence in these successful chimeric antigen receptors highlight the importance of CD137 in oncoimmunology.

  3. Role of CD137 signaling in dengue virus-mediated apoptosis

    International Nuclear Information System (INIS)

    Highlights: → For the first time the role of CD137 in dengue virus (DENV) infection. → Induction of DENV-mediated apoptosis by CD137 signaling. → Sensitization to CD137-mediated apoptosis by dengue virus capsid protein (DENV C). → Nuclear localization of DENV C is required for CD137-mediated apoptosis. -- Abstract: Hepatic dysfunction is a well recognized feature of dengue virus (DENV) infection. However, molecular mechanisms of hepatic injury are still poorly understood. A complex interaction between DENV and the host immune response contributes to DENV-mediated tissue injury. DENV capsid protein (DENV C) physically interacts with the human death domain-associated protein Daxx. A double substitution mutation in DENV C (R85A/K86A) abrogates Daxx interaction, nuclear localization and apoptosis. Therefore we compared the expression of cell death genes between HepG2 cells expressing DENV C and DENV C (R85A/K86A) using a real-time PCR array. Expression of CD137, which is a member of the tumor necrosis factor receptor family, increased significantly in HepG2 cells expressing DENV C compared to HepG2 cells expressing DENV C (R85A/K86A). In addition, CD137-mediated apoptotic activity in HepG2 cells expressing DENV C was significantly increased by anti-CD137 antibody compared to that of HepG2 cells expressing DENV C (R85A/K86A). In DENV-infected HepG2 cells, CD137 mRNA and CD137 positive cells significantly increased and CD137-mediated apoptotic activity was increased by anti-CD137 antibody. This work is the first to demonstrate the contribution of CD137 signaling to DENV-mediated apoptosis.

  4. Role of CD137 signaling in dengue virus-mediated apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Nagila, Amar [Medical Molecular Biology Unit, Office for Research and Development, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Department of Biochemistry, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Netsawang, Janjuree [Faculty of Medical Technology, Rangsit University, Bangkok (Thailand); Srisawat, Chatchawan [Department of Biochemistry, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Noisakran, Sansanee [Dengue Hemorrhagic Fever Research Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok (Thailand); Medical Biotechnology Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Bangkok (Thailand); Morchang, Atthapan; Yasamut, Umpa [Medical Molecular Biology Unit, Office for Research and Development, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Department of Immunology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok (Thailand); Puttikhunt, Chunya [Dengue Hemorrhagic Fever Research Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok (Thailand); Medical Biotechnology Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Bangkok (Thailand); Kasinrerk, Watchara [Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai (Thailand); Biomedical Technology Research Center, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency at Chiang Mai University, Chiang Mai (Thailand); and others

    2011-07-08

    Highlights: {yields} For the first time the role of CD137 in dengue virus (DENV) infection. {yields} Induction of DENV-mediated apoptosis by CD137 signaling. {yields} Sensitization to CD137-mediated apoptosis by dengue virus capsid protein (DENV C). {yields} Nuclear localization of DENV C is required for CD137-mediated apoptosis. -- Abstract: Hepatic dysfunction is a well recognized feature of dengue virus (DENV) infection. However, molecular mechanisms of hepatic injury are still poorly understood. A complex interaction between DENV and the host immune response contributes to DENV-mediated tissue injury. DENV capsid protein (DENV C) physically interacts with the human death domain-associated protein Daxx. A double substitution mutation in DENV C (R85A/K86A) abrogates Daxx interaction, nuclear localization and apoptosis. Therefore we compared the expression of cell death genes between HepG2 cells expressing DENV C and DENV C (R85A/K86A) using a real-time PCR array. Expression of CD137, which is a member of the tumor necrosis factor receptor family, increased significantly in HepG2 cells expressing DENV C compared to HepG2 cells expressing DENV C (R85A/K86A). In addition, CD137-mediated apoptotic activity in HepG2 cells expressing DENV C was significantly increased by anti-CD137 antibody compared to that of HepG2 cells expressing DENV C (R85A/K86A). In DENV-infected HepG2 cells, CD137 mRNA and CD137 positive cells significantly increased and CD137-mediated apoptotic activity was increased by anti-CD137 antibody. This work is the first to demonstrate the contribution of CD137 signaling to DENV-mediated apoptosis.

  5. CD137 is induced by the CD40 signal on chronic lymphocytic leukemia B cells and transduces the survival signal via NF-κB activation.

    Directory of Open Access Journals (Sweden)

    Yukana Nakaima

    Full Text Available CD137 is a member of the tumor necrosis factor receptor family that is expressed on activated T cells. This molecule provides a co-stimulatory signal that enhances the survival, and differentiation of cells, and has a crucial role in the development of CD8 cytotoxic T cells and anti-tumor immunity. Here we report that CD137 expression is also induced on normal or malignant human B cells by CD40 ligation by its ligand CD154. This CD137 induction was more prominent in chronic lymphocytic leukemia (CLL cells than in other types of B cells. CD137 stimulation on B cells by its ligand induced the nuclear translocation of p52 (a non-canonical NF-κB factor. In agreement with this finding, expression of the survival factor BCL-XL was upregulated. Consequently, the CD137 signal augmented the survival of CD154-stimulated CLL B cells in vitro. This unexpected induction of CD137 on B cells by CD40 signal may influence the clinical course of CLL.

  6. Hypoxia-induced soluble CD137 in malignant cells blocks CD137L-costimulation as an immune escape mechanism

    Science.gov (United States)

    Labiano, Sara; Palazón, Asis; Bolaños, Elixabet; Azpilikueta, Arantza; Sánchez-Paulete, Alfonso R.; Morales-Kastresana, Aizea; Quetglas, Jose I.; Perez-Gracia, José L.; Gúrpide, Alfonso; Rodriguez-Ruiz, Maria; Aznar, M. Angela; Jure-Kunkel, Maria; Berraondo, Pedro; Melero, Ignacio

    2016-01-01

    ABSTRACT Hypoxia is a common feature in solid tumors that has been implicated in immune evasion. Previous studies from our group have shown that hypoxia upregulates the co-stimulatory receptor CD137 on activated T lymphocytes and on vascular endothelial cells. In this study, we show that exposure of mouse and human tumor cell lines to hypoxic conditions (1% O2) promotes CD137 transcription. However, the resulting mRNA is predominantly an alternatively spliced form that encodes for a soluble variant, lacking the transmembrane domain. Accordingly, soluble CD137 (sCD137) is detectable by ELISA in the supernatant of hypoxia-exposed cell lines and in the serum of tumor-bearing mice. sCD137, as secreted by tumor cells, is able to bind to CD137-Ligand (CD137L). Our studies on primed T lymphocytes in co-culture with stable transfectants for CD137L demonstrate that tumor-secreted sCD137 prevents co-stimulation of T lymphocytes. Such an effect results from preventing the interaction of CD137L with the transmembrane forms of CD137 expressed on T lymphocytes undergoing activation. Indeed, silencing CD137 with shRNA renders more immunogenic tumor-cell variants upon inoculation to immunocompetent mice but which readily grafted on immunodeficient or CD8+ T-cell-depleted mice. These mechanisms are interpreted as a molecular strategy deployed by tumors to repress lymphocyte co-stimulation via CD137/CD137L. PMID:26942078

  7. CD137 facilitates the resolution of acute DSS-induced colonic inflammation in mice.

    Directory of Open Access Journals (Sweden)

    Julia M Martínez Gómez

    Full Text Available BACKGROUND: CD137 and its ligand (CD137L are potent immunoregulatory molecules that influence activation, proliferation, differentiation and cell death of leukocytes. Expression of CD137 is upregulated in the lamina propria cells of Crohn's disease patients. Here, the role of CD137 in acute Dextran-Sodium-Sulfate (DSS-induced colitis in mice was examined. METHODS: We induced acute large bowel inflammation (colitis via DSS administration in CD137(-/- and wild-type (WT mice. Colitis severity was evaluated by clinical parameters (weight loss, cytokine secretion in colon segment cultures, and scoring of histological inflammatory parameters. Additionally, populations of lamina propria mononuclear cells (LPMNC and intraepithelial lymphocytes (IEL were characterized by flow cytometry. In a subset of mice, resolution of intestinal inflammation was evaluated 3 and 7 days after withdrawal of DSS. RESULTS: We found that both CD137(-/- and WT mice demonstrated a similar degree of inflammation after 5 days of DSS exposure. However, the resolution of colonic inflammation was impaired in the absence of CD137. This was accompanied by a higher histological score of inflammation, and increased release of the pro-inflammatory mediators granulocyte macrophage colony-stimulating factor (GM-CSF, CXCL1, IL-17 and IFN-γ. Further, there were significantly more neutrophils among the LPMNC of CD137(-/- mice, and reduced numbers of macrophages among the IEL. CONCLUSION: We conclude that CD137 plays an essential role in the resolution of acute DSS-induced intestinal inflammation in mice.

  8. Anti-CD137 monoclonal antibodies and adoptive T cell therapy: a perfect marriage?

    Science.gov (United States)

    Weigelin, Bettina; Bolaños, Elixabet; Rodriguez-Ruiz, Maria E; Martinez-Forero, Ivan; Friedl, Peter; Melero, Ignacio

    2016-05-01

    CD137(4-1BB) costimulation and adoptive T cell therapy strongly synergize in terms of achieving maximal efficacy against experimental cancers. These costimulatory biological functions of CD137 have been exploited by means of introducing the CD137 signaling domain in clinically successful chimeric antigen receptors and to more efficiently expand T cells in culture. In addition, immunomagnetic sorting of CD137-positive T cells among tumor-infiltrating lymphocytes selects for the fittest antitumor T lymphocytes for subsequent cultures. In mouse models, co-infusion of both agonist antibodies and T cells attains marked synergistic effects that result from more focused and intense cytolytic activity visualized under in vivo microscopy and from more efficient entrance of T cells into the tumor through the vasculature. These several levels of dynamic interaction between adoptive T cell therapy and CD137 offer much opportunity to raise the efficacy of current cancer immunotherapies.

  9. CD137 Facilitates the Resolution of Acute DSS-Induced Colonic Inflammation in Mice

    Science.gov (United States)

    Martínez Gómez, Julia M.; Chen, Lieping; Schwarz, Herbert; Karrasch, Thomas

    2013-01-01

    Background CD137 and its ligand (CD137L) are potent immunoregulatory molecules that influence activation, proliferation, differentiation and cell death of leukocytes. Expression of CD137 is upregulated in the lamina propria cells of Crohn’s disease patients. Here, the role of CD137 in acute Dextran-Sodium-Sulfate (DSS)-induced colitis in mice was examined. Methods We induced acute large bowel inflammation (colitis) via DSS administration in CD137−/− and wild-type (WT) mice. Colitis severity was evaluated by clinical parameters (weight loss), cytokine secretion in colon segment cultures, and scoring of histological inflammatory parameters. Additionally, populations of lamina propria mononuclear cells (LPMNC) and intraepithelial lymphocytes (IEL) were characterized by flow cytometry. In a subset of mice, resolution of intestinal inflammation was evaluated 3 and 7 days after withdrawal of DSS. Results We found that both CD137−/− and WT mice demonstrated a similar degree of inflammation after 5 days of DSS exposure. However, the resolution of colonic inflammation was impaired in the absence of CD137. This was accompanied by a higher histological score of inflammation, and increased release of the pro-inflammatory mediators granulocyte macrophage colony-stimulating factor (GM-CSF), CXCL1, IL-17 and IFN-γ. Further, there were significantly more neutrophils among the LPMNC of CD137−/− mice, and reduced numbers of macrophages among the IEL. Conclusion We conclude that CD137 plays an essential role in the resolution of acute DSS-induced intestinal inflammation in mice. PMID:24023849

  10. Functional expression of CD137 (4-1BB) on T helper follicular cells

    Science.gov (United States)

    Alfaro, Carlos; Echeveste, Jose I; Rodriguez-Ruiz, Maria E; Solorzano, Jose L; Perez-Gracia, Jose L; Idoate, Miguel A; Lopez-Picazo, Jose M; Sanchez-Paulete, Alfonso R; Labiano, Sara; Rouzaut, Ana; Oñate, Carmen; Aznar, Angela; Lozano, Maria D; Melero, Ignacio

    2015-01-01

    CD137 (4-1BB) is a surface protein initially discovered to mark activated T lymphocytes. However, its broader expression pattern also encompasses activated NK cells, B cells and myeloid cells, including mature dendritic cells. In this study, we have immunostained for CD137 on paraffin-embedded lymphoid tissues including tonsils, lymph nodes, ectopic tertiary lymphoid tissue in Hashimoto thyroiditis and cancer. Surprisingly, immunostaining mainly decorated intrafollicular lymphocytes in the tissues analyzed, with only scattered staining in interfollicular areas. Moreover, pathologic lymphoid follicles in follicular lymphoma and tertiary lymphoid tissue associated with non-small cell lung cancer showed a similar pattern of immunostaining. Multispectral fluorescence cytometry demonstrated that CD137 expression was restricted to CD4+ CXCR5+ follicular T helper lymphocytes (TFH cells) in tonsils and lymph nodes. Short-term culture of lymph node cell suspensions in the presence of either an agonistic anti-CD137 monoclonal antibody (mAb) or CD137-ligand stimulated the functional upregulation of TFH cells in 3 out of 6 cases, as indicated by CD40L surface expression and cytokine production. As a consequence, immunostimulatory monoclonal antibodies targeting CD137 (such as urelumab and PF-05082566) should be expected to primarily act on this lymphocyte subset, thus modifying ongoing humoral immune responses in patients with autoimmune disease and cancer. PMID:26587331

  11. CD137-mediated immunotherapy for allergic asthma

    OpenAIRE

    Polte, Tobias; Foell, Juergen; Werner, Christoph; Hoymann, Heinz-Gerd; Braun, Armin; Burdach, Stefan; Mittler, Robert S.; Hansen, Gesine

    2006-01-01

    The prevalence of asthma continues to increase. Asthma is caused by a Th2 cell–driven immune response. Its optimal treatment remains a challenge, and a sufficient immunotherapeutic approach to treating asthma has yet to be found. Using a murine asthma model, we show that a single injection of an anti-CD137 (4-1BB) mAb prevents the development of airway hyperreactivity, eosinophilic airway inflammation, excessive mucus production, and elevated IgE during the observation period of 7 weeks. Most...

  12. Rationale for anti-CD137 cancer immunotherapy.

    Science.gov (United States)

    Makkouk, Amani; Chester, Cariad; Kohrt, Holbrook E

    2016-02-01

    The consideration of the complex interplay between the tumour microenvironment (TME) and the immune response is the key for designing effective immunotherapies. Therapeutic strategies that harness co-stimulatory receptors have recently gained momentum in the clinic. One such strategy with promising clinical applications is the targeting of CD137, a member of the tumour necrosis factor receptor superfamily. Its expression on both innate and adaptive immune cells, coupled with its unique ability to potentiate antitumour responses through modulating the TME and to ameliorate autoimmune responses, has established it as an appealing target. In this review, we will discuss the various CD137-targeted immunotherapeutics that have reached clinical development, with a focus on recent advances and novel modalities such as CD137 chimeric antigen receptors and CD137 bispecific antibodies. We will also highlight the effect of CD137 targeting on the TME and discuss the importance of probing TME changes for predicting and testing the efficacy of CD137-mediated immunotherapy.

  13. Targeting CD137 enhances the efficacy of cetuximab

    Science.gov (United States)

    Kohrt, Holbrook E.; Colevas, A. Dimitrios; Houot, Roch; Weiskopf, Kipp; Goldstein, Matthew J.; Lund, Peder; Mueller, Antonia; Sagiv-Barfi, Idit; Marabelle, Aurelien; Lira, Ruth; Troutner, Emily; Richards, Lori; Rajapaska, Amanda; Hebb, Jonathan; Chester, Cariad; Waller, Erin; Ostashko, Anton; Weng, Wen-Kai; Chen, Lieping; Czerwinski, Debra; Fu, Yang-Xin; Sunwoo, John; Levy, Ronald

    2014-01-01

    Treatment with cetuximab, an EGFR-targeting IgG1 mAb, results in beneficial, yet limited, clinical improvement for patients with head and neck (HN) cancer as well as colorectal cancer (CRC) patients with WT KRAS tumors. Antibody-dependent cell-mediated cytotoxicity (ADCC) by NK cells contributes to the efficacy of cetuximab. The costimulatory molecule CD137 (4-1BB) is expressed following NK and memory T cell activation. We found that isolated human NK cells substantially increased expression of CD137 when exposed to cetuximab-coated, EGFR-expressing HN and CRC cell lines. Furthermore, activation of CD137 with an agonistic mAb enhanced NK cell degranulation and cytotoxicity. In multiple murine xenograft models, including EGFR-expressing cancer cells, HN cells, and KRAS-WT and KRAS-mutant CRC, combined cetuximab and anti-CD137 mAb administration was synergistic and led to complete tumor resolution and prolonged survival, which was dependent on the presence of NK cells. In patients receiving cetuximab, the level of CD137 on circulating and intratumoral NK cells was dependent on postcetuximab time and host FcyRIIIa polymorphism. Interestingly, the increase in CD137-expressing NK cells directly correlated to an increase in EGFR-specific CD8+ T cells. These results support development of a sequential antibody approach against EGFR-expressing malignancies that first targets the tumor and then the host immune system. PMID:24837434

  14. Agonists of Co-stimulation in Cancer Immunotherapy Directed Against CD137, OX40, GITR, CD27, CD28, and ICOS.

    Science.gov (United States)

    Sanmamed, Miguel F; Pastor, Fernando; Rodriguez, Alfonso; Perez-Gracia, Jose Luis; Rodriguez-Ruiz, Maria E; Jure-Kunkel, Maria; Melero, Ignacio

    2015-08-01

    T and natural killer (NK) lymphocytes are considered the main effector players in the immune response against tumors. Full activation of T and NK lymphocytes requires the coordinated participation of several surface receptors that meet their cognate ligands through structured transient cell-to-cell interactions known as immune synapses. In the case of T cells, the main route of stimulation is driven by antigens as recognized in the form of short polypeptides associated with major histocompatibility complex (MHC) antigen-presenting molecules. However, the functional outcome of T-cell stimulation towards clonal expansion and effector function acquisition is contingent on the contact of additional surface receptor-ligand pairs and on the actions of cytokines in the milieu. While some of those interactions are inhibitory, others are activating and are collectively termed co-stimulatory receptors. The best studied belong to either the immunoglobulin superfamily or the tumor necrosis factor-receptor (TNFR) family. Co-stimulatory receptors include surface moieties that are constitutively expressed on resting lymphocytes such as CD28 or CD27 and others whose expression is induced upon recent previous antigen priming, ie, CD137, GITR, OX40, and ICOS. Ligation of these glycoproteins with agonist antibodies actively conveys activating signals to the lymphocyte. Those signals, acting through a potentiation of the cellular immune response, give rise to anti-tumor effects in mouse models. Anti-CD137 antibodies are undergoing clinical trials with evidence of clinical activity and anti-OX40 monoclonal antibodies (mAbs) induce interesting immunomodulation effects in humans. Antibodies anti-CD27 and GITR have recently entered clinical trials. The inherent dangers of these immunomodulation strategies are the precipitation of excessive systemic inflammation or/and invigorating silent autoimmunity. Agonist antibodies, recombinant forms of the natural ligands, and polynucleotide

  15. Immunotherapy with Agonistic Anti-CD137: Two Sides of a Coin

    Institute of Scientific and Technical Information of China (English)

    YonglianSun; JonathanH.Chen; YangxinFu

    2004-01-01

    CD137 (4-1BB), a member of the TNF receptor superfamily, is an inducible T cell costimulatory receptor primarily expressed on activated CD4+ and CD8+ T cells. Agonistic monoclonal antibodies (mAbs) against CD137 greatly enhance T cell-mediated immune responses against many types of tumors and viruses. Surprisingly, these agonists also showed therapeutic effects in several autoimmune diseases. These findings suggest that in different disease environments, CD137 engagement with agonist mAb in vivo can diametrically modulate immune response outcomes. Therefore, CD137 agonists represent a promising immunotherapeutic approach to a wide array of disparate immune disorders. However, CD137's potency in modulating immune response necessitates caution when targeting CD137 clinically. Cellular & Molecular Immunology. 2004;1(1):31-36.

  16. Immunotherapy with Agonistic Anti-CD137: Two Sides of a Coin

    Institute of Scientific and Technical Information of China (English)

    Yonglian Sun; Jonathan H.Chen; Yangxin Fu

    2004-01-01

    CD137 (4-1BB), a member of the TNF receptor superfamily, is an inducible T cell costimulatory receptor primarily expressed on activated CD4+ and CD8+ T cells. Agonistic monoclonal antibodies (mAbs) against CD137 greatly enhance T cell-mediated immune responses against many types of tumors and viruses. Surprisingly, these agonists also showed therapeutic effects in several autoimmune diseases. These findings suggest that in different disease environments, CD137 engagement with agonist mAb in vivo can diametrically modulate immune response outcomes. Therefore, CD137 agonists represent a promising immunotherapeutic approach to a wide array of disparate immune disorders. However, CD137's potency in modulating immune response necessitates caution when targeting CD137 clinically. Cellular & Molecular Immunology. 2004;1(1):31-36.

  17. Immunomodulating antibodies in the treatment of metastatic melanoma: the experience with anti-CTLA-4, anti-CD137, and anti-PD1.

    Science.gov (United States)

    Simeone, Ester; Ascierto, Paolo A

    2012-01-01

    Clinical activity of anti-CTLA-4 (cytotoxic T-lymphocyte antigen-4) monoclonal antibodies (mAb) has changed the approaches for the treatment of cancer in terms of patterns of response, duration of response, and adverse event profiles. In fact, antibodies that block the interaction of CTLA-4 with its ligands B7.1 and B7.2 can enhance immune responses, including anti-tumor immunity. Two recent studies using ipilimumab (an anti-CTLA-4 mAb) demonstrated improvements in overall survival in the treatment of advanced melanoma. These studies utilized two different schedules of treatment in different patient categories (first and second line of treatment). However, the results were quite similar despite the different dosage used and the combination with dacarbazine in the first line treatment. Ongoing clinical studies will establish the efficacy of ipilimumab as monotherapy or in combination with other drugs for the treatment of metastatic melanoma and a variety of other cancers. Other antibodies, such as CD137 agonists and PD-1 antagonists, are currently in various stages of pre-clinical and clinical development. Agonist antibodies directed against CD137 (4-1BB) on the surface of antigen-primed T-lymphocytes increase tumor immunity that is curative against some transplantable murine tumors. Programmed death-1 (PD1) is a surface molecule delivering inhibitory signals important to maintain T-cell functional silence against their cognate antigens. Interference with PD1 or its ligand PD-L1 (B7-H1) increases anti-tumor immunity. As a result, human mAbs anti-PD1 and anti-PD-L1 are under clinical development. This paper reviews recent studies in the treatment of advanced melanoma with these types of monoclonal antibodies. Ipilimumab can be considered a cornerstone of a new era in melanoma treatment. However, the aim is to optimize the therapy with anti-CTLA-4 antibodies to define the best schedule for next combination regimens (other immunomodulatory antibodies, BRAF

  18. Intermittent chemotherapy can retain the therapeutic potential of anti-CD137 antibody during the late tumor-bearing state

    Science.gov (United States)

    Tongu, Miki; Harashima, Nanae; Tamada, Koji; Chen, Lieping; Harada, Mamoru

    2015-01-01

    Immunomodulating monoclonal antibodies (mAb) can evoke antitumor T-cell responses, which are attenuated by regulatory T cells (Treg) and myeloid-derived suppressor cells (MDSC). Treatment with cyclophosphamide (CP) and gemcitabine (GEM) can mitigate the immunosuppression by Treg and MDSC, respectively. In the current study, we examined the antitumor effects of a combination of local injection with anti-CD137 mAb and intermittent low-dose chemotherapy using CP and GEM in subcutaneously established CT26 colon carcinoma. Although a significant antitumor effect was observed when local anti-CD137 mAb therapy (5 μg) was started early in the tumor-bearing stage (day 10), no therapeutic efficacy was observed when the mAb therapy was started at a later tumor-bearing stage (day 17). Analyses of the tumor-infiltrating immune cells revealed that the number of Gr-1high/low CD11b+ MDSC started to increase 13 days after tumor inoculation, whereas injection with low-dose (50 mg/kg) CP and GEM mitigated this increase. In addition, although intermittent injections with low-dose CP and GEM on days 10 and 18 suppressed tumor growth significantly, additional local injections of anti-CD137 mAb on days 19, 21, and 23 further augmented the therapeutic efficacy. Cytotoxic T lymphocytes reactive to CT26 and a tumor antigen peptide were induced successfully from the spleen cells of tumor-cured or tumor-stable mice. In a bilateral tumor inoculation model, this combination therapy achieved systemic therapeutic effects and suppressed the growth of mAb-untreated tumors. These results suggest that intermittent immunochemotherapy using CP and GEM could retain the therapeutic potential of anti-CD137 mAb that is normally impaired during the late tumor-bearing stage. Intermittent chemotherapy and anti-CD137 antibody therapy. PMID:25363339

  19. Focusing and sustaining the antitumor CTL effector killer response by agonist anti-CD137 mAb

    Science.gov (United States)

    Weigelin, Bettina; Bolaños, Elixabet; Teijeira, Alvaro; Martinez-Forero, Ivan; Labiano, Sara; Azpilikueta, Arantza; Morales-Kastresana, Aizea; Quetglas, José I.; Wagena, Esther; Sánchez-Paulete, Alfonso Rodríguez; Chen, Lieping; Friedl, Peter; Melero, Ignacio

    2015-01-01

    Cancer immunotherapy is undergoing significant progress due to recent clinical successes by refined adoptive T-cell transfer and immunostimulatory monoclonal Ab (mAbs). B16F10-derived OVA-expressing mouse melanomas resist curative immunotherapy with either adoptive transfer of activated anti-OVA OT1 CTLs or agonist anti-CD137 (4-1BB) mAb. However, when acting in synergistic combination, these treatments consistently achieve tumor eradication. Tumor-infiltrating lymphocytes that accomplish tumor rejection exhibit enhanced effector functions in both transferred OT-1 and endogenous cytotoxic T lymphocytes (CTLs). This is consistent with higher levels of expression of eomesodermin in transferred and endogenous CTLs and with intravital live-cell two-photon microscopy evidence for more efficacious CTL-mediated tumor cell killing. Anti-CD137 mAb treatment resulted in prolonged intratumor persistence of the OT1 CTL-effector cells and improved function with focused and confined interaction kinetics of OT-1 CTL with target cells and increased apoptosis induction lasting up to six days postadoptive transfer. The synergy of adoptive T-cell therapy and agonist anti-CD137 mAb thus results from in vivo enhancement and sustainment of effector functions. PMID:26034288

  20. Stimulation of natural killer cells with a CD137-specific antibody enhances trastuzumab efficacy in xenotransplant models of breast cancer

    Science.gov (United States)

    Kohrt, Holbrook E.; Houot, Roch; Weiskopf, Kipp; Goldstein, Matthew J.; Scheeren, Ferenc; Czerwinski, Debra; Colevas, A. Dimitrios; Weng, Wen-Kai; Clarke, Michael F.; Carlson, Robert W.; Stockdale, Frank E.; Mollick, Joseph A.; Chen, Lieping; Levy, Ronald

    2012-01-01

    Trastuzumab, a monoclonal antibody targeting human epidermal growth factor receptor 2 (HER2; also known as HER-2/neu), is indicated for the treatment of women with either early stage or metastatic HER2+ breast cancer. It kills tumor cells by several mechanisms, including antibody-dependent cellular cytotoxicity (ADCC). Strategies that enhance the activity of ADCC effectors, including NK cells, may improve the efficacy of trastuzumab. Here, we have shown that upon encountering trastuzumab-coated, HER2-overexpressing breast cancer cells, human NK cells become activated and express the costimulatory receptor CD137. CD137 activation, which was dependent on NK cell expression of the FcγRIII receptor, occurred both in vitro and in the peripheral blood of women with HER2-expressing breast cancer after trastuzumab treatment. Stimulation of trastuzumab-activated human NK cells with an agonistic mAb specific for CD137 killed breast cancer cells (including an intrinsically trastuzumab-resistant cell line) more efficiently both in vitro and in vivo in xenotransplant models of human breast cancer, including one using a human primary breast tumor. The enhanced cytotoxicity was restricted to antibody-coated tumor cells. This sequential antibody strategy, combining a tumor-targeting antibody with a second antibody that activates the host innate immune system, may improve the therapeutic effects of antibodies against breast cancer and other HER2-expressing tumors. PMID:22326955

  1. [Expression of CD152 and CD137 on T regulatory cells in rhinitis and bronchial asthma patients].

    Science.gov (United States)

    Rojas-Ramos, Enrique; Martínez-Jiménez, Norma E; Verdejo-Hernández, Billy; Vázquez, Gilberto

    2015-01-01

    Antecedentes: la rinitis alérgica y el asma bronquial son enfermedades inflamatorias cuya prevalencia ha aumentado en las últimas dos décadas. Los estudios de la respuesta inflamatoria en estas enfermedades muestran que las células T reguladoras (Treg) participan en la tolerancia inmunológica. La expresión de CTLA-4 (CD152) se asocia con la actividad funcional de esta población y la expresión de 4-1BB (CD137) tiene un papel controvertido. Objetivo: determinar la existencia de CTLA-4 y 4-1BB en las células Treg en sangre periférica de pacientes con rinitis, asma, o ambas. Material y método: estudio transversal comparativo en el que se reclutaron tres grupos de pacientes: 20 sujetos con rinitis alérgica, 17 sujetos con asma bronquial y 17 con ambos padecimientos. Se formó un grupo control de 19 sujetos sanos. Se analizó la frecuencia de células Treg en sangre periférica y la expresión de CD152 y CD137 en los diferentes grupos mediante citometría de flujo. Resultados: se encontró menor frecuencia estadísticamente significativa de células Treg (CD4+, CD25altas y FoxP3+) en los grupos de sujetos con: asma bronquial (pasma bronquial (pasma bronquial únicamente tuvieron mayor frecuencia de células Treg que expresan CD152 (p asma bronquial y rinitis alérgica y asma bronquial únicamente tienen deficiencia de Treg CD4+, CD25altas y FoxP3+ en sangre periférica y los sujetos con asma bronquial tienen mayor frecuencia de células Treg que expresan CD152 y CD137.

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

    Science.gov (United States)

    Szekeres-Bartho, Julia; Halasz, Melinda; Palkovics, Tamas

    2009-12-01

    Progesterone is indispensable in creating a suitable endometrial environment for implantation, and also for the maintenance of pregnancy. Successful pregnancy depends on an appropriate maternal immune response to the fetus. Along with its endocrine effects, progesterone also acts as an "immunosteroid", by contributing to the establishment of a pregnancy protective immune milieu. Progesterone plays a role in uterine homing of NK cells and upregulates HLA-G gene expression, the ligand for NK inhibitory and activating receptors. At high concentrations, progesterone is a potent inducer of Th2-type cytokines as well as of LIF and M-CSF production by T cells. A protein called progesterone-induced blocking factor (PIBF), by inducing a Th2-dominant cytokine production mediates the immunological effects of progesterone. PIBF binds to a novel type of the IL-4 receptor and signals via the Jak/STAT pathway, to induce a number of genes, that not only affect the immune response, but might also play a role in trophoblast invasiveness. PMID:19880194

  3. Signal processing in the TGF-beta superfamily ligand-receptor network.

    Directory of Open Access Journals (Sweden)

    Jose M G Vilar

    2006-01-01

    Full Text Available The TGF-beta pathway plays a central role in tissue homeostasis and morphogenesis. It transduces a variety of extracellular signals into intracellular transcriptional responses that control a plethora of cellular processes, including cell growth, apoptosis, and differentiation. We use computational modeling to show that coupling of signaling with receptor trafficking results in a highly versatile signal-processing unit, able to sense by itself absolute levels of ligand, temporal changes in ligand concentration, and ratios of multiple ligands. This coupling controls whether the response of the receptor module is transient or permanent and whether or not different signaling channels behave independently of each other. Our computational approach unifies seemingly disparate experimental observations and suggests specific changes in receptor trafficking patterns that can lead to phenotypes that favor tumor progression.

  4. Mechanism of attenuation of leptin signaling under chronic ligand stimulation

    Directory of Open Access Journals (Sweden)

    Bamberg-Lemper Simone

    2010-01-01

    Full Text Available Abstract Background Leptin is an adipocyte-derived hormone that acts via its hypothalamic receptor (LEPRb to regulate energy balance. A downstream effect essential for the weight-regulatory action of leptin is the phosphorylation and activation of the latent transcription factor STAT3 by LEPRb-associated Janus kinases (JAKs. Obesity is typically associated with chronically elevated leptin levels and a decreased ability of LEPRb to activate intracellular signal transduction pathways (leptin resistance. Here we have studied the roles of the intracellular tyrosine residues in the negative feedback regulation of LEPRb-signaling under chronic leptin stimulation. Results Mutational analysis showed that the presence of either Tyr985 and Tyr1077 in the intracellular domain of LEPRb was sufficient for the attenuation of STAT3 phosphorylation, whereas mutation of both tyrosines rendered LEPRb resistant to feedback regulation. Overexpression and RNA interference-mediated downregulation of suppressor of cytokine signaling 3 (SOCS3 revealed that both Tyr985 and Tyr1077 were capable of supporting the negative modulatory effect of SOCS3 in reporter gene assays. In contrast, the inhibitory effect of SOCS1 was enhanced by the presence of Tyr985 but not Tyr1077. Finally, the reduction of the STAT-phosphorylating activity of the LEPRb complex after 2 h of leptin stimulation was not accompanied by the dephosphorylation or degradation of LEPRb or the receptor-associated JAK molecule, but depended on Tyr985 and/or Tyr1077. Conclusions Both Tyr985 and Tyr1077 contribute to the negative regulation of LEPRb signaling. The inhibitory effects of SOCS1 and SOCS3 differ in the dependence on the tyrosine residues in the intracellular domain of LEPRb.

  5. Cell surface receptors for signal transduction and ligand transport: a design principles study.

    Directory of Open Access Journals (Sweden)

    Harish Shankaran

    2007-06-01

    Full Text Available Receptors constitute the interface of cells to their external environment. These molecules bind specific ligands involved in multiple processes, such as signal transduction and nutrient transport. Although a variety of cell surface receptors undergo endocytosis, the systems-level design principles that govern the evolution of receptor trafficking dynamics are far from fully understood. We have constructed a generalized mathematical model of receptor-ligand binding and internalization to understand how receptor internalization dynamics encodes receptor function and regulation. A given signaling or transport receptor system represents a particular implementation of this module with a specific set of kinetic parameters. Parametric analysis of the response of receptor systems to ligand inputs reveals that receptor systems can be characterized as being: i avidity-controlled where the response control depends primarily on the extracellular ligand capture efficiency, ii consumption-controlled where the ability to internalize surface-bound ligand is the primary control parameter, and iii dual-sensitivity where both the avidity and consumption parameters are important. We show that the transferrin and low-density lipoprotein receptors are avidity-controlled, the vitellogenin receptor is consumption-controlled, and the epidermal growth factor receptor is a dual-sensitivity receptor. Significantly, we show that ligand-induced endocytosis is a mechanism to enhance the accuracy of signaling receptors rather than merely serving to attenuate signaling. Our analysis reveals that the location of a receptor system in the avidity-consumption parameter space can be used to understand both its function and its regulation.

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

    OpenAIRE

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

    2016-01-01

    The binding of epidermal growth factor (EGF) to EGF receptor (EGFR) stimulates cell mitogenesis and survival through various signalling cascades. EGF also stimulates rapid EGFR endocytosis and its eventual degradation in lysosomes. The immediate events induced by ligand binding include receptor dimerization, activation of intrinsic tyrosine kinase and autophosphorylation. However, in spite of intensified efforts, the results regarding the roles of these events in EGFR signalling and internali...

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Directory of Open Access Journals (Sweden)

    George Kourouniotis

    2016-07-01

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

  9. Cell surface receptors for signal transduction and ligand transport - a design principles study

    Energy Technology Data Exchange (ETDEWEB)

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

    2007-06-01

    Although many different receptors undergo endocytosis, the system-level design principles that govern the evolution of receptor dynamics are far from fully understood. We have constructed a generalized mathematical model to understand how receptor internalization dynamics encodes receptor function and regulation. Parametric analysis of the response of receptor systems to ligand inputs reveals that receptors can be categorized a being: i) avidity-controlled where the response control depends primarily on the extracelluar ligand capture efficiency, ii) consumption-controlled where the ability to internalize surface-bound ligand is the primary control parameter, and iii) dual-sensitivity where both the avidity and consumption parameters are important. We show that the transferrin and low-density lipoprotein receptors are avidity-controlled, the vitellogenin receptor is consumption-controlled and epidermal growth factor receptor is a dual-sensitivity receptor. Significantly, we show that ligand-induced endocytosis is a mechanism to anhance the accuracy of signaling receptors rather than serving to attenuate signaling. Our analysis reveals that the location of a receptor system in the avidity-consumption parameter space can be used to understand both its function and its regulations.

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

    OpenAIRE

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

    2012-01-01

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

  11. Distal Hydrogen-bonding Interactions in Ligand Sensing and Signaling by Mycobacterium tuberculosis DosS.

    Science.gov (United States)

    Basudhar, Debashree; Madrona, Yarrow; Yukl, Erik T; Sivaramakrishnan, Santhosh; Nishida, Clinton R; Moënne-Loccoz, Pierre; Ortiz de Montellano, Paul R

    2016-07-29

    Mycobacterium tuberculosis DosS is critical for the induction of M. tuberculosis dormancy genes in response to nitric oxide (NO), carbon monoxide (CO), or hypoxia. These environmental stimuli, which are sensed by the DosS heme group, result in autophosphorylation of a DosS His residue, followed by phosphotransfer to an Asp residue of the response regulator DosR. To clarify the mechanism of gaseous ligand recognition and signaling, we investigated the hydrogen-bonding interactions of the iron-bound CO and NO ligands by site-directed mutagenesis of Glu-87 and His-89. Autophosphorylation assays and molecular dynamics simulations suggest that Glu-87 has an important role in ligand recognition, whereas His-89 is essential for signal transduction to the kinase domain, a process for which Arg-204 is important. Mutation of Glu-87 to Ala or Gly rendered the protein constitutively active as a kinase, but with lower autophosphorylation activity than the wild-type in the Fe(II) and the Fe(II)-CO states, whereas the E87D mutant had little kinase activity except for the Fe(II)-NO complex. The H89R mutant exhibited attenuated autophosphorylation activity, although the H89A and R204A mutants were inactive as kinases, emphasizing the importance of these residues in communication to the kinase core. Resonance Raman spectroscopy of the wild-type and H89A mutant indicates the mutation does not alter the heme coordination number, spin state, or porphyrin deformation state, but it suggests that interdomain interactions are disrupted by the mutation. Overall, these results confirm the importance of the distal hydrogen-bonding network in ligand recognition and communication to the kinase domain and reveal the sensitivity of the system to subtle differences in the binding of gaseous ligands. PMID:27235395

  12. Small Molecule Inhibition of Ligand-Stimulated RAGE-DIAPH1 Signal Transduction

    Science.gov (United States)

    Manigrasso, Michaele B.; Pan, Jinhong; Rai, Vivek; Zhang, Jinghua; Reverdatto, Sergey; Quadri, Nosirudeen; DeVita, Robert J.; Ramasamy, Ravichandran; Shekhtman, Alexander; Schmidt, Ann Marie

    2016-01-01

    The receptor for advanced glycation endproducts (RAGE) binds diverse ligands linked to chronic inflammation and disease. NMR spectroscopy and x-ray crystallization studies of the extracellular domains of RAGE indicate that RAGE ligands bind by distinct charge- and hydrophobicity-dependent mechanisms. The cytoplasmic tail (ct) of RAGE is essential for RAGE ligand-mediated signal transduction and consequent modulation of gene expression and cellular properties. RAGE signaling requires interaction of ctRAGE with the intracellular effector, mammalian diaphanous 1 or DIAPH1. We screened a library of 58,000 small molecules and identified 13 small molecule competitive inhibitors of ctRAGE interaction with DIAPH1. These compounds, which exhibit in vitro and in vivo inhibition of RAGE-dependent molecular processes, present attractive molecular scaffolds for the development of therapeutics against RAGE-mediated diseases, such as those linked to diabetic complications, Alzheimer’s disease, and chronic inflammation, and provide support for the feasibility of inhibition of protein-protein interaction (PPI). PMID:26936329

  13. Ligand-Occupied Integrin Internalization Links Nutrient Signaling to Invasive Migration

    Directory of Open Access Journals (Sweden)

    Elena Rainero

    2015-01-01

    Full Text Available Integrin trafficking is key to cell migration, but little is known about the spatiotemporal organization of integrin endocytosis. Here, we show that α5β1 integrin undergoes tensin-dependent centripetal movement from the cell periphery to populate adhesions located under the nucleus. From here, ligand-engaged α5β1 integrins are internalized under control of the Arf subfamily GTPase, Arf4, and are trafficked to nearby late endosomes/lysosomes. Suppression of centripetal movement or Arf4-dependent endocytosis disrupts flow of ligand-bound integrins to late endosomes/lysosomes and their degradation within this compartment. Arf4-dependent integrin internalization is required for proper lysosome positioning and for recruitment and activation of mTOR at this cellular subcompartment. Furthermore, nutrient depletion promotes subnuclear accumulation and endocytosis of ligand-engaged α5β1 integrins via inhibition of mTORC1. This two-way regulatory interaction between mTORC1 and integrin trafficking in combination with data describing a role for tensin in invasive cell migration indicate interesting links between nutrient signaling and metastasis.

  14. High density and ligand affinity confer ultrasensitive signal detection by a guanylyl cyclase chemoreceptor.

    Science.gov (United States)

    Pichlo, Magdalena; Bungert-Plümke, Stefanie; Weyand, Ingo; Seifert, Reinhard; Bönigk, Wolfgang; Strünker, Timo; Kashikar, Nachiket Dilip; Goodwin, Normann; Müller, Astrid; Pelzer, Patric; Van, Qui; Enderlein, Jörg; Klemm, Clementine; Krause, Eberhard; Trötschel, Christian; Poetsch, Ansgar; Kremmer, Elisabeth; Kaupp, U Benjamin; Körschen, Heinz G; Collienne, Ursel

    2014-08-18

    Guanylyl cyclases (GCs), which synthesize the messenger cyclic guanosine 3',5'-monophosphate, control several sensory functions, such as phototransduction, chemosensation, and thermosensation, in many species from worms to mammals. The GC chemoreceptor in sea urchin sperm can decode chemoattractant concentrations with single-molecule sensitivity. The molecular and cellular underpinnings of such ultrasensitivity are not known for any eukaryotic chemoreceptor. In this paper, we show that an exquisitely high density of 3 × 10(5) GC chemoreceptors and subnanomolar ligand affinity provide a high ligand-capture efficacy and render sperm perfect absorbers. The GC activity is terminated within 150 ms by dephosphorylation steps of the receptor, which provides a means for precise control of the GC lifetime and which reduces "molecule noise." Compared with other ultrasensitive sensory systems, the 10-fold signal amplification by the GC receptor is surprisingly low. The hallmarks of this signaling mechanism provide a blueprint for chemical sensing in small compartments, such as olfactory cilia, insect antennae, or even synaptic boutons.

  15. High density and ligand affinity confer ultrasensitive signal detection by a guanylyl cyclase chemoreceptor.

    Science.gov (United States)

    Pichlo, Magdalena; Bungert-Plümke, Stefanie; Weyand, Ingo; Seifert, Reinhard; Bönigk, Wolfgang; Strünker, Timo; Kashikar, Nachiket Dilip; Goodwin, Normann; Müller, Astrid; Pelzer, Patric; Van, Qui; Enderlein, Jörg; Klemm, Clementine; Krause, Eberhard; Trötschel, Christian; Poetsch, Ansgar; Kremmer, Elisabeth; Kaupp, U Benjamin; Körschen, Heinz G; Collienne, Ursel

    2014-08-18

    Guanylyl cyclases (GCs), which synthesize the messenger cyclic guanosine 3',5'-monophosphate, control several sensory functions, such as phototransduction, chemosensation, and thermosensation, in many species from worms to mammals. The GC chemoreceptor in sea urchin sperm can decode chemoattractant concentrations with single-molecule sensitivity. The molecular and cellular underpinnings of such ultrasensitivity are not known for any eukaryotic chemoreceptor. In this paper, we show that an exquisitely high density of 3 × 10(5) GC chemoreceptors and subnanomolar ligand affinity provide a high ligand-capture efficacy and render sperm perfect absorbers. The GC activity is terminated within 150 ms by dephosphorylation steps of the receptor, which provides a means for precise control of the GC lifetime and which reduces "molecule noise." Compared with other ultrasensitive sensory systems, the 10-fold signal amplification by the GC receptor is surprisingly low. The hallmarks of this signaling mechanism provide a blueprint for chemical sensing in small compartments, such as olfactory cilia, insect antennae, or even synaptic boutons. PMID:25135936

  16. A biased ligand for OXE-R uncouples Gα and Gβγ signaling within a heterotrimer

    DEFF Research Database (Denmark)

    Blättermann, Stefanie; Peters, Lucas; Ottersbach, Philipp Aaron;

    2012-01-01

    Differential targeting of heterotrimeric G protein versus β-arrestin signaling are emerging concepts in G protein-coupled receptor (GPCR) research and drug discovery, and biased engagement by GPCR ligands of either β-arrestin or G protein pathways has been disclosed. Herein we report on a new...... mechanism of ligand bias to titrate the signaling specificity of a cell-surface GPCR. Using a combination of biomolecular and virtual screening, we identified the small-molecule modulator Gue1654, which inhibits Gβγ but not Gα signaling triggered upon activation of Gα(i)-βγ by the chemoattractant receptor...... OXE-R in both recombinant and human primary cells. Gue1654 does not interfere nonspecifically with signaling directly at or downstream of Gβγ. This hitherto unappreciated mechanism of ligand bias at a GPCR highlights both a new paradigm for functional selectivity and a potentially new strategy...

  17. Statistical removal of background signals from high-throughput 1H NMR line-broadening ligand-affinity screens

    International Nuclear Information System (INIS)

    NMR ligand-affinity screens are vital to drug discovery, are routinely used to screen fragment-based libraries, and used to verify chemical leads from high-throughput assays and virtual screens. NMR ligand-affinity screens are also a highly informative first step towards identifying functional epitopes of unknown proteins, as well as elucidating the biochemical functions of protein–ligand interaction at their binding interfaces. While simple one-dimensional 1H NMR experiments are capable of indicating binding through a change in ligand line shape, they are plagued by broad, ill-defined background signals from protein 1H resonances. We present an uncomplicated method for subtraction of protein background in high-throughput ligand-based affinity screens, and show that its performance is maximized when phase-scatter correction is applied prior to subtraction

  18. Statistical removal of background signals from high-throughput {sup 1}H NMR line-broadening ligand-affinity screens

    Energy Technology Data Exchange (ETDEWEB)

    Worley, Bradley; Sisco, Nicholas J.; Powers, Robert, E-mail: rpowers3@unl.edu [University of Nebraska-Lincoln, Department of Chemistry (United States)

    2015-09-15

    NMR ligand-affinity screens are vital to drug discovery, are routinely used to screen fragment-based libraries, and used to verify chemical leads from high-throughput assays and virtual screens. NMR ligand-affinity screens are also a highly informative first step towards identifying functional epitopes of unknown proteins, as well as elucidating the biochemical functions of protein–ligand interaction at their binding interfaces. While simple one-dimensional {sup 1}H NMR experiments are capable of indicating binding through a change in ligand line shape, they are plagued by broad, ill-defined background signals from protein {sup 1}H resonances. We present an uncomplicated method for subtraction of protein background in high-throughput ligand-based affinity screens, and show that its performance is maximized when phase-scatter correction is applied prior to subtraction.

  19. ASIC3 Channels Integrate Agmatine and Multiple Inflammatory Signals through the Nonproton Ligand Sensing Domain

    Directory of Open Access Journals (Sweden)

    Cao Hui

    2010-12-01

    Full Text Available Abstract Background Acid-sensing ion channels (ASICs have long been known to sense extracellular protons and contribute to sensory perception. Peripheral ASIC3 channels represent natural sensors of acidic and inflammatory pain. We recently reported the use of a synthetic compound, 2-guanidine-4-methylquinazoline (GMQ, to identify a novel nonproton sensing domain in the ASIC3 channel, and proposed that, based on its structural similarity with GMQ, the arginine metabolite agmatine (AGM may be an endogenous nonproton ligand for ASIC3 channels. Results Here, we present further evidence for the physiological correlation between AGM and ASIC3. Among arginine metabolites, only AGM and its analog arcaine (ARC activated ASIC3 channels at neutral pH in a sustained manner similar to GMQ. In addition to the homomeric ASIC3 channels, AGM also activated heteromeric ASIC3 plus ASIC1b channels, extending its potential physiological relevance. Importantly, the process of activation by AGM was highly sensitive to mild acidosis, hyperosmolarity, arachidonic acid (AA, lactic acid and reduced extracellular Ca2+. AGM-induced ASIC3 channel activation was not through the chelation of extracellular Ca2+ as occurs with increased lactate, but rather through a direct interaction with the newly identified nonproton ligand sensing domain. Finally, AGM cooperated with the multiple inflammatory signals to cause pain-related behaviors in an ASIC3-dependent manner. Conclusions Nonproton ligand sensing domain might represent a novel mechanism for activation or sensitization of ASIC3 channels underlying inflammatory pain-sensing under in vivo conditions.

  20. Ligand-dependent inhibition of beta-catenin/TCF signaling by androgen receptor.

    Science.gov (United States)

    Chesire, Dennis R; Isaacs, William B

    2002-12-01

    Beta-catenin signaling may contribute to prostate cancer (CaP) progression. Although beta-catenin is known to upregulate T cell factor (TCF) target gene expression in CaP cells, recent evidence demonstrates its capacity to enhance ligand-dependent androgen receptor (AR) function. Thus, we wished to further understand the interaction between these two pathways. We find in both CaP cells (CWR22-Rv1, LAPC-4, DU145) and non-CaP cells (HEK-293, TSU, SW480, HCT-116) that beta-catenin/TCF-related transcription (CRT), as measured by activation of a synthetic promoter and that of cyclin D1, is inhibited by androgen treatment. This inhibition is AR-dependent, as it only occurs in cells expressing AR endogenously or transiently, and is abrogated by AR antagonists. Additional analyses convey that the ligand-dependent nature of CRT suppression depends on transactivation-competent AR in the nucleus, but not on indirect effects stemming from AR target gene expression. Given the recent work identifying an AR/beta-catenin interaction, and from our finding that liganded AR does not prompt gross changes in the constitutive nuclear localization of TCF4 or mutant beta-catenin, we hypothesized that transcription factor (i.e. AR and TCF) competition for beta-catenin recruitment may explain, in part, androgen-induced suppression of CRT. To address this idea, we expressed an AR mutant lacking its DNA-binding domain (DBD). This receptor could not orchestrate ligand-dependent CRT repression, thereby providing support for those recent data implicating the AR DBD/LBD as necessary for beta-catenin interaction. Further supporting this hypothesis, TCF/LEF over-expression counteracts androgen-induced suppression of CRT, and requires beta-catenin binding activity to do so. Interestingly, TCF4 over-expression potently antagonizes AR function; however, this inhibition may occur independently of beta-catenin/TCF4 interaction. These results from TCF4 over-expression analyses, taken together, provide

  1. Signal transduction pathways in the pentameric ligand-gated ion channels.

    Directory of Open Access Journals (Sweden)

    David Mowrey

    Full Text Available The mechanisms of allosteric action within pentameric ligand-gated ion channels (pLGICs remain to be determined. Using crystallography, site-directed mutagenesis, and two-electrode voltage clamp measurements, we identified two functionally relevant sites in the extracellular (EC domain of the bacterial pLGIC from Gloeobacter violaceus (GLIC. One site is at the C-loop region, where the NQN mutation (D91N, E177Q, and D178N eliminated inter-subunit salt bridges in the open-channel GLIC structure and thereby shifted the channel activation to a higher agonist concentration. The other site is below the C-loop, where binding of the anesthetic ketamine inhibited GLIC currents in a concentration dependent manner. To understand how a perturbation signal in the EC domain, either resulting from the NQN mutation or ketamine binding, is transduced to the channel gate, we have used the Perturbation-based Markovian Transmission (PMT model to determine dynamic responses of the GLIC channel and signaling pathways upon initial perturbations in the EC domain of GLIC. Despite the existence of many possible routes for the initial perturbation signal to reach the channel gate, the PMT model in combination with Yen's algorithm revealed that perturbation signals with the highest probability flow travel either via the β1-β2 loop or through pre-TM1. The β1-β2 loop occurs in either intra- or inter-subunit pathways, while pre-TM1 occurs exclusively in inter-subunit pathways. Residues involved in both types of pathways are well supported by previous experimental data on nAChR. The direct coupling between pre-TM1 and TM2 of the adjacent subunit adds new insight into the allosteric signaling mechanism in pLGICs.

  2. Immunostimulation by OX40 Ligand Transgenic Ewing Sarcoma Cells.

    Science.gov (United States)

    Reuter, Dajana; Staege, Martin S; Kühnöl, Caspar D; Föll, Jürgen

    2015-01-01

    Interleukin-2 (IL-2) transgenic Ewing sarcoma cells can induce tumor specific T and NK cell responses and reduce tumor growth in vivo and in vitro. Nevertheless, the efficiency of this stimulation is not high enough to inhibit tumor growth completely. In addition to recognition of the cognate antigen, optimal T-cell stimulation requires signals from so-called co-stimulatory molecules. Several members of the tumor necrosis factor superfamily have been identified as co-stimulatory molecules that can augment antitumor immune responses. OX40 (CD134) and OX40 ligand (OX40L = CD252; also known as tumor necrosis factor ligand family member 4) is one example of such receptor/ligand pair with co-stimulatory function. In the present investigation, we generated OX40L transgenic Ewing sarcoma cells and tested their immunostimulatory activity in vitro. OX40L transgenic Ewing sarcoma cells showed preserved expression of Ewing sarcoma-associated (anti)gens including lipase member I, cyclin D1 (CCND1), cytochrome P450 family member 26B1 (CYP26B1), and the Ewing sarcoma breakpoint region 1-friend leukemia virus integration 1 (EWSR1-FLI1) oncogene. OX40L-expressing tumor cells showed a trend for enhanced immune stimulation against Ewing sarcoma cells in combination with IL-2 and stimulation of CD137. Our data suggest that inclusion of the OX40/OX40L pathway of co-stimulation might improve immunotherapy strategies for the treatment of Ewing sarcoma. PMID:26579494

  3. Heterologous desensitization of T cell functions by CCR5 and CXCR4 ligands: inhibition of cellular signaling, adhesion and chemotaxis.

    Science.gov (United States)

    Hecht, Iris; Cahalon, Liora; Hershkoviz, Rami; Lahat, Adi; Franitza, Suzanne; Lider, Ofer

    2003-01-01

    T cells migrate into inflamed sites through the extracellular matrix (ECM) in response to chemotactic areas and are then simultaneously or sequentially exposed to multiple chemotactic ligands. We examined the responses of human peripheral blood T cells, present in an ECM-like context, to combinatorial signaling transduced by SDF-1alpha (CXCL12), and two CCR5 ligands, RANTES (CCL5) and MIP-1beta (CCL4). Separately, these chemokines, at G protein-coupled receptor (GPCR)-stimulating concentrations, induced T cell adhesion to fibronectin (FN) and T cell chemotaxis. However, the pro-adhesive and pro-migratory capacities of SDF-1alpha and RANTES or MIP-1beta were mutually suppressed by the simultaneous or sequential exposure of the cells to these CCR5 or CXCR4 ligands. This cross-talk did not involve the internalization of the SDF-1alpha receptor, CXCR4, but rather, a decrease in phosphorylation of ERK and Pyk-2, as well as inhibition of Ca(2+) mobilization. Strikingly, early CXCR4 signaling of phosphatidylinositol-3-kinase, detected by SDF-1alpha-induced AKT phosphorylation, was insensitive to RANTES-CCR5 signals. Accordingly, early chemotaxis to SDF-1alpha was not susceptible to CCR5 occupancy, whereas late stages of T cell chemotaxis were markedly down-regulated. This is an example of a specialized functional desensitization of heterologous chemokine receptors that induces GPCR interference with T cell adhesion to ECM ligands and chemotaxis within chemokine-rich extravascular contexts. PMID:12502723

  4. Engineering the melanocortin-4 receptor to control constitutive and ligand-mediated G(S signaling in vivo.

    Directory of Open Access Journals (Sweden)

    Supriya Srinivasan

    Full Text Available The molecular and functional diversity of G protein-coupled receptors is essential to many physiological processes. However, this diversity presents a significant challenge to understanding the G protein-mediated signaling events that underlie a specific physiological response. To increase our understanding of these processes, we sought to gain control of the timing and specificity of G(s signaling in vivo. We used naturally occurring human mutations to develop two G(s-coupled engineered receptors that respond solely to a synthetic ligand (RASSLs. Our G(s-coupled RASSLs are based on the melanocortin-4 receptor, a centrally expressed receptor that plays an important role in the regulation of body weight. These RASSLs are not activated by the endogenous hormone alpha-melanocyte-stimulating hormone but respond potently to a selective synthetic ligand, tetrahydroisoquinoline. The RASSL variants reported here differ in their intrinsic basal activities, allowing the separation of the effects of basal signaling from ligand-mediated activation of the G(s pathway in vivo. These RASSLs can be used to activate G(s signaling in any tissue, but would be particularly useful for analyzing downstream events that mediate body weight regulation in mice. Our study also demonstrates the use of human genetic variation for protein engineering.

  5. Structural insights of homotypic interaction domains in the ligand-receptor signal transduction of tumor necrosis factor (TNF)

    Science.gov (United States)

    Park, Young-Hoon; Jeong, Mi Suk; Jang, Se Bok

    2016-01-01

    Several members of tumor necrosis factor receptor (TNFR) superfamily that these members activate caspase-8 from death-inducing signaling complex (DISC) in TNF ligand-receptor signal transduction have been identified. In the extrinsic pathway, apoptotic signal transduction is induced in death domain (DD) superfamily; it consists of a hexahelical bundle that contains 80 amino acids. The DD superfamily includes about 100 members that belong to four subfamilies: death domain (DD), caspase recruitment domain (CARD), pyrin domain (PYD), and death effector domain (DED). This superfamily contains key building blocks: with these blocks, multimeric complexes are formed through homotypic interactions. Furthermore, each DD-binding event occurs exclusively. The DD superfamily regulates the balance between death and survival of cells. In this study, the structures, functions, and unique features of DD superfamily members are compared with their complexes. By elucidating structural insights of DD superfamily members, we investigate the interaction mechanisms of DD domains; these domains are involved in TNF ligand-receptor signaling. These DD superfamily members play a pivotal role in the development of more specific treatments of cancer. [BMB Reports 2016; 49(3): 159-166] PMID:26615973

  6. Chemogenomic analysis of G-protein coupled receptors and their ligands deciphers locks and keys governing diverse aspects of signalling.

    Directory of Open Access Journals (Sweden)

    Jörg D Wichard

    Full Text Available Understanding the molecular mechanism of signalling in the important super-family of G-protein-coupled receptors (GPCRs is causally related to questions of how and where these receptors can be activated or inhibited. In this context, it is of great interest to unravel the common molecular features of GPCRs as well as those related to an active or inactive state or to subtype specific G-protein coupling. In our underlying chemogenomics study, we analyse for the first time the statistical link between the properties of G-protein-coupled receptors and GPCR ligands. The technique of mutual information (MI is able to reveal statistical inter-dependence between variations in amino acid residues on the one hand and variations in ligand molecular descriptors on the other. Although this MI analysis uses novel information that differs from the results of known site-directed mutagenesis studies or published GPCR crystal structures, the method is capable of identifying the well-known common ligand binding region of GPCRs between the upper part of the seven transmembrane helices and the second extracellular loop. The analysis shows amino acid positions that are sensitive to either stimulating (agonistic or inhibitory (antagonistic ligand effects or both. It appears that amino acid positions for antagonistic and agonistic effects are both concentrated around the extracellular region, but selective agonistic effects are cumulated between transmembrane helices (TMHs 2, 3, and ECL2, while selective residues for antagonistic effects are located at the top of helices 5 and 6. Above all, the MI analysis provides detailed indications about amino acids located in the transmembrane region of these receptors that determine G-protein signalling pathway preferences.

  7. Immuno-stimulation by OX40 ligand transgenic Ewing sarcoma cells

    Directory of Open Access Journals (Sweden)

    Dajana eReuter

    2015-10-01

    Full Text Available Interleukin-2 (IL-2 transgenic Ewing sarcoma cells can induce tumor specific T and NK cell responses and reduce tumor growth in vivo and in vitro. Nevertheless, the efficiency of this stimulation is not high enough to inhibit tumor growth completely. In addition to recognition of the cognate antigen, optimal T cell stimulation requires signals from so-called co-stimulatory molecules. Several members of the tumor necrosis factor superfamily (TNFSF have been identified as co-stimulatory molecules that can augment anti-tumor immune responses. OX40 (CD134 and OX40 ligand (OX40L = CD252; also known as tumor necrosis factor ligand family member 4 is one example for such receptor/ligand pair with co-stimulatory function. In the present investigation we generated OX40L transgenic Ewing sarcoma cells and tested their immuno-stimulatory activity in vitro. OX40L transgenic Ewing sarcoma cells showed preserved expression of Ewing sarcoma associated (antigens including lipase member I (LIPI, cyclin D1 (CCND1, cytochrome P450 family member 26B1 (CYP26B1 and the Ewing sarcoma breakpoint region 1-friend leukemia virus integration 1 (EWSR1-FLI1 oncogene. OX40L expressing tumor cells showed a trend for enhanced immune stimulation against Ewing sarcoma cells in combination with IL-2 and stimulation of CD137. Our data suggest that inclusion of the OX40/OX40L pathway of co-stimulation might improve immunotherapy strategies for treatment of Ewing sarcoma.

  8. Signal Transduction at the Domain Interface of Prokaryotic Pentameric Ligand-Gated Ion Channels

    Science.gov (United States)

    Bertozzi, Carlo; Zimmermann, Iwan; Engeler, Sibylle; Hilf, Ricarda J. C.; Dutzler, Raimund

    2016-01-01

    Pentameric ligand-gated ion channels are activated by the binding of agonists to a site distant from the ion conduction path. These membrane proteins consist of distinct ligand-binding and pore domains that interact via an extended interface. Here, we have investigated the role of residues at this interface for channel activation to define critical interactions that couple conformational changes between the two structural units. By characterizing point mutants of the prokaryotic channels ELIC and GLIC by electrophysiology, X-ray crystallography and isothermal titration calorimetry, we have identified conserved residues that, upon mutation, apparently prevent activation but not ligand binding. The positions of nonactivating mutants cluster at a loop within the extracellular domain connecting β-strands 6 and 7 and at a loop joining the pore-forming helix M2 with M3 where they contribute to a densely packed core of the protein. An ionic interaction in the extracellular domain between the turn connecting β-strands 1 and 2 and a residue at the end of β-strand 10 stabilizes a state of the receptor with high affinity for agonists, whereas contacts of this turn to a conserved proline residue in the M2-M3 loop appear to be less important than previously anticipated. When mapping residues with strong functional phenotype on different channel structures, mutual distances are closer in conducting than in nonconducting conformations, consistent with a potential role of contacts in the stabilization of the open state. Our study has revealed a pattern of interactions that are crucial for the relay of conformational changes from the extracellular domain to the pore region of prokaryotic pentameric ligand-gated ion channels. Due to the strong conservation of the interface, these results are relevant for the entire family. PMID:26943937

  9. Comparative analysis of species-specific ligand recognition in Toll-like receptor 8 signaling: a hypothesis.

    Directory of Open Access Journals (Sweden)

    Rajiv Gandhi Govindaraj

    Full Text Available Toll-like receptors (TLRs play a central role in the innate immune response by recognizing conserved structural patterns in a variety of microbes. TLRs are classified into six families, of which TLR7 family members include TLR7, 8, and 9, which are localized to endolysosomal compartments recognizing viral infection in the form of foreign nucleic acids. In our current study, we focused on TLR8, which has been shown to recognize different types of ligands such as viral or bacterial ssRNA as well as small synthetic molecules. The primary sequences of rodent and non-rodent TLR8s are similar, but the antiviral compound (R848 that activates the TLR8 pathway is species-specific. Moreover, the factors underlying the receptor's species-specificity remain unknown. To this end, comparative homology modeling, molecular dynamics simulations refinement, automated docking and computational mutagenesis studies were employed to probe the intermolecular interactions between this anti-viral compound and TLR8. Furthermore, comparative analyses of modeled TLR8 (rodent and non-rodent structures have shown that the variation mainly occurs at LRR14-15 (undefined region; hence, we hypothesized that this variation may be the primary reason for the exhibited species-specificity. Our hypothesis was further bolstered by our docking studies, which clearly showed that this undefined region was in close proximity to the ligand-binding site and thus may play a key role in ligand recognition. In addition, the interface between the ligand and TLR8s varied depending upon the amino acid charges, free energy of binding, and interaction surface. Therefore, our current work provides a hypothesis for previous in vivo studies in the context of TLR signaling.

  10. Examining the critical roles of human CB2 receptor residues Valine 3.32 (113) and Leucine 5.41 (192) in ligand recognition and downstream signaling activities.

    Science.gov (United States)

    Alqarni, Mohammed; Myint, Kyaw Zeyar; Tong, Qin; Yang, Peng; Bartlow, Patrick; Wang, Lirong; Feng, Rentian; Xie, Xiang-Qun

    2014-09-26

    We performed molecular modeling and docking to predict a putative binding pocket and associated ligand-receptor interactions for human cannabinoid receptor 2 (CB2). Our data showed that two hydrophobic residues came in close contact with three structurally distinct CB2 ligands: CP-55,940, SR144528 and XIE95-26. Site-directed mutagenesis experiments and subsequent functional assays implicated the roles of Valine residue at position 3.32 (V113) and Leucine residue at position 5.41 (L192) in the ligand binding function and downstream signaling activities of the CB2 receptor. Four different point mutations were introduced to the wild type CB2 receptor: V113E, V113L, L192S and L192A. Our results showed that mutation of Val113 with a Glutamic acid and Leu192 with a Serine led to the complete loss of CB2 ligand binding as well as downstream signaling activities. Substitution of these residues with those that have similar hydrophobic side chains such as Leucine (V113L) and Alanine (L192A), however, allowed CB2 to retain both its ligand binding and signaling functions. Our modeling results validated by competition binding and site-directed mutagenesis experiments suggest that residues V113 and L192 play important roles in ligand binding and downstream signaling transduction of the CB2 receptor.

  11. Cell Signaling in Tenocytes: Response to Load and Ligands in Health and Disease.

    Science.gov (United States)

    Wall, Michelle E; Dyment, Nathaniel A; Bodle, Josie; Volmer, Jon; Loboa, Elizabeth; Cederlund, Anna; Fox, Ann M; Banes, Albert J

    2016-01-01

    Signaling in tenocytes during development, homeostasis and injury involves multiple and redundant pathways. Given that tendons transmit mechanical forces from muscle to bone to effect movement, a key function for tenocytes is the detection of and response to mechanical stimulation. Mechanotransduction involves matrix-integrin-cytoskeleton to nucleus signaling, gap junction intercellular communication, changes in intracellular calcium (Ca(2+)), activation of receptors and their pathways, and responses to biochemical factors such as hormones, growth factors, adenosine triphosphate (ATP) and its derivatives, and neuromodulators. The primary cilium also plays a key role in the detection of mechanical signals. During development, transforming growth factor-β (TGF-β), bone morphogenetic protein (BMP), and hedgehog (Hh) signaling modulate tendon differentiation and formation. The response to injury is complex and varied involving not only inflammatory mediators such as interleukin-1β but also mechanosensing. This chapter reviews the signaling pathways tenocytes use during mechanotransduction, development and in response to injury. PMID:27535250

  12. Reduced retinoid signaling in the skin after systemic retinoid-X receptor ligand treatment in mice with potential relevance for skin disorders.

    Science.gov (United States)

    Mihály, Johanna; Gericke, Janine; Aydemir, Gamze; Weiss, Kathrin; Carlsen, Harald; Blomhoff, Rune; Garcia, Javier; Rühl, Ralph

    2012-01-01

    Retinoid-X receptor (RXR)- and retinoic acid receptor (RAR)-mediated signaling is induced by retinoic acids (RA), which are involved in the regulation of skin permeability, differentiation and immune response. Dysregulation of retinoid signaling is present in various skin disorders. Topically and systemically administered synthetic RAR or RXR agonists might influence retinoid-mediated signaling in the skin of RARE reporter animals and gene expression analysis for retinoid, skin homeostasis and skin inflammation marker genes and local retinoid concentrations. Mice were treated orally and topically with synthetic ligands and bioimaging, QRT-PCR and retinoid analysis were performed. Topical application of the synthetic RAR ligand AM580 significantly enhanced retinoid signaling in skin while topical application of the RXR ligand LG268 did not influence retinoic acid receptor response elements (RARE)-mediated signaling. Systemic treatments with LG268 decreased the expression of genes involved in skin homeostasis, RA synthesis and skin RA concentrations, while it increased various markers for skin inflammation and RA degradation, which corresponds to decreased skin RARE signaling. We conclude from these observations that increased systemic concentrations of an RXR -ligand may be one reason for reduced retinoid signaling, -reduced all-trans RA levels in the skin, reduced epidermal homeostasis and increased skin inflammation marker expression with potential relevance for various skin disorders, like atopic dermatitis.

  13. Matched sizes of activating and inhibitory receptor/ligand pairs are required for optimal signal integration by human natural killer cells.

    Directory of Open Access Journals (Sweden)

    Karsten Köhler

    Full Text Available It has been suggested that receptor-ligand complexes segregate or co-localise within immune synapses according to their size, and this is important for receptor signaling. Here, we set out to test the importance of receptor-ligand complex dimensions for immune surveillance of target cells by human Natural Killer (NK cells. NK cell activation is regulated by integrating signals from activating receptors, such as NKG2D, and inhibitory receptors, such as KIR2DL1. Elongating the NKG2D ligand MICA reduced its ability to trigger NK cell activation. Conversely, elongation of KIR2DL1 ligand HLA-C reduced its ability to inhibit NK cells. Whereas normal-sized HLA-C was most effective at inhibiting activation by normal-length MICA, only elongated HLA-C could inhibit activation by elongated MICA. Moreover, HLA-C and MICA that were matched in size co-localised, whereas HLA-C and MICA that were different in size were segregated. These results demonstrate that receptor-ligand dimensions are important in NK cell recognition, and suggest that optimal integration of activating and inhibitory receptor signals requires the receptor-ligand complexes to have similar dimensions.

  14. A Saccharomyces cerevisiae Assay System to Investigate Ligand/AdipoR1 Interactions That Lead to Cellular Signaling

    KAUST Repository

    Aouida, Mustapha

    2013-06-07

    Adiponectin is a mammalian hormone that exerts anti-diabetic, anti-cancer and cardioprotective effects through interaction with its major ubiquitously expressed plasma membrane localized receptors, AdipoR1 and AdipoR2. Here, we report a Saccharomyces cerevisiae based method for investigating agonist-AdipoR interactions that is amenable for high-throughput scale-up and can be used to study both AdipoRs separately. Agonist-AdipoR1 interactions are detected using a split firefly luciferase assay based on reconstitution of firefly luciferase (Luc) activity due to juxtaposition of its N- and C-terminal fragments, NLuc and CLuc, by ligand induced interaction of the chimeric proteins CLuc-AdipoR1 and APPL1-NLuc (adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif 1-NLuc) in a S. cerevisiae strain lacking the yeast homolog of AdipoRs (Izh2p). The assay monitors the earliest known step in the adiponectin-AdipoR anti-diabetic signaling cascade. We demonstrate that reconstituted Luc activity can be detected in colonies or cells using a CCD camera and quantified in cell suspensions using a microplate reader. AdipoR1-APPL1 interaction occurs in absence of ligand but can be stimulated specifically by agonists such as adiponectin and the tobacco protein osmotin that was shown to have AdipoR-dependent adiponectin-like biological activity in mammalian cells. To further validate this assay, we have modeled the three dimensional structures of receptor-ligand complexes of membrane-embedded AdipoR1 with cyclic peptides derived from osmotin or osmotin-like plant proteins. We demonstrate that the calculated AdipoR1-peptide binding energies correlate with the peptides\\' ability to behave as AdipoR1 agonists in the split luciferase assay. Further, we demonstrate agonist-AdipoR dependent activation of protein kinase A (PKA) signaling and AMP activated protein kinase (AMPK) phosphorylation in S. cerevisiae, which are homologous to

  15. A Saccharomyces cerevisiae assay system to investigate ligand/AdipoR1 interactions that lead to cellular signaling.

    Directory of Open Access Journals (Sweden)

    Mustapha Aouida

    Full Text Available Adiponectin is a mammalian hormone that exerts anti-diabetic, anti-cancer and cardioprotective effects through interaction with its major ubiquitously expressed plasma membrane localized receptors, AdipoR1 and AdipoR2. Here, we report a Saccharomyces cerevisiae based method for investigating agonist-AdipoR interactions that is amenable for high-throughput scale-up and can be used to study both AdipoRs separately. Agonist-AdipoR1 interactions are detected using a split firefly luciferase assay based on reconstitution of firefly luciferase (Luc activity due to juxtaposition of its N- and C-terminal fragments, NLuc and CLuc, by ligand induced interaction of the chimeric proteins CLuc-AdipoR1 and APPL1-NLuc (adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif 1-NLuc in a S. cerevisiae strain lacking the yeast homolog of AdipoRs (Izh2p. The assay monitors the earliest known step in the adiponectin-AdipoR anti-diabetic signaling cascade. We demonstrate that reconstituted Luc activity can be detected in colonies or cells using a CCD camera and quantified in cell suspensions using a microplate reader. AdipoR1-APPL1 interaction occurs in absence of ligand but can be stimulated specifically by agonists such as adiponectin and the tobacco protein osmotin that was shown to have AdipoR-dependent adiponectin-like biological activity in mammalian cells. To further validate this assay, we have modeled the three dimensional structures of receptor-ligand complexes of membrane-embedded AdipoR1 with cyclic peptides derived from osmotin or osmotin-like plant proteins. We demonstrate that the calculated AdipoR1-peptide binding energies correlate with the peptides' ability to behave as AdipoR1 agonists in the split luciferase assay. Further, we demonstrate agonist-AdipoR dependent activation of protein kinase A (PKA signaling and AMP activated protein kinase (AMPK phosphorylation in S. cerevisiae, which are

  16. Modeling signal propagation mechanisms and ligand-based conformational dynamics of the Hsp90 molecular chaperone full-length dimer.

    Directory of Open Access Journals (Sweden)

    Giulia Morra

    2009-03-01

    Full Text Available Hsp90 is a molecular chaperone essential for protein folding and activation in normal homeostasis and stress response. ATP binding and hydrolysis facilitate Hsp90 conformational changes required for client activation. Hsp90 plays an important role in disease states, particularly in cancer, where chaperoning of the mutated and overexpressed oncoproteins is important for function. Recent studies have illuminated mechanisms related to the chaperone function. However, an atomic resolution view of Hsp90 conformational dynamics, determined by the presence of different binding partners, is critical to define communication pathways between remote residues in different domains intimately affecting the chaperone cycle. Here, we present a computational analysis of signal propagation and long-range communication pathways in Hsp90. We carried out molecular dynamics simulations of the full-length Hsp90 dimer, combined with essential dynamics, correlation analysis, and a signal propagation model. All-atom MD simulations with timescales of 70 ns have been performed for complexes with the natural substrates ATP and ADP and for the unliganded dimer. We elucidate the mechanisms of signal propagation and determine "hot spots" involved in interdomain communication pathways from the nucleotide-binding site to the C-terminal domain interface. A comprehensive computational analysis of the Hsp90 communication pathways and dynamics at atomic resolution has revealed the role of the nucleotide in effecting conformational changes, elucidating the mechanisms of signal propagation. Functionally important residues and secondary structure elements emerge as effective mediators of communication between the nucleotide-binding site and the C-terminal interface. Furthermore, we show that specific interdomain signal propagation pathways may be activated as a function of the ligand. Our results support a "conformational selection model" of the Hsp90 mechanism, whereby the protein may

  17. Regulated ADAM17-dependent EGF family ligand release by substrate-selecting signaling pathways.

    Science.gov (United States)

    Dang, Michelle; Armbruster, Nicole; Miller, Miles A; Cermeno, Efrain; Hartmann, Monika; Bell, George W; Root, David E; Lauffenburger, Douglas A; Lodish, Harvey F; Herrlich, Andreas

    2013-06-11

    Ectodomain cleavage of cell-surface proteins by A disintegrin and metalloproteinases (ADAMs) is highly regulated, and its dysregulation has been linked to many diseases. ADAM10 and ADAM17 cleave most disease-relevant substrates. Broad-spectrum metalloprotease inhibitors have failed clinically, and targeting the cleavage of a specific substrate has remained impossible. It is therefore necessary to identify signaling intermediates that determine substrate specificity of cleavage. We show here that phorbol ester or angiotensin II-induced proteolytic release of EGF family members may not require a significant increase in ADAM17 protease activity. Rather, inducers activate a signaling pathway using PKC-α and the PKC-regulated protein phosphatase 1 inhibitor 14D that is required for ADAM17 cleavage of TGF-α, heparin-binding EGF, and amphiregulin. A second pathway involving PKC-δ is required for neuregulin (NRG) cleavage, and, indeed, PKC-δ phosphorylation of serine 286 in the NRG cytosolic domain is essential for induced NRG cleavage. Thus, signaling-mediated substrate selection is clearly distinct from regulation of enzyme activity, an important mechanism that offers itself for application in disease.

  18. Disintegrins: integrin selective ligands which activate integrin-coupled signaling and modulate leukocyte functions

    Directory of Open Access Journals (Sweden)

    Barja-Fidalgo C.

    2005-01-01

    Full Text Available Extracellular matrix proteins and cell adhesion receptors (integrins play essential roles in the regulation of cell adhesion and migration. Interactions of integrins with the extracellular matrix proteins lead to phosphorylation of several intracellular proteins such as focal adhesion kinase, activating different signaling pathways responsible for the regulation of a variety of cell functions, including cytoskeleton mobilization. Once leukocytes are guided to sites of infection, inflammation, or antigen presentation, integrins can participate in the initiation, maintenance, or termination of the immune and inflammatory responses. The modulation of neutrophil activation through integrin-mediated pathways is important in the homeostatic control of the resolution of inflammatory states. In addition, during recirculation, T lymphocyte movement through distinct microenvironments is mediated by integrins, which are critical for cell cycle, differentiation and gene expression. Disintegrins are a family of low-molecular weight, cysteine-rich peptides first identified in snake venom, usually containing an RGD (Arg-Gly-Asp motif, which confers the ability to selectively bind to integrins, inhibiting integrin-related functions in different cell systems. In this review we show that, depending on the cell type and the microenvironment, disintegrins are able to antagonize the effects of integrins or to act agonistically by activating integrin-mediated signaling. Disintegrins have proven useful as tools to improve the understanding of the molecular events regulated by integrin signaling in leukocytes and prototypes in order to design therapies able to interfere with integrin-mediated effects.

  19. The Dipole Potential Modifies the Clustering and Ligand Binding Affinity of ErbB Proteins and Their Signaling Efficiency

    Science.gov (United States)

    Kovács, Tamás; Batta, Gyula; Hajdu, Tímea; Szabó, Ágnes; Váradi, Tímea; Zákány, Florina; Csomós, István; Szöllősi, János; Nagy, Peter

    2016-01-01

    Although activation of the ErbB family of receptor tyrosine kinases (ErbB1-4) is driven by oligomerization mediated by intermolecular interactions between the extracellular, the kinase and the transmembrane domains, the transmembrane domain has been largely neglected in this regard. The largest contributor to the intramembrane electric field, the dipole potential, alters the conformation of transmembrane peptides, but its effect on ErbB proteins is unknown. Here, we show by Förster resonance energy transfer (FRET) and number and brightness (N&B) experiments that the epidermal growth factor (EGF)-induced increase in the homoassociation of ErbB1 and ErbB2 and their heteroassociation are augmented by increasing the dipole potential. These effects were even more pronounced for ErbB2 harboring an activating Val → Glu mutation in the transmembrane domain (NeuT). The signaling capacity of ErbB1 and ErbB2 was also correlated with the dipole potential. Since the dipole potential decreased the affinity of EGF to ErbB1, the augmented growth factor-induced effects at an elevated dipole potential were actually induced at lower receptor occupancy. We conclude that the dipole potential plays a permissive role in the clustering of ErbB receptors and that the effects of lipid rafts on ligand binding and receptor signaling can be partially attributed to the dipole potential. PMID:27775011

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

    Institute of Scientific and Technical Information of China (English)

    Jianjun Jiang; Chi Zhang; Xuelu Wang

    2013-01-01

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

  1. Structural insights into a novel interkingdom signaling circuit by cartography of the ligand-binding sites of the homologous quorum sensing LuxR-family.

    Science.gov (United States)

    Covaceuszach, Sonia; Degrassi, Giuliano; Venturi, Vittorio; Lamba, Doriano

    2013-01-01

    Recent studies have identified a novel interkingdom signaling circuit, via plant signaling molecules, and a bacterial sub-family of LuxR proteins, bridging eukaryotes and prokaryotes. Indeed pivotal plant-bacteria interactions are regulated by the so called Plant Associated Bacteria (PAB) LuxR solo regulators that, although closely related to the quorum sensing (QS) LuxR family, do not bind or respond to canonical quorum sensing N-acyl homoserine lactones (AHLs), but only to specific host plant signal molecules. The large body of structural data available for several members of the QS LuxR family complexed with different classes of ligands (AHLs and other compounds), has been exploited to dissect the cartography of their regulatory domains through structure-based multiple sequence alignments, structural superimposition and a comparative analysis of the contact residues involved in ligand binding. In the absence of experimentally determined structures of members of the PAB LuxR solos subfamily, an homology model of its prototype OryR is presented, aiming to elucidate the architecture of its ligand-binding site. The obtained model, in combination with the cartography of the regulatory domains of the homologous QS LuxRs, provides novel insights into the 3D structure of its ligand-binding site and unveils the probable molecular determinants responsible for differences in selectivity towards specific host plant signal molecules, rather than to canonical QS compounds.

  2. Alzheimer’s disease diagnosis by detecting exogenous fluorescent signal of ligand bound to beta amyloid in the lens of human eye: an exploratory study

    Directory of Open Access Journals (Sweden)

    Charles eKerbage

    2013-05-01

    Full Text Available We report results of a clinical exploratory human trial involving 10 participants using a combination of a fluorescent ligand and a laser scanning device, SAPPHIRE System, as an aid in the diagnosis of Probable Alzheimer’s disease (AD. To the best of our knowledge, this is the first time that such a technique has been used in vivo of a human lens. The primary goal of the clinical trial, in addition to safety assessment, was to evaluate efficacy of the system. By detecting specific fluorescent signature of ligand bound beta amyloid in the supranucleus region of the human lens, a two-fold differentiation factor between AD patients and Control groups is achieved. Data from our studies indicates that deeper regions of the supranucleus provide the highest measures of ligand bound fluorescence signal from both controls and AD patients. Further studies are underway involving a larger population for statistical evaluation of the method.

  3. Characterization of the pharmacology, signal transduction and internalization of the fluorescent PACAP ligand, fluor-PACAP, on NIH/3T3 cells expressing PAC1.

    Science.gov (United States)

    Germano, P M; Stalter, J; Le, S V; Wu, M; Yamaguchi, D J; Scott, D; Pisegna, J R

    2001-06-01

    Fluor-PACAP, a fluorescent derivative of PACAP-27, has been confirmed to share a high affinity for PAC1 receptors transfected into NIH/3T3 cells and to have comparable pharmacological characteristics to the unconjugated, native form. Through competitive binding with 125I-PACAP-27, the two ligands exhibited similar dose- dependent inhibition. Additional examination of the efficacy of activating adenylyl cyclase revealed that both ligands analogously stimulated the production of cyclic AMP. Furthermore, PAC1 internalization visualized by our Fluor-PACAP, is compareable to that performed with the radioligand, 125I-PACAP-27, with maximal internalization achieved within thirty minutes. Thus, Fluor-PACAP exhibits intracellular signaling abilities homologous to the native ligand.

  4. Chemokine signaling involving chemokine (C-C motif) ligand 2 plays a role in descending pain facilitation

    Institute of Scientific and Technical Information of China (English)

    Wei Guo; Hu Wang; Shiping Zou; Ronald Dubner; Ke Ren

    2012-01-01

    Objective Despite accumulating evidence on a role of immune cells and their associated chemicals in mechanisms of pain,few studies have addressed the potential role of chemokines in the descending facilitation of persistent pain.The present study was undertaken to test the hypothesis that the chemokine (C-C motif) ligand 2 (CCL2) (commonly known as monocyte chemoattractant protein-1) signaling in the rostral ventromedial medulla (RVM),a pivotal structure in brainstem pain modulatory circuitry,is involved in descending pain facilitation in rats.Methods An L5 spinal nerve ligation (SNL) was produced in rats under pentobarbital anesthesia.Western blot and immunohistochemistry were used to detect the expression levels of CCL2 and CCL2 receptor (CCR2),and examine their distributions compared with the neuronal marker NeuN as well as glial markers glial fibrillary acidic protein (GFAP,astroglial) and CD11b (microglial),respectively.Results SNL induced an increase in CCL2 expression in the RVM,and this returned to the control level at 4 weeks after injury.The induced CCL2 colocalized with NeuN,but not with GFAP and CD11b.CCR2 was also upregulated by SNL in the RVM,and this increase lasted for at least 4 weeks.CCR2 was colocalized with CD1 1b but not GFAP.Few RVM neurons also exhibited CCR2 staining.Neutralizing CCL2 with an anti-CCL2 antibody (0.2-20 ng) or injecting RS-102895 (0.1-10 pmol),a CCR2b chemokine receptor antagonist,into the RVM on day 1 after SNL,significantly attenuated the established thermal and mechanical hypersensitivity.In addition,injection of recombinant rat CCL2 (0.03-3pmol) into the RVM induced dose-dependent hyperalgesia,which was prevented by pretreatment with RS-102895 (10pmol).Interleukin-1β (IL-1β),a potent inducer of neuronal CCL2,was also selectively upregulated in RVM reactive astrocytes.Injection of IL-1β (120 fmol) into the RVM induced behavioral hyperalgesia,which was blocked by RS-102895(10 pmol).However,an IL-1 receptor antagonist (3

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

    NARCIS (Netherlands)

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

    2013-01-01

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

  6. Model of the initiation of signal transduction by ligands in a cell culture: Simulation of molecules near a plane membrane comprising receptors

    Science.gov (United States)

    Plante, Ianik; Cucinotta, Francis A.

    2011-11-01

    Cell communication is a key mechanism in tissue responses to radiation. Several molecules are implicated in radiation-induced signaling between cells, but their contributions to radiation risk are poorly understood. Meanwhile, Green's functions for diffusion-influenced reactions have appeared in the literature, which are applied to describe the diffusion of molecules near a plane membrane comprising bound receptors with the possibility of reversible binding of a ligand and activation of signal transduction proteins by the ligand-receptor complex. We have developed Brownian dynamics algorithms to simulate particle histories in this system which can accurately reproduce the theoretical distribution of distances of a ligand from the membrane, the number of reversibly bound particles, and the number of receptor complexes activating signaling proteins as a function of time, regardless of the number of time steps used for the simulation. These simulations will be of great importance to model interactions at low doses where stochastic effects induced by a small number of molecules or interactions come into play.

  7. Response of SCP-2L domain of human MFE-2 to ligand removal: binding site closure and burial of peroxisomal targeting signal.

    Science.gov (United States)

    Lensink, M F; Haapalainen, A M; Hiltunen, J K; Glumoff, T; Juffer, A H

    2002-10-11

    In the study of the structure and function relationship of human MFE-2, we have investigated the dynamics of human MFE-2SCP-2L (hSCP-2L) and its response to ligand removal. A comparison was made with homologous rabbit SCP-2. Breathing and a closing motion are found, identifiable with an adjustment in size and a closing off of the binding pocket. Crucial residues for structural integrity have been identified. Particularly mobile areas of the protein are loop 1 that is connecting helices A and C in space, and helix D, next to the entrance of the pocket. In hSCP-2L, the binding pocket gets occupied by Phe93, which is making a tight hydrophobic contact with Trp36. In addition, it is found that the C-terminal peroxisomal targeting signal (PTS1) that is solvent exposed in the complexed structure becomes buried when no ligand is present. Moreover, an anti-correlation exists between burial of PTS1 and the size of the binding pocket. The results are in accordance with plant nsLTPs, where a similar accommodation of binding pocket size was found after ligand binding/removal. Furthermore, the calculations support the suggestion of a ligand-assisted targeting mechanism.

  8. Mycobacteria-responsive sonic hedgehog signaling mediates programmed death-ligand 1- and prostaglandin E2-induced regulatory T cell expansion.

    Science.gov (United States)

    Holla, Sahana; Stephen-Victor, Emmanuel; Prakhar, Praveen; Sharma, Meenu; Saha, Chaitrali; Udupa, Vibha; Kaveri, Srinivas V; Bayry, Jagadeesh; Balaji, Kithiganahalli Narayanaswamy

    2016-01-01

    CD4(+)CD25(+)FoxP3(+) regulatory T cells (Tregs) are exploited by mycobacteria to subvert the protective host immune responses. The Treg expansion in the periphery requires signaling by professional antigen presenting cells and in particularly dendritic cells (DC). However, precise molecular mechanisms by which mycobacteria instruct Treg expansion via DCs are not established. Here we demonstrate that mycobacteria-responsive sonic hedgehog (SHH) signaling in human DCs leads to programmed death ligand-1 (PD-L1) expression and cyclooxygenase (COX)-2-catalyzed prostaglandin E2 (PGE2) that orchestrate mycobacterial infection-induced expansion of Tregs. While SHH-responsive transcription factor GLI1 directly arbitrated COX-2 transcription, specific microRNAs, miR-324-5p and miR-338-5p, which target PD-L1 were downregulated by SHH signaling. Further, counter-regulatory roles of SHH and NOTCH1 signaling during mycobacterial-infection of human DCs was also evident. Together, our results establish that Mycobacterium directs a fine-balance of host signaling pathways and molecular regulators in human DCs to expand Tregs that favour immune evasion of the pathogen. PMID:27080341

  9. NOD1 and NOD2 receptors in mrigal (Cirrhinus mrigala): Inductive expression and downstream signalling in ligand stimulation and bacterial infections

    Indian Academy of Sciences (India)

    Banikalyan Swain; Madhubanti Basu; Mrinal Samanta

    2013-09-01

    Nucleotide binding and oligomerization domain (NOD)1 and NOD2 are important cytoplasmic pattern recognition receptors (PRRs) and key members of the NOD-like receptor (NLR) family. They sense a wide range of bacteria or their products and play a key role in inducing innate immunity. This report describes the role of NOD1 and NOD2 receptors signalling in innate immunity in the Indian major carp, mrigal (Cirrhinus mrigala). Tissue-specific expression analysis of NOD1 and NOD2 genes by quantitative real-time PCR (qRT-PCR) revealed their wide distribution in various organs/tissues. In the untreated fish, the highest expression of NOD1 and NOD2 was detected in liver and blood, respectively. Stimulation with NOD1- and NOD2-specific ligands, i.e. iE-DAP and MDP, activated NOD1 and NOD2 receptor signalling in vivo and in vitro resulting in significant ( < 0.05) induction of downstream signalling molecule RICK, and the effector molecules IL-1, IL-8 and IFN- in the treated group as compared to their controls. In response to both Gram-positive and Gram-negative bacterial infections, NOD1 and NOD2 receptors signalling were activated and IL-1, IL-8 and IFN- were induced. These findings highlight the important role of NOD receptors in eliciting innate immune response during the pathogenic invasion to the fish.

  10. Linking ligand perception by PEPR pattern recognition receptors to cytosolic Ca2+ elevation and downstream immune signaling in plants.

    Science.gov (United States)

    Ma, Yi; Walker, Robin K; Zhao, Yichen; Berkowitz, Gerald A

    2012-11-27

    Little is known about molecular steps linking perception of pathogen invasion by cell surface sentry proteins acting as pattern recognition receptors (PRRs) to downstream cytosolic Ca(2+) elevation, a critical step in plant immune signaling cascades. Some PRRs recognize molecules (such as flagellin) associated with microbial pathogens (pathogen-associated molecular patterns, PAMPs), whereas others bind endogenous plant compounds (damage-associated molecular patterns, DAMPs) such as peptides released from cells upon attack. This work focuses on the Arabidopsis DAMPs plant elicitor peptides (Peps) and their receptors, PEPR1 and PEPR2. Pep application causes in vivo cGMP generation and downstream signaling that is lost when the predicted PEPR receptor guanylyl cyclase (GC) active site is mutated. Pep-induced Ca(2+) elevation is attributable to cGMP activation of a Ca(2+) channel. Some differences were identified between Pep/PEPR signaling and the Ca(2+)-dependent immune signaling initiated by the flagellin peptide flg22 and its cognate receptor Flagellin-sensing 2 (FLS2). FLS2 signaling may have a greater requirement for intracellular Ca(2+) stores and inositol phosphate signaling, whereas Pep/PEPR signaling requires extracellular Ca(2+). Maximal FLS2 signaling requires a functional Pep/PEPR system. This dependence was evidenced as a requirement for functional PEPR receptors for maximal flg22-dependent Ca(2+) elevation, H(2)O(2) generation, defense gene [WRKY33 and Plant Defensin 1.2 (PDF1.2)] expression, and flg22/FLS2-dependent impairment of pathogen growth. In a corresponding fashion, FLS2 loss of function impaired Pep signaling. In addition, a role for PAMP and DAMP perception in bolstering effector-triggered immunity (ETI) is reported; loss of function of either FLS2 or PEPR receptors impaired the hypersensitive response (HR) to an avirulent pathogen.

  11. Nuclear expression of FLT1 and its ligand PGF in FUS-DDIT3 carrying myxoid liposarcomas suggests the existence of an intracrine signaling loop

    Directory of Open Access Journals (Sweden)

    Andersson Carola

    2010-06-01

    effect of FUS-DDIT3 expression in MLS. This could be a consequence of the ability of FUS-DDIT3 to hijack parts of normal adipose tissue development and reprogram primary cells to a liposarcoma-like phenotype. The findings of nuclear FLT1 protein and expression of corresponding ligands in MLS and normal tissues may have implications for tissue homeostasis and tumor development through auto- or intracrine signaling.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2012-03-09

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

  13. Actions of translocator protein ligands on neutrophil adhesion and motility induced by G-protein coupled receptor signaling.

    Science.gov (United States)

    de Lima, Camila Bento; Tamura, Eduardo K; Montero-Melendez, Trindad; Palermo-Neto, João; Perretti, Mauro; Markus, Regina P; Farsky, Sandra Helena Poliselli

    2012-01-13

    The 18 kDa translocator protein (TSPO) also known as the peripheral benzodiazepine receptor (PBR), mediates the transportation of cholesterol and anions from the outer to the inner mitochondrial membrane in different cells types. Although recent evidences indicate a potential role for TSPO in the development of inflammatory processes, the mechanisms involved have not been elucidated. The present study investigated the ability of the specific TSPO ligands, the isoquinoline carboxamide PK11195 and benzodiazepine Ro5-4864, on neutrophil recruitment promoted by the N-formylmethionyl-leucyl-phenylalanine peptide (fMLP), an agonist of G-protein coupled receptor (GPCR). Pre-treatment with Ro5-4864 abrograted fMLP-induced leukocyte-endothelial interactions in mesenteric postcapillary venules in vivo. Moreover, in vitro Ro5-4864 treatment prevented fMLP-induced: (i) L-selectin shedding and overexpression of PECAM-1 on the neutrophil cell surface; (ii) neutrophil chemotaxis and (iii) enhancement of intracellular calcium cations (iCa(+2)). Intriguingly, the two latter effects were augmented by cell treatment with PK11195. An allosteric agonist/antagonist relation may be suggested, as the effects of Ro5-4864 on fMLP-stimulated neutrophils were reverted by simultaneous treatment with PK11195. Taken together, these data highlight TSPO as a modulator of pathways of neutrophil adhesion and locomotion induced by GPCR, connecting TSPO actions and the onset of an innate inflammatory response. PMID:22209795

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

    Directory of Open Access Journals (Sweden)

    Christina Koers-Wunrau

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

  15. Delphinidin, a dietary anthocyanidin, inhibits platelet-derived growth factor ligand/receptor (PDGF/PDGFR) signaling.

    Science.gov (United States)

    Lamy, Sylvie; Beaulieu, Edith; Labbé, David; Bédard, Valérie; Moghrabi, Albert; Barrette, Stéphane; Gingras, Denis; Béliveau, Richard

    2008-05-01

    Most cancers are dependent on the growth of tumor blood vessels and inhibition of tumor angiogenesis may thus provide an efficient strategy to retard or block tumor growth. Recently, tumor vascular targeting has expanded to include not only endothelial cells (ECs) but also smooth muscle cells (SMCs), which contribute to a mature and functional vasculature. We have reported previously that delphinidin, a major biologically active constituent of berries, inhibits the vascular endothelial growth factor-induced phosphorylation of vascular endothelial growth factor receptor-2 and blocks angiogenesis in vitro and in vivo. In the present study, we show that delphinidin also inhibits activation of the platelet-derived growth factor (PDGF)-BB receptor-beta [platelet-derived growth factor receptor-beta (PDGFR-beta)] in SMC and that this inhibition may contribute to its antitumor effect. The inhibitory effect of delphinidin on PDGFR-beta was very rapid and led to the inhibition of PDGF-BB-induced activation of extracellular signal-regulated kinase (ERK)-1/2 signaling and of the chemotactic motility of SMC, as well as the differentiation and stabilization of EC and SMC into capillary-like tubular structures in a three-dimensional coculture system. Using an anthocyan-rich extract of berries, we show that berry extracts were able to suppress the synergistic induction of vessel formation by basic fibroblast growth factor-2 and PDGF-BB in the mouse Matrigel plug assay. Oral administration of the berry extract also significantly retarded tumor growth in a lung carcinoma xenograft model. Taken together, these results provide new insight into the molecular mechanisms underlying the antiangiogenic activity of delphinidin that will be helpful for the development of dietary-based chemopreventive strategies. PMID:18339683

  16. Abnormal structural luteolysis in ovaries of the senescence accelerated mouse (SAM): expression of Fas ligand/Fas-mediated apoptosis signaling molecules in luteal cells.

    Science.gov (United States)

    Kiso, Minako; Manabe, Noboru; Komatsu, Kohji; Shimabe, Munetake; Miyamoto, Hajime

    2003-12-01

    Senescence accelerated mouse-prone (SAMP) mice with a shortened life span show accelerated changes in many of the signs of aging and a shorter reproductive life span than SAM-resistant (SAMR) controls. We previously showed that functional regression (progesterone dissimilation) occurs in abnormally accumulated luteal bodies (aaLBs) of SAMP mice, but structural regression of luteal cells in aaLB is inhibited. A deficiency of luteal cell apoptosis causes the abnormal accumulation of LBs in SAMP ovaries. In the present study, to show the abnormality of Fas ligand (FasL)/Fas-mediated apoptosis signal transducing factors in the aaLBs of the SAMP ovaries, we assessed the changes in the expression of FasL, Fas, caspase-8 and caspase-3 mRNAs by reverse transcription-polymerase chain reaction, and in the expression and localization of FasL, Fas and activated caspase-3 proteins by Western blotting and immunohistochemistry, respectively, during the estrus cycle/luteolysis. These mRNAs and proteins were expressed in normal LBs of both SAMP and SAMR ovaries, but not at all or only in trace amounts in aaLBs of SAMP, indicating that structural regression is inhibited by blockage of the expression of these transducing factors in luteal cells of aaLBs in SAMP mice. PMID:14967896

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

    Science.gov (United States)

    Tarnow, Patrick; Tralau, Tewes; Luch, Andreas

    2016-08-01

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

  18. Pattern recognition receptor signaling in human dendritic cells is enhanced by ICOS ligand and modulated by the Crohn's disease ICOSLG risk allele.

    Science.gov (United States)

    Hedl, Matija; Lahiri, Amit; Ning, Kaida; Cho, Judy H; Abraham, Clara

    2014-05-15

    Inflammatory bowel disease (IBD) is characterized by dysregulated intestinal immune homeostasis and cytokine secretion. Multiple loci are associated with IBD, but a functional explanation is missing for most. Here we found that pattern-recognition receptor (PRR)-induced cytokine secretion was diminished in human monocyte-derived dendritic cells (MDDC) from rs7282490 ICOSLG GG risk carriers. Homotypic interactions between the costimulatory molecule ICOS and the ICOS ligand on MDDCs amplified nucleotide-binding oligomerization domain 2 (NOD2)-initiated cytokine secretion. This amplification required arginine residues in the ICOSL cytoplasmic tail that recruited the adaptor protein RACK1 and the kinases PKC and JNK leading to PKC, MAPK, and NF-κB activation. MDDC from rs7282490 GG risk-carriers had reduced ICOSL expression and PRR-initiated signaling and this loss-of-function ICOSLG risk allele associated with an ileal Crohn's disease phenotype, similar to polymorphisms in NOD2. Taken together, ICOSL amplifies PRR-initiated outcomes, which might contribute to immune homeostasis.

  19. MEK1/2 inhibition enhances the radiosensitivity of cancer cells by downregulating survival and growth signals mediated by EGFR ligands.

    Science.gov (United States)

    Chung, Eun Joo; Urick, Mary Ellen; Kurshan, Naamit; Shield, William; Asano, Hiroaki; Smith, Paul D; Scroggins, Bradley S; Burkeen, Jeffrey; Citrin, Deborah E

    2013-06-01

    The inhibition of the Ras/mitogen-activated protein kinase (Ras/MAPK) pathway through the suppression of mutated Ras or MAPK/extracellular signal-regulated kinase 1/2 (MEK1/2) has been shown to sensitize tumor cells to ionizing radiation (IR). The molecular mechanisms of this sensitization however, are not yet fully understood. In this study, we investigated the role of transforming growth factor-α (TGF-α) in the radiosensitizing effects of selumetinib, a selective inhibitor of MEK1/2. The expression of epidermal growth factor receptor (EGFR) ligands was assessed by ELISA in both Ras wild-type and Ras mutant cells that were exposed to radiation with or without selumetinib. The effects of selumetinib on the TGF-α/EGFR signaling cascade in response to radiation were examined by western blot analysis, clonogenic assay and by determing the yield of mitotic catastrophe. The treatment of cells with selumetinib reduced the basal and IR-induced secretion of TGF-α in both Ras wild-type and Ras mutant cell lines in vitro and in vivo. The reduction of TGF-α secretion was accompanied with a reduction in phosphorylated tumor necrosis factor-α converting enzyme (TACE) in the cells treated with selumetinib with or without IR. The treatment of cells with selumetinib with or without IR inhibited the phosphorylation of EGFR and checkpoint kinase 2 (Chk2), and reduced the expression of survivin. Supplementation with exogenous TGF-α partially rescued the selumetinib-treated cells from IR-induced cell death, restored EGFR and Chk2 phosphorylation and increased survivin expression. These data suggest that the inhibition of MEK1/2 with selumetinib may provide a mechanism to sensitize tumor cells to IR in a fashion that prevents the activation of the TGF-α autocrine loop following IR.

  20. Dimerization of melanocortin receptor 1 (MC1R) and MC5R creates a ligand-dependent signal modulation: Potential participation in physiological color change in the flounder.

    Science.gov (United States)

    Kobayashi, Yuki; Hamamoto, Akie; Takahashi, Akiyoshi; Saito, Yumiko

    2016-05-01

    Vertebrates produce α-melanocyte-stimulating hormone (α-MSH), which contains an N-terminal acetyl group, and desacetyl-α-MSH, which does not contain an N-terminal acetyl group. In teleosts and amphibians, α-MSH-related peptides stimulate pigment dispersion via melanocortin receptors 1-5 (MC1R-MC5R), which are members of the G-protein-coupled receptor (GPCR) family. We previously reported an interesting phenomenon associated with physiological color changes in the skin of a flatfish, barfin flounder (bf). Specifically, pigments in xanthophores expressing only the bfMC5R gene were dispersed by both α-MSH and desacetyl-α-MSH, whereas those in melanophores expressing both the bfMC1R and bfMC5R genes were dispersed by desacetyl-α-MSH, but not by α-MSH. In this study, we examined whether heterodimers of bfMC1R and bfMC5R can act as significant inhibitory receptors for the N-terminal acetylation of α-MSH in mammalian Chinese hamster ovary cells. Immunofluorescence analyses showed that bfMC1R and bfMC5R were localized together at the plasma membrane when expressed in the same cells. Indeed, after coexpression of Flag-bfMC1R and HA-bfMC5R, immunoprecipitation with anti-Flag antibodies resulted in the presence of anti-HA immunoreactivity in the precipitate, and vice versa. Importantly, cyclic AMP assays showed that cotransfection of bfMC1R with bfMC5R inhibited the cyclic AMP accumulation induced by α-MSH to a greater extent than that observed after transfection of bfMC1R alone. Of note, this inhibitory response was not caused by desacetyl-α-MSH. Thus, we show a ligand-dependent signaling through functional heterodimerization of MC1R and MC5R in mammalian cells. The ligand-selective receptor complex also provide the first mechanistic implication that may play a role in the control of color change in teleosts. PMID:27080548

  1. The CXC chemokine receptor encoded by herpesvirus saimiri, ECRF3, shows ligand-regulated signaling through Gi, Gq, and G12/13 proteins but constitutive signaling only through Gi and G12/13 proteins

    DEFF Research Database (Denmark)

    Rosenkilde, Mette M; McLean, Katherine A; Holst, Peter J;

    2004-01-01

    bound the so-called ELR (Glu-Leu-Arg) CXC chemokines (125)I-CXCL1/GRO alpha, (125)I-CXCL6/GCP-2, and (125)I-CXCL8/interleukin-8 with high affinity; but in contrast to ORF74 from HHV8, it did not bind the non-ELR CXC chemokine (125)I-CXCL10/IP10. Interestingly, the B(max) value for CXCL6/GCP-2 was 3-fold...... higher than the capacity for maximal binding of CXCL1/GRO alpha to ECRF3 and 85-fold higher than that of CXCL8/interleukin-8, despite similar affinities. Like ORF74 from HHV8, ECRF3 activated a broad range of pathways (G(q), G(i), and G(12/13) as well as the cAMP response element-binding protein, NF......-kappa B, NFAT, and serum response element transcription factors) in a ligand-regulated manner, with CXCL6/GCP-2 being the most potent and efficacious agonist. ECRF3 signaled constitutively through G(i) and G(12/13), but surprisingly not through G(q). At the level of transcription factor activation...

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

    Science.gov (United States)

    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...

  3. The role of the class A scavenger receptors, SR-A and MARCO, in the immune system. Part 1. The structure of receptors, their ligand binding repertoires and ability to initiate intracellular signaling

    Directory of Open Access Journals (Sweden)

    Szczepan Józefowski

    2012-02-01

    Full Text Available  Recognition of pathogens by innate immune cells is mediated by pattern recognition receptors (PRR, which include scavenger receptors (SR. The class A SR, SR-A/CD204 and MARCO, are characterized by the presence of collagenous and SR cysteine-rich domains in their extracellular portions. Both receptors are expressed mainly on macrophages and dendritic cells. Thanks to their ability to bind to a wide range of polyanionic ligands, the class A SR may participate in numerous functions of these cells, such as endocytosis, and adhesion to extracellular matrix and to other cells. Among SR-A ligands are oxidized lipoproteins and β-amyloid fibrils, which link SR-A to the pathogenesis of arteriosclerosis and Alzheimer’s disease. Despite the demonstration of class A SR involvement in so many processes, the lack of selective ligands precluded reaching definite conclusions concerning their signaling abilities. Using specific receptor ligation with antibodies, we showed that SR-A and MARCO trigger intracellular signaling, modulating pro-inflammatory and microbicidal activities of macrophages. Surprisingly, despite similarities in structure and ligand binding repertoires, SR-A and MARCO exert opposite effects on interleukin-12 (IL-12 production in macrophages. SR-A ligation also stimulated H2O2 and IL-10 production, but had no effect on the release of several other cytokines. These limited effects of specific SR-A ligation contrast with generalized enhancement of immune responses observed in SR-A-deficient mice. Recent studies have revealed that many of these effects of SR-A deficiency may be caused by compensatory changes in the expression of other receptors and/or disinhibition of signal transduction from receptors belonging to the Toll/IL-1R family, rather than by the loss of the receptor function of SR-A.

  4. Crystallization and preliminary crystal structure analysis of the ligand-binding domain of PqsR (MvfR), the Pseudomonas quinolone signal (PQS) responsive quorum-sensing transcription factor of Pseudomonas aeruginosa

    International Nuclear Information System (INIS)

    The ligand-binding domain of the transcription factor PqsR from P. aeruginosa has been crystallized and initial phases have been obtained using SAD data from seleno-l-methionine-labelled crystals. The opportunistic bacterial pathogen Pseudomonas aeruginosa employs three transcriptional regulators, LasR, RhlR and PqsR, to control the transcription of a large subset of its genes in a cell-density-dependent process known as quorum sensing. Here, the recombinant production, crystallization and structure solution of the ligand-binding domain of PqsR (MvfR), the LysR-type transcription factor that responds to the Pseudomonas quinolone signal (PQS), a quinolone-based quorum-sensing signal that is unique to P. aeruginosa and possibly a small number of other bacteria, is reported. PqsR regulates the expression of many virulence genes and may therefore be an interesting drug target. The ligand-binding domain (residues 91–319) was produced as a fusion with SUMO, and hexagonal-shaped crystals of purified PqsR-91–319 were obtained using the vapour-diffusion method. Crystallization in the presence of a PQS precursor allowed data collection to 3.25 Å resolution on a synchrotron beamline, and initial phases have been obtained using single-wavelength anomalous diffraction data from seleno-l-methionine-labelled crystals, revealing the space group to be P6522, with unit-cell parameters a = b = 116–120, c = 115–117 Å

  5. Ylide Ligands

    OpenAIRE

    Esteban P. Urriolabeitia

    2010-01-01

    The use of ylides of P, N, As, or S as ligands toward transition metals is still a very active research area in organometallic chemistry. This fact is mainly due to the nucleophilic character of the ylides and to their particular bonding properties and coordination modes. They can behave as monodentate or bidentate chelate or bridging species, they can be used as chiral auxiliary reagents, and they are interesting reaction intermediates or useful starting materials in a wide ...

  6. K-RAS(V12) Induces Autocrine Production of EGFR Ligands and Mediates Radioresistance Through EGFR-Dependent Akt Signaling and Activation of DNA-PKcs

    International Nuclear Information System (INIS)

    Purpose: It is known that postirradiation survival of tumor cells presenting mutated K-RAS is mediated through autocrine activation of epidermal growth factor receptor (EGFR). In this study the molecular mechanism of radioresistance of cells overexpressing mutated K-RAS(V12) was investigated. Methods and Materials: Head-and-neck cancer cells (FaDu) presenting wild-type K-RAS were transfected with empty vector or vector expressing mutated K-RAS(V12). The effect of K-RAS(V12) on autocrine production of EGFR ligands, activation of EGFR downstream pathways, DNA damage repair, and postirradiation survival was analyzed. Results: Conditioned medium collected from K-RAS(V12)–transfected cells enhanced activation of the phosphatidylinositol-3-kinase–Akt pathway and increased postirradiation survival of wild-type K-RAS parental cells when compared with controls. These effects were reversed by amphiregulin (AREG)–neutralizing antibody. In addition, secretion of the EGFR ligands AREG and transforming growth factor α was significantly increased upon overexpression of K-RAS(V12). Expression of mutated K-RAS(V12) resulted in an increase in radiation-induced DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phosphorylation at S2056. This increase was accompanied by increased repair of DNA double-strand breaks. Abrogation of DNA-PKcs phosphorylation by serum depletion or AREG-neutralizing antibody underscored the role of autocrine production of EGFR ligands, namely, AREG, in regulating DNA-PKcs activation in K-RAS mutated cells. Conclusions: These data indicate that radioresistance of K-RAS mutated tumor cells is at least in part due to constitutive production of EGFR ligands, which mediate enhanced repair of DNA double-strand breaks through the EGFR–phosphatidylinositol-3-kinase–Akt cascade.

  7. Inhibition of signaling between human CXCR4 and zebrafish ligands by the small molecule IT1t impairs the formation of triple-negative breast cancer early metastases in a zebrafish xenograft model

    Directory of Open Access Journals (Sweden)

    Claudia Tulotta

    2016-02-01

    Full Text Available Triple-negative breast cancer (TNBC is a highly aggressive and recurrent type of breast carcinoma that is associated with poor patient prognosis. Because of the limited efficacy of current treatments, new therapeutic strategies need to be developed. The CXCR4-CXCL12 chemokine signaling axis guides cell migration in physiological and pathological processes, including breast cancer metastasis. Although targeted therapies to inhibit the CXCR4-CXCL12 axis are under clinical experimentation, still no effective therapeutic approaches have been established to block CXCR4 in TNBC. To unravel the role of the CXCR4-CXCL12 axis in the formation of TNBC early metastases, we used the zebrafish xenograft model. Importantly, we demonstrate that cross-communication between the zebrafish and human ligands and receptors takes place and human tumor cells expressing CXCR4 initiate early metastatic events by sensing zebrafish cognate ligands at the metastatic site. Taking advantage of the conserved intercommunication between human tumor cells and the zebrafish host, we blocked TNBC early metastatic events by chemical and genetic inhibition of CXCR4 signaling. We used IT1t, a potent CXCR4 antagonist, and show for the first time its promising anti-tumor effects. In conclusion, we confirm the validity of the zebrafish as a xenotransplantation model and propose a pharmacological approach to target CXCR4 in TNBC.

  8. Structure of the unique SEFIR domain from human interleukin 17 receptor A reveals a composite ligand-binding site containing a conserved α-helix for Act1 binding and IL-17 signaling

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Bing [Oklahoma State University, Stillwater, OK 74078 (United States); Liu, Caini; Qian, Wen [Lerner Research Institute, Cleveland Clinic, 9500 Euclid Avenue, Cleveland, OH 44195 (United States); Han, Yue [Oklahoma State University, Stillwater, OK 74078 (United States); Li, Xiaoxia, E-mail: lix@ccf.org [Lerner Research Institute, Cleveland Clinic, 9500 Euclid Avenue, Cleveland, OH 44195 (United States); Deng, Junpeng, E-mail: lix@ccf.org [Oklahoma State University, Stillwater, OK 74078 (United States)

    2014-05-01

    Crystal structure of the SEFIR domain from human IL-17 receptor A provides new insights into IL-17 signaling. Interleukin 17 (IL-17) cytokines play a crucial role in mediating inflammatory and autoimmune diseases. A unique intracellular signaling domain termed SEFIR is found within all IL-17 receptors (IL-17Rs) as well as the key adaptor protein Act1. SEFIR-mediated protein–protein interaction is a crucial step in IL-17 cytokine signaling. Here, the 2.3 Å resolution crystal structure of the SEFIR domain of IL-17RA, the most commonly shared receptor for IL-17 cytokine signaling, is reported. The structure includes the complete SEFIR domain and an additional α-helical C-terminal extension, which pack tightly together to form a compact unit. Structural comparison between the SEFIR domains of IL-17RA and IL-17RB reveals substantial differences in protein topology and folding. The uniquely long insertion between strand βC and helix αC in IL-17RA SEFIR is mostly well ordered, displaying a helix (αCC′{sub ins}) and a flexible loop (CC′). The DD′ loop in the IL-17RA SEFIR structure is much shorter; it rotates nearly 90° with respect to the counterpart in the IL-17RB SEFIR structure and shifts about 12 Å to accommodate the αCC′{sub ins} helix without forming any knots. Helix αC was identified as critical for its interaction with Act1 and IL-17-stimulated gene expression. The data suggest that the heterotypic SEFIR–SEFIR association via helix αC is a conserved and signature mechanism specific for IL-17 signaling. The structure also suggests that the downstream motif of IL-17RA SEFIR together with helix αC could provide a composite ligand-binding surface for recruiting Act1 during IL-17 signaling.

  9. Regulation of Ligand and Shear Stress-induced Insulin-like Growth Factor 1 (IGF1) Signaling by the Integrin Pathway.

    Science.gov (United States)

    Tahimic, Candice G T; Long, Roger K; Kubota, Takuo; Sun, Maggie Yige; Elalieh, Hashem; Fong, Chak; Menendez, Alicia T; Wang, Yongmei; Vilardaga, Jean-Pierre; Bikle, Daniel D

    2016-04-01

    Mechanical loading of the skeleton, as achieved during daily movement and exercise, preserves bone mass and stimulates bone formation, whereas skeletal unloading from prolonged immobilization leads to bone loss. A functional interplay between the insulin-like growth factor 1 receptor (IGF1R), a major player in skeletal development, and integrins, mechanosensors, is thought to regulate the anabolic response of osteogenic cells to mechanical load. The mechanistic basis for this cross-talk is unclear. Here we report that integrin signaling regulates activation of IGF1R and downstream targets in response to both IGF1 and a mechanical stimulus. In addition, integrins potentiate responsiveness of IGF1R to IGF1 and mechanical forces. We demonstrate that integrin-associated kinases, Rous sarcoma oncogene (SRC) and focal adhesion kinase (FAK), display distinct actions on IGF1 signaling; FAK regulates IGF1R activation and its downstream effectors, AKT and ERK, whereas SRC controls signaling downstream of IGF1R. These findings linked to our observation that IGF1 assembles the formation of a heterocomplex between IGF1R and integrin β3 subunit indicate that the regulation of IGF1 signaling by integrins proceeds by direct receptor-receptor interaction as a possible means to translate biomechanical forces into osteoanabolic signals. PMID:26865633

  10. IL-1beta-induced pro-apoptotic signalling is facilitated by NCAM/FGF receptor signalling and inhibited by the C3d ligand in the INS-1E rat beta cell line

    DEFF Research Database (Denmark)

    Petersen, L G; Størling, J; Heding, P;

    2006-01-01

    AIMS/HYPOTHESIS: IL-1beta released from immune cells induces beta cell pro-apoptotic signalling via mitogen-activated protein kinases (MAPKs) and nuclear factor-kappaB (NF-kappaB). In neurons, the neural cell adhesion molecule (NCAM) signals to several elements involved in IL-1beta-induced pro......-apoptotic signalling in beta cells. Pancreatic beta cells express NCAM, but its biological effects in these cells are unclear. The aim of this study was to investigate whether there is cross-talk between NCAM signalling and cytokine-induced pro-apoptotic signalling. MATERIALS AND METHODS: Western blotting was used...... to investigate levels of NCAM and inducible nitric oxide synthase, phosphorylation of Src and MAPKs, and cleavage of caspase-3. MAPK activity was investigated with an in vitro kinase assay. Apoptosis was detected by cleaved caspase-3 and a Cell Death Detection ELISA(plus) assay. NCAM-induced fibroblast growth...

  11. Rosetta and the Design of Ligand Binding Sites.

    Science.gov (United States)

    Moretti, Rocco; Bender, Brian J; Allison, Brittany; Meiler, Jens

    2016-01-01

    Proteins that bind small molecules (ligands) can be used as biosensors, signal modulators, and sequestering agents. When naturally occurring proteins for a particular target ligand are not available, artificial proteins can be computationally designed. We present a protocol based on RosettaLigand to redesign an existing protein pocket to bind a target ligand. Starting with a protein structure and the structure of the ligand, Rosetta can optimize both the placement of the ligand in the pocket and the identity and conformation of the surrounding sidechains, yielding proteins that bind the target compound. PMID:27094285

  12. Real time analysis of β2-adrenoceptor-mediated signaling kinetics in Human Primary Airway Smooth Muscle Cells reveals both ligand and dose dependent differences

    Directory of Open Access Journals (Sweden)

    Hall Ian P

    2011-07-01

    Full Text Available Abstract Background β2-adrenoceptor agonists elicit bronchodilator responses by binding to β2-adrenoceptors on airway smooth muscle (ASM. In vivo, the time between drug administration and clinically relevant bronchodilation varies significantly depending on the agonist used. Our aim was to utilise a fluorescent cyclic AMP reporter probe to study the temporal profile of β2-adrenoceptor-mediated signaling induced by isoproterenol and a range of clinically relevant agonists in human primary ASM (hASM cells by using a modified Epac protein fused to CFP and a variant of YFP. Methods Cells were imaged in real time using a spinning disk confocal system which allowed rapid and direct quantification of emission ratio imaging following direct addition of β2-adrenoceptor agonists (isoproterenol, salbutamol, salmeterol, indacaterol and formoterol into the extracellular buffer. For pharmacological comparison a radiolabeling assay for whole cell cyclic AMP formation was used. Results Temporal analysis revealed that in hASM cells the β2-adrenoceptor agonists studied did not vary significantly in the onset of initiation. However, once a response was initiated, significant differences were observed in the rate of this response with indacaterol and isoproterenol inducing a significantly faster response than salmeterol. Contrary to expectation, reducing the concentration of isoproterenol resulted in a significantly faster initiation of response. Conclusions We conclude that confocal imaging of the Epac-based probe is a powerful tool to explore β2-adrenoceptor signaling in primary cells. The ability to analyse the kinetics of clinically used β2-adrenoceptor agonists in real time and at a single cell level gives an insight into their possible kinetics once they have reached ASM cells in vivo.

  13. An agonist sensitive, quick and simple cell-based signaling assay for determination of ligands mimicking prostaglandin E2 or E1 activity through subtype EP1 receptor: Suitable for high throughput screening

    Directory of Open Access Journals (Sweden)

    Ruan Ke-He

    2011-02-01

    fractionation and high performance liquid chromatography. We, however, demonstrated that the active ingredients of the herbal extracts can be tested before purification using an agonist sensitive, quick and simple cell-based signaling assay for ligands mimicking the agonists, PGE1 and PGE2.

  14. A response calculus for immobilized T cell receptor ligands

    DEFF Research Database (Denmark)

    Andersen, P S; Menné, C; Mariuzza, R A;

    2001-01-01

    determine the level of T cell activation. When fitted to T cell responses against purified ligands immobilized on plastic surfaces, the 2D-affinity model adequately simulated changes in cellular activation as a result of varying ligand affinity and ligand density. These observations further demonstrated......To address the molecular mechanism of T cell receptor (TCR) signaling, we have formulated a model for T cell activation, termed the 2D-affinity model, in which the density of TCR on the T cell surface, the density of ligand on the presenting surface, and their corresponding two-dimensional affinity...... the importance of receptor cross-linking density in determining TCR signaling. Moreover, it was found that the functional two-dimensional affinity of TCR ligands was affected by the chemical composition of the ligand-presenting surface. This makes it possible that cell-bound TCR ligands, despite their low...

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

    Science.gov (United States)

    Baek, Jong Min; Park, Sun-Hyang; Cheon, Yoon-Hee; Ahn, Sung-Jun; Lee, Myeung Su; Oh, Jaemin; Kim, Ju-Young

    2015-05-29

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

  16. Designer TGFβ superfamily ligands with diversified functionality.

    Directory of Open Access Journals (Sweden)

    George P Allendorph

    Full Text Available Transforming Growth Factor--beta (TGFβ superfamily ligands, including Activins, Growth and Differentiation Factors (GDFs, and Bone Morphogenetic Proteins (BMPs, are excellent targets for protein-based therapeutics because of their pervasiveness in numerous developmental and cellular processes. We developed a strategy termed RASCH (Random Assembly of Segmental Chimera and Heteromer, to engineer chemically-refoldable TGFβ superfamily ligands with unique signaling properties. One of these engineered ligands, AB208, created from Activin-βA and BMP-2 sequences, exhibits the refolding characteristics of BMP-2 while possessing Activin-like signaling attributes. Further, we find several additional ligands, AB204, AB211, and AB215, which initiate the intracellular Smad1-mediated signaling pathways more strongly than BMP-2 but show no sensitivity to the natural BMP antagonist Noggin unlike natural BMP-2. In another design, incorporation of a short N-terminal segment from BMP-2 was sufficient to enable chemical refolding of BMP-9, without which was never produced nor refolded. Our studies show that the RASCH strategy enables us to expand the functional repertoire of TGFβ superfamily ligands through development of novel chimeric TGFβ ligands with diverse biological and clinical values.

  17. Polypharmacology of dopamine receptor ligands.

    Science.gov (United States)

    Butini, S; Nikolic, K; Kassel, S; Brückmann, H; Filipic, S; Agbaba, D; Gemma, S; Brogi, S; Brindisi, M; Campiani, G; Stark, H

    2016-07-01

    Most neurological diseases have a multifactorial nature and the number of molecular mechanisms discovered as underpinning these diseases is continuously evolving. The old concept of developing selective agents for a single target does not fit with the medical need of most neurological diseases. The development of designed multiple ligands holds great promises and appears as the next step in drug development for the treatment of these multifactorial diseases. Dopamine and its five receptor subtypes are intimately involved in numerous neurological disorders. Dopamine receptor ligands display a high degree of cross interactions with many other targets including G-protein coupled receptors, transporters, enzymes and ion channels. For brain disorders like Parkinsońs disease, schizophrenia and depression the dopaminergic system, being intertwined with many other signaling systems, plays a key role in pathogenesis and therapy. The concept of designed multiple ligands and polypharmacology, which perfectly meets the therapeutic needs for these brain disorders, is herein discussed as a general ligand-based concept while focusing on dopaminergic agents and receptor subtypes in particular. PMID:27234980

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2015-05-29

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

  19. Metal-ligand cooperation.

    Science.gov (United States)

    Khusnutdinova, Julia R; Milstein, David

    2015-10-12

    Metal-ligand cooperation (MLC) has become an important concept in catalysis by transition metal complexes both in synthetic and biological systems. MLC implies that both the metal and the ligand are directly involved in bond activation processes, by contrast to "classical" transition metal catalysis where the ligand (e.g. phosphine) acts as a spectator, while all key transformations occur at the metal center. In this Review, we will discuss examples of MLC in which 1) both the metal and the ligand are chemically modified during bond activation and 2) bond activation results in immediate changes in the 1st coordination sphere involving the cooperating ligand, even if the reactive center at the ligand is not directly bound to the metal (e.g. via tautomerization). The role of MLC in enabling effective catalysis as well as in catalyst deactivation reactions will be discussed. PMID:26436516

  20. Dynamic Presentation of Immobilized Ligands Regulated through Biomolecular Recognition

    OpenAIRE

    Liu, Bo; Liu, Yang; Riesberg, Jeremiah J.; Shen, Wei

    2010-01-01

    To mimic the dynamic regulation of signaling ligands immobilized on extracellular matrices or on the surfaces of neighboring cells for guidance of cell behavior and fate selection, we have harnessed biomolecular recognition in combination with polymer engineering to create dynamic surfaces on which the accessibility of immobilized ligands to cell surface receptors can be reversibly interconverted under physiological conditions. The cell-adhesive RGD peptide is chosen as a model ligand. RGD is...

  1. Ligand modeling and design

    Energy Technology Data Exchange (ETDEWEB)

    Hay, B.P. [Pacific Northwest National Lab., Richland, WA (United States)

    1997-10-01

    The purpose of this work is to develop and implement a molecular design basis for selecting organic ligands that would be used in the cost-effective removal of specific radionuclides from nuclear waste streams. Organic ligands with metal ion specificity are critical components in the development of solvent extraction and ion exchange processes that are highly selective for targeted radionuclides. The traditional approach to the development of such ligands involves lengthy programs of organic synthesis and testing, which in the absence of reliable methods for screening compounds before synthesis, results in wasted research effort. The author`s approach breaks down and simplifies this costly process with the aid of computer-based molecular modeling techniques. Commercial software for organic molecular modeling is being configured to examine the interactions between organic ligands and metal ions, yielding an inexpensive, commercially or readily available computational tool that can be used to predict the structures and energies of ligand-metal complexes. Users will be able to correlate the large body of existing experimental data on structure, solution binding affinity, and metal ion selectivity to develop structural design criteria. These criteria will provide a basis for selecting ligands that can be implemented in separations technologies through collaboration with other DOE national laboratories and private industry. The initial focus will be to select ether-based ligands that can be applied to the recovery and concentration of the alkali and alkaline earth metal ions including cesium, strontium, and radium.

  2. Ligand modeling and design

    Energy Technology Data Exchange (ETDEWEB)

    Hay, B. [Pacific Northwest Lab., Richland, WA (United States)

    1996-10-01

    The purpose of this work is to develop and implement a molecular design basis for selecting organic ligands that would be used tin applications for the cost-effective removal of specific radionuclides from nuclear waste streams.

  3. Gut microbiota regulates NKG2D ligand expression on intestinal epithelial cells

    DEFF Research Database (Denmark)

    Hansen, Camilla Hartmann Friis; Holm, Thomas L.; Krych, Lukasz;

    2013-01-01

    Intestinal epithelial cells (IECs) are one of a few cell types in the body with constitutive surface expression of natural killer group 2 member D (NKG2D) ligands, although the magnitude of ligand expression by IECs varies. Here, we investigated whether the gut microbiota regulates the NKG2D ligand...... that the constitutive levels of NKG2D ligand expression on IECs are regulated by microbial signaling in the gut and further disfavor the intuitive notion that IEC NKG2D ligand expression is caused by low-grade immune reaction against commensal bacteria. It is more likely that constitutively high IEC NKG2D ligand...

  4. Stress stimulation and signal pathways of osteoprotegerin, receptor activator of nuclear factor kappa B and its ligand in bone metabolism%应力刺激与骨保护蛋白、核因子κB受体活化因子及其配体骨代谢信号通路

    Institute of Scientific and Technical Information of China (English)

    王丹; 鲍捷; 王国祥

    2012-01-01

    背景:骨保护蛋白、核因子κB 受体活化因子及其配体(osteoprote-gerin/ ligand of receptor activator of NF-κB/receptoractivator of NF-κB,OPG/RANKL/RANK)骨代谢信号通路是对应力敏感的通路之一.不同性质的运动会产生不同的机械应力刺激,影响骨代谢信号通路.目的:观察不同性质的运动对OPG/RANKL/RANK 信号通路的影响.方法:由第一作者于2000/2011 通过计算机检索 CNKI,HighWire 和Elsevier 数据库中关于" 应力刺激与OPG/RANKL/RANK"的相关的论文报告.以"应力刺激,OPG/RANKL/RANK"或"应力刺激,骨代谢"为检索词进行检索.选择的文章内容与应力刺激对信号通路的影响有关,选择相关近期发表的文献或者是发表在权威期刊的文献.共检索到215 篇文章,按纳入和排除标准对文献进行筛选,共纳入31 篇文章.结果与结论:运动对骨骼不断产生机械应力刺激,这种机械应力刺激可以通过影响成骨细胞和破骨细胞的OPG/RNAKL/RANK 信号调节系统而调节骨组织代谢.但是相关文献中的研究结果不一致,有待进一步的研究.%BACKGROUND: Signal pathway of osteoprotegerin, receptor activator of nuclear factor kappa B and its ligand in bone metabolismis one of the accesses to cope with the stress sensitivity. Different types of the exercise can produce different types of stressstimulation, and influence the signal pathways in bone metabolism.OBJECTIVE: To explore the influence of different types of exercise on the signal pathways of osteoprotegerin, receptor activator ofnuclear factor kappa B and its ligand in bone metabolism.METHODS: An online search of CNKI, HighWire and Elsevier databases was performed for articles related to "stress stimulation,osteoprotegerin, receptor activator of nuclear factor kappa B and its ligand". Articles published from 2000 to 2011 with thekeywords of "stress stimulation, osteoprotegerin, receptor activator of nuclear factor kappa B and its ligand" or

  5. Aryl hydrocarbon receptor ligand effects in RBL2H3 cells

    DEFF Research Database (Denmark)

    Maaetoft-Udsen, Kristina; Shimoda, Lori M. N.; Frøkiær, Hanne;

    2012-01-01

    (KA), Resveratrol, indolmycin, and violacein, affect mast cell activation and signaling. These ligands were tested on calcium signaling, degranulation, and gene expression. The data show that AHR is present in three model mast cell lines, and that various known and putative AHR ligands regulate gene...

  6. Organotellurium ligands - designing and complexation reactions

    Indian Academy of Sciences (India)

    Ajai K Singh

    2002-08-01

    A variety of tellurium ligands has been designed and studied for their complexation reactions in the last decade. Of these hybrid telluroethers, halotellurium ligands and polytellurides are the most notable ones. RTe- and polytelluride ions have also been used to design clusters. Ligation of ditelluroethers and several hybrid telluroethers is extensively studied in our laboratories. The ditelluroether ligand RTeCH2TeR (where R = 4-MeOC6H4) (1), similar to dppm [1,2-bis(diphenylphosphino) methane], has been synthesized in good yield (∼80 %) by reacting CHCl3 with RTe- (generated in situ by borohydride reduction of R2Te2). Iodine reacts with 1 to give tetra-iodo derivative, which has intermolecular Te$\\cdots$I interactions resulting in a macro structure containing rectangular Te-I$\\cdots$Te bridges. 1 readily forms four membered rings with Pd(II) and Ru(II). On the formation of this chelate ring, the signal in 125Te NMR spectra shifts significantly upfield (50-60 ppm). The bridging mode of 1 has been shown in [Ru(-cymene)Cl2](-1)[Ru(-cymene)Cl2]. The hybrid telluroether ligands explored are of the types (Te, S), (Te, N) and (Te, O). The tellurium donor site has strong trans influence, which is manifested more strongly in square planar complexes of palladium(II). The morpholine N-donor site has been found to have weaker donor characteristics in (Te, N) ligands than pyridine and alkylamine donor sites of analogous ligands. The singlet oxygen readily oxidises the coordinated Te. This oxidation follows first order kinetics. The complexation reaction of RuCl3.H2O with N-[2-(4-methoxyphenyltelluro)ethyl]phthalimide (2) results in a novel (Te, N, O)-heterocycle, Te-chloro,Te-anisyl-1a-aza-4-oxa-3-tellura-1H, 2H, 4aH-9 fluorenone. The (Te, O) ligands can be used as hemilabile ligands, the oxygen atom temporarily protects the vacant coordination site before the arrival of the substrate. The chelate shifts observed in 125Te NMR spectra of metal complexes of Te-ligands have

  7. AMPA receptor ligands

    DEFF Research Database (Denmark)

    Strømgaard, Kristian; Mellor, Ian

    2004-01-01

    polyamines are known to modulate the function of these receptors in vivo. In this study, recent developments in the medicinal chemistry of polyamine-based ligands are given, particularly focusing on the use of solid-phase synthesis (SPS) as a tool for the facile generation of libraries of polyamine toxin...

  8. KLIFS: a knowledge-based structural database to navigate kinase-ligand interaction space.

    Science.gov (United States)

    van Linden, Oscar P J; Kooistra, Albert J; Leurs, Rob; de Esch, Iwan J P; de Graaf, Chris

    2014-01-23

    Protein kinases regulate the majority of signal transduction pathways in cells and have become important targets for the development of designer drugs. We present a systematic analysis of kinase-ligand interactions in all regions of the catalytic cleft of all 1252 human kinase-ligand cocrystal structures present in the Protein Data Bank (PDB). The kinase-ligand interaction fingerprints and structure database (KLIFS) contains a consistent alignment of 85 kinase ligand binding site residues that enables the identification of family specific interaction features and classification of ligands according to their binding modes. We illustrate how systematic mining of kinase-ligand interaction space gives new insights into how conserved and selective kinase interaction hot spots can accommodate the large diversity of chemical scaffolds in kinase ligands. These analyses lead to an improved understanding of the structural requirements of kinase binding that will be useful in ligand discovery and design studies.

  9. Agonists and Antagonists of TGF-β Family Ligands.

    Science.gov (United States)

    Chang, Chenbei

    2016-08-01

    The discovery of the transforming growth factor β (TGF-β) family ligands and the realization that their bioactivities need to be tightly controlled temporally and spatially led to intensive research that has identified a multitude of extracellular modulators of TGF-β family ligands, uncovered their functions in developmental and pathophysiological processes, defined the mechanisms of their activities, and explored potential modulator-based therapeutic applications in treating human diseases. These studies revealed a diverse repertoire of extracellular and membrane-associated molecules that are capable of modulating TGF-β family signals via control of ligand availability, processing, ligand-receptor interaction, and receptor activation. These molecules include not only soluble ligand-binding proteins that were conventionally considered as agonists and antagonists of TGF-β family of growth factors, but also extracellular matrix (ECM) proteins and proteoglycans that can serve as "sink" and control storage and release of both the TGF-β family ligands and their regulators. This extensive network of soluble and ECM modulators helps to ensure dynamic and cell-specific control of TGF-β family signals. This article reviews our knowledge of extracellular modulation of TGF-β growth factors by diverse proteins and their molecular mechanisms to regulate TGF-β family signaling.

  10. Analysis of macromolecules, ligands and macromolecule-ligand complexes

    Science.gov (United States)

    Von Dreele, Robert B.

    2008-12-23

    A method for determining atomic level structures of macromolecule-ligand complexes through high-resolution powder diffraction analysis and a method for providing suitable microcrystalline powder for diffraction analysis are provided. In one embodiment, powder diffraction data is collected from samples of polycrystalline macromolecule and macromolecule-ligand complex and the refined structure of the macromolecule is used as an approximate model for a combined Rietveld and stereochemical restraint refinement of the macromolecule-ligand complex. A difference Fourier map is calculated and the ligand position and points of interaction between the atoms of the macromolecule and the atoms of the ligand can be deduced and visualized. A suitable polycrystalline sample of macromolecule-ligand complex can be produced by physically agitating a mixture of lyophilized macromolecule, ligand and a solvent.

  11. Ligand-Receptor Interactions

    CERN Document Server

    Bongrand, Pierre

    2008-01-01

    The formation and dissociation of specific noncovalent interactions between a variety of macromolecules play a crucial role in the function of biological systems. During the last few years, three main lines of research led to a dramatic improvement of our understanding of these important phenomena. First, combination of genetic engineering and X ray cristallography made available a simultaneous knowledg of the precise structure and affinity of series or related ligand-receptor systems differing by a few well-defined atoms. Second, improvement of computer power and simulation techniques allowed extended exploration of the interaction of realistic macromolecules. Third, simultaneous development of a variety of techniques based on atomic force microscopy, hydrodynamic flow, biomembrane probes, optical tweezers, magnetic fields or flexible transducers yielded direct experimental information of the behavior of single ligand receptor bonds. At the same time, investigation of well defined cellular models raised the ...

  12. Ligand exclusion on acetylcholinesterase.

    Science.gov (United States)

    Berman, H A; Leonard, K

    1990-11-27

    This paper examines covalent reactivity of AchE with respect to cationic and uncharged methylphosphonates and substrates in the absence and presence of cationic ligands selective for the active center and the peripheral anionic site. The organophosphorus inhibitors are enantiomeric alkyl methylphosphonothioates (1-5) containing cycloheptyl and isopropyl phosphono ester groups and S-methyl, S-n-pentyl, and S-[beta-(trimethylammonio)ethyl] leaving groups; these agents differ in their configuration about phosphorus and their steric, hydrophobic, and electrostatic characteristics. The synthetic substrates examined are acetylthiocholine, p-nitrophenyl acetate, and 7-acetoxy-4-methylcoumarin (7AMC). Antagonism of the methylphosphonothioate reaction by cationic ligands is strongly dependent on the nature of both the cation and the methylphosphonate but independent of the configuration about phosphorus. While all cations cause linear mixed inhibition of acetylthiocholine hydrolysis, there are observed a variety of inhibition patterns of 7AMC and p-nitrophenyl acetate hydrolysis that are distinctly nonlinear, as well as patterns in which the reciprocal plots intersect in the upper right quadrant. Strong antagonism of cationic (methylphosphonyl)thiocholines correlates very well with linear inhibition of acetylthiocholine. Ligands that cause only negligible antagonism of the uncharged methylphosphonates display nonlinear inhibition of uncharged substrates. These relationships, since they are most pronounced for peripheral site ligands and are strongly dependent on the charge carried by the reactant, suggest that the peripheral anionic site alters enzyme reactivity through an electrostatic interaction with the net negative active center. Such behavior indicates a potential role for the peripheral anionic site in conserving AchE catalytic efficiency within a narrow range of values. PMID:2271673

  13. Saturation-Transfer Difference (STD) NMR: A Simple and Fast Method for Ligand Screening and Characterization of Protein Binding

    Science.gov (United States)

    Viegas, Aldino; Manso, Joao; Nobrega, Franklin L.; Cabrita, Eurico J.

    2011-01-01

    Saturation transfer difference (STD) NMR has emerged as one of the most popular ligand-based NMR techniques for the study of protein-ligand interactions. The success of this technique is a consequence of its robustness and the fact that it is focused on the signals of the ligand, without any need of processing NMR information about the receptor…

  14. Serum concentrations of Flt-3 ligand in rheumatic diseases.

    Science.gov (United States)

    Nakamura, Kayo; Nakatsuka, Noriko; Jinnin, Masatoshi; Makino, Takamitsu; Kajihara, Ikko; Makino, Katsunari; Honda, Noritoshi; Inoue, Kuniko; Fukushima, Satoshi; Ihn, Hironobu

    2015-10-01

    Fms-like tyrosine kinase 3 (Flt-3) is a cytokine receptor expressed on the surface of bone-marrow progenitor of hematopoietic cells. Flt-3 ligands are produced by peripheral blood mononuclear cells, and found in various human body fluids. Flt-3 signal is involved in the regulation of vessel formation as well as B cell differentiation, suggesting that Flt-3 signal contributes to the pathogenesis of vascular abnormalities and immune dysregulation in rheumatic diseases. The aim of the present study is to examine serum Flt-3 ligand levels in patients with various rheumatic diseases, and to evaluate the possibility that serum Flt-3 ligand levels can be a useful disease marker. Sera were obtained from 20 dermatomyositis (DM) patients, 36 systemic sclerosis (SSc) patients, 10 systemic lupus erythematosus (SLE) patients, 10 scleroderma spectrum disorder (SSD) patients, 4 mixed connective tissue disease (MCTD) patients, and 12 normal subjects. Flt-3 ligand levels were determined with ELISA. Serum Flt-3 ligand levels were significantly elevated in patients with DM, SSc, SSD and MCTD compared to those in normal subjects. DM patients with elevated Flt-3 ligand levels were accompanied with significantly increased CRP levels and increased frequency of heliotrope rash than those with normal levels. In addition, SSc patients with elevated Flt-3 ligand levels showed significantly reduced frequency of nailfold bleeding. Serum Flt-3 ligand levels can be a marker of cutaneous manifestation in DM and a marker of microangiopathy in SSc. Clarifying the role of Flt-3 ligand in rheumatic diseases may lead to further understanding of these diseases and new therapeutic approaches. PMID:26559027

  15. Bexarotene ligand pharmaceuticals.

    Science.gov (United States)

    Hurst, R E

    2000-12-01

    Bexarotene (LGD-1069), from Ligand, was the first retinoid X receptor (RXR)-selective, antitumor retinoid to enter clinical trials. The company launched the drug for the treatment of cutaneous T-cell lymphoma (CTCL), as Targretin capsules, in the US in January 2000 [359023]. The company filed an NDA for Targretin capsules in June 1999, and for topical gel in December 1999 [329011], [349982] specifically for once-daily oral administration for the treatment of patients with early-stage CTCL who have not tolerated other therapies, patients with refractory or persistent early stage CTCL and patients with refractory advanced stage CTCL. The FDA approved Targretin capsules at the end of December 1999 for once-daily oral treatment of all stages of CTCL in patients refractory to at least one prior systemic therapy, at an initial dose of 300 mg/m2/day. After an NDA was submitted in December 1999 for Targretin gel, the drug received Priority Review status for use as a treatment of cutaneous lesions in patients with stage IA, IB or IIA CTCL [354836]. The FDA issued an approvable letter in June 2000, and granted marketing clearance for CTCL in the same month [370687], [372768], [372769], [373279]. Ligand had received Orphan Drug designation for this indication [329011]. At the request of the FDA, Ligand agreed to carry out certain post-approval phase IV and pharmacokinetic studies [351604]. The company filed an MAA with the EMEA for Targretin Capsules to treat lymphoma in November 1999 [348944]. The NDA for Targretin gel is based on a multicenter phase III trial that was conducted in the US, Canada, Europe and Australia involving 50 patients and a multicenter phase I/II clinical program involving 67 patients. Targretin gel was evaluated for the treatment of patients with early stage CTCL (IA-IIA) who were refractory to, intolerant to, or reached a response plateau for at least 6 months on at least two prior therapies. Efficacy results exceeded the protocol-defined response

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

    Directory of Open Access Journals (Sweden)

    Joshua D W Teo

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

  17. Molecular path for ligand search

    Institute of Scientific and Technical Information of China (English)

    Tao Lu; Yuan Yuan Qiao; Pan Wen Shen

    2011-01-01

    A ligand is a small molecule bind to several residues of a receptor. We adapt the concept of molecular path for effective ligand search with its contacting residues. Additionally, we allow wild type definitions on atoms and bonds of molecular paths for fuzzy algorithms on structural match. We choose hydrogen bond interactions to characterize the binding mode of a ligand by several proper molecular paths and use them to query the deposited ligands in PDBe that interact with their residues in the same way. Expression of molecular path and format of database entries are described with examples. Our molecular path provides a new approach to explore the ligand-receptor interactions and to provide structural framework reference on new ligand design.

  18. Macrocyclic G-quadruplex ligands

    DEFF Research Database (Denmark)

    Nielsen, M C; Ulven, Trond

    2010-01-01

    G-quadruplex stabilizing compounds have recently received increased interest due to their potential application as anticancer therapeutics. A significant number of structurally diverse G-quadruplex ligands have been developed. Some of the most potent and selective ligands currently known are...... macrocyclic structures which have been modeled after the natural product telomestatin or from porphyrin-based ligands discovered in the late 1990s. These two structural classes of G-quadruplex ligands are reviewed here with special attention to selectivity and structure-activity relationships, and with focus...

  19. Ligand-selective activation of heterologously-expressed mammalian olfactory receptor.

    Science.gov (United States)

    Ukhanov, K; Bobkov, Y; Corey, E A; Ache, B W

    2014-10-01

    Mammalian olfactory receptors (ORs) appear to have the capacity to couple to multiple G protein-coupled signaling pathways in a ligand-dependent selective manner. To better understand the mechanisms and molecular range of such ligand selectivity, we expressed the mouse eugenol OR (mOR-EG) in HEK293T cells together with Gα15 to monitor activation of the phospholipase-C (PLC) signaling pathway and/or Gαolf to monitor activation of the adenylate cyclase (AC) signaling pathway, resulting in intracellular Ca(2+) release and/or Ca(2+) influx through a cyclic nucleotide-gated channel, respectively. PLC-dependent responses differed dynamically from AC-dependent responses, allowing them to be distinguished when Gα15 and Gαolf were co-expressed. The dynamic difference in readout was independent of the receptor, the heterologous expression system, and the ligand concentration. Of 17 reported mOR-EG ligands tested, including eugenol, its analogs, and structurally dissimilar compounds (mousse cristal, nootkatone, orivone), some equally activated both signaling pathways, some differentially activated both signaling pathways, and some had no noticeable effect even at 1-5mM. Our findings argue that mOR-EG, when heterologously expressed, can couple to two different signaling pathways in a ligand selective manner. The challenge now is to determine the potential of mOR-EG, and perhaps other ORs, to activate multiple signaling pathways in a ligand selective manner in native ORNs. PMID:25149566

  20. CLE Peptides in Plants: Proteolytic Processing,Structure-Activity Relationship, and Ligand-Receptor Interaction

    Institute of Scientific and Technical Information of China (English)

    Xiaoming Gao; Yongfeng Guo

    2012-01-01

    Ligand-receptor signaling initiated by the CLAVATA3/ENDOSPERM SURROUNDING REGION (CLE) family peptides is critical in regulating cell division and differentiation in meristematic tissues in plants.Biologically active CLE peptides are released from precursor proteins via proteolytic processing.The mature form of CLE ligands consists of 12-13 amino acids with several post-translational modifications.This review summarizes recent progress toward understanding the proteolytic activities that cleave precursor proteins to release CLE peptides,the molecular structure and function of mature CLE ligands,and interactions between CLE ligands and corresponding leucine-rich repeat (LRR) receptor-like kinases (RLKs).

  1. Inductive expression of toll-like receptor 5 (TLR5) and associated downstream signaling molecules following ligand exposure and bacterial infection in the Indian major carp, mrigal (Cirrhinus mrigala).

    Science.gov (United States)

    Basu, M; Swain, B; Maiti, N K; Routray, P; Samanta, M

    2012-01-01

    Toll-like receptors (TLRs) are one of the key components of innate immunity. Among various types of TLRs, TLR5 is involved in recognizing bacterial flagellin and after binding, it triggers myeloid differentiation primary response gene 88 (MyD88)-dependent signaling pathway to induce pro-inflammatory cytokines. In this report, we analyzed the expression profile of TLR5 and its associated downstream signaling molecules like MyD88 and tumor necrosis factor (TNF) receptor-associated factor (TRAF) 6 in the Indian major carp (IMC), mrigal (Cirrhinus mrigala) which is highly commercially important fish species in the Indian subcontinent. Ontogeny analysis of TLR5, MyD88 and TRAF6 revealed constitutive expression of these genes in all embryonic developmental stages, and highlighted the importance of embryonic innate immune defense system in fish. Tissue specific expression analysis of these genes by quantitative real-time PCR (qRT-PCR) revealed their wide distribution in various organs and tissues; highest expression of TLR5 and MyD88 was in liver and TRAF6 was in kidney. Modulation of TLR5, MyD88 and TRAF6 gene expression, and the induction of interleukin (IL)-8 and TNF-α were analyzed in various organs by qRT-PCR following flagellin stimulation, and Aeromonas hydrophila and Edwardsiella tarda infection. In the treated fish, majority of the tested tissues exhibited significant induction of these genes, although with varied intensity among the tissues and with the types of treatments. Among the examined tissues, a significant relationship of TLR5 induction, MyD88 and TRAF6 up-regulation, and enhanced expression of IL-8 and TNF-α gene transcripts was observed in the blood and intestine of both flagellin stimulated and bacteria infected fish. These findings may indicate the involvement of TLR5 in inducing IL-8 and TNF-α, and suggest the important role of TLR5 in augmenting innate immunity in fish in response to pathogenic invasion. This study will enrich the information

  2. Post-Synapse Model Cell for Synaptic Glutamate Receptor (GluR-Based Biosensing: Strategy and Engineering to Maximize Ligand-Gated Ion-Flux Achieving High Signal-to-Noise Ratio

    Directory of Open Access Journals (Sweden)

    Tetsuya Haruyama

    2012-01-01

    Full Text Available Cell-based biosensing is a “smart” way to obtain efficacy-information on the effect of applied chemical on cellular biological cascade. We have proposed an engineered post-synapse model cell-based biosensors to investigate the effects of chemicals on ionotropic glutamate receptor (GluR, which is a focus of attention as a molecular target for clinical neural drug discovery. The engineered model cell has several advantages over native cells, including improved ease of handling and better reproducibility in the application of cell-based biosensors. However, in general, cell-based biosensors often have low signal-to-noise (S/N ratios due to the low level of cellular responses. In order to obtain a higher S/N ratio in model cells, we have attempted to design a tactic model cell with elevated cellular response. We have revealed that the increase GluR expression level is not directly connected to the amplification of cellular responses because the saturation of surface expression of GluR, leading to a limit on the total ion influx. Furthermore, coexpression of GluR with a voltage-gated potassium channel increased Ca2+ ion influx beyond levels obtained with saturating amounts of GluR alone. The construction of model cells based on strategy of amplifying ion flux per individual receptors can be used to perform smart cell-based biosensing with an improved S/N ratio.

  3. Exosomes in developmental signalling.

    Science.gov (United States)

    McGough, Ian John; Vincent, Jean-Paul

    2016-07-15

    In order to achieve coordinated growth and patterning during development, cells must communicate with one another, sending and receiving signals that regulate their activities. Such developmental signals can be soluble, bound to the extracellular matrix, or tethered to the surface of adjacent cells. Cells can also signal by releasing exosomes - extracellular vesicles containing bioactive molecules such as RNA, DNA and enzymes. Recent work has suggested that exosomes can also carry signalling proteins, including ligands of the Notch receptor and secreted proteins of the Hedgehog and WNT families. Here, we describe the various types of exosomes and their biogenesis. We then survey the experimental strategies used so far to interfere with exosome formation and critically assess the role of exosomes in developmental signalling. PMID:27436038

  4. Galaxy7TM: flexible GPCR-ligand docking by structure refinement.

    Science.gov (United States)

    Lee, Gyu Rie; Seok, Chaok

    2016-07-01

    G-protein-coupled receptors (GPCRs) play important physiological roles related to signal transduction and form a major group of drug targets. Prediction of GPCR-ligand complex structures has therefore important implications to drug discovery. With previously available servers, it was only possible to first predict GPCR structures by homology modeling and then perform ligand docking on the model structures. However, model structures generated without explicit consideration of specific ligands of interest can be inaccurate because GPCR structures can be affected by ligand binding. The Galaxy7TM server, freely accessible at http://galaxy.seoklab.org/7TM, improves an input GPCR structure by simultaneous ligand docking and flexible structure refinement using GALAXY methods. The server shows better performance in both ligand docking and GPCR structure refinement than commonly used programs AutoDock Vina and Rosetta MPrelax, respectively. PMID:27131365

  5. Role of the T cell receptor ligand affinity in T cell activation by bacterial superantigens

    DEFF Research Database (Denmark)

    Andersen, P S; Geisler, C; Buus, S;

    2001-01-01

    (SEC3) with up to a 150-fold increase in TCR affinity. By stimulating T cells with SEC3 molecules immobilized onto plastic surfaces, we demonstrate that increasing the affinity of the SEC3/TCR interaction caused a proportional increase in the ability of SEC3 to activate T cells. Thus, the potency......Similar to native peptide/MHC ligands, bacterial superantigens have been found to bind with low affinity to the T cell receptor (TCR). It has been hypothesized that low ligand affinity is required to allow optimal TCR signaling. To test this, we generated variants of Staphylococcus enterotoxin C3...... correlation between ligand affinity and ligand potency indicating that it is the density of receptor-ligand complexes in the T cell contact area that determines TCR signaling strength....

  6. Determinants governing ligand specificity of the Vibrio harveyi LuxN quorum-sensing receptor.

    Science.gov (United States)

    Ke, Xiaobo; Miller, Laura C; Bassler, Bonnie L

    2015-01-01

    Quorum sensing is a process of bacterial cell-cell communication that relies on the production, release and receptor-driven detection of extracellular signal molecules called autoinducers. The quorum-sensing bacterium Vibrio harveyi exclusively detects the autoinducer N-((R)-3-hydroxybutanoyl)-L-homoserine lactone (3OH-C4 HSL) via the two-component receptor LuxN. To discover the principles underlying the exquisite selectivity LuxN has for its ligand, we identified LuxN mutants with altered specificity. LuxN uses three mechanisms to verify that the bound molecule is the correct ligand: in the context of the overall ligand-binding site, His210 validates the C3 modification, Leu166 surveys the chain-length and a strong steady-state kinase bias imposes an energetic hurdle for inappropriate ligands to elicit signal transduction. Affinities for the LuxN kinase on and kinase off states underpin whether a ligand will act as an antagonist or an agonist. Mutations that bias LuxN to the agonized, kinase off, state are clustered in a region adjacent to the ligand-binding site, suggesting that this region acts as the switch that triggers signal transduction. Together, our analyses illuminate how a histidine sensor kinase differentiates between ligands and exploits those differences to regulate its signaling activity.

  7. Ligand Identification Scoring Algorithm (LISA)

    Science.gov (United States)

    Zheng, Zheng; Merz, Kenneth M.

    2011-01-01

    A central problem in de novo drug design is determining the binding affinity of a ligand with a receptor. A new scoring algorithm is presented that estimates the binding affinity of a protein-ligand complex given a three-dimensional structure. The method, LISA (Ligand Identification Scoring Algorithm), uses an empirical scoring function to describe the binding free energy. Interaction terms have been designed to account for van der Waals (VDW) contacts, hydrogen bonding, desolvation effects and metal chelation to model the dissociation equilibrium constants using a linear model. Atom types have been introduced to differentiate the parameters for VDW, H-bonding interactions and metal chelation between different atom pairs. A training set of 492 protein-ligand complexes was selected for the fitting process. Different test sets have been examined to evaluate its ability to predict experimentally measured binding affinities. By comparing with other well known scoring functions, the results show that LISA has advantages over many existing scoring functions in simulating protein-ligand binding affinity, especially metalloprotein-ligand binding affinity. Artificial Neural Network (ANN) was also used in order to demonstrate that the energy terms in LISA are well designed and do not require extra cross terms. PMID:21561101

  8. Memetic algorithms for ligand expulsion from protein cavities

    CERN Document Server

    Rydzewski, Jakub

    2015-01-01

    Ligand diffusion through a protein interior is a fundamental process governing biological signaling and enzymatic catalysis. A complex topology of channels in proteins leads often to difficulties in modeling ligand escape pathways by classical molecular dynamics simulations. In this paper two novel memetic methods for searching the exit paths and cavity space exploration are proposed: Memory Enhanced Random Acceleration (MERA) Molecular Dynamics and Immune Algorithm (IA). In MERA, a pheromone concept is introduced to optimize an expulsion force. In IA, hybrid learning protocols are exploited to predict ligand exit paths. They are tested on three protein channels with increasing complexity: M2 muscarinic GPCR receptor, enzyme nitrile hydratase and heme-protein cytochrome P450cam. In these cases, the memetic methods outperform Simulated Annealing and Random Acceleration Molecular Dynamics. The proposed algorithms are general and appropriate in all problems where an accelerated transport of an object through a n...

  9. Advances Towards The Discovery of GPR55 Ligands.

    Science.gov (United States)

    Morales, Paula; Jagerovic, Nadine

    2016-01-01

    The G-protein-coupled receptor 55 (GPR55) was identified in 1999. It was proposed as a novel member of the endocannabinoid system due to the fact that some endogenous, plant-derived and synthetic cannabinoid ligands act on GPR55. However, the complexity of the cellular downstream signaling pathways related to GPR55 activation delayed the discovery of selective GPR55 ligands. It was only a few years ago that the high throughput screening of libraries of pharmaceutical companies and governmental organizations allowed to identify selective GPR55 agonists and antagonists. Since then, several GPR55 modulator scaffolds have been reported. The relevance of GPR55 has been explored in diverse physiological and pathological processes revealing its role in inflammation, neuropathic pain, bone physiology, diabetes and cancer. Considering GPR55 as a new promising therapeutic target, there is a clear need for new selective and potent GPR55 modulators. This review will address a current structural update of GPR55 ligands.

  10. An NMR-Based Structural Rationale for Contrasting Stoichiometry and Ligand Binding Site(s) in Fatty Acid-binding Proteins†

    OpenAIRE

    He, Yan; Estephan, Rima; Yang, Xiaomin; Vela, Adriana; Wang, Hsin; Bernard, Cédric; Stark, Ruth E.

    2011-01-01

    Liver fatty acid-binding protein (LFABP) is a 14-kDa cytosolic polypeptide, differing from other family members in number of ligand binding sites, diversity of bound ligands, and transfer of fatty acid(s) to membranes primarily via aqueous diffusion rather than direct collisional interactions. Distinct two-dimensional 1H-15N NMR signals indicative of slowly exchanging LFABP assemblies formed during stepwise ligand titration were exploited, without solving the protein-ligand complex structures...

  11. Integrin receptors and ligand-gated channels.

    Science.gov (United States)

    Morini, Raffaella; Becchetti, Andrea

    2010-01-01

    Plastic expression of different integrin subunits controls the different stages of neural development, whereas in the adult integrins regulate synaptic stability. Evidence of integrin-channel crosstalk exists for ionotropic glutamate receptors. As is often the case in other tissues, integrin engagement regulates channel activity through complex signaling pathways that often include tyrosine phosphorylation cascades. The specific pathways recruited by integrin activation depend on cerebral region and cell type. In turn, ion channels control integrin expression onto the plasma membrane and their ligand binding affinity. The most extensive studies concern the hippocampus and suggest implications for neuronal circuit plasticity. The physiological relevance of these findings depends on whether adhesion molecules, aside from determining tissue stability, contribute to synaptogenesis and the responsiveness of mature synapses, thus contributing to long-term circuit consolidation. Little evidence is available for other ligand-gated channels, with the exception of nicotinic receptors. These exert a variety of functions in neurons and non neural tissue, both in development and in the adult, by regulating cell cycle, synaptogenesis and synaptic circuit refinement. Detailed studies in epidermal keratinocytes have shed some light on the possible mechanisms through which ACh can regulate cell motility, which may be of general relevance for morphogenetic processes. As to the control of mature synapses, most results concern the integrinic control of nicotinic receptors in the neuromuscular junction. Following this lead, a few studies have addressed similar topics in adult cerebral synapses. However, pursuing and interpreting these results in the brain is especially difficult because of the complexity of the nicotinic roles and the widespread contribution of nonsynaptic, paracrine transmission. From a pathological point of view, considering the well-known contribution of both

  12. Minireview: More than just a hammer: ligand "bias" and pharmaceutical discovery.

    Science.gov (United States)

    Luttrell, Louis M

    2014-03-01

    Conventional orthosteric drug development programs targeting G protein-coupled receptors (GPCRs) have focused on the concepts of agonism and antagonism, in which receptor structure determines the nature of the downstream signal and ligand efficacy determines its intensity. Over the past decade, the emerging paradigms of "pluridimensional efficacy" and "functional selectivity" have revealed that GPCR signaling is not monolithic, and that ligand structure can "bias" signal output by stabilizing active receptor states in different proportions than the native ligand. Biased ligands are novel pharmacologic entities that possess the unique ability to qualitatively change GPCR signaling, in effect creating "new receptors" with distinct efficacy profiles driven by ligand structure. The promise of biased agonism lies in this ability to engender "mixed" effects not attainable using conventional agonists or antagonists, promoting therapeutically beneficial signals while antagonizing deleterious ones. Indeed, arrestin pathway-selective agonists for the type 1 parathyroid hormone and angiotensin AT1 receptors, and G protein pathway-selective agonists for the GPR109A nicotinic acid and μ-opioid receptors, have demonstrated unique, and potentially therapeutic, efficacy in cell-based assays and preclinical animal models. Conversely, activating GPCRs in "unnatural" ways may lead to downstream biological consequences that cannot be predicted from prior knowledge of the actions of the native ligand, especially in the case of ligands that selectively activate as-yet poorly characterized G protein-independent signaling networks mediated via arrestins. Although much needs to be done to realize the clinical potential of functional selectivity, biased GPCR ligands nonetheless appear to be important new additions to the pharmacologic toolbox. PMID:24433041

  13. IDENTIFICATION OF SPECIFIC PEPTIDE LIGANDS FOR B-LYMPHOMA CELL AND ITS EFFECT ON TYROSINE PHOSPHORYLATION AND CELL APOPTOSIS

    Institute of Scientific and Technical Information of China (English)

    宋良文; 马宪梅; 崔雪梅; 李扬; 王晓民

    2004-01-01

    Objective To search novel method for diagnosis and therapy of B-lymphoma, specific small molecular peptide ligands against binding site of tumor cells were screened and its effects on signal transduction and cell apoptosis were tested. Methods Specific peptide ligands were screened by binding with site of human B lymphoma cell (OC1LY8) using peptide-bead libraries. The identified peptides were characterized with responsible cells by rebinding test. The role of tyrosine phosphorylation of peptide ligand was tested by Western blot;and its apoptosispromoting role was observed by confocal fluorescent microscope. Results Specific peptide ligand was able to bind specifically to site on cell surface and enter into cytoplasm. Tetrameric peptide ligand was able to strongly trigger signal transduction resulting in tyrosine phosphorylation and cellular apoptosis in OC1LY8 cell line.Conclusion Screened peptide ligand can effectively bind with OC1LY8 cell, stimulate cellular tyrosine phosphorylation and induce cellular apoptosis.

  14. NKG2D ligands mediate immunosurveillance of senescent cells.

    Science.gov (United States)

    Sagiv, Adi; Burton, Dominick G A; Moshayev, Zhana; Vadai, Ezra; Wensveen, Felix; Ben-Dor, Shifra; Golani, Ofra; Polic, Bojan; Krizhanovsky, Valery

    2016-02-01

    Cellular senescence is a stress response mechanism that limits tumorigenesis and tissue damage. Induction of cellular senescence commonly coincides with an immunogenic phenotype that promotes self-elimination by components of the immune system, thereby facilitating tumor suppression and limiting excess fibrosis during wound repair. The mechanisms by which senescent cells regulate their immune surveillance are not completely understood. Here we show that ligands of an activating Natural Killer (NK) cell receptor (NKG2D), MICA and ULBP2 are consistently up-regulated following induction of replicative senescence, oncogene-induced senescence and DNA damage - induced senescence. MICA and ULBP2 proteins are necessary for efficient NK-mediated cytotoxicity towards senescent fibroblasts. The mechanisms regulating the initial expression of NKG2D ligands in senescent cells are dependent on a DNA damage response, whilst continuous expression of these ligands is regulated by the ERK signaling pathway. In liver fibrosis, the accumulation of senescent activated stellate cells is increased in mice lacking NKG2D receptor leading to increased fibrosis. Overall, our results provide new insights into the mechanisms regulating the expression of immune ligands in senescent cells and reveal the importance of NKG2D receptor-ligand interaction in protecting against liver fibrosis. PMID:26878797

  15. A Fluid Membrane-Based Soluble Ligand Display System for Live CellAssays

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Jwa-Min; Nair, Pradeep N.; Neve, Richard M.; Gray, Joe W.; Groves, Jay T.

    2005-10-14

    Cell communication modulates numerous biological processes including proliferation, apoptosis, motility, invasion and differentiation. Correspondingly, there has been significant interest in the development of surface display strategies for the presentation of signaling molecules to living cells. This effort has primarily focused on naturally surface-bound ligands, such as extracellular matrix components and cell membranes. Soluble ligands (e.g. growth factors and cytokines) play an important role in intercellular communications, and their display in a surface-bound format would be of great utility in the design of array-based live cell assays. Recently, several cell microarray systems that display cDNA, RNAi, or small molecules in a surface array format were proven to be useful in accelerating high-throughput functional genetic studies and screening therapeutic agents. These surface display methods provide a flexible platform for the systematic, combinatorial investigation of genes and small molecules affecting cellular processes and phenotypes of interest. In an analogous sense, it would be an important advance if one could display soluble signaling ligands in a surface assay format that allows for systematic, patterned presentation of soluble ligands to live cells. Such a technique would make it possible to examine cellular phenotypes of interest in a parallel format with soluble signaling ligands as one of the display parameters. Herein we report a ligand-modified fluid supported lipid bilayer (SLB) assay system that can be used to functionally display soluble ligands to cells in situ (Figure 1A). By displaying soluble ligands on a SLB surface, both solution behavior (the ability to become locally enriched by reaction-diffusion processes) and solid behavior (the ability to control the spatial location of the ligands in an open system) could be combined. The method reported herein benefits from the naturally fluid state of the supported membrane, which allows

  16. Combined Treatment With Peroxisome Proliferator-Activated Receptor (PPAR) Gamma Ligands and Gamma Radiation Induces Apoptosis by PPARγ-Independent Up-Regulation of Reactive Oxygen Species-Induced Deoxyribonucleic Acid Damage Signals in Non-Small Cell Lung Cancer Cells

    Energy Technology Data Exchange (ETDEWEB)

    Han, Eun Jong; Im, Chang-Nim; Park, Seon Hwa [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Moon, Eun-Yi [Department of Bioscience and Biotechnology, Sejong University, Seoul (Korea, Republic of); Hong, Sung Hee, E-mail: gobrian@kcch.re.kr [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2013-04-01

    Purpose: To investigate possible radiosensitizing activities of the well-known peroxisome proliferator-activated receptor (PPAR)γ ligand ciglitazone and novel PPARγ ligands CAY10415 and CAY10506 in non-small cell lung cancer (NSCLC) cells. Methods and Materials: Radiosensitivity was assessed using a clonogenic cell survival assay. To investigate the mechanism underlying PPARγ ligand-induced radiosensitization, the subdiploid cellular DNA fraction was analyzed by flow cytometry. Activation of the caspase pathway by combined PPARγ ligands and γ-radiation treatment was detected by immunoblot analysis. Reactive oxygen species (ROS) were measured using 2,7-dichlorodihydrofluorescein diacetate and flow cytometry. Results: The 3 PPARγ ligands induced cell death and ROS generation in a PPARγ-independent manner, enhanced γ-radiation–induced apoptosis and caspase-3–mediated poly (ADP-ribose) polymerase (PARP) cleavage in vitro. The combined PPARγ ligand/γ-radiation treatment triggered caspase-8 activation, and this initiator caspase played an important role in the combination-induced apoptosis. Peroxisome proliferator-activated receptor-γ ligands may enhance the γ-radiation-induced DNA damage response, possibly by increasing γ-H2AX expression. Moreover, the combination treatment significantly increased ROS generation, and the ROS scavenger N-acetylcysteine inhibited the combined treatment-induced ROS generation and apoptotic cell death. Conclusions: Taken together, these results indicated that the combined treatment of PPARγ ligands and γ-radiation synergistically induced DNA damage and apoptosis, which was regulated by ROS.

  17. Visualization of Metal-to-Ligand and Ligand-to-Ligand Charge Transfer in Metal-Ligand Complexes

    Institute of Scientific and Technical Information of China (English)

    Yong Ding; Jian-xiu Guo; Xiang-si Wang; Sha-sha Liu; Feng-cai Ma

    2009-01-01

    Three methods including the atomic resolved density of state, charge difference density, and the transition density matrix are used to visualize metal to ligand charge transfer (MLCT) in ruthenium(Ⅱ) ammine complex. The atomic resolved density of state shows that there is density of Ru on the HOMOs. All the density is localized on the ammine, which reveals that the excited electrons in the Ru complex are delocalized over the ammine ligand. The charge difference density shows that all the holes are localized on the Ru and the electrons on the ammine. The localization explains the MLCT on excitation. The transition density matrix shows that there is electron-hole coherence between Ru and ammine. These methods are also used to examine the MLCT in Os(bpy)(p0p)Cl ("Osp0p"; bpy=2,2'-bipyridyl; p0p=4,4'-bipyridyl) and the ligand-to-ligand charge transfer (LLCT) in Alq3. The calculated results show that these methods are powerful to examine MLCT and LLCT in the metal-ligand system.

  18. Why mercury prefers soft ligands

    Energy Technology Data Exchange (ETDEWEB)

    Riccardi, Demian M [ORNL; Guo, Hao-Bo [ORNL; Gu, Baohua [ORNL; Parks, Jerry M [ORNL; Summers, Anne [University of Georgia, Athens, GA; Miller, S [University of California, San Francisco; Liang, Liyuan [ORNL; Smith, Jeremy C [ORNL

    2013-01-01

    Mercury (Hg) is a major global pollutant arising from both natural and anthropogenic sources. Defining the factors that determine the relative affinities of different ligands for the mercuric ion, Hg2+, is critical to understanding its speciation, transformation, and bioaccumulation in the environment. Here, we use quantum chemistry to dissect the relative binding free energies for a series of inorganic anion complexes of Hg2+. Comparison of Hg2+ ligand interactions in the gaseous and aqueous phases shows that differences in interactions with a few, local water molecules led to a clear periodic trend within the chalcogenide and halide groups and resulted in the well-known experimentally observed preference of Hg2+ for soft ligands such as thiols. Our approach establishes a basis for understanding Hg speciation in the biosphere.

  19. Inside job: ligand-receptor pharmacology beneath the plasma membrane

    Science.gov (United States)

    Babcock, Joseph J; Li, Min

    2013-01-01

    Most drugs acting on the cell surface receptors are membrane permeable and thus able to engage their target proteins in different subcellular compartments. However, these drugs' effects on cell surface receptors have historically been studied on the plasma membrane alone. Increasing evidence suggests that small molecules may also modulate their targeted receptors through membrane trafficking or organelle-localized signaling inside the cell. These additional modes of interaction have been reported for functionally diverse ligands of GPCRs, ion channels, and transporters. Such intracellular drug-target engagements affect cell surface expression. Concurrent intracellular and cell surface signaling may also increase the complexity and therapeutic opportunities of small molecule modulation. Here we discuss examples of ligand-receptor interactions that are present in both intra- and extracellular sites, and the potential therapeutic opportunities presented by this phenomenon. PMID:23685953

  20. Inside job: ligand-receptor pharmacology beneath the plasma membrane

    Institute of Scientific and Technical Information of China (English)

    Joseph J BABCOCK; Min LI

    2013-01-01

    Most drugs acting on the cell surface receptors are membrane permeable and thus able to engage their target proteins in different subcellular compartments.However,these drugs' effects on cell surface receptors have historically been studied on the plasma membrane alone.Increasing evidence suggests that small molecules may also modulate their targeted receptors through membrane trafficking or organelle-localized signaling inside the cell.These additional modes of interaction have been reported for functionally diverse ligands of GPCRs,ion channels,and transporters.Such intracellular drug-target engagements affect cell surface expression.Concurrent intracellular and cell surface signaling may also increase the complexity and therapeutic opportunities of small molecule modulation.Here we discuss examples of ligand-receptor interactions that are present in both intra- and extracellular sites,and the potential therapeutic opportunities presented by this phenomenon.

  1. Fringe-mediated extension of O-linked fucose in the ligand-binding region of Notch1 increases binding to mammalian Notch ligands.

    Science.gov (United States)

    Taylor, Paul; Takeuchi, Hideyuki; Sheppard, Devon; Chillakuri, Chandramouli; Lea, Susan M; Haltiwanger, Robert S; Handford, Penny A

    2014-05-20

    The Notch signaling pathway is essential for many aspects of development, cell fate determination, and tissue homeostasis. Notch signaling can be modulated by posttranslational modifications to the Notch receptor, which are known to alter both ligand binding and receptor activation. We have modified the ligand-binding region (EGF domains 11-13) of human Notch1 (hN1) with O-fucose and O-glucose glycans and shown by flow cytometry and surface plasmon resonance that the Fringe-catalyzed addition of GlcNAc to the O-fucose at T466 in EGF12 substantially increases binding to Jagged1 and Delta-like 1 (DLL1) ligands. We have subsequently determined the crystal structures of EGF domains 11-13 of hN1 modified with either the O-fucose monosaccharide or the GlcNAc-fucose disaccharide at T466 of EGF12 and observed no change in backbone structure for each variant. Collectively, these data demonstrate a role for GlcNAc in modulating the ligand-binding site in hN1 EGF12, resulting in an increased affinity of this region for ligands Jagged1 and DLL1. We propose that this finding explains the Fringe-catalyzed enhancement of Notch-Delta signaling observed in flies and humans, but suggest that the inhibitory effect of Fringe on Jagged/Serrate mediated signaling involves other regions of Notch.

  2. Probing heterotrimeric G protein activation: applications to biased ligands.

    OpenAIRE

    Denis, Colette; Saulière, Aude; Galandrin, Ségolène; Sénard, Jean-Michel; Galés, Céline

    2012-01-01

    Cell surface G protein-coupled receptors (GPCRs) drive numerous signaling pathways involved in the regulation of a broad range of physiologic processes. Today, they represent the largest target for modern drugs development with potential application in all clinical fields. Recently, the concept of “ligand-directed trafficking” has led to a conceptual revolution in pharmacological theory, thus opening new avenues for drug discovery. Accordingly, GPCRs do not function as simple on-off switch bu...

  3. A race for RAGE ligands.

    Science.gov (United States)

    Schleicher, Erwin D

    2010-08-01

    In experimental animals a causal involvement of the multiligand receptor for advanced glycation end products (RAGE) in the development of diabetic vascular complications has been demonstrated. However, the nature of RAGE ligands present in patients with diabetic nephropathy has not yet been defined; this leaves open the relevance of the RAGE system to the human disease.

  4. Lipoteichoic acid induces unique inflammatory responses when compared to other toll-like receptor 2 ligands.

    Directory of Open Access Journals (Sweden)

    Elizabeth M Long

    Full Text Available Toll-like receptors (TLRs recognize evolutionarily-conserved molecular patterns originating from invading microbes. In this study, we were interested in determining if microbial ligands, which use distinct TLR2-containing receptor complexes, represent unique signals to the cell and can thereby stimulate unique cellular responses. Using the TLR2 ligands, R-FSL1, S-FSL1, Pam2CSK4, Pam3CSK4, and lipoteichoic acid (LTA, we demonstrate that these ligands activate NF-kappaB and MAP Kinase pathways with ligand-specific differential kinetics in murine macrophages. Most strikingly, LTA stimulation of these pathways was substantially delayed when compared with the other TLR2 ligands. These kinetics differences were associated with a delay in the LTA-induced expression of a subset of genes as compared with another TLR2 ligand, R-FSL1. However, this did not translate to overall differences in gene expression patterns four hours following stimulation with different TLR2 ligands. We extended this study to evaluate the in vivo responses to distinct TLR2 ligands using a murine model of acute inflammation, which employs intravital microscopy to monitor leukocyte recruitment into the cremaster muscle. We found that, although R-FSL1, S-FSL1, Pam2CSK4, and Pam3CSK4 were all able to stimulate robust leukocyte recruitment in vivo, LTA remained functionally inert in this in vivo model. Therefore distinct TLR2 ligands elicit unique cellular responses, as evidenced by differences in the kinetic profiles of signaling and gene expression responses in vitro, as well as the physiologically relevant differences in the in vivo responses to these ligands.

  5. SELECTIVITY AND SPECIFICITY OF SPHINGOSINE 1-PHOSPHATE RECEPTOR LIGANDS: ‘OFF-TARGETS’ OR COMPLEX PHARMACOLOGY?

    Directory of Open Access Journals (Sweden)

    Nigel John Pyne

    2011-05-01

    Full Text Available A recent perspective published in frontiers of Pharmacology by Salomone and Waeber (2011 discussed the selectivity and specificity of sphingosine 1-phosphate (S1P receptor ligands. This perspective surveyed the use of various S1P receptor ligands and attempted to reconcile a number of inconsistencies in the predicted biological outcomes: these were interpreted as ‘off-target’ effects. Therefore the perspective cautioned against the use of these S1P receptor ligands. Here we highlight the complex pharmacology of S1P receptors, which along with ‘inside-out’ signalling might provide an alternative explanation for ‘off-target’ effects.

  6. plasticity of TGF-β signaling

    Directory of Open Access Journals (Sweden)

    Cellière Geraldine

    2011-11-01

    Full Text Available Abstract Background The family of TGF-β ligands is large and its members are involved in many different signaling processes. These signaling processes strongly differ in type with TGF-β ligands eliciting both sustained or transient responses. Members of the TGF-β family can also act as morphogen and cellular responses would then be expected to provide a direct read-out of the extracellular ligand concentration. A number of different models have been proposed to reconcile these different behaviours. We were interested to define the set of minimal modifications that are required to change the type of signal processing in the TGF-β signaling network. Results To define the key aspects for signaling plasticity we focused on the core of the TGF-β signaling network. With the help of a parameter screen we identified ranges of kinetic parameters and protein concentrations that give rise to transient, sustained, or oscillatory responses to constant stimuli, as well as those parameter ranges that enable a proportional response to time-varying ligand concentrations (as expected in the read-out of morphogens. A combination of a strong negative feedback and fast shuttling to the nucleus biases signaling to a transient rather than a sustained response, while oscillations were obtained if ligand binding to the receptor is weak and the turn-over of the I-Smad is fast. A proportional read-out required inefficient receptor activation in addition to a low affinity of receptor-ligand binding. We find that targeted modification of single parameters suffices to alter the response type. The intensity of a constant signal (i.e. the ligand concentration, on the other hand, affected only the strength but not the type of the response. Conclusions The architecture of the TGF-β pathway enables the observed signaling plasticity. The observed range of signaling outputs to TGF-β ligand in different cell types and under different conditions can be explained with

  7. Olfactory signaling in insects.

    Science.gov (United States)

    Wicher, Dieter

    2015-01-01

    The detection of volatile chemical information in insects is performed by three types of olfactory receptors, odorant receptors (ORs), specific gustatory receptor (GR) proteins for carbon dioxide perception, and ionotropic receptors (IRs) which are related to ionotropic glutamate receptors. All receptors form heteromeric assemblies; an OR complex is composed of an odor-specific OrX protein and a coreceptor (Orco). ORs and GRs have a 7-transmembrane topology as for G protein-coupled receptors, but they are inversely inserted into the membrane. Ligand-gated ion channels (ionotropic receptors) and ORs operate as IRs activated by volatile chemical cues. ORs are evolutionarily young receptors, and they first appear in winged insects and seem to be evolved to allow an insect to follow sparse odor tracks during flight. In contrast to IRs, the ORs can be sensitized by repeated subthreshold odor stimulation. This process involves metabotropic signaling. Pheromone receptors are especially sensitive and require an accessory protein to detect the lipid-derived pheromone molecules. Signaling cascades involved in pheromone detection depend on intensity and duration of stimuli and underlie a circadian control. Taken together, detection and processing of volatile information in insects involve ionotropic as well as metabotropic mechanisms. Here, I review the cellular signaling events associated with detection of cognate ligands by the different types of odorant receptors.

  8. Synergistic Effects of PPARγ Ligands and Retinoids in Cancer Treatment

    Directory of Open Access Journals (Sweden)

    Masahito Shimizu

    2008-01-01

    Full Text Available Peroxisome proliferator-activated receptors (PPARs are members of the nuclear receptor superfamily. The activation of PPARs by their specific ligands is regarded as one of the promising strategies to inhibit cancer cell growth. However, recent clinical trials targeting several common cancers showed no beneficial effect when PPAR ligands are used as a monotherapy. Retinoid X receptors (RXRs, which play a critical role in normal cell proliferation as a master regulator for nuclear receptors, preferentially form heterodimers with PPARs. A malfunction of RXRα due to phosphorylation by the Ras/MAPK signaling pathway is associated with the development of certain types of human malignancies. The activation of PPARγ/RXR heterodimer by their respective ligands synergistically inhibits cell growth, while inducing apoptosis in human colon cancer cells when the phosphorylation of RXRα was inhibited. We herein review the synergistic antitumor effects produced by the combination of the PPAR, especially PPARγ, ligands plus other agents, especially retinoids, in a variety of human cancers. We also focus on the phosphorylation of RXRα because the inhibition of RXRα phosphorylation and the restoration of its physiological function may activate PPAR/RXR heterodimer and, therefore, be a potentially effective and critical strategy for the inhibition of cancer cell growth.

  9. Fluorescent ligands for studying neuropeptide receptors by confocal microscopy

    Directory of Open Access Journals (Sweden)

    Beaudet A.

    1998-01-01

    Full Text Available This paper reviews the use of confocal microscopy as it pertains to the identification of G-protein coupled receptors and the study of their dynamic properties in cell cultures and in mammalian brain following their tagging with specific fluorescent ligands. Principles that should guide the choice of suitable ligands and fluorophores are discussed. Examples are provided from the work carried out in the authors' laboratory using custom synthetized fluoresceinylated or BODIPY-tagged bioactive peptides. The results show that confocal microscopic detection of specifically bound fluorescent ligands permits high resolution appraisal of neuropeptide receptor distribution both in cell culture and in brain sections. Within the framework of time course experiments, it also allows for a dynamic assessment of the internalization and subsequent intracellular trafficking of bound fluorescent molecules. Thus, it was found that neurotensin, somatostatin and mu- and delta-selective opioid peptides are internalized in a receptor-dependent fashion and according to receptor-specific patterns into their target cells. In the case of neurotensin, this internalization process was found to be clathrin-mediated, to proceed through classical endosomal pathways and, in neurons, to result in a mobilization of newly formed endosomes from neural processes to nerve cell bodies and from the periphery of cell bodies towards the perinuclear zone. These mechanisms are likely to play an important role for ligand inactivation, receptor regulation and perhaps also transmembrane signaling.

  10. Ligand chain length conveys thermochromism.

    Science.gov (United States)

    Ganguly, Mainak; Panigrahi, Sudipa; Chandrakumar, K R S; Sasmal, Anup Kumar; Pal, Anjali; Pal, Tarasankar

    2014-08-14

    Thermochromic properties of a series of non-ionic copper compounds have been reported. Herein, we demonstrate that Cu(II) ion with straight-chain primary amine (A) and alpha-linolenic (fatty acid, AL) co-jointly exhibit thermochromic properties. In the current case, we determined that thermochromism becomes ligand chain length-dependent and at least one of the ligands (A or AL) must be long chain. Thermochromism is attributed to a balanced competition between the fatty acids and amines for the copper(II) centre. The structure-property relationship of the non-ionic copper compounds Cu(AL)2(A)2 has been substantiated by various physical measurements along with detailed theoretical studies based on time-dependent density functional theory. It is presumed from our results that the compound would be a useful material for temperature-sensor applications. PMID:24943491

  11. Controlled-deactivation cannabinergic ligands.

    Science.gov (United States)

    Sharma, Rishi; Nikas, Spyros P; Paronis, Carol A; Wood, Jodianne T; Halikhedkar, Aneetha; Guo, Jason Jianxin; Thakur, Ganesh A; Kulkarni, Shashank; Benchama, Othman; Raghav, Jimit Girish; Gifford, Roger S; Järbe, Torbjörn U C; Bergman, Jack; Makriyannis, Alexandros

    2013-12-27

    We report an approach for obtaining novel cannabinoid analogues with controllable deactivation and improved druggability. Our design involves the incorporation of a metabolically labile ester group at the 2'-position on a series of (-)-Δ(8)-THC analogues. We have sought to introduce benzylic substituents α to the ester group which affect the half-lives of deactivation through enzymatic activity while enhancing the affinities and efficacies of individual ligands for the CB1 and CB2 receptors. The 1'-(S)-methyl, 1'-gem-dimethyl, and 1'-cyclobutyl analogues exhibit remarkably high affinities for both CB receptors. The novel ligands are susceptible to enzymatic hydrolysis by plasma esterases in a controllable manner, while their metabolites are inactive at the CB receptors. In further in vitro and in vivo experiments key analogues were shown to be potent CB1 receptor agonists and to exhibit CB1-mediated hypothermic and analgesic effects.

  12. Privileged chiral ligands and catalysts

    CERN Document Server

    Zhou, Qi-Lin

    2011-01-01

    This ultimate ""must have"" and long awaited reference for every chemist working in the field of asymmetric catalysis starts with the core structure of the catalysts, explaining why a certain ligand or catalyst is so successful. It describes in detail the history, the basic structural characteristics, and the applications of these ""privileged catalysts"". A novel concept that gives readers a much deeper insight into the topic.

  13. Sphingosine signaling and atherogenesis

    DEFF Research Database (Denmark)

    Xu, Cang-bao; Hansen-Schwartz, Jacob; Edvinsson, Lars

    2004-01-01

    Sphingosine-1-phosphate (S1P) has diverse biological functions acting inside cells as a second messenger to regulate cell proliferation and survival, and extracellularly, as a ligand for a group of G protein-coupled receptors (GPCRs) named the endothelial differentiation gene (EDG) family. Five......+ mobilization, and expression of adhesion molecules. The formation of an atherosclerotic lesion occurs through activation of cellular events that include monocyte adhesion to the endothelium and vascular smooth muscle cell (VSMC) migration and proliferation. Thus, S1P signaling may play an important role...

  14. Radioiodinated ligands for dopamine receptors

    International Nuclear Information System (INIS)

    The dopamine receptor system is important for normal brain function; it is also the apparent action site for various neuroleptic drugs for the treatment of schizophrenia and other metal disorders. In the past few years radioiodinated ligands for single photon emission tomography (SPECT) have been successfully developed and tested in humans: [123I]TISCH for D1 dopamine receptors; [123I]IBZM, epidepride, IBF and FIDA2, four iodobenzamide derivatives, for D2/D3 dopamine receptors. In addition, [123I]β-CIT (RTI-55) and IPT, cocaine derivatives, for the dopamine reuptake site are potentially useful for diagnosis of loss of dopamine neurons. The first iodinated ligand, (R)trans-7-OH-PIPAT, for D3 dopamine receptors, was synthesized and characterized with cloned cell lines (Spodoptera frugiperda, Sf9) expressing the D2 and D3 dopamine receptors and with rat basal forebrain membrane preparations. Most of the known iodobenzamides displayed similar potency in binding to both D2 and D3 dopamine receptors expressed in the cell lines. Initial studies appear to suggest that by fine tuning the structures it may be possible to develop agents specific for D2 and D3 dopamine receptors. It is important to investigate D2/D3 selectivity for this series of potent ligands

  15. EDA Signaling and Skin Appendage Development

    OpenAIRE

    Cui, Chang-Yi; Schlessinger, David

    2006-01-01

    The same morphogenetic signals are often involved in the development of different organs. For developing skin appendages, a model for tissue-specific regulation of signaling is provided by the EDA pathway, which accesses the otherwise ubiquitous NFκB transcription factors. EDA signaling is mediated by ectodysplasin, EDAR and EDARADD, which form a new TNF ligand-receptor-adaptor family that is restricted to skin appendages in vertebrates from fish to human. The critical function of the pathway...

  16. Benchmarking the Predictive Power of Ligand Efficiency Indices in QSAR.

    Science.gov (United States)

    Cortes-Ciriano, Isidro

    2016-08-22

    Compound physicochemical properties favoring in vitro potency are not always correlated to desirable pharmacokinetic profiles. Therefore, using potency (i.e., IC50) as the main criterion to prioritize candidate drugs at early stage drug discovery campaigns has been questioned. Yet, the vast majority of the virtual screening models reported in the medicinal chemistry literature predict the biological activity of compounds by regressing in vitro potency on topological or physicochemical descriptors. Two studies published in this journal showed that higher predictive power on external molecules can be achieved by using ligand efficiency indices as the dependent variable instead of a metric of potency (IC50) or binding affinity (Ki). The present study aims at filling the shortage of a thorough assessment of the predictive power of ligand efficiency indices in QSAR. To this aim, the predictive power of 11 ligand efficiency indices has been benchmarked across four algorithms (Gradient Boosting Machines, Partial Least Squares, Random Forest, and Support Vector Machines), two descriptor types (Morgan fingerprints, and physicochemical descriptors), and 29 data sets collected from the literature and ChEMBL database. Ligand efficiency metrics led to the highest predictive power on external molecules irrespective of the descriptor type or algorithm used, with an R(2)test difference of ∼0.3 units and a this difference ∼0.4 units when modeling small data sets and a normalized RMSE decrease of >0.1 units in some cases. Polarity indices, such as SEI and NSEI, led to higher predictive power than metrics based on molecular size, i.e., BEI, NBEI, and LE. LELP, which comprises a polarity factor (cLogP) and a size parameter (LE) constantly led to the most predictive models, suggesting that these two properties convey a complementary predictive signal. Overall, this study suggests that using ligand efficiency indices as the dependent variable might be an efficient strategy to model

  17. Interleukin-24 inhibits tumor cell metastasis through (C-X-C motif) ligand 12/(C-X-C motif) receptor 4 signal axis%白细胞介素-24通过CXC趋化因子配体12/CXC趋化因子受体4信号轴抑制肿瘤细胞转移

    Institute of Scientific and Technical Information of China (English)

    杨俊波; 黄晓洁

    2015-01-01

    目的 探讨白细胞介素(IL)-24在体外实验中通过抑制CXC趋化因子配体12/CXC趋化因子受体4 (CXCL12/CXCR4)信号轴而抑制肺癌细胞迁移和侵袭.方法 以慢病毒介导IL-24基因转染H1299细胞为研究对象,研究IL-24对CXCL12/CXCR4信号轴的抑制作用.采用Western blot和Transwell细胞迁移及细胞侵袭实验评价其生物学效应.结果 IL-24在H1299-IL24细胞中的表达明显抑制了CXCR4 mRNA及蛋白的表达(P<0.05).IL-24通过下调CXCR4的表达明显抑制了肿瘤细胞的迁移和侵袭,H1299-IL24细胞的迁移数量较对照细胞下降了约30%,侵袭能力比较对照组下降30% ~40%,此外IL-24联合CXCR4抑制剂AMD3100使用时,表现出对肿瘤细胞迁移能力更强的抑制能力(P<0.05).结论 IL-24通过抑制CXCL12/CXCR4信号通路,从而抑制肿瘤细胞的迁移和侵袭.此外,当IL-24联合CXCR4抑制剂使用时,表现出更强的对抗肿瘤转移的能力.%Objective To investigate whetheinterleukin-24 (IL-24) could inhibithe chemokine (C-X-motif) ligand (CXCL)-12/chemokine (C-X-motif) recepto(CXCR)-4 signaling pathway and suppreslung cancecell migration and invasion in vitro.MethodThe lung cancecell line H1299 wastably transfected with recombinanlentiviral vectorharboring open reading frame (ORF) of IL-24 and used in the presenstudy to determine the inhibitory effectof IL-24 on CXCL12/CXCR4 axis.The inhibitory effectof IL-24 on CXCL12/CXCR4 were assessed by Western blotting.Biological function wastudied using cell migration and invasion assays.ResultIL-24 expression in the H1299-IL24 cell line resulted in reduced CXCR4 expression on both mRNand protein level(P < 0.05).Functional studieshowed thaIL-24 inhibited tumocell migration (30% decrease) and invasion (30%-40% decrease, P <0.05).Finally, IL-24, when combined with CXCR4 inhibito(AMD3100), demonstrated enhanced inhibitory activity on tumocell migration (P < 0.05).Conclusion IL-24 interferewith the CXCL12

  18. Ligand placement based on prior structures: the guided ligand-replacement method

    Energy Technology Data Exchange (ETDEWEB)

    Klei, Herbert E. [Lawrence Berkeley National Laboratory, Berkeley, CA 94720 (United States); Bristol-Myers Squibb, Princeton, NJ 08543-4000 (United States); Moriarty, Nigel W., E-mail: nwmoriarty@lbl.gov; Echols, Nathaniel [Lawrence Berkeley National Laboratory, Berkeley, CA 94720 (United States); Terwilliger, Thomas C. [Los Alamos National Laboratory, Los Alamos, NM 87545-0001 (United States); Baldwin, Eric T. [Bristol-Myers Squibb, Princeton, NJ 08543-4000 (United States); Natural Discovery LLC, Princeton, NJ 08542-0096 (United States); Pokross, Matt; Posy, Shana [Bristol-Myers Squibb, Princeton, NJ 08543-4000 (United States); Adams, Paul D. [Lawrence Berkeley National Laboratory, Berkeley, CA 94720 (United States); University of California at Berkeley, Berkeley, CA 94720-1762 (United States)

    2014-01-01

    A new module, Guided Ligand Replacement (GLR), has been developed in Phenix to increase the ease and success rate of ligand placement when prior protein-ligand complexes are available. The process of iterative structure-based drug design involves the X-ray crystal structure determination of upwards of 100 ligands with the same general scaffold (i.e. chemotype) complexed with very similar, if not identical, protein targets. In conjunction with insights from computational models and assays, this collection of crystal structures is analyzed to improve potency, to achieve better selectivity and to reduce liabilities such as absorption, distribution, metabolism, excretion and toxicology. Current methods for modeling ligands into electron-density maps typically do not utilize information on how similar ligands bound in related structures. Even if the electron density is of sufficient quality and resolution to allow de novo placement, the process can take considerable time as the size, complexity and torsional degrees of freedom of the ligands increase. A new module, Guided Ligand Replacement (GLR), was developed in Phenix to increase the ease and success rate of ligand placement when prior protein–ligand complexes are available. At the heart of GLR is an algorithm based on graph theory that associates atoms in the target ligand with analogous atoms in the reference ligand. Based on this correspondence, a set of coordinates is generated for the target ligand. GLR is especially useful in two situations: (i) modeling a series of large, flexible, complicated or macrocyclic ligands in successive structures and (ii) modeling ligands as part of a refinement pipeline that can automatically select a reference structure. Even in those cases for which no reference structure is available, if there are multiple copies of the bound ligand per asymmetric unit GLR offers an efficient way to complete the model after the first ligand has been placed. In all of these applications, GLR

  19. Stress Sensitive Signal Pathway Osteoprotegerin/Receptor Activator of Nuclear Factor-κB ligand/Receptor Activator of Nuclear Fac-tor-κB and Osteoporosis (review)%骨保护素/核因子-κB受体活化因子配体/核因子-κB受体活化因子信号通路与骨质疏松的研究进展

    Institute of Scientific and Technical Information of China (English)

    李盛村; 鲍捷; 王静; 王国祥

    2014-01-01

    Exercise is benefic for osteoporosis, without clear molecular biology mechanism. The osteoprotegerin/receptor activator of nuclear factor-κB ligand/receptor activator of nuclear factor-κB (OPG/RANKL/RANK) signal pathway contributes to osteoporosis, which can be mediated by mechanical force. Research progress on osteoporosis and the stress sensitive signal pathway OPG/RANKL/RANK were reviewed in this paper.%运动对骨质疏松症有积极作用,但其治疗的分子生物学机制仍未清楚。骨保护素/核因子-κB受体活化因子配体/核因子-κB受体活化因子(OPG/RANKL/RANK)信号通路的发现,有助于骨质疏松症的治疗。机械应力可以调节OPG/RANKL/RANK信号通路,参与预防和治疗骨质疏松进程。本文就应力敏感信号通路OPG/RANKL/RANK与骨质疏松的关系进行综述。

  20. Engineering and optimization of an allosteric biosensor protein for peroxisome proliferator-activated receptor γ ligands.

    Science.gov (United States)

    Li, Jingjing; Gierach, Izabela; Gillies, Alison R; Warden, Charles D; Wood, David W

    2011-11-15

    The peroxisome proliferator-activated receptor gamma (PPARγ or PPARG) belongs to the nuclear receptor superfamily, and is a potential drug target for a variety of diseases. In this work, we constructed a series of bacterial biosensors for the identification of functional PPARγ ligands. These sensors entail modified Escherichia coli cells carrying a four-domain fusion protein, comprised of the PPARγ ligand binding domain (LBD), an engineered mini-intein domain, the E. coli maltose binding protein (MBD), and a thymidylate synthase (TS) reporter enzyme. E. coli cells expressing this protein exhibit hormone ligand-dependent growth phenotypes. Unlike our published estrogen (ER) and thyroid receptor (TR) biosensors, the canonical PPARγ biosensor cells displayed pronounced growth in the absence of ligand. They were able to distinguish agonists and antagonists, however, even in the absence of agonist. To improve ligand sensitivity of this sensor, we attempted to engineer and optimize linker peptides flanking the PPARγ LBD insertion point. Truncation of the original linkers led to decreased basal growth and significantly enhanced ligand sensitivity of the PPARγ sensor, while substitution of the native linkers with optimized G(4)S (Gly-Gly-Gly-Gly-Ser) linkers further increased the sensitivity. Our studies demonstrate that the properties of linkers, especially the C-terminal linker, greatly influence the efficiency and fidelity of the allosteric signal induced by ligand binding. Our work also suggests an approach to increase allosteric behavior in this multidomain sensor protein, without modification of the functional LBD. PMID:21893405

  1. PANP is a novel O-glycosylated PILRα ligand expressed in neural tissues

    International Nuclear Information System (INIS)

    Research highlights: → A Novel molecule, PANP, was identified to be a PILRα ligand. → Sialylated O-glycan structures on PANP were required for PILRα recognition. → Transcription of PANP was mainly observed in neural tissues. → PANP seems to be involved in immune regulation as a ligand for PILRα. -- Abstract: PILRα is an immune inhibitory receptor possessing an immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic domain enabling it to deliver inhibitory signals. Binding of PILRα to its ligand CD99 is involved in immune regulation; however, whether there are other PILRα ligands in addition to CD99 is not known. Here, we report that a novel molecule, PILR-associating neural protein (PANP), acts as an additional ligand for PILRα. Transcription of PANP was mainly observed in neural tissues. PILRα-Ig fusion protein bound cells transfected with PANP and the transfectants stimulated PILRα reporter cells. Specific O-glycan structures on PANP were found to be required for PILR recognition of this ligand. These results suggest that PANP is involved in immune regulation as a ligand of the PILRα.

  2. Revealing a steroid receptor ligand as a unique PPARγagonist

    Institute of Scientific and Technical Information of China (English)

    Shengchen Lin; Ying Han; Yuzhe Shi; Hui Rong; Songyang Zheng; Shikan Jin; Shu-Yong Lin; Sheng-Cai Lin; Yong Li

    2012-01-01

    Peroxisome proliferator-activated receptor gamma (PPARγ) regulates metabolic homeostasis and is a molecular target for anti-diabetic drugs.We report here the identification of a steroid receptor ligand,RU-486,as an unexpected PPARγ agonist,thereby uncovering a novel signaling route for this steroid drug.Similar to rosiglitazone,RU486 modulates the expression of key PPARγ target genes and promotes adipocyte differentiation,but with a lower adipogenic activity.Structural and functional studies of receptor-ligand interactions reveal the molecular basis for a unique binding mode for RU-486 in the PPARγ ligand-binding pocket with distinctive properties and epitopes,providing the molecular mechanisms for the discrimination of RU-486 from thiazolidinediones (TZDs) drugs.Our findings together indicate that steroid compounds may represent an alternative approach for designing non-TZD PPARγligands in the treatment of insulin resistance.

  3. Revealing a steroid receptor ligand as a unique PPAR[gamma] agonist

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Shengchen; Han, Ying; Shi, Yuzhe; Rong, Hui; Zheng, Songyang; Jin, Shikan; Lin, Shu-Yong; Lin, Sheng-Cai; Li, Yong (Pitt); (Xiamen)

    2012-06-28

    Peroxisome proliferator-activated receptor gamma (PPAR{gamma}) regulates metabolic homeostasis and is a molecular target for anti-diabetic drugs. We report here the identification of a steroid receptor ligand, RU-486, as an unexpected PPAR{gamma} agonist, thereby uncovering a novel signaling route for this steroid drug. Similar to rosiglitazone, RU-486 modulates the expression of key PPAR{gamma} target genes and promotes adipocyte differentiation, but with a lower adipogenic activity. Structural and functional studies of receptor-ligand interactions reveal the molecular basis for a unique binding mode for RU-486 in the PPAR{gamma} ligand-binding pocket with distinctive properties and epitopes, providing the molecular mechanisms for the discrimination of RU-486 from thiazolidinediones (TZDs) drugs. Our findings together indicate that steroid compounds may represent an alternative approach for designing non-TZD PPAR{gamma} ligands in the treatment of insulin resistance.

  4. Ligand photo-isomerization triggers conformational changes in iGluR2 ligand binding domain.

    Directory of Open Access Journals (Sweden)

    Tino Wolter

    Full Text Available Neurological glutamate receptors bind a variety of artificial ligands, both agonistic and antagonistic, in addition to glutamate. Studying their small molecule binding properties increases our understanding of the central nervous system and a variety of associated pathologies. The large, oligomeric multidomain membrane protein contains a large and flexible ligand binding domains which undergoes large conformational changes upon binding different ligands. A recent application of glutamate receptors is their activation or inhibition via photo-switchable ligands, making them key systems in the emerging field of optochemical genetics. In this work, we present a theoretical study on the binding mode and complex stability of a novel photo-switchable ligand, ATA-3, which reversibly binds to glutamate receptors ligand binding domains (LBDs. We propose two possible binding modes for this ligand based on flexible ligand docking calculations and show one of them to be analogues to the binding mode of a similar ligand, 2-BnTetAMPA. In long MD simulations, it was observed that transitions between both binding poses involve breaking and reforming the T686-E402 protein hydrogen bond. Simulating the ligand photo-isomerization process shows that the two possible configurations of the ligand azo-group have markedly different complex stabilities and equilibrium binding modes. A strong but slow protein response is observed after ligand configuration changes. This provides a microscopic foundation for the observed difference in ligand activity upon light-switching.

  5. Estrogen Signaling and Cardiovascular Disease

    OpenAIRE

    Murphy, Elizabeth

    2011-01-01

    Estrogen has pleiotropic effects on the cardiovascular system. The mechanisms by which estrogen confers these pleiotropic effects on cardiovascular function is under active investigation. Until a decade ago, all estrogen signaling was thought to occur by estrogen binding to nuclear estrogen receptors (ERα and ERβ), which bind to DNA and function as ligand activated transcription factors. Estrogen binding to the receptor alters gene expression thereby altering cell function. In 2000 estrogen w...

  6. Ruthenium Cumulenylidene Complexes Bearing Heteroscorpionate Ligands

    OpenAIRE

    Strinitz, Frank

    2014-01-01

    In previous work of the BURZLAFF group, the design of suitable N,N,O ligands for a wide variety of applications ranging from catalysis to bioinorganic model compounds has been extensively investigated. Especially the methyl substituted bis(3,5-dimethylpyrazol-1-yl) acetate (bdmpza) ligand has shown manifold chemistry, comparable to the anionic cyclopentadienyl (Cp) and hydridotris(pyrazol-1-yl)borato (Tp) ligand. In the first part of this thesis the new tricarbonylmanganese(I) complexes be...

  7. Trafficking of epidermal growth factor receptor ligands in polarized epithelial cells.

    Science.gov (United States)

    Singh, Bhuminder; Coffey, Robert J

    2014-01-01

    A largely unilamellar epithelial layer lines body cavities and organ ducts such as the digestive tract and kidney tubules. This polarized epithelium is composed of biochemically and functionally separate apical and basolateral surfaces. The epidermal growth factor receptor (EGFR) signaling pathway is a critical regulator of epithelial homeostasis and is perturbed in a number of epithelial disorders. It is underappreciated that in vivo EGFR signaling is most often initiated by cell-surface delivery and processing of one of seven transmembrane ligands, resulting in release of the soluble form that binds EGFR. In polarized epithelial cells, EGFR is restricted largely to the basolateral surface, and apical or basolateral ligand delivery therefore has important biological consequences. In vitro approaches have been used to study the biosynthesis, cell-surface delivery, proteolytic processing, and release of soluble EGFR ligands in polarized epithelial cells. We review these results, discuss their relevance to normal physiology, and demonstrate the pathophysiological consequences of aberrant trafficking. These studies have uncovered a rich diversity of apico-basolateral trafficking mechanisms among the EGFR ligands, provided insights into the pathogenesis of an inherited magnesium-wasting disorder of the kidney (isolated renal hypomagnesemia), and identified a new mode of EGFR ligand signaling via exosomes. PMID:24215440

  8. Clinical Use of PPARγ Ligands in Cancer

    Directory of Open Access Journals (Sweden)

    Jennifer L. Hatton

    2008-01-01

    Full Text Available The role of PPARγ in adipocyte differentiation has fueled intense interest in the function of this steroid nuclear receptor for regulation of malignant cell growth and differentiation. Given the antiproliferative and differentiating effects of PPARγ ligands on liposarcoma cells, investigation of PPARγ expression and ligand activation in other solid tumors such as breast, colon, and prostate cancers ensued. The anticancer effects of PPARγ ligands in cell culture and rodent models of a multitude of tumor types suggest broad applicability of these agents to cancer therapy. This review focuses on the clinical use of PPARγ ligands, specifically the thiazolidinediones, for the treatment and prevention of cancer.

  9. Expression and function of Delta-like ligand 4 in a rat model of retinopathy of prematurity

    Institute of Scientific and Technical Information of China (English)

    Shaoyang Shi; Xun Li; You Li; Cunwen Pei; Hongwei Yang; Xiaolong Chen

    2013-01-01

    The Delta-like ligand 4/Notch signaling pathway was shown to participate in the process of retinal development and angiogenesis. However, the function of the Delta-like ligand 4/Notch signaling pathway in retinopathy of prematurity requires further study. Retinopathy of prematurity was induced in 5-day-old Sprague-Dawley rats exposed to hyperoxia for 7 days, and then returned to room air. Reverse transcription-PCR and western blot revealed that Delta-like ligand 4 levels decreased at postnatal day 12 and increased at postnatal day 17 in retinopathy of prematurity rats. Flat-mounted adenosine diphosphatase stained retina and hematoxylin-eosin stained retinal tissue slices showed that the clock hour scores and the nuclei counts in retinopathy of prematurity rats were significantly different compared to normal control rats. After retinopathy of prematurity rats were intravitreally injected with Delta-like ligand 4 monoclonal antibody to inhibit the Delta-like ligand 4/Notch signaling pathway, there was a significant increase in the severity of retinal neovascularization (clock hours) in the intravitreally injected eyes. The nuclei count was highly correlated with the clock hour score. These results suggest that Delta-like ligand 4/Notch signaling plays an essential role in the process of physiological and pathological angiogenesis in the retina.

  10. The sigma receptor as a ligand-regulated auxiliary potassium channel subunit.

    Science.gov (United States)

    Aydar, Ebru; Palmer, Christopher P; Klyachko, Vitaly A; Jackson, Meyer B

    2002-04-25

    The sigma receptor is a novel protein that mediates the modulation of ion channels by psychotropic drugs through a unique transduction mechanism depending neither on G proteins nor protein phosphorylation. The present study investigated sigma receptor signal transduction by reconstituting responses in Xenopus oocytes. Sigma receptors modulated voltage-gated K+ channels (Kv1.4 or Kv1.5) in different ways in the presence and absence of ligands. Association between Kv1.4 channels and sigma receptors was demonstrated by coimmunoprecipitation. These results indicate a novel mechanism of signal transduction dependent on protein-protein interactions. Domain accessibility experiments suggested a structure for the sigma receptor with two cytoplasmic termini and two membrane-spanning segments. The ligand-independent effects on channels suggest that sigma receptors serve as auxiliary subunits to voltage-gated K+ channels with distinct functional interactions, depending on the presence or absence of ligand.

  11. Stochastic description of the ligand-receptor interaction of biologically active substances at extremely low doses.

    Science.gov (United States)

    Gurevich, Konstantin G; Agutter, Paul S; Wheatley, Denys N

    2003-04-01

    Signalling molecules can be effective at extraordinarily low concentrations (down to attomolar levels). To handle such cases, probabilistic methods have been used to describe the formal kinetics of action of biologically active substances in these low doses, although it has been necessary to review what is meant by such a term. The mean numbers of transformed/degraded molecules and their dispersions were calculated for the possible range of ligand-receptor binding schemes. We used both analytical equations and numerical simulations to calculate the coefficients of variation (ratio of standard deviation to mean) and demonstrated that the distribution of the coefficient is highly dependent on the reaction scheme. It may, therefore, be used as an additional factor for discriminating between cooperative and noncooperative models of ligand-receptor interaction over extreme ranges of ligand dilution. The relevance to signalling behaviour is discussed.

  12. Macrocyclic ligands for uranium complexation

    International Nuclear Information System (INIS)

    A highly preorganized 24-macrocycle containing biuret, thiobiuret and pyridine subunits has been prepared by high dilution ring-closure procedures. Intermediate products to this macrocycle have been utilized to extend this synthetic route to include further representatives where solubility and stability will be influenced by substituent variation. A 1:1 complex has been formed from uranyl acetate and a quinquepyridine derivative, this representing a new type of ligand for the uranyl ion. A very convenient synthetic procedure that will allow the incorporation of these macrocycles into polymeric systems has been developed for the introduction of a vinyl substituent into the 4-position of the pyridine ring. Using triflate, vinyltributyltin and Pd0 chemistry, this procedure should make a variety of substituted 4-vinylpyridines available for the first time. 3 refs

  13. Synthesis and characterization of mixed ligand chiral nanoclusters

    OpenAIRE

    Güven, Zekiye Pelin; Guven, Zekiye Pelin; Üstbaş, Burçin; Ustbas, Burcin; Harkness, Kellen M.; Coşkun, Hikmet; Coskun, Hikmet; Joshi, Chakra P.; Besong, Tabot M. D.; Stellacci, Francesco; Bakr, Osman M.; Akbulut, Özge; Akbulut, Ozge

    2015-01-01

    Chiral mixed ligand silver nanoclusters were synthesized in the presence of a chiral and an achiral ligand. The ratio of the ligands was changed to track the formation of these clusters. While the chiral ligand lead to nanoparticles, Presence of the achiral ligand induced the formation of nanoclusters with chiral properties.

  14. Rhodium olefin complexes of diiminate type ligands

    NARCIS (Netherlands)

    Willems, Sander Theodorus Hermanus

    2003-01-01

    The mono-anionic beta-diiminate ligand (ArNC(CH3)CHC(CH3)NAr) on several previous occasions proved useful in stabilising low coordination numbers for both early and late transition metals. In this thesis the reactivity of the rhodium olefin complexes of one of these beta-diiminate ligands (Ar = 2,6-

  15. Flexible Ligand Docking Using Differential Evolution

    DEFF Research Database (Denmark)

    Thomsen, René

    2003-01-01

    the most favorable energetic conformation among the large space of possible protein-ligand complexes. Stochastic search methods, such as evolutionary algorithms (EAs), can be used to sample large search spaces effectively and is one of the preferred methods for flexible ligand docking. The differential...

  16. Ligand binding to G protein-coupled receptors in tethered cell membranes

    DEFF Research Database (Denmark)

    Martinez, Karen L.; Meyer, Bruno H.; Hovius, Ruud;

    2003-01-01

    G protein-coupled receptors (GPCRs) constitute a large class of seven transmembrane proteins, which bind selectively agonists or antagonists with important consequences for cellular signaling and function. Comprehension of the molecular details of ligand binding is important for the understanding...

  17. Ligand Binding Kinetics of the Quorum Sensing Regulator PqsR

    DEFF Research Database (Denmark)

    Welch, Martin; Hodgkinson, James T.; Gross, Jeremy;

    2013-01-01

    The Pseudomonas aeruginosa quinolone signal (PQS) is a quorum sensing molecule that plays an important role in regulating the virulence of this organism. We have purified the ligand binding domain of the receptor PqsRLBD for PQS and have used Förster resonance energy transfer fluorimetry and...

  18. Mechanistic Studies of Vertebrate Hedgehog Signaling

    OpenAIRE

    Tukachinsky, Hanna

    2012-01-01

    Metazoans use Hedgehog signaling to direct many stages of embryonic development, and deregulation of this pathway is implicated in many types of cancer. I investigated several steps of Hedgehog pathway transduction that were poorly understood in mechanistic terms. The mature Hedgehog ligand is produced by a self-proteolysis reaction that covalently attaches a cholesterol molecule to the signaling half of the protein. I showed that the catalytic cysteine forms a disulfide bridge that is essent...

  19. Hedgehog Cholesterolysis: Specialized Gatekeeper to Oncogenic Signaling

    OpenAIRE

    Callahan, Brian P.; Chunyu Wang

    2015-01-01

    Discussions of therapeutic suppression of hedgehog (Hh) signaling almost exclusively focus on receptor antagonism; however, hedgehog’s biosynthesis represents a unique and potentially targetable aspect of this oncogenic signaling pathway. Here, we review a key biosynthetic step called cholesterolysis from the perspectives of structure/function and small molecule inhibition. Cholesterolysis, also called cholesteroylation, generates cholesterol-modified Hh ligand via autoprocessing of a hedgeho...

  20. Dysprosium complexes with the tetraphenylporphyrin macrocyclic ligand

    International Nuclear Information System (INIS)

    In this report, the results obtained on the synthesis, characterization and study of the chemical behavior of dysprosium complex with the acetylacetone chelating agent (Hacac) and the tetraphenylporphyrin macrocyclic ligand (H2TFP) are given. Based on the literature but according to our necessities and interest, the appropriate methodology settled down from the synthesis of prime matters until the obtaining and characterization of the products. The acetyl acetonate complex was obtained of mono hydrated dysprosium [Dy(acac)3. H20] and trihydrated [Dy(acac)3 .3 H20], the mono tetra phenyl porphyrinate [Dy(TFP)(acac). 2 ac] the double sandwich of the dysprosium porphyrinate [Dy(TFP)2] and the triple sandwich of the dysprosium porphyrinate [Dy(TFP)3. 2 TCB] (TCB = trichlorobenzene). Its were characterized by their melting points, solubility, IR, UV, TGA and DTA both first and besides the techniques already mentioned for NMR'H, RPE and Magnetic susceptibility the three last complexes. From the spectroscopic point of view, IR and RPE its suggested the existence of a complex of inverse mixed valence [Dy(TFP)2- (TFP) 1-] for the Dy(TFP)2 as a result of the existence of the free radical (TFP' 1- and that it was not in none of the other porphyrin compounds. In the NMR'H spectra of the compounds were not observed signals in the region from 0 to 10 ppm that which shows that the dysprosium complexes in special those of the porphyrin type are highly paramagnetic and its could be used as displacement reagents, creators of images and contrast agents of great utility in these days in studies of NMR, technique today by today used in medical diagnoses. (Author)

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

    OpenAIRE

    Resat Haluk; Wiley H Steven; Shankaran Harish

    2007-01-01

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

  2. Androgenic regulation of hedgehog signaling pathway components in prostate cancer cells

    OpenAIRE

    Chen, Mengqian; Tanner, Matthew; Levine, Alice C.; Levina, Elina; Ohouo, Patrice; Buttyan, Ralph

    2009-01-01

    Hedgehog signaling is thought to play a role in several human cancers including prostate cancer. Although prostate cancer cells express many of the gene products involved in hedgehog signaling, these cells are refractory to the canonical signaling effects of exogenous hedgehog ligands or to activated Smoothened, the hedgehog-regulated mediator of Gli transcriptional activation. Here, we show that the expression of hedgehog ligands and some hedgehog target genes are regulated by androgen in th...

  3. Coordinate unsaturation with fluorinated ligands

    Energy Technology Data Exchange (ETDEWEB)

    Rack, J.L.; Hurlburt, P.K.; Anderson, O.P.; Strauss, S.H. [Colorado State Univ., Ft. Collins, CO (United States)

    1993-12-31

    The preparation and characterization of Zn(OTeF{sub 5}){sub 2} has resulted in a model compound with which to explore the concept of coordinative unsaturation. The coordination of solvents of varying donicity and dielectric constant to the Zn(II) ions in Zn(OTeF{sub 5}){sub 2} was studied by vapor phase monometry, NMR and IR spectroscopy, conductimetry, and X-Ray crystallography. The structures of [Zn(C{sub 6}H{sub 5}NO{sub 2}){sub 2}(OTeF{sub 5})2]2 and Zn(C{sub 6}H{sub 5}NO{sub 2}){sub 3}(OTEF{sub 5}){sub 2} demonstrate the electronic flexibility of some weakly coordinating solvents in that nitrobenzene can function as either an {eta}{sup 1}O or {eta}{sup 2}O,O`-ligand. The dependence of the number of bound solvent molecules and the degree of OTeF{sub 5}{minus} dissociation on solvent donor number and dielectric constant will be presented.

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

    Directory of Open Access Journals (Sweden)

    Anne eSteen

    2014-06-01

    Full Text Available Biased signaling or functional selectivity occurs when a 7TM receptor preferentially activates one of several available pathways. It can be divided into three distinct forms: ligand bias, receptor bias, and tissue or cell bias, where it is mediated by different ligands (on the same receptor, different receptors (with the same ligand or different tissues or cells (for the same ligand-receptor pair. Most often biased signaling is differentiated into G protein-dependent and β-arrestin-dependent signaling. Yet, it may also cover signaling differences within these groups. Moreover, it may not be absolute, i.e. full versus no activation. Here we discuss biased signaling in the chemokine system, including the structural basis for biased signaling in chemokine receptors, as well as in class A 7TM receptors in general. This includes overall helical movements and the contributions of micro-switches based on recently published 7TM crystals and molecular dynamics studies. All three forms of biased signaling are abundant in the chemokine system. This challenges our understanding of classic redundancy inevitably ascribed to this system, where multiple chemokines bind to the same receptor and where a single chemokine may bind to several receptors – in both cases with the same functional outcome. The ubiquitous biased signaling confer a hitherto unknown specificity to the chemokine system with a complex interaction pattern that is better described as promiscuous with context-defined roles and different functional outcomes in a ligand-, receptor- or cell/tissue-defined manner. As the low number of successful drug development plans implies, there are great difficulties in targeting chemokine receptors; in particular with regard to receptor antagonists as anti-inflammatory drugs. Un-defined and putative non-selective targeting of the complete cellular signaling system could be the underlying cause of lack of success. Therefore, biased ligands could be the

  5. The clathrin-binding motif and the J-domain of Drosophila Auxilin are essential for facilitating Notch ligand endocytosis

    Directory of Open Access Journals (Sweden)

    Chang Henry C

    2008-05-01

    Full Text Available Abstract Background Ligand endocytosis plays a critical role in regulating the activity of the Notch pathway. The Drosophila homolog of auxilin (dAux, a J-domain-containing protein best known for its role in the disassembly of clathrin coats from clathrin-coated vesicles, has recently been implicated in Notch signaling, although its exact mechanism remains poorly understood. Results To understand the role of auxilin in Notch ligand endocytosis, we have analyzed several point mutations affecting specific domains of dAux. In agreement with previous work, analysis using these stronger dAux alleles shows that dAux is required for several Notch-dependent processes, and its function during Notch signaling is required in the signaling cells. In support of the genetic evidences, the level of Delta appears elevated in dAux deficient cells, suggesting that the endocytosis of Notch ligand is disrupted. Deletion analysis shows that the clathrin-binding motif and the J-domain, when over-expressed, are sufficient for rescuing dAux phenotypes, implying that the recruitment of Hsc70 to clathrin is a critical role for dAux. However, surface labeling experiment shows that, in dAux mutant cells, Delta accumulates at the cell surface. In dAux mutant cells, clathrin appears to form large aggregates, although Delta is not enriched in these aberrant clathrin-positive structures. Conclusion Our data suggest that dAux mutations inhibit Notch ligand internalization at an early step during clathrin-mediated endocytosis, before the disassembly of clathrin-coated vesicles. Further, the inhibition of ligand endocytosis in dAux mutant cells possibly occurs due to depletion of cytosolic pools of clathrin via the formation of clathrin aggregates. Together, our observations argue that ligand endocytosis is critical for Notch signaling and auxilin participates in Notch signaling by facilitating ligand internalization.

  6. 4-1BB Signaling Enhances Primary and Secondary Population Expansion of CD8+ T Cells by Maximizing Autocrine IL-2/IL-2 Receptor Signaling.

    Directory of Open Access Journals (Sweden)

    Ho S Oh

    Full Text Available 4-1BB (CD137, a member of the tumor necrosis factor receptor superfamily (TNFRSF, is primarily expressed on activated T cells and is known to enhance proliferation of T cells, prevent activation-induced cell death, and promote memory formation of CD8+ T cells. In particular, it is well acknowledged that 4-1BB triggering preferentially enhances the expansion of CD8+ T cells rather than CD4+ T cells, but the underlying mechanism remains unclear. Here we found that 4-1BB triggering markedly increased IL-2Rα (CD25 and IL-2 expressions of CD8+ T cells but minimally for CD4+ T cells. Proliferation of CD8+ T cells was moderately enhanced by direct 4-1BB triggering in the absence of signaling through IL-2Rα/IL-2 interactions, but further promoted in the presence of IL-2Rα/IL-2 interactions. Among the TNFRSF members including OX40, GITR, CD30, and CD27, 4-1BB was superior in the ability to induce IL-2Rα expression on CD8+ T cells. When the primary and secondary expansions of CD8+ T cells in vivo were examined by adoptively transferring OVA-specific CD8+ T cells along with the treatment with agonistic anti-4-1BB and/or antagonistic anti-CD25 F(ab'2 mAb, 4-1BB triggering enhanced both primary and secondary expansion of CD8+ T cells in vivo, and the 4-1BB effects were moderately suppressed in primary expansion while completely abolished in secondary expansion of OVA-specific CD8+ T cells by blocking IL-2Rα. These results suggest that 4-1BB-mediated increases of IL-2Rα and IL-2 prolong the effects of transient TCR- and 4-1BB-mediated signaling in CD8+ T cells, and that 4-1BB triggering preferentially enhances the expansion of CD8+ T cells through the amplification of autocrine IL-2/IL-2R signaling loop.

  7. 4-1BB Signaling Enhances Primary and Secondary Population Expansion of CD8+ T Cells by Maximizing Autocrine IL-2/IL-2 Receptor Signaling.

    Science.gov (United States)

    Oh, Ho S; Choi, Beom K; Kim, Young H; Lee, Don G; Hwang, Sunhee; Lee, Myoung J; Park, Sang H; Bae, Yong-Soo; Kwon, Byoung S

    2015-01-01

    4-1BB (CD137), a member of the tumor necrosis factor receptor superfamily (TNFRSF), is primarily expressed on activated T cells and is known to enhance proliferation of T cells, prevent activation-induced cell death, and promote memory formation of CD8+ T cells. In particular, it is well acknowledged that 4-1BB triggering preferentially enhances the expansion of CD8+ T cells rather than CD4+ T cells, but the underlying mechanism remains unclear. Here we found that 4-1BB triggering markedly increased IL-2Rα (CD25) and IL-2 expressions of CD8+ T cells but minimally for CD4+ T cells. Proliferation of CD8+ T cells was moderately enhanced by direct 4-1BB triggering in the absence of signaling through IL-2Rα/IL-2 interactions, but further promoted in the presence of IL-2Rα/IL-2 interactions. Among the TNFRSF members including OX40, GITR, CD30, and CD27, 4-1BB was superior in the ability to induce IL-2Rα expression on CD8+ T cells. When the primary and secondary expansions of CD8+ T cells in vivo were examined by adoptively transferring OVA-specific CD8+ T cells along with the treatment with agonistic anti-4-1BB and/or antagonistic anti-CD25 F(ab')2 mAb, 4-1BB triggering enhanced both primary and secondary expansion of CD8+ T cells in vivo, and the 4-1BB effects were moderately suppressed in primary expansion while completely abolished in secondary expansion of OVA-specific CD8+ T cells by blocking IL-2Rα. These results suggest that 4-1BB-mediated increases of IL-2Rα and IL-2 prolong the effects of transient TCR- and 4-1BB-mediated signaling in CD8+ T cells, and that 4-1BB triggering preferentially enhances the expansion of CD8+ T cells through the amplification of autocrine IL-2/IL-2R signaling loop. PMID:25962156

  8. Ligand inducible assembly of a DNA tetrahedron.

    Science.gov (United States)

    Dohno, Chikara; Atsumi, Hiroshi; Nakatani, Kazuhiko

    2011-03-28

    Here we show that a small synthetic ligand can be used as a key building component for DNA nanofabrication. Using naphthyridinecarbamate dimer (NCD) as a molecular glue for DNA hybridization, we demonstrate NCD-triggered formation of a DNA tetrahedron.

  9. Chemistry of Marine Ligands and Siderophores

    OpenAIRE

    Vraspir, Julia M.; Butler, Alison

    2009-01-01

    Marine microorganisms are presented with unique challenges to obtain essential metal ions required to survive and thrive in the ocean. The production of organic ligands to complex transition metal ions is one strategy to both facilitate uptake of specific metals, such as iron, and to mitigate the potential toxic effects of other metal ions, such as copper. A number of important trace metal ions are complexed by organic ligands in seawater, including iron, cobalt, nickel, copper, zinc, and cad...

  10. Fas ligand deficiency in HIV disease

    OpenAIRE

    Sieg, Scott; Smith, Dawn; Yildirim, Zafer; Kaplan, David

    1997-01-01

    Apoptosis is postulated to be involved as an anti-viral immune mechanism by killing infected cells before viral replication has occurred. The Fas–Fas ligand interaction is a powerful regulator of T cell apoptosis and could potentially act as a potent anti-viral immune mechanism against T cell tropic virus such as human immunodeficiency virus (HIV). We investigated the status of Fas ligand in peripheral blood mononuclear cells (PBMCs) obtained from persons infected with HIV. We found that mono...

  11. Construction of dinuclear complexes using multidentate ligands

    Energy Technology Data Exchange (ETDEWEB)

    Sampson, C.L

    2000-04-01

    This work details the synthesis of novel copper(I), copper(II), nickel(II) and zinc(II) dinuclear complexes. Attempts have been made to control the co-ordination architectures of the metal centres by using bis-bidentate and tridentate chelating N,S- and N-donor ligands to generate dinuclear systems. The ligands were both symmetrically and asymmetrically disubstituted pyridazine-based and pyridine-based ligands consisting of a mixture of N-only and mixed N,S-donors. The study using the pyridazine-based ligands continues previous research in our group using 3,6-bis disubstituted pyridazine-based ligands to form complexes with copper(l) and copper(II). The pyridazine-based ligands have been seen to be bis-bidentate upon co-ordination of copper. The pyridazine-based ligands could be envisaged to generate dinuclear complexes by directly bridging between two metal ions. This study involved the formation of copper(l), nickel(II) and zinc(II) complexes with these ligands. The structural properties of two particular complexes have been explored using X-ray crystallography and spectroscopic techniques. Pyridine-based ligands have also been used previously in our group as tridentate chelating ligands. They have been seen to form dinuclear complexes with copper(I) and copper(II) when reacted with an additional bridging ligand e.g. 4,4'-bipyridine. This provides an alternative method for generating dinuclear complexes. Chapter 1 presents an introduction to the area of supramolecular chemistry from which we can learn the principles of polymer formation and them 'in reverse' to generate discrete dinuclear systems. Chapter 2 details the synthesis of the pyridazine and pyridine-based ligands including a detailed nmr study of the ligands. Since the ligands were synthesised using cyclic thioamides as terminal groups it has been found that thiol-thione tautomerisation occurred during synthesis giving rise to two possible ligand conformations. The nmr study has been used

  12. LigandRFs: random forest ensemble to identify ligand-binding residues from sequence information alone

    KAUST Repository

    Chen, Peng

    2014-12-03

    Background Protein-ligand binding is important for some proteins to perform their functions. Protein-ligand binding sites are the residues of proteins that physically bind to ligands. Despite of the recent advances in computational prediction for protein-ligand binding sites, the state-of-the-art methods search for similar, known structures of the query and predict the binding sites based on the solved structures. However, such structural information is not commonly available. Results In this paper, we propose a sequence-based approach to identify protein-ligand binding residues. We propose a combination technique to reduce the effects of different sliding residue windows in the process of encoding input feature vectors. Moreover, due to the highly imbalanced samples between the ligand-binding sites and non ligand-binding sites, we construct several balanced data sets, for each of which a random forest (RF)-based classifier is trained. The ensemble of these RF classifiers forms a sequence-based protein-ligand binding site predictor. Conclusions Experimental results on CASP9 and CASP8 data sets demonstrate that our method compares favorably with the state-of-the-art protein-ligand binding site prediction methods.

  13. Synthesis and characterization of mixed ligand chiral nanoclusters

    KAUST Repository

    Guven, Zekiye P.

    2016-06-22

    Chiral mixed ligand silver nanoclusters were synthesized in the presence of a chiral and an achiral ligand. While the chiral ligand led mostly to the formation of nanoparticles, the presence of the achiral ligand drastically increased the yield of nanoclusters with enhanced chiral properties. © 2016 The Royal Society of Chemistry.

  14. Computation of Rate Constants for Diffusion of Small Ligands to and from Buried Protein Active Sites.

    Science.gov (United States)

    Wang, P-H; De Sancho, D; Best, R B; Blumberger, J

    2016-01-01

    The diffusion of ligands to actives sites of proteins is essential to enzyme catalysis and many cellular signaling processes. In this contribution we review our recently developed methodology for calculation of rate constants for diffusion and binding of small molecules to buried protein active sites. The diffusive dynamics of the ligand obtained from molecular dynamics simulation is coarse grained and described by a Markov state model. Diffusion and binding rate constants are then obtained either from the reactive flux formalism or by fitting the time-dependent population of the Markov state model to a phenomenological rate law. The method is illustrated by applications to diffusion of substrate and inhibitors in [NiFe] hydrogenase, CO-dehydrogenase, and myoglobin. We also discuss a recently developed sensitivity analysis that allows one to identify hot spots in proteins, where mutations are expected to have the strongest effects on ligand diffusion rates. PMID:27497172

  15. A yeast surface display system for the discovery of ligands that trigger cell activation.

    Science.gov (United States)

    Cho, B K; Kieke, M C; Boder, E T; Wittrup, K D; Kranz, D M

    1998-11-01

    Opposing cells often communicate signalling events using multivalent interactions between receptors present on their cell surface. For example, T cells are typically activated when the T cell receptor (TCR) and its associated costimulatory molecules are multivalently engaged by the appropriate ligands present on an antigen presenting cell. In this report, yeast expressing high cell-surface levels of a TCR ligand (a recombinant antibody to the TCR Vbeta domain) were shown to act as 'pseudo' antigen presenting cells and induce T cell activation as monitored by increased levels of CD25 and CD69 and by downregulation of cell surface TCR. Similar levels of T cell activation could occur even when a 30-fold excess of irrelevant yeast was present, suggesting that such a yeast display system, by virtue of its ability to present ligands multivalently, may be used in highly sensitive procedures to identify novel polypeptides that interact multivalently with cell surface receptors and thereby trigger specific cellular responses.

  16. Ligand binding by PDZ domains

    DEFF Research Database (Denmark)

    Chi, Celestine N.; Bach, Anders; Strømgaard, Kristian;

    2012-01-01

    The postsynaptic density protein-95/disks large/zonula occludens-1 (PDZ) protein domain family is one of the most common protein-protein interaction modules in mammalian cells, with paralogs present in several hundred human proteins. PDZ domains are found in most cell types, but neuronal proteins......, for example, are particularly rich in these domains. The general function of PDZ domains is to bring proteins together within the appropriate cellular compartment, thereby facilitating scaffolding, signaling, and trafficking events. The many functions of PDZ domains under normal physiological as well...

  17. Retinoic acid signalling in thymocytes regulates T cell development

    DEFF Research Database (Denmark)

    Wendland, Kerstin; Sitnik, Katarzyna Maria; Kotarsky, Knut;

    The Vitamin A derivative retinoic acid (RA) works as a ligand for a family of nuclearRA receptors (RARα, RARβ and RARγ) which form heterodimers with retinoid Xreceptors (RXR). These complexes function as ligand-activated transcription factors,recognizing specific RA responsive elements in the reg......RARα. This blocks RA signalling in developing thymocytes from the DN3/4 stageonwards and thus allows us to study the role of RA in T cell development...

  18. The hedgehog/Gli signaling paradigm in prostate cancer

    OpenAIRE

    Chen, Mengqian; Carkner, Richard; Buttyan, Ralph

    2011-01-01

    Hedgehog is a ligand-activated signaling pathway that regulates Gli-mediated transcription. Although most noted for its role as an embryonic morphogen, hyperactive hedgehog also causes human skin and brain malignancies. The hedgehog-related gene anomalies found in these tumors are rarely found in prostate cancer. Yet surveys of human prostate tumors show concordance of high expression of hedgehog ligands and Gli2 that correlate with the potential for metastasis and therapy-resistant behavior....

  19. 4-1BB signaling breaks the tolerance of maternal CD8+ T cells that are reactive with alloantigens.

    Directory of Open Access Journals (Sweden)

    Kwang H Kim

    Full Text Available 4-1BB (CD137, TNFRSF9, a member of the activation-induced tumor necrosis factor receptor family, is a powerful T-cell costimulatory molecule. It generally enhances CD8(+ T responses and even breaks the tolerance of CD8(+ T cells in an antigen-specific manner. In the present study we found that it was expressed in the placentas of pregnant mice and that its expression coincided with that of the immunesuppressive enzyme indoleamine 2,3-dioxygenase (IDO. Therefore, we investigated whether 4-1BB signaling is involved in fetal rejection using agonistic anti-4-1BB mAb and 4-1BB-deficient mice. Treatment with agonistic anti-4-1BB mAb markedly increased the rate of rejection of allogeneic but not syngeneic fetuses, and this was primarily dependent on CD8(+ T cells. Complement component 3 (C3 seemed to be the effector molecule because 4-1BB triggering resulting in accumulation of C3 in the placenta, and this accumulation was also reversed by anti-CD8 mAb treatment. These findings demonstrate that 4-1BB triggering breaks the tolerance of CD8(+ T cells to alloantigens in the placenta. Moreover, triggering 4-1BB protected the pregnant mice from Listeria monocytogenes (LM infection, but led to rejection of semi-allogeneic fetuses. Therefore, given the cross-recognition of alloantigen by pathogen-reactive CD8(+ T cells, the true function of 4-1BB may be to reverse the hypo-responsiveness of pathogen-reactive CD8(+ T cells in the placenta in cases of infection, even if that risks losing the fetus.

  20. Signal processing by the endosomal system.

    Science.gov (United States)

    Villaseñor, Roberto; Kalaidzidis, Yannis; Zerial, Marino

    2016-04-01

    Cells need to decode chemical or physical signals from their environment in order to make decisions on their fate. In the case of signalling receptors, ligand binding triggers a cascade of chemical reactions but also the internalization of the activated receptors in the endocytic pathway. Here, we highlight recent studies revealing a new role of the endosomal network in signal processing. The diversity of entry pathways and endosomal compartments is exploited to regulate the kinetics of receptor trafficking, and interactions with specific signalling adaptors and effectors. By governing the spatio-temporal distribution of signalling molecules, the endosomal system functions analogously to a digital-analogue computer that regulates the specificity and robustness of the signalling response.

  1. Potential applications for sigma receptor ligands in cancer diagnosis and therapy.

    Science.gov (United States)

    van Waarde, Aren; Rybczynska, Anna A; Ramakrishnan, Nisha K; Ishiwata, Kiichi; Elsinga, Philip H; Dierckx, Rudi A J O

    2015-10-01

    Sigma receptors (sigma-1 and sigma-2) represent two independent classes of proteins. Their endogenous ligands may include the hallucinogen N,N-dimethyltryptamine (DMT) and sphingolipid-derived amines which interact with sigma-1 receptors, besides steroid hormones (e.g., progesterone) which bind to both sigma receptor subpopulations. The sigma-1 receptor is a ligand-regulated molecular chaperone with various ion channels and G-protein-coupled membrane receptors as clients. The sigma-2 receptor was identified as the progesterone receptor membrane component 1 (PGRMC1). Although sigma receptors are over-expressed in tumors and up-regulated in rapidly dividing normal tissue, their ligands induce significant cell death only in tumor tissue. Sigma ligands may therefore be used to selectively eradicate tumors. Multiple mechanisms appear to underlie cell killing after administration of sigma ligands, and the signaling pathways are dependent both on the type of ligand and the type of tumor cell. Recent evidence suggests that the sigma-2 receptor is a potential tumor and serum biomarker for human lung cancer and an important target for inhibiting tumor invasion and cancer progression. Current radiochemical efforts are focused on the development of subtype-selective radioligands for positron emission tomography (PET) imaging. Right now, the mostpromising tracers are [18F]fluspidine and [18F]FTC-146 for sigma-1 receptors and [11C]RHM-1 and [18F]ISO-1 for the sigma-2 subtype. Nanoparticles coupled to sigma ligands have shown considerable potential for targeted delivery of antitumor drugs in animal models of cancer, but clinical studies exploring this strategy in cancer patients have not yet been reported. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers. PMID:25173780

  2. Effects of Na/K-ATPase and its ligands on bone marrow stromal cell differentiation

    Directory of Open Access Journals (Sweden)

    Moustafa Sayed

    2014-07-01

    Full Text Available Endogenous ligands of Na/K-ATPase have been demonstrated to increase in kidney dysfunction and heart failure. It is also reported that Na/K-ATPase signaling function effects stem cell differentiation. This study evaluated whether Na/K-ATPase activation through its ligands and associated signaling functions affect bone marrow stromal cells (BMSCs, also known as bone marrow-derived mesenchymal stem cells differentiation capacity. BMSCs were isolated from male Sprague–Dawley rats and cultured in minimal essential medium alpha (MEM-α supplemented with 15% Fetal Bovine serum (FBS. The results showed that marinobufagenin (MBG, a specific Na/K-ATPase ligand, potentiated rosiglitazone-induced adipogenesis in these BMSCs. Meanwhile, it attenuated BMSC osteogenesis. Mechanistically, MBG increased CCAAT/enhancer binding protein alpha (C/EBPα protein expression through activation of an extracellular regulated kinase (ERK signaling pathway, which leads to enhanced rosiglitazone-induced adipogenesis. Inhibition of ERK activation by U0126 blocks the effect of MBG on C/EBPα expression and on rosiglitazone-induced adipogenesis. Reciprocally, MBG reduced runt-related transcription factor 2 (RunX2 expression, which resulted in the inhibition of osteogenesis induced by β-glycerophosphate/ascorbic acid. MBG also potentiated rosiglitazone-induced adipogenesis in 3T3-L1 cells and in mouse BMSCs. These results suggest that Na/K-ATPase and its signaling functions are involved in the regulation of BMSCs differentiation.

  3. Immobilisation of ligands by radio-derivatized polymers; Immobilisering av ligander med radioderiverte polymerer

    Energy Technology Data Exchange (ETDEWEB)

    Varga, J.M.; Fritsch, P.

    1995-01-30

    The invention relates to radio-derivatized polymers and a method of producing them by contacting non-polymerizable conjugands with radiolysable polymers in the presence of irradiation. The resulting radio-derivatized polymers can be further linked with ligand of organic or inorganic nature to immobilize such ligands. 2 figs., 5 tabs.

  4. Luminescent solutions and powders of new samarium complexes with N,N',O,O'-chelating ligands

    Science.gov (United States)

    Kharcheva, Anastasia V.; Nikolskiy, Kirill S.; Borisova, Nataliya E.; Ivanov, Alexey V.; Reshetova, Marina D.; Yuzhakov, Viktor I.; Patsaeva, Svetlana V.

    2016-04-01

    Imaging techniques in biology and medicine are crucial tools to obtain information on structural and functional properties of living cells and organisms. To fulfill the requirements associated with application of these techniques it appears necessary to design markers with specific characteristics. Luminescent complexes of trivalent lanthanide ions with chelating ligands are of increasing importance in biomedical applications because of their millisecond luminescence lifetime, narrow emission band, high signal-to-noise ratio and minimal photodamage to biological samples. In order to extend the available emission wavelength range the luminescent samarium chelates are highly desirable. In this study the ligands with diamides of 2,2'-bipyridin-6,6'-dicarboxylic acid were used to improve photophysical characteristics of samarium complexes. We report the luminescence characteristics of samarium complexes with novel ligands. All complexes exhibited the characteristic emission of Sm (III) ion with the lines at 565, 597, 605, 645 and 654 nm, the intensity strongly depended on the ligand. Absorption and luminescence excitation spectra of Sm (III) complexes showed main peaks in the UV range demonstrating lanthanide coordination to the ligand. The absolute lumenescence quantum yield was measured for solutions in acetonitrile with excitation at 350 nm. The largest luminescence quantum yield was found for the samarium complex Bipy 6MePy Sm (3%) being much higher that for samarium complexes reported in the literature earlier. These results prove as well that samarium chelates are potential markers for multiparametric imaging techniques.

  5. Tumor cells prevent mouse dendritic cell maturation induced by TLR ligands.

    Science.gov (United States)

    Idoyaga, Juliana; Moreno, José; Bonifaz, Laura

    2007-08-01

    Tumor cells can evade the immune system through several mechanisms, one of which is to block DC maturation. It has been suggested that signaling via Toll-like receptors (TLR) may be involved in the induction of prophylactic anti-cancer immunity and in the treatment of established tumors. In the present study we found that high numbers of tumor cells interfere with BMDC activation induced by the TLR ligands LPS and poly IC. Tumor cells blocked TLR3- and TLR4-mediated induction of MHCII and the co-stimulatory molecules CD40 and CD86, as well as the cytokines IL-12, TNF-alpha and IL-6. Importantly, tumor cells induced inhibitory molecules (B7-DC, B7-H1 and CD80) on spleen DC in vivo and on BMDC, even in the presence of TLR ligands. Moreover, after a long exposure with tumor cells, purified BMDC were unable to respond to a second challenge with TLR ligands. The failure of tumor exposed-BMDC to express co-stimulatory molecules and cytokines in the presence of TLR ligands has implications for the future development of DC-based cancer immune therapies using TLR ligands as adjuvants for the activation of DC.

  6. An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions.

    Science.gov (United States)

    Syedbasha, Mohameedyaseen; Linnik, Janina; Santer, Deanna; O'Shea, Daire; Barakat, Khaled; Joyce, Michael; Khanna, Nina; Tyrrell, D Lorne; Houghton, Michael; Egli, Adrian

    2016-01-01

    A comprehensive understanding of signaling pathways requires detailed knowledge regarding ligand-receptor interaction. This article describes two fast and reliable point-by-point protocols of enzyme-linked immunosorbent assays (ELISAs) for the investigation of ligand-receptor interactions: the direct ligand-receptor interaction assay (LRA) and the competition LRA. As a case study, the ELISA based analysis of the interaction between different lambda interferons (IFNLs) and the alpha subunit of their receptor (IL28RA) is presented: the direct LRA is used for the determination of dissociation constants (KD values) between receptor and IFN ligands, and the competition LRA for the determination of the inhibitory capacity of an oligopeptide, which was designed to compete with the IFNLs at their receptor binding site. Analytical steps to estimate KD and half maximal inhibitory concentration (IC50) values are described. Finally, the discussion highlights advantages and disadvantages of the presented method and how the results enable a better molecular understanding of ligand-receptor interactions.

  7. Ligands for Ionotropic Glutamate Receptors

    Science.gov (United States)

    Swanson, Geoffrey T.; Sakai, Ryuichi

    Marine-derived small molecules and peptides have played a central role in elaborating pharmacological specificities and neuronal functions of mammalian ionotropic glutamate receptors (iGluRs), the primary mediators of excitatory syn-aptic transmission in the central nervous system (CNS). As well, the pathological sequelae elicited by one class of compounds (the kainoids) constitute a widely-used animal model for human mesial temporal lobe epilepsy (mTLE). New and existing molecules could prove useful as lead compounds for the development of therapeutics for neuropathologies that have aberrant glutamatergic signaling as a central component. In this chapter we discuss natural source origins and pharmacological activities of those marine compounds that target ionotropic glutamate receptors.

  8. International Union of Basic and Clinical Pharmacology. XC. multisite pharmacology: recommendations for the nomenclature of receptor allosterism and allosteric ligands.

    Science.gov (United States)

    Christopoulos, Arthur; Changeux, Jean-Pierre; Catterall, William A; Fabbro, Doriano; Burris, Thomas P; Cidlowski, John A; Olsen, Richard W; Peters, John A; Neubig, Richard R; Pin, Jean-Philippe; Sexton, Patrick M; Kenakin, Terry P; Ehlert, Frederick J; Spedding, Michael; Langmead, Christopher J

    2014-10-01

    Allosteric interactions play vital roles in metabolic processes and signal transduction and, more recently, have become the focus of numerous pharmacological studies because of the potential for discovering more target-selective chemical probes and therapeutic agents. In addition to classic early studies on enzymes, there are now examples of small molecule allosteric modulators for all superfamilies of receptors encoded by the genome, including ligand- and voltage-gated ion channels, G protein-coupled receptors, nuclear hormone receptors, and receptor tyrosine kinases. As a consequence, a vast array of pharmacologic behaviors has been ascribed to allosteric ligands that can vary in a target-, ligand-, and cell-/tissue-dependent manner. The current article presents an overview of allostery as applied to receptor families and approaches for detecting and validating allosteric interactions and gives recommendations for the nomenclature of allosteric ligands and their properties.

  9. A new class of PN3-pincer ligands for metal–ligand cooperative catalysis

    KAUST Repository

    Li, Huaifeng

    2014-12-01

    Work on a new class of PN3-pincer ligands for metal-ligand cooperative catalysis is reviewed. While the field of the pyridine-based PN3-transition metal pincer complexes is still relatively young, many important applications of these complexes have already emerged. In several cases, the PN3-pincer complexes for metal-ligand cooperative catalysis result in significantly improved or unprecedented activities. The synthesis and coordination chemistry of PN3-pincer ligands are briefly summarized first to cover the synthetic routes for their preparation, followed by a focus review on their applications in catalysis. A specific emphasis is placed on the later section about the role of PN3-pincer ligands\\' dearomatization-rearomatization steps during the catalytic cycles. The mechanistic insights from density functional theory (DFT) calculations are also discussed.

  10. Sliding tethered ligands add topological interactions to the toolbox of ligand-receptor design

    Science.gov (United States)

    Bauer, Martin; Kékicheff, Patrick; Iss, Jean; Fajolles, Christophe; Charitat, Thierry; Daillant, Jean; Marques, Carlos M.

    2015-09-01

    Adhesion in the biological realm is mediated by specific lock-and-key interactions between ligand-receptor pairs. These complementary moieties are ubiquitously anchored to substrates by tethers that control the interaction range and the mobility of the ligands and receptors, thus tuning the kinetics and strength of the binding events. Here we add sliding anchoring to the toolbox of ligand-receptor design by developing a family of tethered ligands for which the spacer can slide at the anchoring point. Our results show that this additional sliding degree of freedom changes the nature of the adhesive contact by extending the spatial range over which binding may sustain a significant force. By introducing sliding tethered ligands with self-regulating length, this work paves the way for the development of versatile and reusable bio-adhesive substrates with potential applications for drug delivery and tissue engineering.

  11. Ligand-specific conformational changes in the alpha1 glycine receptor ligand-binding domain

    DEFF Research Database (Denmark)

    Pless, Stephan Alexander; Lynch, Joseph W

    2009-01-01

    indicate that channel opening is accompanied by conformational rearrangements in both beta-sheets. In an attempt to resolve ligand-dependent movements in the ligand-binding domain, we employed voltage-clamp fluorometry on alpha1 glycine receptors to compare changes mediated by the agonist, glycine...... in the inner beta-sheet and pre-M1 domain that may be important for activation, desensitization, or both. In contrast, most labeled residues in loops C and F yielded fluorescence changes identical in magnitude for glycine and strychnine. A notable exception was H201C in loop C. This labeled residue responded...... differently to glycine and strychnine, thus underlining the importance of loop C in ligand discrimination. These results provide an important step toward mapping the domains crucial for ligand discrimination in the ligand-binding domain of glycine receptors and possibly other Cys loop receptors....

  12. A review of monoamine transporter-ligand interactions.

    Science.gov (United States)

    Immadisetty, Kalyan; Madura, Jeffry D

    2013-12-01

    Transporters of the monoamines serotonin, dopamine, and norepinephrine are plasma membrane proteins belonging to the neurotransmitter sodium symporter family (NSS). Monoamine transporters (MATs) by facilitating reuptake of neurotransmitters from the synapse into the presynaptic nerve terminal, regulate neurotransmitter chemical signaling and maintain homeostasis. MATs are targets for several psychostimulants such as cocaine and amphetamine; and also for drugs treating several psychiatric disorders such as depression, attention deficit hyperactivity disorder, Parkinson's disease, and schizophrenia. Since, currently available treatment has several limitations and side effects, novel treatment is highly desired. Efforts to develop better treatment have been hampered by the lack of crystal structures for MATs. However, leucine transporter (LeuTAa), a bacterial protein from Aquifex aeolicus, belonging to the same NSS family as MATs has recently been crystallized. LeuTAa is used as a template to develop homology models of MATs, which facilitates understanding of the structure, function and pharmacology of MATs. Experimental methods for drug discovery demand a significant amount of time, effort and money. Efficient utilization of computational techniques hastens the process of drug discovery and also significantly reduces the cost. Assessing the binding affinity of drugs to the receptors is a key aspect of drug design. Free energy calculations compliment the experiment by quantitatively assessing the affinity of ligands to receptors. These methods are highly beneficial in the lead identification and optimization stages of rational drug design. We review the currently available free energy methods to treat protein-ligand interactions along with several free energy studies performed on MATs. PMID:24138394

  13. Single-Molecule Imaging of Cellular Signaling

    Science.gov (United States)

    De Keijzer, Sandra; Snaar-Jagalska, B. Ewa; Spaink, Herman P.; Schmidt, Thomas

    Single-molecule microscopy is an emerging technique to understand the function of a protein in the context of its natural environment. In our laboratory this technique has been used to study the dynamics of signal transduction in vivo. A multitude of signal transduction cascades are initiated by interactions between proteins in the plasma membrane. These cascades start by binding a ligand to its receptor, thereby activating downstream signaling pathways which finally result in complex cellular responses. To fully understand these processes it is important to study the initial steps of the signaling cascades. Standard biological assays mostly call for overexpression of the proteins and high concentrations of ligand. This sets severe limits to the interpretation of, for instance, the time-course of the observations, given the large temporal spread caused by the diffusion-limited binding processes. Methods and limitations of single-molecule microscopy for the study of cell signaling are discussed on the example of the chemotactic signaling of the slime-mold Dictyostelium discoideum. Single-molecule studies, as reviewed in this chapter, appear to be one of the essential methodologies for the full spatiotemporal clarification of cellular signaling, one of the ultimate goals in cell biology.

  14. Divergent Label-free Cell Phenotypic Pharmacology of Ligands at the Overexpressed β2-Adrenergic Receptors

    Science.gov (United States)

    Ferrie, Ann M.; Sun, Haiyan; Zaytseva, Natalya; Fang, Ye

    2014-01-01

    We present subclone sensitive cell phenotypic pharmacology of ligands at the β2-adrenergic receptor (β2-AR) stably expressed in HEK-293 cells. The parental cell line was transfected with green fluorescent protein (GFP)-tagged β2-AR. Four stable subclones were established and used to profile a library of sixty-nine AR ligands. Dynamic mass redistribution (DMR) profiling resulted in a pharmacological activity map suggesting that HEK293 endogenously expresses functional Gi-coupled α2-AR and Gs-coupled β2-AR, and the label-free cell phenotypic activity of AR ligands are subclone dependent. Pathway deconvolution revealed that the DMR of epinephrine is originated mostly from the remodeling of actin microfilaments and adhesion complexes, to less extent from the microtubule networks and receptor trafficking, and certain agonists displayed different efficacy towards the cAMP-Epac pathway. We demonstrate that receptor signaling and ligand pharmacology is sensitive to the receptor expression level, and the organization of the receptor and its signaling circuitry.

  15. Linking aptamer-ligand binding and expression platform folding in riboswitches: prospects for mechanistic modeling and design.

    Science.gov (United States)

    Aboul-ela, Fareed; Huang, Wei; Abd Elrahman, Maaly; Boyapati, Vamsi; Li, Pan

    2015-01-01

    The power of riboswitches in regulation of bacterial metabolism derives from coupling of two characteristics: recognition and folding. Riboswitches contain aptamers, which function as biosensors. Upon detection of the signaling molecule, the riboswitch transduces the signal into a genetic decision. The genetic decision is coupled to refolding of the expression platform, which is distinct from, although overlapping with, the aptamer. Early biophysical studies of riboswitches focused on recognition of the ligand by the aptamer-an important consideration for drug design. A mechanistic understanding of ligand-induced riboswitch RNA folding can further enhance riboswitch ligand design, and inform efforts to tune and engineer riboswitches with novel properties. X-ray structures of aptamer/ligand complexes point to mechanisms through which the ligand brings together distal strand segments to form a P1 helix. Transcriptional riboswitches must detect the ligand and form this P1 helix within the timescale of transcription. Depending on the cell's metabolic state and cellular environmental conditions, the folding and genetic outcome may therefore be affected by kinetics of ligand binding, RNA folding, and transcriptional pausing, among other factors. Although some studies of isolated riboswitch aptamers found homogeneous, prefolded conformations, experimental, and theoretical studies point to functional and structural heterogeneity for nascent transcripts. Recently it has been shown that some riboswitch segments, containing the aptamer and partial expression platforms, can form binding-competent conformers that incorporate an incomplete aptamer secondary structure. Consideration of the free energy landscape for riboswitch RNA folding suggests models for how these conformers may act as transition states-facilitating rapid, ligand-mediated aptamer folding. PMID:26361734

  16. Structural studies of Ca2+-ATPase ligand and regulatory complexes

    DEFF Research Database (Denmark)

    Drachmann, Nikolaj Düring

    2015-01-01

    against their concentration gradient upon ATP hydrolysis. The ion gradients are used to drive several key cellular processes, like the action potential in nerve tissue, acidification of the gastric juice, cell signalling and muscle contraction. The Ca2+-ATPase is an important part of mammalian cells...... choline lipids with different aliphatic chain length and saturation show three specific lipid binding sites. The four different lipids analysed bind to the same binding sites with varying degrees of disorder. The study contributes to understanding the complex interplay between the surrounding membrane......-of-concept Ca2+ bound crystal form, indicated that the information content of SFX data is higher than synchrotron data, and ligands and ions can be detected with low redundant data. The data of the E2 stabilised form was processed to 5 Å resolution, and it was possible to extract useful anomalous data showing...

  17. Flexible Ligand Docking Using Evolutionary Algorithms

    DEFF Research Database (Denmark)

    Thomsen, Rene

    2003-01-01

    search spaces effectively and is one of the commonly used methods for flexible ligand docking. During the last decade, several EAs using different variation operators have been introduced, such as the ones provided with the AutoDock program. In this paper we evaluate the performance of different EA......The docking of ligands to proteins can be formulated as a computational problem where the task is to find the most favorable energetic conformation among the large space of possible protein–ligand complexes. Stochastic search methods such as evolutionary algorithms (EAs) can be used to sample large...... settings such as choice of variation operators, population size, and usage of local search. The comparison is performed on a suite of six docking problems previously used to evaluate the performance of search algorithms provided with the AutoDock program package. The results from our investigation confirm...

  18. Effects of PPARγ Ligands on Leukemia

    Directory of Open Access Journals (Sweden)

    Yoko Tabe

    2012-01-01

    Full Text Available Peroxisome proliferator-activated receptors (PPARs and retinoic acid receptors (RARs, members of the nuclear receptor superfamily, are transcription factors that regulate a variety of important cellular functions. PPARs form heterodimers retinoid X receptor (RXR, an obligate heterodimeric partner for other nuclear receptors. Several novel links between retinoid metabolism and PPAR responses have been identified, and activation of PPAR/RXR expression has been shown to increase response to retinoids. PPARγ has emerged as a key regulator of cell growth and survival, whose activity is modulated by a number of synthetic and natural ligands. While clinical trials in cancer patients with thiazolidinediones (TZD have been disappointing, novel structurally different PPARγ ligands, including triterpenoids, have entered clinical arena as therapeutic agents for epithelial and hematopoietic malignancies. Here we shall review the antitumor advances of PPARγ, alone and in combination with RARα ligands in control of cell proliferation, differentiation, and apoptosis and their potential therapeutic applications in hematological malignancies.

  19. Ligand sphere conversions in terminal carbide complexes

    DEFF Research Database (Denmark)

    Morsing, Thorbjørn Juul; Reinholdt, Anders; Sauer, Stephan P. A.;

    2016-01-01

    Metathesis is introduced as a preparative route to terminal carbide complexes. The chloride ligands of the terminal carbide complex [RuC(Cl)2(PCy3)2] (RuC) can be exchanged, paving the way for a systematic variation of the ligand sphere. A series of substituted complexes, including the first...... example of a cationic terminal carbide complex, [RuC(Cl)(CH3CN)(PCy3)2]+, is described and characterized by NMR, MS, X-ray crystallography, and computational studies. The experimentally observed irregular variation of the carbide 13C chemical shift is shown to be accurately reproduced by DFT, which also...... demonstrates that details of the coordination geometry affect the carbide chemical shift equally as much as variations in the nature of the auxiliary ligands. Furthermore, the kinetics of formation of the sqaure pyramidal dicyano complex, trans-[RuC(CN)2(PCy3)2], from RuC has been examined and the reaction...

  20. Mutual inactivation of Notch receptors and ligands facilitates developmental patterning.

    Directory of Open Access Journals (Sweden)

    David Sprinzak

    2011-06-01

    Full Text Available Developmental patterning requires juxtacrine signaling in order to tightly coordinate the fates of neighboring cells. Recent work has shown that Notch and Delta, the canonical metazoan juxtacrine signaling receptor and ligand, mutually inactivate each other in the same cell. This cis-interaction generates mutually exclusive sending and receiving states in individual cells. It generally remains unclear, however, how this mutual inactivation and the resulting switching behavior can impact developmental patterning circuits. Here we address this question using mathematical modeling in the context of two canonical pattern formation processes: boundary formation and lateral inhibition. For boundary formation, in a model motivated by Drosophila wing vein patterning, we find that mutual inactivation allows sharp boundary formation across a broader range of parameters than models lacking mutual inactivation. This model with mutual inactivation also exhibits robustness to correlated gene expression perturbations. For lateral inhibition, we find that mutual inactivation speeds up patterning dynamics, relieves the need for cooperative regulatory interactions, and expands the range of parameter values that permit pattern formation, compared to canonical models. Furthermore, mutual inactivation enables a simple lateral inhibition circuit architecture which requires only a single downstream regulatory step. Both model systems show how mutual inactivation can facilitate robust fine-grained patterning processes that would be difficult to implement without it, by encoding a difference-promoting feedback within the signaling system itself. Together, these results provide a framework for analysis of more complex Notch-dependent developmental systems.

  1. Supramolecular architectures constructed using angular bipyridyl ligands

    International Nuclear Information System (INIS)

    This work details the synthesis and characterization of a series of coordination frameworks that are formed using bidentate angular N-donor ligands. Pyrimidine was reacted with metal(ll) nitrate salts. Reactions using Cd(NO3)2 receive particular focus and the analogous reactions using the linear ligand, pyrazine, were studied for comparison. In all cases, two-dimensional coordination networks were prepared. Structural diversity is observed for the Cd(ll) centres including metal-nitrate bridging. In contrast, first row transition metal nitrates form isostructural one-dimensional chains with only the bridging N-donor ligands generating polymeric propagation. The angular ligand, 2,4-bis(4-pyridyl)-1,3,5-triazine (dpt), was reacted with Cd(NO3)2 and Zn(NO3)2. Whereas Zn(NO3)2 compounds exhibit solvent mediated polymorphism, a range of structures were obtained for the reactions with Cd(NO3)2, including the first example of a doubly parallel interpenetrated 4.82 net. 4,7-phenanthroline, was reacted with various metal(ll) nitrates as well as cobalt(ll) and copper(ll) halides. The ability of 4,7-phenanthroline to act as both a N-donor ligand and a hydrogen bond acceptor has been discussed. Reactions of CuSCN with pyrimidine yield an unusual three-dimensional structure in which polymeric propagation is not a result of ligand bridging. The reaction of CuSCN with dpt yielded structural supramolecular isomers. (author)

  2. Emerging Roles for CSF-1 Receptor and its Ligands in the Nervous System.

    Science.gov (United States)

    Chitu, Violeta; Gokhan, Şölen; Nandi, Sayan; Mehler, Mark F; Stanley, E Richard

    2016-06-01

    The colony-stimulating factor-1 receptor (CSF-1R) kinase regulates tissue macrophage homeostasis, osteoclastogenesis, and Paneth cell development. However, recent studies in mice have revealed that CSF-1R signaling directly controls the development and maintenance of microglia, and cell autonomously regulates neuronal differentiation and survival. While the CSF-1R-cognate ligands, CSF-1 and interleukin-34 (IL-34) compete for binding to the CSF-1R, they are expressed in a largely non-overlapping manner by mature neurons. The recent identification of a dominantly inherited, adult-onset, progressive dementia associated with inactivating mutations in the CSF-1R highlights the importance of CSF-1R signaling in the brain. We review the roles of the CSF-1R and its ligands in microglial and neural development and function, and their relevance to our understanding of neurodegenerative disease.

  3. Multilayered proteomics reveals molecular switches dictating ligand-dependent EGFR trafficking

    DEFF Research Database (Denmark)

    Francavilla, Chiara; Papetti, Moreno; Rigbolt, Kristoffer T G;

    2016-01-01

    A fascinating conundrum in cell signaling is how stimulation of the same receptor tyrosine kinase with distinct ligands generates specific outcomes. To decipher the functional selectivity of EGF and TGF-α, which induce epidermal growth factor receptor (EGFR) degradation and recycling, respectively...... identified RAB7 phosphorylation and RCP recruitment to EGFR as switches for EGF and TGF-α outputs, controlling receptor trafficking, signaling duration, proliferation, and migration. By manipulating RCP levels or phosphorylation of RAB7 in EGFR-positive cancer cells, we were able to switch a TGF......, we devised an integrated multilayered proteomics approach (IMPA). We analyzed dynamic changes in the receptor interactome, ubiquitinome, phosphoproteome, and late proteome in response to both ligands in human cells by quantitative MS and identified 67 proteins regulated at multiple levels. We...

  4. Cationic ruthenium alkylidene catalysts bearing phosphine ligands

    OpenAIRE

    Endo, Koji; Grubbs, Robert H.

    2016-01-01

    The discovery of highly active catalysts and the success of ionic liquid immobilized systems have accelerated attention to a new class of cationic metathesis catalysts. We herein report the facile syntheses of cationic ruthenium catalysts bear-ing bulky phosphine ligands. Simple ligand exchange using silver(I) salts of non-coordinating or weakly coordinating anions pro-vided either PPh3 or chelating Ph2P(CH2)nPPh2 (n = 2 or 3) ligated cationic catalysts. The structures of these newly reported...

  5. Ligand Intermediates in Metal-Catalyzed Reactions

    Energy Technology Data Exchange (ETDEWEB)

    Gladysz, John A.

    1999-07-31

    The longest-running goal of this project has been the synthesis, isolation, and physical chemical characterization of homogeneous transition metal complexes containing ligand types believed to be intermediates in the metal-catalyzed conversion of CO/H{sub 2}, CO{sub 2}, CH{sub 4}, and similar raw materials to organic fuels, feedstocks, etc. In the current project period, complexes that contain unusual new types of C{sub x}(carbide) and C{sub x}O{sub y} (carbon oxide) ligands have been emphasized. A new program in homogeneous fluorous phase catalysis has been launched as described in the final report.

  6. Efficient chemoenzymatic synthesis of chiral pincer ligands.

    Science.gov (United States)

    Felluga, Fulvia; Baratta, Walter; Fanfoni, Lidia; Pitacco, Giuliana; Rigo, Pierluigi; Benedetti, Fabio

    2009-05-01

    Chiral, nonracemic pincer ligands based on the 6-phenyl-2-aminomethylpyridine and 2-aminomethylbenzo[h]quinoline scaffolds were obtained by a chemoenzymatic approach starting from 2-pyridyl and 2-benzoquinolyl ethanone. In the enantiodifferentiating step, secondary alcohols of opposite absolute configuration were obtained by a baker's yeast reduction of the ketones and by lipase-mediated dynamic kinetic resolution of the racemic alcohols. Their transformation into homochiral 1-methyl-1-heteroarylethanamines occurred without loss of optical purity, giving access to pincer ligands used in enantioselective catalysis.

  7. Epidermal growth factor receptor ligands as new extracellular targets for the metastasis-promoting S100A4 protein

    DEFF Research Database (Denmark)

    Klingelhöfer, Jörg; Møller, Henrik D.; Sumer, Eren U;

    2009-01-01

    that the most abundant peptide mimicked the F/YCC motif present in the epidermal growth factor domain of ErbB receptor ligands. S100A4 selectively interacted with a number of epidermal growth factor receptor (EGFR) ligands, demonstrating highest affinity for amphiregulin. Importantly, we found that S100A4...... stimulated EGFR/ErbB2 receptor signaling and enhanced the amphiregulin-mediated proliferation of mouse embryonic fibroblasts. S100A4-neutralizing antibodies, as well as EGFR- and ErbB2 receptor-specific tyrosine kinase inhibitors, blocked these effects. The present results suggest that extracellular S100A4...... regulates tumor progression by interacting with EGFR ligands, thereby enhancing EGFR/ErbB2 receptor signaling and cell proliferation. Structured digital abstract: * MINT-7256556: EGF (uniprotkb:P01133) binds (MI:0407) to S100A4 (uniprotkb:P26447) by far western blotting (MI:0047) * MINT-7256512: BC...

  8. FIST: a sensory domain for diverse signal transduction pathways in prokaryotes and ubiquitin signaling in eukaryotes

    Energy Technology Data Exchange (ETDEWEB)

    Borziak, Kirill [ORNL; Jouline, Igor B [ORNL

    2007-01-01

    Motivation: Sensory domains that are conserved among Bacteria, Archaea and Eucarya are important detectors of common signals detected by living cells. Due to their high sequence divergence, sensory domains are difficult to identify. We systematically look for novel sensory domains using sensitive profile-based searches initi-ated with regions of signal transduction proteins where no known domains can be identified by current domain models. Results: Using profile searches followed by multiple sequence alignment, structure prediction, and domain architecture analysis, we have identified a novel sensory domain termed FIST, which is present in signal transduction proteins from Bacteria, Archaea and Eucarya. Remote similarity to a known ligand-binding fold and chromosomal proximity of FIST-encoding genes to those coding for proteins involved in amino acid metabolism and transport suggest that FIST domains bind small ligands, such as amino acids.

  9. Abundance of Flt3 and its ligand in astrocytic tumors

    Directory of Open Access Journals (Sweden)

    Eßbach C

    2013-05-01

    Full Text Available C Eßbach,1 N Andrae,1 D Pachow,1 J-P Warnke,2 A Wilisch-Neumann,1 E Kirches,1 C Mawrin11Department of Neuropathology, Otto-von-Guericke University, Magdeburg, 2Department of Neurosurgery, Paracelsus Hospital, Zwickau, GermanyBackground: Molecular targeted therapies for astrocytic tumors are the subject of growing research interest, due to the limited response of these tumors, especially glioblastoma multiforme, to conventional chemotherapeutic regimens. Several of these approaches exploit the inhibition of receptor tyrosine kinases. To date, it has not been elucidated if fms-like tyrosine kinase-3 (Flt3 and its natural ligand (Flt3L are expressed in astrocytic tumors, although some of the clinically intended small-molecule receptor tyrosine kinase inhibitors affect Flt3, while others do not. More importantly, the recent proof of principle for successful stimulation of the immune system against gliomas in preclinical models via local Flt3L application requires elucidation of this receptor tyrosine kinase pathway in these tumors in more detail. This therapy is based on recruitment of Flt3-positive dendritic cells, but may be corroborated by activity of this signaling pathway in glioma cells.Methods: Receptor and ligand expression was analyzed by real-time polymerase chain reaction in 31 astrocytic tumors (six diffuse and 11 anaplastic astrocytomas, 14 glioblastomas derived from patients of both genders and in glioblastoma cell lines. The two most common activating mutations of the Flt3 gene, ie, internal tandem duplication and D835 point mutation, were assessed by specific polymerase chain reaction.Results: A relatively high abundance of Flt3L mRNA (4%–6% of the reference, β2 microglobulin could be demonstrated in all tumor samples. Flt3 expression could generally be demonstrated by 40 specific polymerase chain reaction cycles and gel electrophoresis in 87% of the tumors, including all grades, although the small quantities of the receptor did

  10. Differential role of ICAM ligands in determination of human memory T cell differentiation

    Directory of Open Access Journals (Sweden)

    Mitchell Dennis

    2007-01-01

    Full Text Available Abstract Background Leukocyte Function Antigen-1 (LFA-1 is a primary adhesion molecule that plays important roles in T cell activation, leukocyte recirculation, and trans-endothelial migration. By applying a multivariate intracellular phospho-proteomic analysis, we demonstrate that LFA-1 differentially activates signaling molecules. Results Signal intensity was dependent on both ICAM ligand and LFA-1 concentration. In the presence of CD3 and CD28 stimulation, ICAM-2 and ICAM-3 decreased TGFβ1 production more than ICAM-1. In long-term differentiation experiments, stimulation with ICAM-3, CD3, and CD28 generated IFNγ producing CD4+CD45RO+CD62L-CD11aBrightCD27- cells that had increased expression of intracellular BCL2, displayed distinct chemokine receptor profiles, and exhibited distinct migratory characteristics. Only CD3/CD28 with ICAM-3 generated CD4+CD45RO+CD62L-CD11aBrightCD27- cells that were functionally responsive to chemotaxis and exhibited higher frequencies of cells that signaled to JNK and ERK1/2 upon stimulation with MIP3α. Furthermore, these reports identify that the LFA-1 receptor, when presented with multiple ligands, can result in distinct T cell differentiation states and suggest that the combinatorial integration of ICAM ligand interactions with LFA-1 have functional consequences for T cell biology. Conclusion Thus, the ICAM ligands, differentially modulate LFA-1 signaling in T cells and potentiate the development of memory human T cells in vitro. These findings are of importance in a mechanistic understanding of memory cell differentiation and ex vivo generation of memory cell subsets for therapeutic applications.

  11. Ligand-gated chloride channels are receptors for biogenic amines in C. elegans

    OpenAIRE

    Ringstad, Niels; Abe, Namiko; Horvitz, H. Robert

    2009-01-01

    Biogenic amines such as serotonin and dopamine are intercellular signaling molecules that function widely as neurotransmitters and neuromodulators. We have identified in the nematode Caenorhabditis elegans three ligand-gated chloride channels that are receptors for biogenic amines: LGC-53 is a high-affinity dopamine receptor, LGC-55 is a high-affinity tyramine receptor, and LGC-40 is a low-affinity serotonin receptor that is also gated by choline and acetylcholine. lgc-55 mutants are defectiv...

  12. The identification of histidine ligands to cytochrome a in cytochrome c oxidase

    OpenAIRE

    Martin, Craig T.; Scholes, Charles P.; Chan, Sunney I.

    1985-01-01

    A histidine auxotroph of Saccharomyces cerevisiae has been used to metabolically incorporate [1,3-15N2] histidine into yeast cytochrome c oxidase. Electron nuclear double resonance (ENDOR) spectroscopy of cytochrome a in the [15N]histidine-substituted enzyme reveals an ENDOR signal which can be assigned to hyperfine coupling of a histidine 15N with the low-spin heme, thereby unambiguously identifying histidine as an axial ligand to this cytochrome. Comparison of this result with similar ENDOR...

  13. Nitrosamines as nicotinic receptor ligands.

    Science.gov (United States)

    Schuller, Hildegard M

    2007-05-30

    Nitrosamines are carcinogens formed in the mammalian organism from amine precursors contained in food, beverages, cosmetics and drugs. The potent carcinogen, NNK, and the weaker carcinogen, NNN, are nitrosamines formed from nicotine. Metabolites of the nitrosamines react with DNA to form adducts responsible for genotoxic effects. We have identified NNK as a high affinity agonist for the alpha7 nicotinic acetylcholine receptor (alpha7nAChR) whereas NNN bound with high affinity to epibatidine-sensitive nAChRs. Diethylnitrosamine (DEN) bound to both receptors but with lower affinity. High levels of the alpha7nAChR were expressed in human small cell lung cancer (SCLC) cell lines and in hamster pulmonary neuroendocrine cells (PNECs), which serve as a model for the cell of origin of human SCLC. Exposure of SCLC or PNECs to NNK or nicotine increased expression of the alpha7nAChR and caused influx of Ca(2+), activation of PKC, Raf-1, ERK1/2, and c-myc, resulting in the stimulation of cell proliferation. Signaling via the alpha7nAChR was enhanced when cells were maintained in an environment of 10-15% CO(2) similar to that in the diseased lung. Hamsters with hyperoxia-induced pulmonary fibrosis developed neuroendocrine lung carcinomas similar to human SCLC when treated with NNK, DEN, or nicotine. The development of the NNK-induced tumors was prevented by green tea or theophylline. The beta-adrenergic receptor agonist, isoproterenol or theophylline blocked NNK-induced cell proliferation in vitro. NNK and nicotine-induced hyperactivity of the alpha7nAChR/RAF/ERK1/2 pathway thus appears to play a crucial role in the development of SCLC in smokers and could be targeted for cancer prevention. PMID:17459420

  14. Activation of epidermal growth factor receptor by metal-ligand complexes decreases levels of extracellular amyloid beta peptide.

    Science.gov (United States)

    Price, Katherine A; Filiz, Gulay; Caragounis, Aphrodite; Du, Tai; Laughton, Katrina M; Masters, Colin L; Sharples, Robyn A; Hill, Andrew F; Li, Qiao-Xin; Donnelly, Paul S; Barnham, Kevin J; Crouch, Peter J; White, Anthony R

    2008-01-01

    The epidermal growth factor receptor is a receptor tyrosine kinase expressed in a range of tissues and cell-types. Activation of the epidermal growth factor receptor by a number of ligands induces downstream signalling that modulates critical cell functions including growth, survival and differentiation. Abnormal epidermal growth factor receptor expression and activation is also involved in a number of cancers. In addition to its cognate ligands, the epidermal growth factor receptor can be activated by metals such as zinc (Zn) and copper (Cu). Due to the important role of these metals in a number of diseases including neurodegenerative disorders, therapeutic approaches are being developed based on the use of lipid permeable metal-complexing molecules. While these agents are showing promising results in animal models and clinical trials, little is known about the effects of metal-ligand complexes on cell signalling pathways. In this study, we investigated the effects of clioquinol (CQ)-metal complexes on activation of epidermal growth factor receptor. We show here that CQ-Cu complexes induced potent epidermal growth factor receptor phosphorylation resulting in downstream activation of extracellular signal-regulated kinase. Similar levels of epidermal growth factor receptor activation were observed with alternative lipid permeable metal-ligands including neocuproine and pyrrolidine dithiocarbamate. We found that CQ-Cu complexes induced a significant reduction in the level of extracellular Abeta1-40 in cell culture. Inhibition of epidermal growth factor receptor activation by PD153035 blocked extracellular signal-regulated kinase phosphorylation and restored Abeta1-40 levels. Activation of the epidermal growth factor receptor by CQ-Cu was mediated through up-regulation of src kinase activity by a cognate ligand-independent process involving membrane integrins. These findings provide the first evidence that metal-ligand complexes can activate the epidermal growth

  15. Simultaneous inhibition of epidermal growth factor receptor (EGFR) signaling and enhanced activation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor-mediated apoptosis induction by an scFv : sTRAIL fusion protein with specificity for human EGFR

    NARCIS (Netherlands)

    Bremer, E; Samplonius, DF; van Genne, L; Dijkstra, MH; Kroesen, BJ; de Leij, LFMH; Helfrich, W

    2005-01-01

    Epidermal growth factor receptor (EGFR) signaling inhibition by monoclonal antibodies and EGFR-specific tyrosine kinase inhibitors has shown clinical efficacy in cancer by restoring susceptibility of tumor cells to therapeutic apoptosis induction. Tumor necrosis factor-related apoptosis-inducing lig

  16. An in Vitro and in Vivo Investigation of Bivalent Ligands That Display Preferential Binding and Functional Activity for Different Melanocortin Receptor Homodimers.

    Science.gov (United States)

    Lensing, Cody J; Freeman, Katie T; Schnell, Sathya M; Adank, Danielle N; Speth, Robert C; Haskell-Luevano, Carrie

    2016-04-14

    Pharmacological probes for the melanocortin receptors have been utilized for studying various disease states including cancer, sexual function disorders, Alzheimer's disease, social disorders, cachexia, and obesity. This study focused on the design and synthesis of bivalent ligands to target melanocortin receptor homodimers. Lead ligands increased binding affinity by 14- to 25-fold and increased cAMP signaling potency by 3- to 5-fold compared to their monovalent counterparts. Unexpectedly, different bivalent ligands showed preferences for particular melanocortin receptor subtypes depending on the linker that connected the binding scaffolds, suggesting structural differences between the various dimer subtypes. Homobivalent compound 12 possessed a functional profile that was unique from its monovalent counterpart providing evidence of the discrete effects of bivalent ligands. Lead compound 7 significantly decreased feeding in mice after intracerebroventricular administration. To the best of our knowledge, this is the first report of a melanocortin bivalent ligand's in vivo physiological effects. PMID:26959173

  17. Characterization and mucosal responses of interleukin 17 family ligand and receptor genes in channel catfish Ictalurus punctatus

    Science.gov (United States)

    Interleukin (IL) 17 family cytokines are important mediators of mucosal immune responses, tightly regulated by signals from the complex milieu of pathogenic and commensal microbes, epithelial cells and innate and adaptive leukocytes found at tissue barriers. In mammals, IL17 ligand expression has be...

  18. Regulation of NKG2D-ligand cell surface expression by intracellular calcium after HDAC-inhibitor treatment

    DEFF Research Database (Denmark)

    Jensen, Helle; Hagemann-Jensen, Michael Henrik; Lauridsen, Felicia Kathrine Bratt;

    2013-01-01

    surface expression of ULBP2, but not MICA/B, was sensitive to treatment calmidazolium and trifluoperazine, two agents known to block calcium signaling. siRNA-mediated knock-down of the calcium-regulated proteins calmodulin or calpain did however not affect NKG2D-ligand cell surface expression on Jurkat T...

  19. A versatile dinucleating ligand containing sulfonamide groups

    DEFF Research Database (Denmark)

    Sundberg, Jonas; Witt, Hannes; Cameron, Lisa;

    2014-01-01

    Copper, iron, and gallium coordination chemistries of the new pentadentate bis-sulfonamide ligand 2,6-bis(N-2-pyridylmethylsulfonamido)-4-methylphenol (psmpH3) were investigated. PsmpH3 is capable of varying degrees of deprotonation, and notably, complexes containing the fully trideprotonated...

  20. Constitutive and ligand-induced TCR degradation

    DEFF Research Database (Denmark)

    von Essen, Marina; Bonefeld, Charlotte Menné; Siersma, Volkert;

    2004-01-01

    divergent models for TCR down-regulation and degradation have been suggested. The aims of this study were to determine the rate constants for constitutive and ligand-induced TCR degradation and to determine whether the TCR subunits segregate or are processed as an intact unit during TCR down-regulation and...

  1. Ligand iron catalysts for selective hydrogenation

    Science.gov (United States)

    Casey, Charles P.; Guan, Hairong

    2010-11-16

    Disclosed are iron ligand catalysts for selective hydrogenation of aldehydes, ketones and imines. A catalyst such as dicarbonyl iron hydride hydroxycyclopentadiene) complex uses the OH on the five member ring and hydrogen linked to the iron to facilitate hydrogenation reactions, particularly in the presence of hydrogen gas.

  2. Metal speciation dynamics in colloidal ligand dispersions

    NARCIS (Netherlands)

    Pinheiro, J.P.; Minor, M.; Leeuwen, van H.P.

    2005-01-01

    In this work we propose a dynamic metal speciation theory for colloidal systems in which the complexing ligands are localized on the surface of the particles; i.e., there is spatial heterogeneity of binding sites within the sample volume. The differences between the complex formation and dissociatio

  3. Supramolecular architectures constructed using angular bipyridyl ligands

    CERN Document Server

    Barnett, S A

    2003-01-01

    This work details the synthesis and characterization of a series of coordination frameworks that are formed using bidentate angular N-donor ligands. Pyrimidine was reacted with metal(ll) nitrate salts. Reactions using Cd(NO sub 3) sub 2 receive particular focus and the analogous reactions using the linear ligand, pyrazine, were studied for comparison. In all cases, two-dimensional coordination networks were prepared. Structural diversity is observed for the Cd(ll) centres including metal-nitrate bridging. In contrast, first row transition metal nitrates form isostructural one-dimensional chains with only the bridging N-donor ligands generating polymeric propagation. The angular ligand, 2,4-bis(4-pyridyl)-1,3,5-triazine (dpt), was reacted with Cd(NO sub 3) sub 2 and Zn(NO sub 3) sub 2. Whereas Zn(NO sub 3) sub 2 compounds exhibit solvent mediated polymorphism, a range of structures were obtained for the reactions with Cd(NO sub 3) sub 2 , including the first example of a doubly parallel interpenetrated 4.8 sup...

  4. Follicle-restricted compartmentalization of transforming growth factor beta superfamily ligands in the feline ovary.

    Science.gov (United States)

    Bristol, Sarah K; Woodruff, Teresa K

    2004-03-01

    Ovarian follicular development, follicle selection, and the process of ovulation remain poorly understood in most species. Throughout reproductive life, follicle fate is balanced between growth and apoptosis. These opposing forces are controlled by numerous endocrine, paracrine, and autocrine factors, including the ligands represented by the transforming growth factor beta (TGFbeta) superfamily. TGFbeta, activin, inhibin, bone morphometric protein (BMP), and growth differentiation factor 9 (GDF-9) are present in the ovary of many animals; however, no comprehensive analysis of the localization of each ligand or its receptors and intracellular signaling molecules during folliculogenesis has been done. The domestic cat is an ideal model for studying ovarian follicle dynamics due to an abundance of all follicle populations, including primordial stage, and the amount of readily available tissue following routine animal spaying. Additionally, knowledge of the factors involved in feline follicular development could make an important impact on in vitro maturation/in vitro fertilization (IVM/IVF) success for endangered feline species. Thus, the presence and position of TGFbeta superfamily members within the feline ovary have been evaluated in all stages of follicular development by immunolocalization. The cat inhibin alpha subunit protein is present in all follicle stages but increases in intensity within the mural granulosa cells in large antral follicles. The inhibin betaA and betaB subunit proteins, in addition to the activin type I (ActRIB) and activin type II receptor (ActRIIB), are produced in primordial and primary follicle granulosa cells. Additionally, inhibin betaA subunit is detected in the theca cells from secondary through large antral follicle size classes. GDF-9 is restricted to the oocyte of preantral and antral follicles, whereas the type II BMP receptor (BMP-RII) protein is predominantly localized to primordial- and primary-stage follicles. TGFbeta1, 2

  5. Ligand-induced Coupling versus Receptor Pre-association: Cellular automaton simulations of FGF-2 binding

    OpenAIRE

    Gopalakrishnan, Manoj; Forsten-Williams, Kimberly; Tauber, Uwe C.

    2003-01-01

    The binding of basic fibroblast growth factor (FGF-2) to its cell surface receptor (CSR) and subsequent signal transduction is known to be enhanced by Heparan Sulfate Proteoglycans (HSPGs). HSPGs bind FGF-2 with low affinity and likely impact CSR-mediated signaling via stabilization of FGF-2-CSR complexes via association with both the ligand and the receptor. What is unknown is whether HSPG associates with CSR in the absence of FGF-2. In this paper, we determine conditions by which pre-associ...

  6. Chemotaxis receptor complexes: from signaling to assembly.

    Directory of Open Access Journals (Sweden)

    Robert G Endres

    2007-07-01

    Full Text Available Complexes of chemoreceptors in the bacterial cytoplasmic membrane allow for the sensing of ligands with remarkable sensitivity. Despite the excellent characterization of the chemotaxis signaling network, very little is known about what controls receptor complex size. Here we use in vitro signaling data to model the distribution of complex sizes. In particular, we model Tar receptors in membranes as an ensemble of different sized oligomer complexes, i.e., receptor dimers, dimers of dimers, and trimers of dimers, where the relative free energies, including receptor modification, ligand binding, and interaction with the kinase CheA determine the size distribution. Our model compares favorably with a variety of signaling data, including dose-response curves of receptor activity and the dependence of activity on receptor density in the membrane. We propose that the kinetics of complex assembly can be measured in vitro from the temporal response to a perturbation of the complex free energies, e.g., by addition of ligand.

  7. Substrate coated with receptor and labelled ligand for assays

    International Nuclear Information System (INIS)

    Improvements in the procedures for assaying ligands are described. The assay consists of a polystyrene tube on which receptors are present for both the ligand to be assayed and a radioactively labelled form of the ligand. The receptors on the bottom portion of the tube are also coated with labelled ligands, thus eliminating the necessity for separate addition of the labelled ligand and sample during an assay. Examples of ligands to which this method is applicable include polypeptides, nucleotides, nucleosides and proteins. Specific examples are given in which the ligand to be assayed is digoxin, the labelled form of the ligand is 3-0-succinyl digoxyigenin tyrosine (125I) and the receptor is digoxin antibody. (U.K.)

  8. The Role of the Enterohepatic Circulation of Bile Salts and Nuclear Hormone Receptors in the Regulation of Cholesterol Homeostasis: Bile Salts as Ligands for Nuclear Hormone Receptors

    OpenAIRE

    Redinger, Richard N.

    2003-01-01

    The coordinated effect of lipid activated nuclear hormone receptors; liver X receptor (LXR), bound by oxysterol ligands and farnesoid X receptor (FXR), bound by bile acid ligands, act as genetic transcription factors to cause feed-forward cholesterol catabolism to bile acids and feedback repression of bile acid synthesis, respectively. It is the coordinated action of LXR and FXR, each dimerized to retinoid X receptor, that signal nuclear DNA response elements to encode proteins that prevent e...

  9. Role of ligand-ligand vs. core-core interactions in gold nanoclusters.

    Science.gov (United States)

    Milowska, Karolina Z; Stolarczyk, Jacek K

    2016-05-14

    The controlled assembly of ligand-coated gold nanoclusters (NCs) into larger structures paves the way for new applications ranging from electronics to nanomedicine. Here, we demonstrate through rigorous density functional theory (DFT) calculations employing novel functionals accounting for van der Waals forces that the ligand-ligand interactions determine whether stable assemblies can be formed. The study of NCs with different core sizes, symmetry forms, ligand lengths, mutual crystal orientations, and in the presence of a solvent suggests that core-to-core van der Waals interactions play a lesser role in the assembly. The dominant interactions originate from combination of steric effects, augmented by ligand bundling on NC facets, and related to them changes in electronic properties induced by neighbouring NCs. We also show that, in contrast to standard colloidal theory approach, DFT correctly reproduces the surprising experimental trends in the strength of the inter-particle interaction observed when varying the length of the ligands. The results underpin the importance of understanding NC interactions in designing gold NCs for a specific function. PMID:27097887

  10. Structural studies of P-type ATPase–ligand complexes using an X-ray free-electron laser

    Directory of Open Access Journals (Sweden)

    Maike Bublitz

    2015-07-01

    Full Text Available Membrane proteins are key players in biological systems, mediating signalling events and the specific transport of e.g. ions and metabolites. Consequently, membrane proteins are targeted by a large number of currently approved drugs. Understanding their functions and molecular mechanisms is greatly dependent on structural information, not least on complexes with functionally or medically important ligands. Structure determination, however, is hampered by the difficulty of obtaining well diffracting, macroscopic crystals. Here, the feasibility of X-ray free-electron-laser-based serial femtosecond crystallography (SFX for the structure determination of membrane protein–ligand complexes using microcrystals of various native-source and recombinant P-type ATPase complexes is demonstrated. The data reveal the binding sites of a variety of ligands, including lipids and inhibitors such as the hallmark P-type ATPase inhibitor orthovanadate. By analyzing the resolution dependence of ligand densities and overall model qualities, SFX data quality metrics as well as suitable refinement procedures are discussed. Even at relatively low resolution and multiplicity, the identification of ligands can be demonstrated. This makes SFX a useful tool for ligand screening and thus for unravelling the molecular mechanisms of biologically active proteins.

  11. Ligand binding was acquired during evolution of nuclear receptors

    OpenAIRE

    Escriva, Hector; Safi, Rachid; Hänni, Catherine; Langlois, Marie-Claire; Saumitou-Laprade, Pierre; Stehelin, Dominique; Capron, André; Pierce, Raymond; Laudet, Vincent

    1997-01-01

    The nuclear receptor (NR) superfamily comprises, in addition to ligand-activated transcription factors, members for which no ligand has been identified to date. We demonstrate that orphan receptors are randomly distributed in the evolutionary tree and that there is no relationship between the position of a given liganded receptor in the tree and the chemical nature of its ligand. NRs are specific to metazoans, as revealed by a screen of NR-related sequences in early- and non-metazoan organism...

  12. Role of ROBO4 Signalling in Developmental and Pathological Angiogenesis

    Directory of Open Access Journals (Sweden)

    Suresh Singh Yadav

    2014-01-01

    Full Text Available Transmembrane roundabout receptor family members (ROBO1–ROBO4 principally orchestrate the neuronal guidance mechanism of the nervous system. Secreted glycoprotein SLITs are the most appreciated ligands for ROBOs. Recently identified ROBO4 is the key mediator of SLIT-ROBO mediated developmental and pathological angiogenesis. Although SLIT2 has been shown to interact with ROBO4 as ligand, it remains an open question whether this protein is the physiologic partner of ROBO4. The purpose of this review is to summarise how reliable SLIT2 as ligand for ROBO4 is, if not what the other possible mechanisms demonstrated till date for ROBO4 mediated developmental and pathological angiogenesis are. We conclude that ROBO4 is expressed specially in vascular endothelial cells and maintains the vascular integrity via either SLIT2 dependent or SLIT2 independent manner. On the contrary, it promotes the pathological angiogenesis by involving different signalling arm(s/unknown ligand(s. This review explores the interactions SLIT2/ROBO1, SLIT2/ROBO1–ROBO4, ROBO1/ROBO4, and ROBO4/UNC5B which can be promising and potential therapeutic targets for developmental angiogenesis defects and pathological angiogenesis. Finally we have reviewed the ROBO4 signalling pathways and made an effort to elaborate the insight of this signalling as therapeutic target of pathological angiogenesis.

  13. Open-shell organometallics: reactivity at the ligand

    NARCIS (Netherlands)

    W.I. Dzik; B. de Bruin

    2011-01-01

    The purpose of this review is to show that (cooperative) ligand radical reactivity can be effectively employed in synthetic organometallic chemistry and catalysis to achieve selectivity in radical-type transformations. The ‘redox non-innocence’ of ligands, and the controlled reactivity of ‘ligand ra

  14. Triple Bioaffinity Mass Spectrometry Concept for Thyroid Transporter Ligands

    NARCIS (Netherlands)

    Aqai, P.; Fryganas, C.; Mizuguchi, M.; Haasnoot, W.; Nielen, M.W.F.

    2012-01-01

    For the analysis of thyroid transporter ligands, a triple bioaffinity mass spectrometry (BioMS) concept was developed, with the aim at three different analytical objectives: rapid screening of any ligand, confirmation of known ligands in accordance with legislative requirements, and identification o

  15. Integrating structural and mutagenesis data to elucidate GPCR ligand binding

    DEFF Research Database (Denmark)

    Munk, Christian; Harpsøe, Kasper; Hauser, Alexander S;

    2016-01-01

    G protein-coupled receptors (GPCRs) represent the largest family of human membrane proteins, as well as drug targets. A recent boom in GPCR structural biology has provided detailed images of receptor ligand binding sites and interactions on the molecular level. An ever-increasing number of ligands...... elucidate new GPCR ligand binding sites, and ultimately design drugs with tailored pharmacological activity....

  16. Prosocial Signalling

    DEFF Research Database (Denmark)

    Kahsay, Goytom Abraha

    suggested that consumers pay price premium because this sends the signal that the consumer has prosocial preferences and a few empirical studies have documented that reputation plays a key role when consumers choose products containing prosocial components. However, little is known about consumers...... consumer goods and presents empirical evidences from a natural consumption data. This thesis also investigates consumers’ behaviour under a newly introduced pricing system called Pay-What-You-Want (PWYW) and investigates empirically whether reputation signalling can be used as a policy instrument in other...... on the role of social network in facilitating factor input transactions and the role of reputation in reducing enforcement. Finally, the third part consists of one paper which is concerned with investigating the effect of climate change and adaptation policy on agricultural production in Eastern Africa...

  17. Identification of Novel Smoothened Ligands Using Structure-Based Docking

    Science.gov (United States)

    Torosyan, Hayarpi; Parathaman, Pranavan; Irwin, John J.; Shoichet, Brian K.

    2016-01-01

    The seven transmembrane protein Smoothened is required for Hedgehog signaling during embryonic development and adult tissue homeostasis. Inappropriate activation of the Hedgehog signalling pathway leads to cancers such as basal cell carcinoma and medulloblastoma, and Smoothened inhibitors are now available clinically to treat these diseases. However, resistance to these inhibitors rapidly develops thereby limiting their efficacy. The determination of Smoothened crystal structures enables structure-based discovery of new ligands with new chemotypes that will be critical to combat resistance. In this study, we docked 3.2 million available, lead-like molecules against Smoothened, looking for those with high physical complementarity to its structure; this represents the first such campaign against the class Frizzled G-protein coupled receptor family. Twenty-one high-ranking compounds were selected for experimental testing, and four, representing three different chemotypes, were identified to antagonize Smoothened with IC50 values better than 50 μM. A screen for analogs revealed another six molecules, with IC50 values in the low micromolar range. Importantly, one of the most active of the new antagonists continued to be efficacious at the D473H mutant of Smoothened, which confers clinical resistance to the antagonist vismodegib in cancer treatment. PMID:27490099

  18. Auxin signaling

    OpenAIRE

    Quint, Marcel; Gray, William M.

    2006-01-01

    Auxin regulates a host of plant developmental and physiological processes, including embryogenesis, vascular differentiation, organogenesis, tropic growth, and root and shoot architecture. Genetic and biochemical studies carried out over the past decade have revealed that much of this regulation involves the SCFTIR1/AFB-mediated proteolysis of the Aux/IAA family of transcriptional regulators. With the recent finding that the TRANSPORT INHIBITOR RESPONSE1 (TIR1)/AUXIN SIGNALING F-BOX (AFB) pro...

  19. Gas-phase ligand loss and ligand substitution reactions of platinum(II) complexes of tridentate nitrogen donor ligands.

    Science.gov (United States)

    Wee, Sheena; O'Hair, Richard A J; McFadyen, W David

    2004-01-01

    The source of protons associated with the ligand loss channel of HX((n - 1)+) from [Pt(II)(dien)X](n+) (X = Cl, Br and I for n = 1 and X = NC(5)H(5) for n = 2) in the gas phase was investigated by deuterium-labelling studies. The results of these studies indicate that these protons originate from both the amino groups and the carbon backbone of the dien ligand. In some instances (e.g. X = Br and I), the protons lost from the carbon backbone can be even more abundant than the protons lost from the amino groups. The gas-phase substitution reactions of coordinatively saturated [Pt(II)(L(3))L(a)](2+) complexes (L(3) = tpy or dien) were also examined using ion-molecule reactions. The outcome of the ion-molecule reactions depends on both the ancillary ligand (L(3)) as well as the leaving group (L(a)). [Pt(II)(tpy)L(a)](2+) complexes undergo substitution reactions, with a faster rate when L(a) is a good leaving group, while the [Pt(II)(dien)L(a)](2+) complex undergoes a proton transfer reaction. PMID:15164352

  20. Costimulation of IL-2 Production through CD28 Is Dependent on the Size of Its Ligand.

    Science.gov (United States)

    Lim, Hong-Sheng; Cordoba, Shaun-Paul; Dushek, Omer; Goyette, Jesse; Taylor, Alison; Rudd, Christopher E; van der Merwe, P Anton

    2015-12-01

    Optimal T cell activation typically requires engagement of both the TCR and costimulatory receptors, such as CD28. Engagement of CD28 leads to tyrosine phosphorylation of its cytoplasmic region and recruitment of cytoplasmic signaling proteins. Although the exact mechanism of CD28 signal transduction is unknown, CD28 triggering has similarities to the TCR, which was proposed to use the kinetic-segregation (KS) mechanism. The KS model postulates that, when small receptors engage their ligands within areas of close (∼15 nm) contact in the T cell/APC interface, this facilitates phosphorylation by segregating the engaged receptor/ligand complex from receptor protein tyrosine phosphatases with large ectodomains, such as CD45. To test this hypothesis, we examined the effect of elongating the extracellular region of the CD28 ligand, CD80, on its ability to costimulate IL-2 production by primary T cells. CD80 elongation reduced its costimulatory effect without abrogating CD28 binding. Confocal microscopy revealed that elongated CD80 molecules were less well segregated from CD45 at the T cell/APC interface. T cells expressing CD28 harboring a key tyrosine-170 mutation were less sensitive to CD80 elongation. In summary, the effectiveness of CD28 costimulation is inversely proportional to the dimensions of the CD28-CD80 complex. Small CD28-CD80 complex dimensions are required for optimal costimulation by segregation from large inhibitory tyrosine phosphatases. These results demonstrate the importance of ligand dimensions for optimal costimulation of IL-2 production by T cells and suggest that the KS mechanism contributes to CD28 signaling.

  1. A robust ligand exchange approach for preparing hydrophilic, biocompatible photoluminescent quantum dots

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Sujuan; Zhou, Changhua [Key Laboratory for Special Functional Materials of the Ministry of Education, Henan University, Kaifeng 475004 (China); Yuan, Hang [Life Science Division, Graduate School at Shenzhen, Tsinghua University, Shenzhen 518055 (China); Shen, Huaibin [Key Laboratory for Special Functional Materials of the Ministry of Education, Henan University, Kaifeng 475004 (China); Zhao, Wenxiu [Life Science Division, Graduate School at Shenzhen, Tsinghua University, Shenzhen 518055 (China); Ma, Lan, E-mail: malan@sz.tsinghua.edu.cn [Life Science Division, Graduate School at Shenzhen, Tsinghua University, Shenzhen 518055 (China); Li, Lin Song, E-mail: lsli@henu.edu.cn [Key Laboratory for Special Functional Materials of the Ministry of Education, Henan University, Kaifeng 475004 (China)

    2013-08-01

    Graphical abstract: - Highlights: • Aqueous CdSe/ZnS QDs were prepared using polymaleic anhydrides as capping ligand. • Effect of reaction temperature and time were systematically studied in the synthesis process. • Water-soluble QDs exhibited a good stability in physiological relevant environment. • The aqueous QDs were applied as biological probe to detect human embryonic stem cell. - Abstract: This paper describes a robust ligand exchange approach for preparing biocompatible CdSe/ZnS quantum dots (QDs) to make bioprobe for effective cell imaging. In this method, polymaleic anhydride (PMA) ligand are first used to replace original hydrophobic ligand (oleic acid) and form a protection shell with multiple hydrophilic groups to coat and protect CdSe/ZnS QDs. The as-prepared aqueous QDs exhibit small particle size, good colloidal stability in aqueous solutions with a wide range of pH, salt concentrations and under thermal treatment, which are necessary for biological applications. The use of this new class of aqueous QDs for effective cell imaging shows strong fluorescence signal to human embryonic stem cell, which demonstrate that PMA coated QDs are fully satisfied with the requirements of preparing high quality biological probe.

  2. Ligand-stimulated downregulation of the alpha interferon receptor: role of protein kinase D2.

    Science.gov (United States)

    Zheng, Hui; Qian, Juan; Varghese, Bentley; Baker, Darren P; Fuchs, Serge

    2011-02-01

    Alpha interferon (IFN-α) controls homeostasis of hematopoietic stem cells, regulates antiviral resistance, inhibits angiogenesis, and suppresses tumor growth. This cytokine is often used to treat cancers and chronic viral infections. The extent of cellular responses to IFN-α is limited by the IFN-induced ubiquitination and degradation of the IFN-α/β receptor chain 1 (IFNAR1) chain of the cognate receptor. IFNAR1 ubiquitination is facilitated by the βTrcp E3 ubiquitin ligase that is recruited to IFNAR1 upon its degron phosphorylation, which is induced by the ligand. Here we report identification of protein kinase D2 (PKD2) as a kinase that mediates the ligand-inducible phosphorylation of IFNAR1 degron and enables binding of βTrcp to the receptor. Treatment of cells with IFN-α induces catalytic activity of PKD2 and stimulates its interaction with IFNAR1. Expression and kinase activity of PKD2 are required for the ligand-inducible stimulation of IFNAR1 ubiquitination and endocytosis and for accelerated proteolytic turnover of IFNAR1. Furthermore, inhibition or knockdown of PKD2 robustly augments intracellular signaling induced by IFN-α and increases the efficacy of its antiviral effects. The mechanisms of the ligand-inducible elimination of IFNAR1 are discussed, along with the potential medical significance of this regulation. PMID:21173164

  3. Effects of different ligands on epidermal growth factor receptor (EGFR) nuclear translocation.

    Science.gov (United States)

    Faria, Jerusa A Q A; de Andrade, Carolina; Goes, Alfredo M; Rodrigues, Michele A; Gomes, Dawidson A

    2016-09-01

    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. PMID:27462018

  4. Macromolecular Modelling and Docking Simulations for the Discovery of Selective GPER Ligands.

    Science.gov (United States)

    Rosano, Camillo; Ponassi, Marco; Santolla, Maria Francesca; Pisano, Assunta; Felli, Lamberto; Vivacqua, Adele; Maggiolini, Marcello; Lappano, Rosamaria

    2016-01-01

    Estrogens influence multiple physiological processes and are implicated in many diseases as well. Cellular responses to estrogens are mainly mediated by the estrogen receptors (ER)α and ERβ, which act as ligand-activated transcription factors. Recently, a member of the G protein-coupled receptor (GPCR) superfamily, namely GPER/GPR30, has been identified as a further mediator of estrogen signalling in different pathophysiological conditions, including cancer. Today, computational methods are commonly used in all areas of health science research. Among these methods, virtual ligand screening has become an established technique for hit discovery and optimization. The absence of an established three-dimensional structure of GPER promoted studies of structure-based drug design in order to build reliable molecular models of this receptor. Here, we discuss the results obtained through the structure-based virtual ligand screening for GPER, which allowed the identification and synthesis of different selective agonist and antagonist moieties. These compounds led significant advances in our understanding of the GPER function at the cellular, tissue, and organismal levels. In particular, selective GPER ligands were critical toward the evaluation of the role elicited by this receptor in several pathophysiological conditions, including cancer. Considering that structure-based approaches are fundamental in drug discovery, future research breakthroughs with the aid of computer-aided molecular design and chemo-bioinformatics could generate a new class of drugs that, acting through GPER, would be useful in a variety of diseases as well as in innovative anticancer strategies.

  5. Leaching behavior of butanedionedioxime as gold ligand

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Butanedionedioxime, a small organic compound with low-toxicity and good chemical stability, has been proposed as an effective gold ligand in gold extraction. The result of experiment shows that: 1) highly effective gold lixiviantcan be composed of butanedionedioxime (BDM) with many oxidants, especially potassium permanganate; 2)in the leaching system of BD M- K M nO4 the suitable Ox/Lig(ratio of oxidants to gold ligands) tange is 0.20 ~ 0. 50, optimally 0.25 ~0.45 at the pH range of 7 ~ 11; 3) BDM-KMnO4 extraction of gold from an oxide ore is similar to cyanide(cyanide-O2)extraction, but the leaching rate of gold by BDM-KMnO4 is faster than that by cyanide-O2; 4) gold may readily be recov-ered by carbon adsorption and zinc precipitation

  6. Glycomimetic ligands for the human asialoglycoprotein receptor.

    Science.gov (United States)

    Mamidyala, Sreeman K; Dutta, Sanjay; Chrunyk, Boris A; Préville, Cathy; Wang, Hong; Withka, Jane M; McColl, Alexander; Subashi, Timothy A; Hawrylik, Steven J; Griffor, Matthew C; Kim, Sung; Pfefferkorn, Jeffrey A; Price, David A; Menhaji-Klotz, Elnaz; Mascitti, Vincent; Finn, M G

    2012-02-01

    The asialoglycoprotein receptor (ASGPR) is a high-capacity galactose-binding receptor expressed on hepatocytes that binds its native substrates with low affinity. More potent ligands are of interest for hepatic delivery of therapeutic agents. We report several classes of galactosyl analogues with varied substitution at the anomeric, C2-, C5-, and C6-positions. Significant increases in binding affinity were noted for several trifluoromethylacetamide derivatives without covalent attachment to the protein. A variety of new ligands were obtained with affinity for ASGPR as good as or better than that of the parent N-acetylgalactosamine, showing that modification on either side of the key C3,C4-diol moiety is well tolerated, consistent with previous models of a shallow binding pocket. The galactosyl pyranose motif therefore offers many opportunities for the attachment of other functional units or payloads while retaining low-micromolar or better affinity for the ASGPR.

  7. Design principles for ligand-sensing, conformation-switching ribozymes.

    Directory of Open Access Journals (Sweden)

    Xi Chen

    2009-12-01

    Full Text Available Nucleic acid sensor elements are proving increasingly useful in biotechnology and biomedical applications. A number of ligand-sensing, conformational-switching ribozymes (also known as allosteric ribozymes or aptazymes have been generated by some combination of directed evolution or rational design. Such sensor elements typically fuse a molecular recognition domain (aptamer with a catalytic signal generator (ribozyme. Although the rational design of aptazymes has begun to be explored, the relationships between the thermodynamics of aptazyme conformational changes and aptazyme performance in vitro and in vivo have not been examined in a quantitative framework. We have therefore developed a quantitative and predictive model for aptazymes as biosensors in vitro and as riboswitches in vivo. In the process, we have identified key relationships (or dimensionless parameters that dictate aptazyme performance, and in consequence, established equations for precisely engineering aptazyme function. In particular, our analysis quantifies the intrinsic trade-off between ligand sensitivity and the dynamic range of activity. We were also able to determine how in vivo parameters, such as mRNA degradation rates, impact the design and function of aptazymes when used as riboswitches. Using this theoretical framework we were able to achieve quantitative agreement between our models and published data. In consequence, we are able to suggest experimental guidelines for quantitatively predicting the performance of aptazyme-based riboswitches. By identifying factors that limit the performance of previously published systems we were able to generate immediately testable hypotheses for their improvement. The robust theoretical framework and identified optimization parameters should now enable the precision design of aptazymes for biotechnological and clinical applications.

  8. Selective oxoanion separation using a tripodal ligand

    Energy Technology Data Exchange (ETDEWEB)

    Custelcean, Radu; Moyer, Bruce A.; Rajbanshi, Arbin

    2016-02-16

    The present invention relates to urea-functionalized crystalline capsules self-assembled by sodium or potassium cation coordination and by hydrogen-bonding water bridges to selectively encapsulate tetrahedral divalent oxoanions from highly competitive aqueous alkaline solutions and methods using this system for selective anion separations from industrial solutions. The method involves competitive crystallizations using a tripodal tris(urea) functionalized ligand and, in particular, provides a viable approach to sulfate separation from nuclear wastes.

  9. Targeting Selectins and Their Ligands in Cancer.

    Science.gov (United States)

    Natoni, Alessandro; Macauley, Matthew S; O'Dwyer, Michael E

    2016-01-01

    Aberrant glycosylation is a hallmark of cancer cells with increased evidence pointing to a role in tumor progression. In particular, aberrant sialylation of glycoproteins and glycolipids has been linked to increased immune cell evasion, drug evasion, drug resistance, tumor invasiveness, and vascular dissemination, leading to metastases. Hypersialylation of cancer cells is largely the result of overexpression of sialyltransferases (STs). Differentially, humans express twenty different STs in a tissue-specific manner, each of which catalyzes the attachment of sialic acids via different glycosidic linkages (α2-3, α2-6, or α2-8) to the underlying glycan chain. One important mechanism whereby overexpression of STs contributes to an enhanced metastatic phenotype is via the generation of selectin ligands. Selectin ligand function requires the expression of sialyl-Lewis X and its structural isomer sialyl-Lewis A, which are synthesized by the combined action of alpha α1-3-fucosyltransferases, α2-3-sialyltransferases, β1-4-galactosyltranferases, and N-acetyl-β-glucosaminyltransferases. The α2-3-sialyltransferases ST3Gal4 and ST3Gal6 are critical to the generation of functional E- and P-selectin ligands and overexpression of these STs have been linked to increased risk of metastatic disease in solid tumors and poor outcome in multiple myeloma. Thus, targeting selectins and their ligands as well as the enzymes involved in their generation, in particular STs, could be beneficial to many cancer patients. Potential strategies include ST inhibition and the use of selectin antagonists, such as glycomimetic drugs and antibodies. Here, we review ongoing efforts to optimize the potency and selectivity of ST inhibitors, including the potential for targeted delivery approaches, as well as evaluate the potential utility of selectin inhibitors, which are now in early clinical development. PMID:27148485

  10. Targeting Selectins and Their Ligands in Cancer

    Directory of Open Access Journals (Sweden)

    Alessandro eNatoni

    2016-04-01

    Full Text Available Aberrant glycosylation is a hallmark of cancer cells with increased evidence pointing to a role in tumor progression. In particular, aberrant sialylation of glycoproteins and glycolipids have been linked to increased immune cell evasion, drug evasion, drug resistance, tumor invasiveness, and vascular dissemination leading to metastases. Hypersialylation of cancer cells is largely the result of overexpression of sialyltransferases. Humans differentially express twenty different sialyltransferases in a tissue-specific manner, each of which catalyze the attachment of sialic acids via different glycosidic linkages (2-3; 2-6 or 2-8 to the underlying glycan chain. One important mechanism whereby overexpression of sialyltransferases contributes to an enhanced metastatic phenotype is via the generation of selectin ligands. Selectin ligand function requires the expression of sialyl-Lewis X and its structural-isomer sialyl-Lewis A, which are synthesized by the combined action of alpha 1-3-fucosyltransferases, 2-3-sialyltransferases, 1-4-galactosyltranferases, and N-acetyl--glucosaminyltransferases. The α2-3-sialyltransferases ST3Gal4 and ST3Gal6 are critical to the generation of functional E- and P-selectin ligands and overexpression of these sialyltransferases have been linked to increased risk of metastatic disease in solid tumors and poor outcome in multiple myeloma. Thus, targeting selectins and their ligands as well as the enzymes involved in their generation, in particular sialyltransferases, could be beneficial to many cancer patients. Potential strategies include sialyltransferase inhibition and the use of selectin antagonists, such as glycomimetic drugs and antibodies. Here, we review ongoing efforts to optimize the potency and selectivity of sialyltransferase inhibitors, including the potential for targeted delivery approaches, as well as evaluate the potential utility of selectin inhibitors, which are now in early clinical

  11. Syntheses of oxysterol receptors'(LXRs) ligands

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The LXRs' agonist, 24S,25-epoxycholesterol 1, was synthesized stereoselectively (100% d.e.) in 56% overall yield from methyl hyodeoxycholanate 4 in 9 steps with desmosterol acetate 11 as the key intermediate and the modified Sharpless asymmetric dihydroxylation as the key step. The LXR? subtype selective agonist 5α,6α:24S,25-diepoxycho- lesterol 2 and the novel LXRs' ligand 5β,6β:24S,25-diepo- xycholesterol 3 were also synthesized from 1.

  12. EXPRESSION OF Fas LIGAND IN HUMAN COLON CANCER CELL LINES

    Institute of Scientific and Technical Information of China (English)

    张建军; 丁尔迅; 王强; 陈学云; 付志仁

    2001-01-01

    To investigate the expression of Fas ligand in human colon carcinoma cell lines. Methods: A total of six human colon cancer cell lines were examined for the expression of Fas ligand mRNA and cell surface protein by using RT-PCR and flow cytometry respectively. Results: The results showed that Fas ligand mRNA was expressed in all of the six cancer cell lines and Fas ligand cell surface protein was expressed in part of them. Conclusion: These data suggest that Fas ligand was expressed, at least in part, in human colon cancer cell lines and might facilitate to escape from immune surveillance of the host.

  13. Regulation of Hedgehog Signalling Inside and Outside the Cell

    Science.gov (United States)

    Ramsbottom, Simon A.; Pownall, Mary E.

    2016-01-01

    The hedgehog (Hh) signalling pathway is conserved throughout metazoans and plays an important regulatory role in both embryonic development and adult homeostasis. Many levels of regulation exist that control the release, reception, and interpretation of the hedgehog signal. The fatty nature of the Shh ligand means that it tends to associate tightly with the cell membrane, and yet it is known to act as a morphogen that diffuses to elicit pattern formation. Heparan sulfate proteoglycans (HSPGs) play a major role in the regulation of Hh distribution outside the cell. Inside the cell, the primary cilium provides an important hub for processing the Hh signal in vertebrates. This review will summarise the current understanding of how the Hh pathway is regulated from ligand production, release, and diffusion, through to signal reception and intracellular transduction.

  14. The Recognition of Identical Ligands by Unrelated Proteins.

    Science.gov (United States)

    Barelier, Sarah; Sterling, Teague; O'Meara, Matthew J; Shoichet, Brian K

    2015-12-18

    The binding of drugs and reagents to off-targets is well-known. Whereas many off-targets are related to the primary target by sequence and fold, many ligands bind to unrelated pairs of proteins, and these are harder to anticipate. If the binding site in the off-target can be related to that of the primary target, this challenge resolves into aligning the two pockets. However, other cases are possible: the ligand might interact with entirely different residues and environments in the off-target, or wholly different ligand atoms may be implicated in the two complexes. To investigate these scenarios at atomic resolution, the structures of 59 ligands in 116 complexes (62 pairs in total), where the protein pairs were unrelated by fold but bound an identical ligand, were examined. In almost half of the pairs, the ligand interacted with unrelated residues in the two proteins (29 pairs), and in 14 of the pairs wholly different ligand moieties were implicated in each complex. Even in those 19 pairs of complexes that presented similar environments to the ligand, ligand superposition rarely resulted in the overlap of related residues. There appears to be no single pattern-matching "code" for identifying binding sites in unrelated proteins that bind identical ligands, though modeling suggests that there might be a limited number of different patterns that suffice to recognize different ligand functional groups.

  15. NFkappaB Selectivity of Estrogen Receptor Ligands Revealed By Comparative Crystallographic Analyses

    Energy Technology Data Exchange (ETDEWEB)

    Nettles, K.W.; Bruning, J.B.; Gil, G.; Nowak, J.; Sharma, S.K.; Hahm, J.B.; Kulp, K.; Hochberg, R.B.; Zhou, H.; Katzenellenbogen, J.A.; Katzenllenbogen, B.S.; Kim, Y.; Joachmiak, A.; Greene, G.L.

    2009-05-22

    Our understanding of how steroid hormones regulate physiological functions has been significantly advanced by structural biology approaches. However, progress has been hampered by misfolding of the ligand binding domains in heterologous expression systems and by conformational flexibility that interferes with crystallization. Here, we show that protein folding problems that are common to steroid hormone receptors are circumvented by mutations that stabilize well-characterized conformations of the receptor. We use this approach to present the structure of an apo steroid receptor that reveals a ligand-accessible channel allowing soaking of preformed crystals. Furthermore, crystallization of different pharmacological classes of compounds allowed us to define the structural basis of NF{kappa}B-selective signaling through the estrogen receptor, thus revealing a unique conformation of the receptor that allows selective suppression of inflammatory gene expression. The ability to crystallize many receptor-ligand complexes with distinct pharmacophores allows one to define structural features of signaling specificity that would not be apparent in a single structure.

  16. Ligand induced circular dichroism and circularly polarized luminescence in CdSe quantum dots.

    Science.gov (United States)

    Tohgha, Urice; Deol, Kirandeep K; Porter, Ashlin G; Bartko, Samuel G; Choi, Jung Kyu; Leonard, Brian M; Varga, Krisztina; Kubelka, Jan; Muller, Gilles; Balaz, Milan

    2013-12-23

    Chiral thiol capping ligands L- and D-cysteines induced modular chiroptical properties in achiral cadmium selenide quantum dots (CdSe QDs). Cys-CdSe prepared from achiral oleic acid capped CdSe by postsynthetic ligand exchange displayed size-dependent electronic circular dichroism (CD) and circularly polarized luminescence (CPL). Opposite CPL signals were measured for the CdSe QDs capped with D- and L-cysteine. The CD profile and CD anisotropy varied with size of CdSe nanocrystals with largest anisotropy observed for CdSe nanoparticles of 4.4 nm. Magic angle spinning solid state NMR (MAS ssNMR) experiments suggested bidentate interaction between cysteine and the surface of CdSe. Time Dependent Density Functional Theory (TDDFT) calculations verified that attachment of L- and D-cysteine to the surface of model (CdSe)13 nanoclusters induces measurable opposite CD signals for the exitonic band of the nanocluster. The origin of the induced chirality is consistent with the hybridization of highest occupied CdSe molecular orbitals with those of the chiral ligand.

  17. Dispatched and Scube Mediate the Efficient Secretion of the Cholesterol-Modified Hedgehog Ligand

    Directory of Open Access Journals (Sweden)

    Hanna Tukachinsky

    2012-08-01

    Full Text Available The Hedgehog (Hh signaling pathway plays critical roles in metazoan development and in cancer. How the Hh ligand is secreted and spreads to distant cells is unclear, given its covalent modification with a hydrophobic cholesterol molecule, which makes it stick to membranes. We demonstrate that Hh ligand secretion from vertebrate cells is accomplished via two distinct and synergistic cholesterol-dependent binding events, mediated by two proteins that are essential for vertebrate Hh signaling: the membrane protein Dispatched (Disp and a member of the Scube family of secreted proteins. Cholesterol modification is sufficient for a heterologous protein to interact with Scube and to be secreted in a Scube-dependent manner. Disp and Scube recognize different structural aspects of cholesterol similarly to how Niemann-Pick disease proteins 1 and 2 interact with cholesterol, suggesting a hand-off mechanism for transferring Hh from Disp to Scube. Thus, Disp and Scube cooperate to dramatically enhance the secretion and solubility of the cholesterol-modified Hh ligand.

  18. Membrane-displayed peptide ligand activates the pheromone response pathway in Saccharomyces cerevisiae.

    Science.gov (United States)

    Hara, Keisuke; Ono, Takuya; Kuroda, Kouichi; Ueda, Mitsuyoshi

    2012-05-01

    The budding yeast, Saccharomyces cerevisiae, is an attractive host for studying G protein-coupled receptors (GPCRs). We developed a system in which a peptide ligand specific for GPCR is displayed on yeast plasma membrane. The model system described here is based on yeast plasma membrane display of an analogue of α-factor, which is a peptide ligand for Ste2p, the GPCR that activates the yeast pheromone response pathway. α-Factor analogues, containing linkers of varying lengths and produced in yeast cells, became attached to the cell plasma membrane by linking to the glycosylphosphatidylinositol (GPI)-anchored plasma membrane protein Yps1p. We were able to demonstrate that an optimized α-factor analogue activated the pheromone response pathway in S. cerevisiae, as assessed by a fluorescent reporter assay. Furthermore, it was shown that linker length strongly influenced signalling pathway activation. To our knowledge, this is the first report documenting functional signalling by a plasma membrane-displayed ligand in S. cerevisiae.

  19. Ligand-independent canonical Wnt activity in canine mammary tumor cell lines associated with aberrant LEF1 expression.

    Directory of Open Access Journals (Sweden)

    Ana Gracanin

    Full Text Available Pet dogs very frequently develop spontaneous mammary tumors and have been suggested as a good model organism for breast cancer research. In order to obtain an insight into underlying signaling mechanisms during canine mammary tumorigenesis, in this study we assessed the incidence and the mechanism of canonical Wnt activation in a panel of 12 canine mammary tumor cell lines. We show that a subset of canine mammary cell lines exhibit a moderate canonical Wnt activity that is dependent on Wnt ligands, similar to what has been described in human breast cancer cell lines. In addition, three of the tested canine mammary cell lines have a high canonical Wnt activity that is not responsive to inhibitors of Wnt ligand secretion. Tumor cell lines with highly active canonical Wnt signaling often carry mutations in key members of the Wnt signaling cascade. These cell lines, however, carry no mutations in the coding regions of intracellular Wnt pathway components (APC, β-catenin, GSK3β, CK1α and Axin1 and have a functional β-catenin destruction complex. Interestingly, however, the cell lines with high canonical Wnt activity specifically overexpress LEF1 mRNA and the knock-down of LEF1 significantly inhibits TCF-reporter activity. In addition, LEF1 is overexpressed in a subset of canine mammary carcinomas, implicating LEF1 in ligand-independent activation of canonical Wnt signaling in canine mammary tumors. We conclude that canonical Wnt activation may be a frequent event in canine mammary tumors both through Wnt ligand-dependent and novel ligand-independent mechanisms.

  20. Redox-Active-Ligand-Mediated Formation of an Acyclic Trinuclear Ruthenium Complex with Bridging Nitrido Ligands.

    Science.gov (United States)

    Bagh, Bidraha; Broere, Daniël L J; Siegler, Maxime A; van der Vlugt, Jarl Ivar

    2016-07-11

    Coordination of a redox-active pyridine aminophenol ligand to Ru(II) followed by aerobic oxidation generates two diamagnetic Ru(III) species [1 a (cis) and 1 b (trans)] with ligand-centered radicals. The reaction of 1 a/1 b with excess NaN3 under inert atmosphere resulted in the formation of a rare bis(nitrido)-bridged trinuclear ruthenium complex with two nonlinear asymmetrical Ru-N-Ru fragments. The spontaneous reduction of the ligand centered radical in the parent 1 a/1 b supports the oxidation of a nitride (N(3-) ) to half an equivalent of N2 . The trinuclear omplex is reactive toward TEMPO-H, tin hydrides, thiols, and dihydrogen. PMID:27321547

  1. Interaction of calreticulin with CD40 ligand, TRAIL and Fas ligand

    DEFF Research Database (Denmark)

    Duus, K; Pagh, R T; Holmskov, U;

    2007-01-01

    found to bind calreticulin strongly. A low level or no binding was observed for adiponectin, tumour necrosis factor-alpha (TNF-alpha), CD30L, surfactant protein-A and -D and collagen VIII. The interaction with calreticulin required a conformational change in CD40L, TRAIL and FasL and showed the same...... is utilized by many other functionally diverse molecules and in this work the interaction of calreticulin with C1q and structurally similar molecules was investigated. In addition to C1q and MBL, CD40 ligand (CD40L), tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) and Fas ligand (FasL) were...... characteristics as calreticulin's interaction with C1q and MBL: a time-dependent saturable binding to immobilized protein, which was initially sensitive to salt but gradually developed into a salt-insensitive interaction. Thus, the interaction requires a structural change in the interaction partner and leads...

  2. NMR spectroscopic and X-ray crystallographic study of methylcobalt(III) compounds with saturated amine ligands

    DEFF Research Database (Denmark)

    Kofod, Pauli; Harris, Pernille

    2004-01-01

    The C-13 chemical shifts of methylcobalt(III) compounds with saturated amine ligands in cis positions to the methyl group and a monodentate ligand, L = CN-, NH3, NO2, N-3(-), H2O, or OH-, in the trans position are reported. The amine ligands used, 1,2-ethanediamine (en), 1,3-propanediamine (tn), N......,N'-bis(2-aminoethyl)-1,3-propanediamine (2,3,2-tet), N,N'-bis(3-aminopropyl)-1,2-ethanediamine (3,2,3-tet), and 1,4,8,11-tetraazacyclotetradecane (cyclam), all exert an apparent cis influence on the 13 C resonance signal of the coordinated methyl group. In the trans-[Co-(en)(2)(CH3)(L)](n+) series the N-15...

  3. Endocannabinoid Signaling in Modulating Periimplantation Events

    Institute of Scientific and Technical Information of China (English)

    Hong-ying PENG; Qi CHEN; Dong-mei TAN; Yi TAN

    2008-01-01

    Cannabinoids have long been suspected associating with abnormal fetal growth and outcome. However, the molecular mechanisms in which they are involved had long been obscure for several decades. Only recently, after the identification of two types of cannabinoid receptors (CBI-R and CB2-R) and the following discoveries of their corresponding endogenous ligands, the mystery behind those seemingly facts began gradually unveiled. Through a series of landmark research, it is now indicated that the endocannabinoid signaling via the ligand-receptor interaction plays an important role in modulating early development of preimplantation embryo and synchronizing embryo development with uterine receptivity for implantation. Current data suggest that the physiological functions of endocannabinoid signaling as well as their metabolic pathways are potentially very exited areas to be explored further. This review will first introduce the reproductive functions of endocannabinoid signaling from epidemiological and molecular background, then focus on its reciprocal interactions between the embryo and maternal tissues, as well as related metabolic aspects in regards to implantation. It is hoped that further investigation of this physiologically fundamental signaling will generate more exciting information elucidating the complexity of implantation thus lead to a better control of human reproduction.

  4. Landscape of protein-small ligand binding modes.

    Science.gov (United States)

    Kasahara, Kota; Kinoshita, Kengo

    2016-09-01

    Elucidating the mechanisms of specific small-molecule (ligand) recognition by proteins is a long-standing conundrum. While the structures of these molecules, proteins and ligands, have been extensively studied, protein-ligand interactions, or binding modes, have not been comprehensively analyzed. Although methods for assessing similarities of binding site structures have been extensively developed, the methods for the computational treatment of binding modes have not been well established. Here, we developed a computational method for encoding the information about binding modes as graphs, and assessing their similarities. An all-against-all comparison of 20,040 protein-ligand complexes provided the landscape of the protein-ligand binding modes and its relationships with protein- and chemical spaces. While similar proteins in the same SCOP Family tend to bind relatively similar ligands with similar binding modes, the correlation between ligand and binding similarities was not very high (R(2)  = 0.443). We found many pairs with novel relationships, in which two evolutionally distant proteins recognize dissimilar ligands by similar binding modes (757,474 pairs out of 200,790,780 pairs were categorized into this relationship, in our dataset). In addition, there were an abundance of pairs of homologous proteins binding to similar ligands with different binding modes (68,217 pairs). Our results showed that many interesting relationships between protein-ligand complexes are still hidden in the structure database, and our new method for assessing binding mode similarities is effective to find them.

  5. Landscape of protein-small ligand binding modes.

    Science.gov (United States)

    Kasahara, Kota; Kinoshita, Kengo

    2016-09-01

    Elucidating the mechanisms of specific small-molecule (ligand) recognition by proteins is a long-standing conundrum. While the structures of these molecules, proteins and ligands, have been extensively studied, protein-ligand interactions, or binding modes, have not been comprehensively analyzed. Although methods for assessing similarities of binding site structures have been extensively developed, the methods for the computational treatment of binding modes have not been well established. Here, we developed a computational method for encoding the information about binding modes as graphs, and assessing their similarities. An all-against-all comparison of 20,040 protein-ligand complexes provided the landscape of the protein-ligand binding modes and its relationships with protein- and chemical spaces. While similar proteins in the same SCOP Family tend to bind relatively similar ligands with similar binding modes, the correlation between ligand and binding similarities was not very high (R(2)  = 0.443). We found many pairs with novel relationships, in which two evolutionally distant proteins recognize dissimilar ligands by similar binding modes (757,474 pairs out of 200,790,780 pairs were categorized into this relationship, in our dataset). In addition, there were an abundance of pairs of homologous proteins binding to similar ligands with different binding modes (68,217 pairs). Our results showed that many interesting relationships between protein-ligand complexes are still hidden in the structure database, and our new method for assessing binding mode similarities is effective to find them. PMID:27327045

  6. Cross-talk and information transfer in mammalian and bacterial signaling.

    Directory of Open Access Journals (Sweden)

    Samanthe M Lyons

    Full Text Available In mammalian and bacterial cells simple phosphorylation circuits play an important role in signaling. Bacteria have hundreds of two-component signaling systems that involve phosphotransfer between a receptor and a response regulator. In mammalian cells a similar pathway is the TGF-beta pathway, where extracellular TGF-beta ligands activate cell surface receptors that phosphorylate Smad proteins, which in turn activate many genes. In TGF-beta signaling the multiplicity of ligands begs the question as to whether cells can distinguish signals coming from different ligands, but transduced through a small set of Smads. Here we use information theory with stochastic simulations of networks to address this question. We find that when signals are transduced through only one Smad, the cell cannot distinguish between different levels of the external ligands. Increasing the number of Smads from one to two significantly improves information transmission as well as the ability to discriminate between ligands. Surprisingly, both total information transmitted and the capacity to discriminate between ligands are quite insensitive to high levels of cross-talk between the two Smads. Robustness against cross-talk requires that the average amplitude of the signals are large. We find that smaller systems, as exemplified by some two-component systems in bacteria, are significantly much less robust against cross-talk. For such system sizes phosphotransfer is also less robust against cross-talk than phosphorylation. This suggests that mammalian signal transduction can tolerate a high amount of cross-talk without degrading information content. This may have played a role in the evolution of new functionalities from small mutations in signaling pathways, allowed for the development of cross-regulation and led to increased overall robustness due to redundancy in signaling pathways. On the other hand the lack of cross-regulation observed in many bacterial two

  7. Polymer complexes. XLXI. Supramolecular spectral studies on metal-ligand bonding of novel rhodanine sulphadrugs hydrazone

    International Nuclear Information System (INIS)

    Graphical abstract: The ESR spectra of copper complexes in powder form show a broad signal with values in order g|| > g( > ge (2.0023). The value of covalency factor β and orbital reduction factor K accounts for the covalent nature of the complexes. -- Abstract: Novel polymeric complexes with 5-sulphadiazineazo-3-phenyl-2-thioxo-4-thiazolidine (HL1), 5-sulphamethazineazo-3-phenyl-2-thioxo-4-thiazolidine (HL2) and 5-sulphamethoxazoleazo-3-phenyl-2-thioxo-4-thiazolidine (HL3) and various anions were prepared. Structural and spectroscopic properties have been studied on the basis of elemental analyses, infrared spectra, 1HNMR spectra, electronic spectra, magnetic measurements and ESR. IR and 1H NMR studies reveal that the ligands (HLn) exists in the tautomeric enol/hydrazo form in both states with intramolecular hydrogen bonding. The analytical data and the molar conductance measurements of the polymeric complexes reveal that three molecules of the ligand and four/two(Cl/SO4) of the anions are coordinated to the two metal atoms in all complexes. The infrared spectra of the ligands and their polymeric complexes, confirmed that the ligands coordinate to Cu(II) as a neutral and tetradentate via NH(hydrazone), oxygen of the carbonyl group (CO), nitrogen of the NH(3-phenylamine) and thion sulphur (CS) group. All the polymer complexes were found to have magnetic moments corresponding to one unpaired electrons. The ESR (g|| and gperpendicular) and bonding α2 parameters of the copper ion were greatly affected by substituting several groups position of ring of sulphadrug. The ESR spectra of copper complexes in powder form show a broad signal with values in order g|| > gperpendicular > ge (2.0023). The value of covalency factor β and orbital reduction factor K accounts for the covalent nature of the complexes.

  8. Decreased Autocrine EGFR Signaling in Metastatic Breast Cancer Cells Inhibits Tumor Growth in Bone and Mammary Fat Pad

    OpenAIRE

    Nickerson, Nicole K.; Mohammad, Khalid S.; Gilmore, Jennifer L.; Crismore, Erin; Bruzzaniti, Angela; Guise, Theresa A.; Foley, John

    2012-01-01

    Breast cancer metastasis to bone triggers a vicious cycle of tumor growth linked to osteolysis. Breast cancer cells and osteoblasts express the epidermal growth factor receptor (EGFR) and produce ErbB family ligands, suggesting participation of these growth factors in autocrine and paracrine signaling within the bone microenvironment. EGFR ligand expression was profiled in the bone metastatic MDA-MB-231 cells (MDA-231), and agonist-induced signaling was examined in both breast cancer and oste...

  9. Analysis of Killer Cell Immunoglobulin-like Receptor Genes and Their HLA Ligands in Iranian Patients with Ankylosing Spondylitis.

    Science.gov (United States)

    Mahmoudi, Mehdi; Jamshidi, Ahmad Reza; Karami, Jafar; Mohseni, Alireza; Amirzargar, Ali Akbar; Farhadi, Elham; Ahmadzadeh, Nooshin; Nicknam, Mohammad Hossein

    2016-02-01

    Ankylosing Spondylitis (AS) is a chronic rheumatic disease which mainly involves the axial skeleton. It seems that non-HLA genes, as well as HLA-B27 gene, are linked to the etiology of the disease. Recently, it has been documented that KIRs and their HLA ligands are contributed to the Ankylosing Spondylitis. The aim of this study was to evaluate the KIR genes and their HLA ligands in Iranian AS patients and healthy individuals. The present study includes 200 AS patient samples and 200 healthy control samples. KIR genotyping was performed using the polymerase chain reaction sequence-specific primer (PCR-SSP) method to type the presence or absence of the 16 KIR genes, 6 known specific HLA class I ligands and also, two pseudogenes. Two KIR genes (KIR-2DL3 and KIR2DL5), and among the HLA ligands, two HLA ligands (HLA-C2Lys80 and HLA-B27) genes were significantly different between case and control groups. In addition, we found some interesting KIR/HLA compound genotypes, which were associated with AS susceptibility. Our results suggest that the AS patients present more activating and less inhibitory KIR genes with combination of their HLA ligands than healthy controls. Once the balance of signal transduction between activating and inhibitory receptors is disturbed, the ability of NK cells to identify and lyse the targets in immune responses will be compromised. Accordingly, imbalance of activating and inhibitory KIR genes by up-regulating the activation and losing the inhibition of KIRs signaling or combination of both might be one of the important factors which underlying the pathogenesis of AS. PMID:26996109

  10. Molecular mechanisms of glucocorticoid receptor signaling

    Directory of Open Access Journals (Sweden)

    Marta Labeur

    2010-10-01

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

  11. Transmutable nanoparticles with reconfigurable surface ligands

    Science.gov (United States)

    Kim, Youngeun; Macfarlane, Robert J.; Jones, Matthew R.; Mirkin, Chad A.

    2016-02-01

    Unlike conventional inorganic materials, biological systems are exquisitely adapted to respond to their surroundings. Proteins and other biological molecules can process a complex set of chemical binding events as informational inputs and respond accordingly via a change in structure and function. We applied this principle to the design and synthesis of inorganic materials by preparing nanoparticles with reconfigurable surface ligands, where interparticle bonding can be programmed in response to specific chemical cues in a dynamic manner. As a result, a nascent set of “transmutable nanoparticles” can be driven to crystallize along multiple thermodynamic trajectories, resulting in rational control over the phase and time evolution of nanoparticle-based matter.

  12. TGF-β-induced profibrotic signaling is regulated in part by the WNT receptor Frizzled-8

    NARCIS (Netherlands)

    Spanjer, Anita I R; Baarsma, Hoeke A; Oostenbrink, Lisette M; Jansen, Sepp R; Kuipers, Christine C; Lindner, Michael; Postma, Dirkje S; Meurs, Herman; Heijink, Irene H; Gosens, Reinoud; Königshoff, Melanie

    2016-01-01

    TGF-β is important in lung injury and remodeling processes. TGF-β and Wingless/integrase-1 (WNT) signaling are interconnected; however, the WNT ligand-receptor complexes involved are unknown. Thus, we aimed to identify Frizzled (FZD) receptors that mediate TGF-β-induced profibrotic signaling. MRC-5

  13. Computational understanding and experimental characterization of twice-as-smart quadruplex ligands as chemical sensors of bacterial nucleotide second messengers

    Science.gov (United States)

    Zhou, Jie; Roembke, Benjamin T.; Paragi, Gabor; Laguerre, Aurélien; Sintim, Herman O.; Fonseca Guerra, Célia; Monchaud, David

    2016-01-01

    A twice-as-smart ligand is a small molecule that experiences a structural switch upon interaction with its target (i.e., smart ligand) that concomitantly triggers its fluorescence (i.e., smart probe). Prototypes of twice-as-smart ligands were recently developed to track and label G-quadruplexes: these higher-order nucleic acid structures originate in the assembly of four guanine(G)-rich DNA or RNA strands, whose stability is imparted by the formation and the self-assembly of G-quartets. The first prototypes of twice-as-smart quadruplex ligands were designed to exploit the self-association of quartets, being themselves synthetic G-quartets. While their quadruplex recognition capability has been thoroughly documented, some doubts remain about the precise photophysical mechanism that underlies their peculiar spectroscopic properties. Here, we uncovered this mechanism via complete theoretical calculations. Collected information was then used to develop a novel application of twice-as-smart ligands, as efficient chemical sensors of bacterial signaling pathways via the fluorescent detection of naturally occurring extracellular quadruplexes formed by cyclic dimeric guanosine monophosphate (c-di-GMP). PMID:27667717

  14. Chirality Inversion of CdSe and CdS Quantum Dots without Changing the Stereochemistry of the Capping Ligand.

    Science.gov (United States)

    Choi, Jung Kyu; Haynie, Benjamin E; Tohgha, Urice; Pap, Levente; Elliott, K Wade; Leonard, Brian M; Dzyuba, Sergei V; Varga, Krisztina; Kubelka, Jan; Balaz, Milan

    2016-03-22

    L-cysteine derivatives induce and modulate the optical activity of achiral cadmium selenide (CdSe) and cadmium sulfide (CdS) quantum dots (QDs). Remarkably, N-acetyl-L-cysteine-CdSe and L-homocysteine-CdSe as well as N-acetyl-L-cysteine-CdS and L-cysteine-CdS showed "mirror-image" circular dichroism (CD) spectra regardless of the diameter of the QDs. This is an example of the inversion of the CD signal of QDs by alteration of the ligand's structure, rather than inversion of the ligand's absolute configuration. Non-empirical quantum chemical simulations of the CD spectra were able to reproduce the experimentally observed sign patterns and demonstrate that the inversion of chirality originated from different binding arrangements of N-acetyl-L-cysteine and L-homocysteine-CdSe to the QD surface. These efforts may allow the prediction of the ligand-induced chiroptical activity of QDs by calculating the specific binding modes of the chiral capping ligands. Combined with the large pool of available chiral ligands, our work opens a robust approach to the rational design of chiral semiconducting nanomaterials.

  15. Detection of site-specific binding and co-binding of ligands to macromolecules using sup 19 F NMR

    Energy Technology Data Exchange (ETDEWEB)

    Jenkins, B.G. (Massachusetts General Hospital and Harvard Medical School, Charlestown (USA))

    1991-01-01

    Study of ligand-macromolecular interactions by {sup 19}F nuclear magnetic resonance (NMR) spectroscopy affords many opportunities for obtaining molecular biochemical and pharmaceutical information. This is due to the absence of a background fluorine signal, as well as the relatively high sensitivity of {sup 19}F NMR. Use of fluorine-labeled ligands enables one to probe not only binding and co-binding phenomena to macromolecules, but also can provide data on binding constants, stoichiometries, kinetics, and conformational properties of these complexes. Under conditions of slow exchange and macromolecule-induced chemical shifts, multiple {sup 19}F NMR resonances can be observed for free and bound ligands. These shifted resonances are a direct correlate of the concentration of ligand bound in a specific state rather than the global concentrations of bound or free ligand which are usually determined using other techniques such as absorption spectroscopy or equilibrium dialysis. Examples of these interactions are demonstrated both from the literature and from interactions of 5-fluorotryptophan, 5-fluorosalicylic acid, flurbiprofen, and sulindac sulfide with human serum albumin. Other applications of {sup 19}F NMR to study of these interactions in vivo, as well for receptor binding and metabolic tracing of fluorinated drugs and proteins are discussed.

  16. Distribution and dynamics of rat basophilic leukemia immunoglobulin E receptors (FcepsilonRI) on planar ligand-presenting surfaces.

    Science.gov (United States)

    Spendier, Kathrin; Carroll-Portillo, Amanda; Lidke, Keith A; Wilson, Bridget S; Timlin, Jerilyn A; Thomas, James L

    2010-07-21

    There is considerable interest in the signaling mechanisms of immunoreceptors, especially when triggered with membrane-bound ligands. We have quantified the spatiotemporal dynamics of the redistribution of immunoglobulin E-loaded receptors (IgE-FcepsilonRI) on rat basophilic leukemia-2H3 mast cells in contact with fluid and gel-phase membranes displaying ligands for immunoglobulin E, using total internal reflection fluorescence microscopy. To clearly separate the kinetics of receptor redistribution from cell spreading, and to precisely define the initial contact time (+/-50 ms), micropipette cell manipulation was used to bring individual cells into contact with surfaces. On ligand-free surfaces, there are micron-scale heterogeneities in fluorescence that likely reflect regions of the cell that are more closely apposed to the substrate. When ligands are present, receptor clusters form with this same size scale. The initial rate of accumulation of receptors into the clusters is consistent with diffusion-limited trapping with D approximately 10(-1) microm2/s. These results support the hypothesis that clusters form by diffusion to cell-surface contact regions. Over longer timescales (>10 s), individual clusters moved with both diffusive and directed motion components. The dynamics of the cluster motion is similar to the dynamics of membrane fluctuations of cells on ligand-free fluid membranes. Thus, the same cellular machinery may be responsible for both processes.

  17. The hyperanalytic signal

    CERN Document Server

    Bihan, Nicolas Le

    2010-01-01

    The concept of the analytic signal is extended from the case of a real signal with a complex analytic signal to a complex signal with a hypercomplex analytic signal (which we call a hyperanalytic signal) The hyperanalytic signal may be interpreted as an ordered pair of complex signals or as a quaternion signal. The hyperanalytic signal contains a complex orthogonal signal and we show how to obtain this by three methods: a pair of classical Hilbert transforms; a complex Fourier transform; and a quaternion Fourier transform. It is shown how to derive from the hyperanalytic signal a complex envelope and phase using a polar quaternion representation previously introduced by the authors. The complex modulation of a real sinusoidal carrier is shown to generalize the modulation properties of the classical analytic signal. The paper extends the ideas of properness to deterministic complex signals using the hyperanalytic signal. A signal example is presented, with its orthogonal signal, and its complex envelope and ph...

  18. Increased accuracy of ligand sensing by receptor internalization

    CERN Document Server

    Aquino, Gerardo

    2010-01-01

    Many types of cells can sense external ligand concentrations with cell-surface receptors at extremely high accuracy. Interestingly, ligand-bound receptors are often internalized, a process also known as receptor-mediated endocytosis. While internalization is involved in a vast number of important functions for the life of a cell, it was recently also suggested to increase the accuracy of sensing ligand as the overcounting of the same ligand molecules is reduced. Here we show, by extending simple ligand-receptor models to out-of-equilibrium thermodynamics, that internalization increases the accuracy with which cells can measure ligand concentrations in the external environment. Comparison with experimental rates of real receptors demonstrates that our model has indeed biological significance.

  19. Architectural repertoire of ligand-binding pockets on protein surfaces.

    Science.gov (United States)

    Weisel, Martin; Kriegl, Jan M; Schneider, Gisbert

    2010-03-01

    Knowledge of the three-dimensional structure of ligand binding sites in proteins provides valuable information for computer-assisted drug design. We present a method for the automated extraction and classification of ligand binding site topologies, in which protein surface cavities are represented as branched frameworks. The procedure employs a growing neural gas approach for pocket topology assignment and pocket framework generation. We assessed the structural diversity of 623 known ligand binding site topologies based on framework cluster analysis. At a resolution of 5 A only 23 structurally distinct topology groups were formed; this suggests an overall limited structural diversity of ligand-accommodating protein cavities. Higher resolution allowed for identification of protein-family specific pocket features. Pocket frameworks highlight potentially preferred modes of ligand-receptor interactions and will help facilitate the identification of druggable subpockets suitable for ligand affinity and selectivity optimization. PMID:20069621

  20. Predicting Nanocrystal Shape through Consideration of Surface-Ligand Interactions

    KAUST Repository

    Bealing, Clive R.

    2012-03-27

    Density functional calculations for the binding energy of oleic acid-based ligands on Pb-rich {100} and {111} facets of PbSe nanocrystals determine the surface energies as a function of ligand coverage. Oleic acid is expected to bind to the nanocrystal surface in the form of lead oleate. The Wulff construction predicts the thermodynamic equilibrium shape of the PbSe nanocrystals. The equilibrium shape is a function of the ligand surface coverage, which can be controlled by changing the concentration of oleic acid during synthesis. The different binding energy of the ligand on the {100} and {111} facets results in different equilibrium ligand coverages on the facets, and a transition in the equilibrium shape from octahedral to cubic is predicted when increasing the ligand concentration during synthesis. © 2012 American Chemical Society.

  1. SuperLigands – a database of ligand structures derived from the Protein Data Bank

    Directory of Open Access Journals (Sweden)

    Preissner Robert

    2005-05-01

    Full Text Available Abstract Background Currently, the PDB contains approximately 29,000 protein structures comprising over 70,000 experimentally determined three-dimensional structures of over 5,000 different low molecular weight compounds. Information about these PDB ligands can be very helpful in the field of molecular modelling and prediction, particularly for the prediction of protein binding sites and function. Description Here we present an Internet accessible database delivering PDB ligands in the MDL Mol file format which, in contrast to the PDB format, includes information about bond types. Structural similarity of the compounds can be detected by calculation of Tanimoto coefficients and by three-dimensional superposition. Topological similarity of PDB ligands to known drugs can be assessed via Tanimoto coefficients. Conclusion SuperLigands supplements the set of existing resources of information about small molecules bound to PDB structures. Allowing for three-dimensional comparison of the compounds as a novel feature, this database represents a valuable means of analysis and prediction in the field of biological and medical research.

  2. SuperLigands – a database of ligand structures derived from the Protein Data Bank

    Science.gov (United States)

    Michalsky, Elke; Dunkel, Mathias; Goede, Andrean; Preissner, Robert

    2005-01-01

    Background Currently, the PDB contains approximately 29,000 protein structures comprising over 70,000 experimentally determined three-dimensional structures of over 5,000 different low molecular weight compounds. Information about these PDB ligands can be very helpful in the field of molecular modelling and prediction, particularly for the prediction of protein binding sites and function. Description Here we present an Internet accessible database delivering PDB ligands in the MDL Mol file format which, in contrast to the PDB format, includes information about bond types. Structural similarity of the compounds can be detected by calculation of Tanimoto coefficients and by three-dimensional superposition. Topological similarity of PDB ligands to known drugs can be assessed via Tanimoto coefficients. Conclusion SuperLigands supplements the set of existing resources of information about small molecules bound to PDB structures. Allowing for three-dimensional comparison of the compounds as a novel feature, this database represents a valuable means of analysis and prediction in the field of biological and medical research. PMID:15943884

  3. Mixed ligand oxovanadium(IV) complexes with salicylic acid and N,N-bidentate ligands

    International Nuclear Information System (INIS)

    Two mixed-ligand oxovanadium(IV) complexes VO(A)(B) [where H2A=salicylic acid and B=2,2'-bipyridine or 1,10-phenanthroline (hereafter, bipy and phen respectively)] have been synthesized and characterized by magnetic moment and spectral (IR, UV/VIS and EPR) data. The A2- ion acts as a bidentate dinegative ligand while B ligands acts as a neutral bidentate. The magnetic susceptibility values indicate the existence of a small amount of antiferromagnetic interaction. The vanadium atoms in the complexes are hexacoordinated and the coordination sphere is of the type [VO(OO)(NN)], where O atoms are of oxo, carboxylic and phenolic type and N atoms are of pyridine type. The sixth coordination site is occupied by phenolic oxygen of the neighbouring molecule forming a bridge. The vv=o confirms the hexacoordination. All the complexes have dxy1 type axial EPR spectra and they exhibit two ligand field transitions at 740 and 440 nm. (author)

  4. LASSO-ligand activity by surface similarity order: a new tool for ligand based virtual screening.

    Science.gov (United States)

    Reid, Darryl; Sadjad, Bashir S; Zsoldos, Zsolt; Simon, Aniko

    2008-01-01

    Virtual Ligand Screening (VLS) has become an integral part of the drug discovery process for many pharmaceutical companies. Ligand similarity searches provide a very powerful method of screening large databases of ligands to identify possible hits. If these hits belong to new chemotypes the method is deemed even more successful. eHiTS LASSO uses a new interacting surface point types (ISPT) molecular descriptor that is generated from the 3D structure of the ligand, but unlike most 3D descriptors it is conformation independent. Combined with a neural network machine learning technique, LASSO screens molecular databases at an ultra fast speed of 1 million structures in under 1 min on a standard PC. The results obtained from eHiTS LASSO trained on relatively small training sets of just 2, 4 or 8 actives are presented using the diverse directory of useful decoys (DUD) dataset. It is shown that over a wide range of receptor families, eHiTS LASSO is consistently able to enrich screened databases and provides scaffold hopping ability.

  5. Quasielastic neutron scattering study of POSS ligand dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Jalarvo, Niina H [ORNL; Tyagi, Madhusudan [NIST Center for Neutron Research (NCRN), Gaithersburg, MD; Crawford, Michael [DuPont Experimental Station

    2015-01-01

    Polyoligosilsesquioxanes are molecules having cage-like structures composed of silicon and oxygen. These molecules can have a wide variety of functional ligands attached to them. Depending on the nature of the ligand, interesting properties and applications are found. In this work we present results from quasielastic neutron scattering measurements of four different POSS molecules that illustrate the presence of strong coupling between the ligand dynamics and the POSS crystal structures.

  6. The first nitro-substituted heteroscorpionate ligand.

    Science.gov (United States)

    Pellei, Maura; Benetollo, Franco; Lobbia, Giancarlo Gioia; Alidori, Simone; Santini, Carlo

    2005-02-21

    The new dihydridobis(3-nitro-1,2,4-triazolyl)borate ligand, [H2B(tzNO2)2]-, has been synthesized in dimethylacetamide solution, using 3-nitro-1,2,4-triazole and KBH4 through careful temperature control, and characterized as its potassium salt. The zinc(II) and cadmium(II) complexes, {M[H2B(tzNO2)2]Cl(H2O)2}, have been prepared by metathesis of [H2B(tzNO2)2]K with ZnCl2 and CdCl2, respectively. The complexes likely contain a metal core in which the ligand is coordinated to the metal ions in the K2-N,N' or K4-N,N',O,O' fashion. A single-crystal structural characterization is reported for the potassium dihydrobis(3-nitro-1,2,4-triazolyl)borate. The potassium salt is polymeric and shows several K...N and K...O interactions. PMID:15859260

  7. Do organic ligands affect calcite dissolution rates?

    Science.gov (United States)

    Oelkers, Eric H.; Golubev, Sergey V.; Pokrovsky, Oleg S.; Bénézeth, Pascale

    2011-04-01

    Steady state Iceland-spar calcite dissolution rates were measured at 25 °C in aqueous solutions containing 0.1 M NaCl and up to 0.05 M dissolved bicarbonate at pH from 7.9 to 9.1 in the presence of 13 distinct dissolved organic ligands in mixed-flow reactors. The organic ligands considered in this study include those most likely to be present in either (1) aquifers at the conditions pertinent to CO 2 sequestration or (2) soil/early diagenetic environments: acetate, phthalate, citrate, EDTA 4-, succinate, D-glucosaminate, L-glutamate, D-gluconate, 2,4-dihydroxybenzoate, 3,4-dihydroxybenzoate, fumarate, malonate, and gallate. Results show that the presence of extract, humic acid, pectin, and gum xanthan. In no case did the presence of <100 ppm of these organics change calcite dissolution rates by more than a factor of 2.5. Results obtained in this study suggest that the presence of aqueous organic anions negligibly affects calcite forward dissolution rates in most natural environments. Some effect on calcite reactivity may be observed, however, by the presence of organic anions if they change substantially the chemical affinity of the fluid with respect to calcite.

  8. Continuous microfluidic assortment of interactive ligands (CMAIL)

    Science.gov (United States)

    Hsiao, Yi-Hsing; Huang, Chao-Yang; Hu, Chih-Yung; Wu, Yen-Yu; Wu, Chung-Hsiun; Hsu, Chia-Hsien; Chen, Chihchen

    2016-08-01

    Finding an interactive ligand-receptor pair is crucial to many applications, including the development of monoclonal antibodies. Biopanning, a commonly used technique for affinity screening, involves a series of washing steps and is lengthy and tedious. Here we present an approach termed continuous microfluidic assortment of interactive ligands, or CMAIL, for the screening and sorting of antigen-binding single-chain variable antibody fragments (scFv) displayed on bacteriophages (phages). Phages carrying native negative charges on their coat proteins were electrophoresed through a hydrogel matrix functionalized with target antigens under two alternating orthogonal electric fields. During the weak horizontal electric field phase, phages were differentially swept laterally depending on their affinity for the antigen, and all phages were electrophoresed down to be collected during the strong vertical electric field phase. Phages of different affinity were spatially separated, allowing the continuous operation. More than 105 CFU (colony forming unit) antigen-interacting phages were isolated with ~100% specificity from a phage library containing 3 × 109 individual members within 40 minutes of sorting using CMAIL. CMAIL is rapid, sensitive, specific, and does not employ washing, elution or magnetic beads. In conclusion, we have developed an efficient and cost-effective method for isolating and sorting affinity reagents involving phage display.

  9. EDA Signaling and Skin Appendage Development

    Science.gov (United States)

    Cui, Chang-Yi; Schlessinger, David

    2010-01-01

    The same morphogenetic signals are often involved in the development of different organs. For developing skin appendages, a model for tissue-specific regulation of signaling is provided by the EDA pathway, which accesses the otherwise ubiquitous NFκB transcription factors. EDA signaling is mediated by ectodysplasin, EDAR and EDARADD, which form a new TNF ligand-receptor-adaptor family that is restricted to skin appendages in vertebrates from fish to human. The critical function of the pathway was demonstrated in the hereditary genetic disorder Anhidrotic Ectodermal Dysplasia (EDA), which is characterized by defective formation of hair follicles, sweat glands and teeth. The pathway does not appear to initiate the development of the appendages, but is regulated by and regulates the course of further morphogenesis. In mice, transgenic and knockout strains have increasingly revealed features of the mechanism, and suggest possible non-invasive interventions to alleviate EDA deficiency, especially in sweat glands and eyes. PMID:17102627

  10. Steroid receptors and their ligands: Effects on male gamete functions

    Energy Technology Data Exchange (ETDEWEB)

    Aquila, Saveria; De Amicis, Francesca, E-mail: francesca.deamicis@unical.it

    2014-11-01

    In recent years a new picture of human sperm biology is emerging. It is now widely recognized that sperm contain nuclear encoded mRNA, mitochondrial encoded RNA and different transcription factors including steroid receptors, while in the past sperm were considered incapable of transcription and translation. One of the main targets of steroid hormones and their receptors is reproductive function. Expression studies on Progesterone Receptor, estrogen receptor, androgen receptor and their specific ligands, demonstrate the presence of these systems in mature spermatozoa as surface but also as nuclear conventional receptors, suggesting that both systemic and local steroid hormones, through sperm receptors, may influence male reproduction. However, the relationship between the signaling events modulated by steroid hormones and sperm fertilization potential as well as the possible involvement of the specific receptors are still controversial issues. The main line of this review highlights the current research in human sperm biology examining new molecular systems of response to the hormones as well as specific regulatory pathways controlling sperm cell fate and biological functions. Most significant studies regarding the identification of steroid receptors are reported and the mechanistic insights relative to signaling pathways, together with the change in sperm metabolism energy influenced by steroid hormones are discussed.The reviewed evidences suggest important effects of Progesterone, Estrogen and Testosterone and their receptors on spermatozoa and implicate the involvement of both systemic and local steroid action in the regulation of male fertility potential. - Highlights: • One of the main targets of steroid hormones and their receptors is reproductive function. • Pg/PR co-work to stimulate enzymatic activities to sustain a capacitation process. • E2/ERs regulate sperm motility, capacitation and acrosome reaction and act as survival factors. • Androgens

  11. Superior serum half life of albumin tagged TNF ligands

    International Nuclear Information System (INIS)

    Due to their immune stimulating and apoptosis inducing properties, ligands of the TNF family attract increasing interest as therapeutic proteins. A general limitation of in vivo applications of recombinant soluble TNF ligands is their notoriously rapid clearance from circulation. To improve the serum half life of the TNF family members TNF, TWEAK and TRAIL, we genetically fused soluble variants of these molecules to human serum albumin (HSA). The serum albumin-TNF ligand fusion proteins were found to be of similar bioactivity as the corresponding HSA-less counterparts. Upon intravenous injection (i.v.), serum half life of HSA-TNF ligand fusion proteins, as determined by ELISA, was around 15 h as compared to approximately 1 h for all of the recombinant control TNF ligands without HSA domain. Moreover, serum samples collected 6 or 24 h after i.v. injection still contained high TNF ligand bioactivity, demonstrating that there is only limited degradation/inactivation of circulating HSA-TNF ligand fusion proteins in vivo. In a xenotransplantation model, significantly less of the HSA-TRAIL fusion protein compared to the respective control TRAIL protein was required to achieve inhibition of tumor growth indicating that the increased half life of HSA-TNF ligand fusion proteins translates into better therapeutic action in vivo. In conclusion, our data suggest that genetic fusion to serum albumin is a powerful and generally applicable mean to improve bioavailability and in vivo activity of TNF ligands.

  12. Riboswitch Structure: an Internal Residue Mimicking the Purine Ligand

    Energy Technology Data Exchange (ETDEWEB)

    Delfosse, V.; Bouchard, P; Bonneau, E; Dagenais, P; Lemay, J; Lafontaine, D; Legault, P

    2009-01-01

    The adenine and guanine riboswitches regulate gene expression in response to their purine ligand. X-ray structures of the aptamer moiety of these riboswitches are characterized by a compact fold in which the ligand forms a Watson-Crick base pair with residue 65. Phylogenetic analyses revealed a strict restriction at position 39 of the aptamer that prevents the G39-C65 and A39-U65 combinations, and mutational studies indicate that aptamers with these sequence combinations are impaired for ligand binding. In order to investigate the rationale for sequence conservation at residue 39, structural characterization of the U65C mutant from Bacillus subtilis pbuE adenine riboswitch aptamer was undertaken. NMR spectroscopy and X-ray crystallography studies demonstrate that the U65C mutant adopts a compact ligand-free structure, in which G39 occupies the ligand-binding site of purine riboswitch aptamers. These studies present a remarkable example of a mutant RNA aptamer that adopts a native-like fold by means of ligand mimicking and explain why this mutant is impaired for ligand binding. Furthermore, this work provides a specific insight into how the natural sequence has evolved through selection of nucleotide identities that contribute to formation of the ligand-bound state, but ensures that the ligand-free state remains in an active conformation.

  13. Ligand Release Pathways Obtained with WExplore: Residence Times and Mechanisms.

    Science.gov (United States)

    Dickson, Alex; Lotz, Samuel D

    2016-06-23

    The binding of ligands with their molecular receptors is of tremendous importance in biology. Although much emphasis has been placed on characterizing binding sites and bound poses that determine the binding thermodynamics, the pathway by which a ligand binds importantly determines the binding kinetics. The computational study of entire unbiased ligand binding and release pathways is still an emerging field, made possible only recently by advances in computational hardware and sampling methodologies. We have developed one such method (WExplore) that is based on a weighted ensemble of trajectories, which we apply to ligand release for the first time, using a set of three previously characterized interactions between low-affinity ligands and the protein FKBP-12 (FK-506 binding protein). WExplore is found to be more efficient that conventional sampling, even for the nanosecond-scale unbinding events observed here. From a nonequilibrium ensemble of unbinding trajectories, we obtain ligand residence times and release pathways without using biasing forces or a Markovian assumption of transitions between regions. We introduce a set of analysis tools for unbinding transition pathways, including using von Mises-Fisher distributions to model clouds of ligand exit points, which provide a quantitative proxy for ligand surface diffusion. Differences between the transition pathway ensembles of the three ligands are identified and discussed. PMID:27231969

  14. Single-incubation immunoassay for a multivalent ligand

    International Nuclear Information System (INIS)

    In a two-site immunoassay method for a multivalent ligand using a single incubation, the ligand, labelled receptor for the ligand and unlabelled receptor for the ligand covalently bound to a solid-phase support are incubated as a stable suspension to produce a solid and liquid phase. The solid and liquid phases are separated from each other and the labelled receptor in either phase is quantified. The method has particular application as an assay for human thyroid stimulating hormone using purified, radioactively labelled antibodies and unlabelled antibodies covalently bound to hydrolyzed polyacrylamide particles. (author)

  15. Superior serum half life of albumin tagged TNF ligands

    Energy Technology Data Exchange (ETDEWEB)

    Mueller, Nicole [Division of Molecular Internal Medicine, Department of Internal Medicine II, University Hospital Wuerzburg, Roentgenring 11, 97070 Wuerzburg (Germany); Schneider, Britta; Pfizenmaier, Klaus [Institute of Cell Biology and Immunology, University of Stuttgart, Allmandring 31, 70569 Stuttgart (Germany); Wajant, Harald, E-mail: harald.wajant@mail.uni-wuerzburg.de [Division of Molecular Internal Medicine, Department of Internal Medicine II, University Hospital Wuerzburg, Roentgenring 11, 97070 Wuerzburg (Germany)

    2010-06-11

    Due to their immune stimulating and apoptosis inducing properties, ligands of the TNF family attract increasing interest as therapeutic proteins. A general limitation of in vivo applications of recombinant soluble TNF ligands is their notoriously rapid clearance from circulation. To improve the serum half life of the TNF family members TNF, TWEAK and TRAIL, we genetically fused soluble variants of these molecules to human serum albumin (HSA). The serum albumin-TNF ligand fusion proteins were found to be of similar bioactivity as the corresponding HSA-less counterparts. Upon intravenous injection (i.v.), serum half life of HSA-TNF ligand fusion proteins, as determined by ELISA, was around 15 h as compared to approximately 1 h for all of the recombinant control TNF ligands without HSA domain. Moreover, serum samples collected 6 or 24 h after i.v. injection still contained high TNF ligand bioactivity, demonstrating that there is only limited degradation/inactivation of circulating HSA-TNF ligand fusion proteins in vivo. In a xenotransplantation model, significantly less of the HSA-TRAIL fusion protein compared to the respective control TRAIL protein was required to achieve inhibition of tumor growth indicating that the increased half life of HSA-TNF ligand fusion proteins translates into better therapeutic action in vivo. In conclusion, our data suggest that genetic fusion to serum albumin is a powerful and generally applicable mean to improve bioavailability and in vivo activity of TNF ligands.

  16. Membrane Trafficking of Death Receptors: Implications on Signalling

    Directory of Open Access Journals (Sweden)

    Wulf Schneider-Brachert

    2013-07-01

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

  17. Ligand modulated antagonism of PPARgamma by genomic and non-genomic actions of PPARdelta.

    Directory of Open Access Journals (Sweden)

    Mattias C U Gustafsson

    Full Text Available BACKGROUND: Members of the Peroxisome Proliferator Activated Receptor, PPAR, subfamily of nuclear receptors display complex opposing and overlapping functions and a wide range of pharmacological and molecular genetic tools have been used to dissect their specific functions. Non-agonist bound PPARdelta has been shown to repress PPAR Response Element, PPRE, signalling and several lines of evidence point to the importance of PPARdelta repressive actions in both cardiovascular and cancer biology. METHODOLOGY/PRINCIPAL FINDINGS: In this report we have employed transient transfections and luciferase reporter gene technology to study the repressing effects of PPARdelta and two derivatives thereof. We demonstrate for the first time that the classical dominant negative deletion of the Activation Function 2, AF2, domain of PPARdelta show enhanced repression of PPRE signalling in the presence of a PPARdelta agonist. We propose that the mechanism for the phenomenon is increased RXR heterodimerisation and DNA binding upon ligand binding concomitant with transcriptional co-repressor binding. We also demonstrated ligand-dependent dominant negative action of a DNA non-binding derivative of PPARdelta on PPARgamma1 signalling. This activity was abolished upon over-expression of RXRalpha suggesting a role for PPAR/cofactor competition in the absence of DNA binding. CONCLUSIONS/SIGNIFICANCE: These findings are important in understanding the wide spectrum of molecular interactions in which PPARdelta and PPARgamma have opposing biological roles and suggest novel paradigms for the design of different functional classes of nuclear receptor antagonist drugs.

  18. The Notch ligand Delta-like 1 integrates inputs from TGFbeta/Activin and Wnt pathways

    International Nuclear Information System (INIS)

    Unlike the well-characterized nuclear function of the Notch intracellular domain, it has been difficult to identify a nuclear role for the ligands of Notch. Here we provide evidence for the nuclear function of the Notch ligand Delta-like 1 in colon cancer (CC) cells exposed to butyrate. We demonstrate that the intracellular domain of Delta-like 1 (Dll1icd) augments the activity of Wnt signaling-dependent reporters and that of the promoter of the connective tissue growth factor (CTGF) gene. Data suggest that Dll1icd upregulates CTGF promoter activity through both direct and indirect mechanisms. The direct mechanism is supported by co-immunoprecipitation of endogenous Smad2/3 proteins and Dll1 and by chromatin immunoprecipitation analyses that revealed the occupancy of Dll1icd on CTGF promoter sequences containing a Smad binding element. The indirect upregulation of CTGF expression by Dll1 is likely due to the ability of Dll1icd to increase Wnt signaling, a pathway that targets CTGF. CTGF expression is induced in butyrate-treated CC cells and results from clonal growth assays support a role for CTGF in the cell growth-suppressive role of butyrate. In conclusion, integration of the Notch, Wnt, and TGFbeta/Activin signaling pathways is in part mediated by the interactions of Dll1 with Smad2/3 and Tcf4.

  19. Mincle Signaling Promotes Con A Hepatitis.

    Science.gov (United States)

    Greco, Stephanie H; Torres-Hernandez, Alejandro; Kalabin, Aleksandr; Whiteman, Clint; Rokosh, Rae; Ravirala, Sushma; Ochi, Atsuo; Gutierrez, Johana; Salyana, Muhammad Atif; Mani, Vishnu R; Nagaraj, Savitha V; Deutsch, Michael; Seifert, Lena; Daley, Donnele; Barilla, Rocky; Hundeyin, Mautin; Nikifrov, Yuriy; Tejada, Karla; Gelb, Bruce E; Katz, Steven C; Miller, George

    2016-10-01

    Con A hepatitis is regarded as a T cell-mediated model of acute liver injury. Mincle is a C-type lectin receptor that is critical in the immune response to mycobacteria and fungi but does not have a well-defined role in preclinical models of non-pathogen-mediated inflammation. Because Mincle can ligate the cell death ligand SAP130, we postulated that Mincle signaling drives intrahepatic inflammation and liver injury in Con A hepatitis. Acute liver injury was assessed in the murine Con A hepatitis model using C57BL/6, Mincle(-/-), and Dectin-1(-/-) mice. The role of C/EBPβ and hypoxia-inducible factor-1α (HIF-1α) signaling was assessed using selective inhibitors. We found that Mincle was highly expressed in hepatic innate inflammatory cells and endothelial cells in both mice and humans. Furthermore, sterile Mincle ligands and Mincle signaling intermediates were increased in the murine liver in Con A hepatitis. Most significantly, Mincle deletion or blockade protected against Con A hepatitis, whereas Mincle ligation exacerbated disease. Bone marrow chimeric and adoptive transfer experiments suggested that Mincle signaling in infiltrating myeloid cells dictates disease phenotype. Conversely, signaling via other C-type lectin receptors did not alter disease course. Mechanistically, we found that Mincle blockade decreased the NF-κβ-related signaling intermediates C/EBPβ and HIF-1α, both of which are necessary in macrophage-mediated inflammatory responses. Accordingly, Mincle deletion lowered production of nitrites in Con A hepatitis and inhibition of both C/EBPβ and HIF-1α reduced the severity of liver disease. Our work implicates a novel innate immune driver of Con A hepatitis and, more broadly, suggests a potential role for Mincle in diseases governed by sterile inflammation.

  20. Mincle Signaling Promotes Con A Hepatitis.

    Science.gov (United States)

    Greco, Stephanie H; Torres-Hernandez, Alejandro; Kalabin, Aleksandr; Whiteman, Clint; Rokosh, Rae; Ravirala, Sushma; Ochi, Atsuo; Gutierrez, Johana; Salyana, Muhammad Atif; Mani, Vishnu R; Nagaraj, Savitha V; Deutsch, Michael; Seifert, Lena; Daley, Donnele; Barilla, Rocky; Hundeyin, Mautin; Nikifrov, Yuriy; Tejada, Karla; Gelb, Bruce E; Katz, Steven C; Miller, George

    2016-10-01

    Con A hepatitis is regarded as a T cell-mediated model of acute liver injury. Mincle is a C-type lectin receptor that is critical in the immune response to mycobacteria and fungi but does not have a well-defined role in preclinical models of non-pathogen-mediated inflammation. Because Mincle can ligate the cell death ligand SAP130, we postulated that Mincle signaling drives intrahepatic inflammation and liver injury in Con A hepatitis. Acute liver injury was assessed in the murine Con A hepatitis model using C57BL/6, Mincle(-/-), and Dectin-1(-/-) mice. The role of C/EBPβ and hypoxia-inducible factor-1α (HIF-1α) signaling was assessed using selective inhibitors. We found that Mincle was highly expressed in hepatic innate inflammatory cells and endothelial cells in both mice and humans. Furthermore, sterile Mincle ligands and Mincle signaling intermediates were increased in the murine liver in Con A hepatitis. Most significantly, Mincle deletion or blockade protected against Con A hepatitis, whereas Mincle ligation exacerbated disease. Bone marrow chimeric and adoptive transfer experiments suggested that Mincle signaling in infiltrating myeloid cells dictates disease phenotype. Conversely, signaling via other C-type lectin receptors did not alter disease course. Mechanistically, we found that Mincle blockade decreased the NF-κβ-related signaling intermediates C/EBPβ and HIF-1α, both of which are necessary in macrophage-mediated inflammatory responses. Accordingly, Mincle deletion lowered production of nitrites in Con A hepatitis and inhibition of both C/EBPβ and HIF-1α reduced the severity of liver disease. Our work implicates a novel innate immune driver of Con A hepatitis and, more broadly, suggests a potential role for Mincle in diseases governed by sterile inflammation. PMID:27559045

  1. Monitoring β-arrestin recruitment via β-lactamase enzyme fragment complementation: purification of peptide E as a low-affinity ligand for mammalian bombesin receptors.

    Science.gov (United States)

    Ikeda, Yuichi; Kumagai, Hidetoshi; Okazaki, Hiroaki; Fujishiro, Mitsuhiro; Motozawa, Yoshihiro; Nomura, Seitaro; Takeda, Norifumi; Toko, Haruhiro; Takimoto, Eiki; Akazawa, Hiroshi; Morita, Hiroyuki; Suzuki, Jun-ichi; Yamazaki, Tsutomu; Komuro, Issei; Yanagisawa, Masashi

    2015-01-01

    Identification of cognate ligands for G protein-coupled receptors (GPCRs) provides a starting point for understanding novel regulatory mechanisms. Although GPCR ligands have typically been evaluated through the activation of heterotrimeric G proteins, recent studies have shown that GPCRs signal not only through G proteins but also through β-arrestins. As such, monitoring β-arrestin signaling instead of G protein signaling will increase the likelihood of identifying currently unknown ligands, including β-arrestin-biased agonists. Here, we developed a cell-based assay for monitoring ligand-dependent GPCR-β-arrestin interaction via β-lactamase enzyme fragment complementation. Inter alia, β-lactamase is a superior reporter enzyme because of its cell-permeable fluorescent substrate. This substrate makes the assay non-destructive and compatible with fluorescence-activated cell sorting (FACS). In a reporter cell, complementary fragments of β-lactamase (α and ω) were fused to β-arrestin 2 and GPCR, respectively. Ligand stimulation initiated the interaction of these chimeric proteins (β-arrestin-α and GPCR-ω), and this inducible interaction was measured through reconstituted β-lactamase activity. Utilizing this system, we screened various mammalian tissue extracts for agonistic activities on human bombesin receptor subtype 3 (hBRS3). We purified peptide E as a low-affinity ligand for hBRS3, which was also found to be an agonist for the other two mammalian bombesin receptors such as gastrin-releasing peptide receptor (GRPR) and neuromedin B receptor (NMBR). Successful purification of peptide E has validated the robustness of this assay. We conclude that our newly developed system will facilitate the discovery of GPCR ligands. PMID:26030739

  2. Monitoring β-arrestin recruitment via β-lactamase enzyme fragment complementation: purification of peptide E as a low-affinity ligand for mammalian bombesin receptors.

    Directory of Open Access Journals (Sweden)

    Yuichi Ikeda

    Full Text Available Identification of cognate ligands for G protein-coupled receptors (GPCRs provides a starting point for understanding novel regulatory mechanisms. Although GPCR ligands have typically been evaluated through the activation of heterotrimeric G proteins, recent studies have shown that GPCRs signal not only through G proteins but also through β-arrestins. As such, monitoring β-arrestin signaling instead of G protein signaling will increase the likelihood of identifying currently unknown ligands, including β-arrestin-biased agonists. Here, we developed a cell-based assay for monitoring ligand-dependent GPCR-β-arrestin interaction via β-lactamase enzyme fragment complementation. Inter alia, β-lactamase is a superior reporter enzyme because of its cell-permeable fluorescent substrate. This substrate makes the assay non-destructive and compatible with fluorescence-activated cell sorting (FACS. In a reporter cell, complementary fragments of β-lactamase (α and ω were fused to β-arrestin 2 and GPCR, respectively. Ligand stimulation initiated the interaction of these chimeric proteins (β-arrestin-α and GPCR-ω, and this inducible interaction was measured through reconstituted β-lactamase activity. Utilizing this system, we screened various mammalian tissue extracts for agonistic activities on human bombesin receptor subtype 3 (hBRS3. We purified peptide E as a low-affinity ligand for hBRS3, which was also found to be an agonist for the other two mammalian bombesin receptors such as gastrin-releasing peptide receptor (GRPR and neuromedin B receptor (NMBR. Successful purification of peptide E has validated the robustness of this assay. We conclude that our newly developed system will facilitate the discovery of GPCR ligands.

  3. OSM-11 facilitates LIN-12 Notch signaling during Caenorhabditis elegans vulval development.

    Directory of Open Access Journals (Sweden)

    Hidetoshi Komatsu

    2008-08-01

    Full Text Available Notch signaling is critical for cell fate decisions during development. Caenorhabditis elegans and vertebrate Notch ligands are more diverse than classical Drosophila Notch ligands, suggesting possible functional complexities. Here, we describe a developmental role in Notch signaling for OSM-11, which has been previously implicated in defecation and osmotic resistance in C. elegans. We find that complete loss of OSM-11 causes defects in vulval precursor cell (VPC fate specification during vulval development consistent with decreased Notch signaling. OSM-11 is a secreted, diffusible protein that, like previously described C. elegans Delta, Serrate, and LAG-2 (DSL ligands, can interact with the lineage defective-12 (LIN-12 Notch receptor extracellular domain. Additionally, OSM-11 and similar C. elegans proteins share a common motif with Notch ligands from other species in a sequence defined here as the Delta and OSM-11 (DOS motif. osm-11 loss-of-function defects in vulval development are exacerbated by loss of other DOS-motif genes or by loss of the Notch ligand DSL-1, suggesting that DOS-motif and DSL proteins act together to activate Notch signaling in vivo. The mammalian DOS-motif protein Deltalike1 (DLK1 can substitute for OSM-11 in C. elegans development, suggesting that DOS-motif function is conserved across species. We hypothesize that C. elegans OSM-11 and homologous proteins act as coactivators for Notch receptors, allowing precise regulation of Notch receptor signaling in developmental programs in both vertebrates and invertebrates.

  4. Differential effects of EGFR ligands on endocytic sorting of the receptor

    DEFF Research Database (Denmark)

    Roepstorff, Kirstine; Grandal, Michael Vibo; Henriksen, Lasse;

    2009-01-01

    Endocytic downregulation is a pivotal mechanism turning off signalling from the EGF receptor (EGFR). It is well established that whereas EGF binding leads to lysosomal degradation of EGFR, transforming growth factor (TGF)-alpha causes receptor recycling. TGF-alpha therefore leads to continuous...... recycling. EGF leads to lysosomal degradation of the majority but not all EGFRs. Amphiregulin does not target EGFR for lysosomal degradation but causes fast as well as slow EGFR recycling. The Cbl ubiquitin ligases, especially c-Cbl, are responsible for EGFR ubiquitination after stimulation with all ligands...

  5. Structures of pattern recognition receptors reveal molecular mechanisms of autoinhibition, ligand recognition and oligomerization.

    Science.gov (United States)

    Chuenchor, Watchalee; Jin, Tengchuan; Ravilious, Geoffrey; Xiao, T Sam

    2014-02-01

    Pattern recognition receptors (PRRs) are essential sentinels for pathogens or tissue damage and integral components of the innate immune system. Recent structural studies have provided unprecedented insights into the molecular mechanisms of ligand recognition and signal transduction by several PRR families at distinct subcellular compartments. Here we highlight some of the recent discoveries and summarize the common themes that are emerging from these exciting studies. Better mechanistic understanding of the structure and function of the PRRs will improve future prospects of therapeutic targeting of these important innate immune receptors.

  6. Characterizing mixed phosphonic acid ligand capping on CdSe/ZnS quantum dots using ligand exchange and NMR spectroscopy.

    Science.gov (United States)

    Davidowski, Stephen K; Lisowski, Carmen E; Yarger, Jeffery L

    2016-03-01

    The ligand capping of phosphonic acid functionalized CdSe/ZnS core-shell quantum dots (QDs) was investigated with a combination of solution and solid-state (31) P nuclear magnetic resonance (NMR) spectroscopy. Two phosphonic acid ligands were used in the synthesis of the QDs, tetradecylphosphonic acid and ethylphosphonic acid. Both alkyl phosphonic acids showed broad liquid and solid-state (31) P NMR resonances for the bound ligands, indicative of heterogeneous binding to the QD surface. In order to quantify the two ligand populations on the surface, ligand exchange facilitated by phenylphosphonic acid resulted in the displacement of the ethylphosphonic acid and tetradecylphosphonic acid and allowed for quantification of the free ligands using (31) P liquid-state NMR. After washing away the free ligand, two broad resonances were observed in the liquids' (31) P NMR corresponding to the alkyl and aromatic phosphonic acids. The washed samples were analyzed via solid-state (31) P NMR, which confirmed the ligand populations on the surface following the ligand exchange process. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26639792

  7. Modification of diphenylamine-linked bis(oxazoline) ligands: Tuning of electronic effect and rigidity of ligand skeleton

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    The electronic effect of diphenylamine-linked bis(oxazoline) ligands was tuned through introduction of electron-withdrawing bromo and nitro substituents onto the 4 and 4′ position. The variation of the NH bond acidity was determined by the different chemical shifts of NH. The catalytic activity and enantioselectivity of the modified ligands were tested in the asymmetric Friedel-Crafts alkylation of indole with β-nitrostyrene. The effect of ligand skeleton rigidity was also investigated through the synthesis of iminodibenzyl-linked bis(oxazoline) ligands and evaluation of their catalytic activity in Friedel-Crafts alkylation.

  8. Calculating the mean time to capture for tethered ligands and its effect on the chemical equilibrium of bound ligand pairs.

    Science.gov (United States)

    Shen, Lu; Decker, Caitlin G; Maynard, Heather D; Levine, Alex J

    2016-09-01

    We present here the calculation of the mean time to capture of a tethered ligand to the receptor. This calculation is then used to determine the shift in the partitioning between (1) free, (2) singly bound, and (3) doubly bound ligands in chemical equilibrium as a function of the length of the tether. These calculations are used in the research article Fibroblast Growth Factor 2 Dimer with Superagonist in vitro Activity Improves Granulation Tissue Formation During Wound Healing (Decker et al., in press [1]) to explain quantitatively how changes in polymeric linker length in the ligand dimers modifies the efficacy of these molecules relative to that of free ligands. PMID:27408925

  9. Developing a synthetic signal transduction system in plants.

    Science.gov (United States)

    Morey, Kevin J; Antunes, Mauricio S; Albrecht, Kirk D; Bowen, Tessa A; Troupe, Jared F; Havens, Keira L; Medford, June I

    2011-01-01

    One area of focus in the emerging field of plant synthetic biology is the manipulation of systems involved in sensing and response to environmental signals. Sensing and responding to signals, including ligands, typically involves biological signal transduction. Plants use a wide variety of signaling systems to sense and respond to their environment. One of these systems, a histidine kinase (HK) based signaling system, lends itself to manipulation using the tools of synthetic biology. Both plants and bacteria use HKs to relay signals, which in bacteria can involve as few as two proteins (two-component systems or TCS). HK proteins are evolutionarily conserved between plants and bacteria and plant HK components have been shown to be functional in bacteria. We found that this conservation also applies to bacterial HK components which can function in plants. This conservation of function led us to hypothesize that synthetic HK signaling components can be designed and rapidly tested in bacteria. These novel HK signaling components form the foundation for a synthetic signaling system in plants, but typically require modifications such as codon optimization and proper targeting to allow optimal function. We describe the process and methodology of producing a synthetic signal transduction system in plants. We discovered that the bacterial response regulator (RR) PhoB shows HK-dependent nuclear translocation in planta. Using this discovery, we engineered a partial synthetic pathway in which a synthetic promoter (PlantPho) is activated using a plant-adapted PhoB (PhoB-VP64) and the endogenous HK-based cytokinin signaling pathway. Building on this work, we adapted an input or sensing system based on bacterial chemotactic binding proteins and HKs, resulting in a complete eukaryotic signal transduction system. Input to our eukaryotic signal transduction system is provided by a periplasmic binding protein (PBP), ribose-binding protein (RBP). RBP interacts with the membrane

  10. Elucidating the Functional Roles of Spatial Organization in Cross-Membrane Signal Transduction by a Hybrid Simulation Method.

    Science.gov (United States)

    Chen, Jiawen; Xie, Zhong-Ru; Wu, Yinghao

    2016-07-01

    The ligand-binding of membrane receptors on cell surfaces initiates the dynamic process of cross-membrane signal transduction. It is an indispensable part of the signaling network for cells to communicate with external environments. Recent experiments revealed that molecular components in signal transduction are not randomly mixed, but spatially organized into distinctive patterns. These patterns, such as receptor clustering and ligand oligomerization, lead to very different gene expression profiles. However, little is understood about the molecular mechanisms and functional impacts of this spatial-temporal regulation in cross-membrane signal transduction. In order to tackle this problem, we developed a hybrid computational method that decomposes a model of signaling network into two simulation modules. The physical process of binding between receptors and ligands on cell surfaces are simulated by a diffusion-reaction algorithm, while the downstream biochemical reactions are modeled by stochastic simulation of Gillespie algorithm. These two processes are coupled together by a synchronization framework. Using this method, we tested the dynamics of a simple signaling network in which the ligand binding of cell surface receptors triggers the phosphorylation of protein kinases, and in turn regulates the expression of target genes. We found that spatial aggregation of membrane receptors at cellular interfaces is able to either amplify or inhibit downstream signaling outputs, depending on the details of clustering mechanism. Moreover, by providing higher binding avidity, the co-localization of ligands into multi-valence complex modulates signaling in very different ways that are closely related to the binding affinity between ligand and receptor. We also found that the temporal oscillation of the signaling pathway that is derived from genetic feedback loops can be modified by the spatial clustering of membrane receptors. In summary, our method demonstrates the functional

  11. Gamma secretase inhibitors of Notch signaling

    Directory of Open Access Journals (Sweden)

    Olsauskas-Kuprys R

    2013-07-01

    Full Text Available Roma Olsauskas-Kuprys,1 Andrei Zlobin,1 Clodia Osipo1,2 1The Oncology Institute, Cardinal Bernardin Cancer Center, Loyola University Chicago, Chicago, IL, USA; 2Department of Pathology, Cardinal Bernardin Cancer Center, Loyola University Chicago, Chicago, IL, USA Abstract: The numerous processes involved in the etiology of breast cancer such as cell survival, metabolism, proliferation, differentiation, and angiogenesis are currently being elucidated. However, underlying mechanisms that drive breast cancer progression and drug resistance are still poorly understood. As we discuss here in detail, the Notch signaling pathway is an important regulatory component of normal breast development, cell fate of normal breast stem cells, and proliferation and survival of breast cancer initiating cells. Notch exerts a wide range of critical effects through a canonical pathway where it is expressed as a type I membrane precursor heterodimer followed by at least two subsequent cleavages induced by ligand engagement to ultimately release an intracellular form to function as a transcriptional activator. Notch and its ligands are overexpressed in breast cancer, and one method of effectively blocking Notch activity is preventing its cleavage at the cell surface with γ-secretase inhibitors. In the context of Notch signaling, the application of clinically relevant anti-Notch drugs in treatment regimens may contribute to novel therapeutic interventions and promote more effective clinical response in women with breast cancer. Keywords: breast cancer, signaling pathways, γ-secretase, γ-secretase inhibitors, combination breast cancer therapy

  12. Quantitative proteomic assessment of very early cellular signaling events

    DEFF Research Database (Denmark)

    Dengjel, Joern; Akimov, Vyacheslav; Olsen, Jesper V;

    2007-01-01

    Technical limitations have prevented proteomic analyses of events occurring less than 30 s after signal initiation. We developed an automated, continuous quench-flow system allowing quantitative proteomic assessment of very early cellular signaling events (qPACE) with a time resolution of 1 s....... Using this technique, we determined that autophosphorylation of the epidermal growth factor receptor occurs within 1 s after ligand stimulation and is followed rapidly by phosphorylation of the downstream signaling intermediates Src homologous and collagen-like protein and phospholipase C gamma 1....

  13. Functional metal-organic frameworks via ligand doping: influences of ligand charge and steric demand.

    Science.gov (United States)

    Wang, Cheng; Liu, Demin; Xie, Zhigang; Lin, Wenbin

    2014-02-01

    Doping a functional ligand into a known crystalline system built from ligands of similar shape and length provides a powerful strategy to construct functional metal-organic frameworks (MOFs) with desired functionality and structural topology. This mix-and-match approach mimics the widely applied metal ion doping (or solid solution formation) in traditional inorganic materials, such as metal oxides, wherein maintaining charge balance of the doped lattice and ensuring size match between doped metal ions and the parent lattice are key to successful doping. In this work, we prepared three sterically demanding dicarboxylate ligands based on Ir/Ru-phosphors with similar structures and variable charges (-2 to 0), [Ir(ppy)3]-dicarboxylate (L1, ppy is 2-phenylpyridine), [Ir(bpy)(ppy)2](+)-dicarboxylate (L2, bpy is 2,2'-bipyridine), and Ru(bpy)3](2+)-dicarboxylate (L3), and successfully doped them into the known IRMOF-9/-10 structures by taking advantage of matching length between 4,4'-biphenyl dicarboxylate (BPDC) and L1-L3. We systematically investigated the effects of size and charge of the doping ligand on the MOF structures and the ligand doping levels in these MOFs. L1 carries a -2 charge to satisfy the charge requirement of the parent Zn4O(BPDC)3 framework and can be mixed into the IRMOF-9/-10 structure in the whole range of H2L1/H2BPDC ratios from 0 to 1. The steric bulk of L1 induces a phase transition from the interpenetrated IRMOF-9 structure to the non-interpenetrated IRMOF-10 counterpart. L2 and L3 do not match the dinegative charge of BPDC in order to maintain the charge balance for a neutral IRMOF-9/-10 framework and can only be doped into the IRMOF-9 structure to a certain degree. L2 and L3 form a charge-balanced new phase with a neutral framework structure at higher doping levels (>8% For L2 and >6% For L3). This systematic investigation reveals the influences of steric demand and charge balance on ligand doping in MOFs, a phenomenon that has been well

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

    Institute of Scientific and Technical Information of China (English)

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

    2008-01-01

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

  15. Treatment of autoimmune inflammation by a TLR7 ligand regulating the innate immune system.

    Directory of Open Access Journals (Sweden)

    Tomoko Hayashi

    Full Text Available The Toll-like receptors (TLR have been advocated as attractive therapeutic targets because TLR signaling plays dual roles in initiating adaptive immune responses and perpetuating inflammation. Paradoxically, repeated stimulation of bone marrow mononuclear cells with a synthetic TLR7 ligand 9-benzyl-8-hydroxy-2-(2-methoxyethoxy adenine (called 1V136 leads to subsequent TLR hyporesponsiveness. Further studies on the mechanism of action of this pharmacologic agent demonstrated that the TLR7 ligand treatment depressed dendritic cell activation, but did not directly affect T cell function. To verify this mechanism, we utilized experimental allergic encephalitis (EAE as an in vivo T cell dependent autoimmune model. Drug treated SJL/J mice immunized with proteolipid protein (PLP(139-151 peptide had attenuated disease severity, reduced accumulation of mononuclear cells in the central nervous system (CNS, and limited demyelination, without any apparent systemic toxicity. Splenic T cells from treated mice produced less cytokines upon antigenic rechallenge. In the spinal cords of 1V136-treated EAE mice, the expression of chemoattractants was also reduced, suggesting innate immune cell hyposensitization in the CNS. Indeed, systemic 1V136 did penetrate the CNS. These experiments indicated that repeated doses of a TLR7 ligand may desensitize dendritic cells in lymphoid organs, leading to diminished T cell responses. This treatment strategy might be a new modality to treat T cell mediated autoimmune diseases.

  16. Force-Mediated Kinetics of Single P-Selectin/Ligand Complexes Observed by Atomic Force Microscopy

    Science.gov (United States)

    Fritz, Jurgen; Katopodis, Andreas G.; Kolbinger, Frank; Anselmetti, Dario

    1998-10-01

    Leukocytes roll along the endothelium of postcapillary venules in response to inflammatory signals. Rolling under the hydrodynamic drag forces of blood flow is mediated by the interaction between selectins and their ligands across the leukocyte and endothelial cell surfaces. Here we present force-spectroscopy experiments on single complexes of P-selectin and P-selectin glycoprotein ligand-1 by atomic force microscopy to determine the intrinsic molecular properties of this dynamic adhesion process. By modeling intermolecular and intramolecular forces as well as the adhesion probability in atomic force microscopy experiments we gain information on rupture forces, elasticity, and kinetics of the P-selectin/P-selectin glycoprotein ligand-1 interaction. The complexes are able to withstand forces up to 165 pN and show a chain-like elasticity with a molecular spring constant of 5.3 pN nm-1 and a persistence length of 0.35 nm. The dissociation constant (off-rate) varies over three orders of magnitude from 0.02 s-1 under zero force up to 15 s-1 under external applied forces. Rupture force and lifetime of the complexes are not constant, but directly depend on the applied force per unit time, which is a product of the intrinsic molecular elasticity and the external pulling velocity. The high strength of binding combined with force-dependent rate constants and high molecular elasticity are tailored to support physiological leukocyte rolling.

  17. Ligand-dependent localization and function of ORP-VAP complexes at membrane contact sites.

    Science.gov (United States)

    Weber-Boyvat, Marion; Kentala, Henriikka; Peränen, Johan; Olkkonen, Vesa M

    2015-05-01

    Oxysterol-binding protein/OSBP-related proteins (ORPs) constitute a conserved family of sterol/phospholipid-binding proteins with lipid transporter or sensor functions. We investigated the spatial occurrence and regulation of the interactions of human OSBP/ORPs or the S. cerevisiae orthologs, the Osh (OSBP homolog) proteins, with their endoplasmic reticulum (ER) anchors, the VAMP-associated proteins (VAPs), by employing bimolecular fluorescence complementation and pull-down set-ups. The ORP-VAP interactions localize frequently at distinct subcellular sites, shown in several cases to represent membrane contact sites (MCSs). Using established ORP ligand-binding domain mutants and pull-down assays with recombinant proteins, we show that ORP liganding regulates the ORP-VAP association, alters the subcellular targeting of ORP-VAP complexes, or modifies organelle morphology. There is distinct protein specificity in the effects of the mutants on subcellular targeting of ORP-VAP complexes. We provide evidence that complexes of human ORP2 and VAPs at ER-lipid droplet interfaces regulate the hydrolysis of triglycerides and lipid droplet turnover. The data suggest evolutionarily conserved, complex ligand-dependent functions of ORP-VAP complexes at MCSs, with implications for cellular lipid homeostasis and signaling. PMID:25420878

  18. Fatty Acid-binding Proteins Interact with Comparative Gene Identification-58 Linking Lipolysis with Lipid Ligand Shuttling*

    OpenAIRE

    Hofer, Peter; Boeszoermenyi, Andras; Jaeger, Doris; Feiler, Ursula; Arthanari, Haribabu; Mayer, Nicole; Zehender, Fabian; Rechberger, Gerald; Oberer, Monika; Zimmermann, Robert; Lass, Achim; Haemmerle, Guenter; Breinbauer, Rolf; Zechner, Rudolf; Preiss-Landl, Karina

    2015-01-01

    Background: A multiprotein complex designated as lipolysome degrades intracellular triglycerides and contains proteins such as adipose triglyceride lipase (Atgl) and its co-activator Cgi-58. Results: Cgi-58 interacts with fatty acid-binding proteins (Fabps), which impact Atgl-mediated lipolysis and lipid signaling. Conclusion: Fabps modulate Atgl-mediated TG hydrolysis and link lipolysis with intracellular lipid ligand shuttling. Significance: Novel mechanistic insights into the regulation of...

  19. One- and Two-photon Excited Fluorescence of Zinc(Ⅱ), Cadmium(Ⅱ) Complexes Containing Phenothiazine Ligand

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    A new ligand, 10-ethylphenothiazinyl - 3 - yl - methylene thiosemicarbazon (HL) and its complexes ML2 (M=Zn2+, Cd2+), which exhibit intensive two-photon excited (TPE) fluorescence at 800 nm laser pulses in femtosecond regime, were synthesized and characterized.The measured power dependence of the fluorescence signals provided direct evidence for TPE.All of them exhibited a large two-photon absorptive cross section and, more importantly from the application point of view, high photochemical/photothermal stability.

  20. Ligand-Independent Canonical Wnt Activity in Canine Mammary Tumor Cell Lines Associated with Aberrant LEF1 Expression

    Science.gov (United States)

    van Wolferen, Monique E.; Rao, Nagesha A. S.; Grizelj, Juraj; Vince, Silvijo; Hellmen, Eva; Mol, Jan A.

    2014-01-01

    Pet dogs very frequently develop spontaneous mammary tumors and have been suggested as a good model organism for breast cancer research. In order to obtain an insight into underlying signaling mechanisms during canine mammary tumorigenesis, in this study we assessed the incidence and the mechanism of canonical Wnt activation in a panel of 12 canine mammary tumor cell lines. We show that a subset of canine mammary cell lines exhibit a moderate canonical Wnt activity that is dependent on Wnt ligands, similar to what has been described in human breast cancer cell lines. In addition, three of the tested canine mammary cell lines have a high canonical Wnt activity that is not responsive to inhibitors of Wnt ligand secretion. Tumor cell lines with highly active canonical Wnt signaling often carry mutations in key members of the Wnt signaling cascade. These cell lines, however, carry no mutations in the coding regions of intracellular Wnt pathway components (APC, β-catenin, GSK3β, CK1α and Axin1) and have a functional β-catenin destruction complex. Interestingly, however, the cell lines with high canonical Wnt activity specifically overexpress LEF1 mRNA and the knock-down of LEF1 significantly inhibits TCF-reporter activity. In addition, LEF1 is overexpressed in a subset of canine mammary carcinomas, implicating LEF1 in ligand-independent activation of canonical Wnt signaling in canine mammary tumors. We conclude that canonical Wnt activation may be a frequent event in canine mammary tumors both through Wnt ligand-dependent and novel ligand–independent mechanisms. PMID:24887235

  1. Rational design of class I MHC ligands

    Science.gov (United States)

    Rognan, D.; Scapozza, L.; Folkers, G.; Daser, Angelika

    1995-04-01

    From the knowledge of the three-dimensional structure of a class I MHC protein, several non natural peptides were designed in order to either optimize the interactions of one secondary anchor amino acid with its HLA binding pocket or to substitute the non interacting part with spacer residues. All peptides were synthesized and tested for binding to the class I MHC protein in an in vitro reconstitution assay. As predicted, the non natural peptides present an enhanced binding to the HLA-B27 molecule with respect to their natural parent peptides. This study constitutes the first step towards the rational design of non peptidic MHC ligands that should be very promising tools for the selective immunotherapy of autoimmune diseases.

  2. Evidence of non-canonical NOTCH signaling

    DEFF Research Database (Denmark)

    Traustadóttir, Gunnhildur Ásta; Jensen, Charlotte H; Thomassen, Mads;

    2016-01-01

    suggested to interact with NOTCH1 and act as an antagonist. This non-canonical interaction is, however controversial, and evidence for a direct interaction, still lacking in mammals. In this study, we elucidated the putative DLK1-NOTCH1 interaction in a mammalian context. Taking a global approach and using...... this interaction to occur between EGF domains 5 and 6 of DLK1 and EGF domains 10-15 of NOTCH1. Thus, our data provide the first evidence for a direct interaction between DLK1 and NOTCH1 in mammals, and substantiate that non-canonical NOTCH ligands exist, adding to the complexity of NOTCH signaling....

  3. Regulation of Notch Signaling by Glycosylation

    OpenAIRE

    Stanley, Pamela

    2007-01-01

    Notch receptors are ~300 kd cell surface glycoproteins whose activation by Notch ligands regulates cell fate decisions in the metazoa. The extracellular domain of Notch receptors has many epidermal growth factor-like repeats that are glycosylated with O-fucose and O-glucose glycans as well as N-glycans. Disruption of O-fucose glycan synthesis leads to severe Notch signaling defects in Drosophila and mammals. Removal or addition of O-fucose glycan consensus sites on Notch receptors also leads ...

  4. Optimal Overlay of Ligands with Flexible Bonds Using Differential Evolution

    DEFF Research Database (Denmark)

    Kristensen, Thomas Greve; Pedersen, Christian Storm

    2009-01-01

    When designing novel drugs, the need arise to screen large databases of drug candidates (small synthesizable chemical structures) for structures that resemble active ligands, i.e. small chemical structures that are known to react with the target protein. If several active ligands are known one mi...

  5. Organopalladium complexes with bidentate phosphorus and nitrogen containing ligands

    NARCIS (Netherlands)

    Koten, G. van; Graaf, W. de; Harder, Sjoerd; Boersma, J.; Kanters, J.A.

    1988-01-01

    Organopalladium complexes containing the potentially P, N-bidentate ligands o-diphenylphosphino-N,N-dimethylbenzylamine (PN) and o-diphenylphosphino-@a-methyl-N,N-dimethylbenzylamine (PN}*{) have been studied. The palladium(0) complexes Pd(P@?N){3} (P@?N = PN or PN}*{) have been prepared: the ligand

  6. Death receptors and ligands in cervical carcinogenesis : an immunohistochemical study

    NARCIS (Netherlands)

    Reesink-Peters, N; Hougardy, B M T; van den Heuvel, F A J; Ten Hoor, K A; Hollema, H; Boezen, H M; de Vries, E G E; de Jong, S; van der Zee, A G J

    2005-01-01

    OBJECTIVE: Increasing imbalance between proliferation and apoptosis is important in cervical carcinogenesis. The death ligands FasL and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induce apoptosis by binding to their cognate cell-surface death receptors Fas or death receptor (DR)

  7. Functionalized pyrazines as ligands for minor actinide extraction and catalysis

    NARCIS (Netherlands)

    Nikishkin, N.

    2013-01-01

    The research presented in this thesis concerns the design of ligands for a wide range of applications, from nuclear waste treatment to catalysis. The strategies employed to design actinide-selective extractants, for instance, comprise the fine tuning of the ligand electronic properties as well as us

  8. Influence of the platform in multicoordinate ligands for actinide partitioning

    NARCIS (Netherlands)

    Dam, Henk H.; Reinhoudt, David N.; Verboom, Willem

    2007-01-01

    Multicoordinate ligands based on the trityl, C-pivot, and CTV platforms and the ligating groups CMPO, DGA, PICO, and MPMA were synthesized and studied for their extraction properties. The extraction efficiencies of these multicoordinate ligands are largely influenced by the properties of the platfor

  9. Immobilisation of ligands by radio-derivatized polymers

    International Nuclear Information System (INIS)

    The invention relates to radio-derivatized polymers and a method of producing them by contacting non-polymerizable conjugands with radiolysable polymers in the presence of irradiation. The resulting radio-derivatized polymers can be further linked with ligand of organic or inorganic nature to immobilize such ligands. 2 figs., 5 tabs

  10. Improved ligand geometries in crystallographic refinement using AFITT in PHENIX.

    Science.gov (United States)

    Janowski, Pawel A; Moriarty, Nigel W; Kelley, Brian P; Case, David A; York, Darrin M; Adams, Paul D; Warren, Gregory L

    2016-09-01

    Modern crystal structure refinement programs rely on geometry restraints to overcome the challenge of a low data-to-parameter ratio. While the classical Engh and Huber restraints work well for standard amino-acid residues, the chemical complexity of small-molecule ligands presents a particular challenge. Most current approaches either limit ligand restraints to those that can be readily described in the Crystallographic Information File (CIF) format, thus sacrificing chemical flexibility and energetic accuracy, or they employ protocols that substantially lengthen the refinement time, potentially hindering rapid automated refinement workflows. PHENIX-AFITT refinement uses a full molecular-mechanics force field for user-selected small-molecule ligands during refinement, eliminating the potentially difficult problem of finding or generating high-quality geometry restraints. It is fully integrated with a standard refinement protocol and requires practically no additional steps from the user, making it ideal for high-throughput workflows. PHENIX-AFITT refinements also handle multiple ligands in a single model, alternate conformations and covalently bound ligands. Here, the results of combining AFITT and the PHENIX software suite on a data set of 189 protein-ligand PDB structures are presented. Refinements using PHENIX-AFITT significantly reduce ligand conformational energy and lead to improved geometries without detriment to the fit to the experimental data. For the data presented, PHENIX-AFITT refinements result in more chemically accurate models for small-molecule ligands. PMID:27599738

  11. Wnt signaling and stem cell control

    Institute of Scientific and Technical Information of China (English)

    Roel Nusse

    2008-01-01

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

  12. Polypharmacology: in silico methods of ligand design and development.

    Science.gov (United States)

    McKie, Samuel A

    2016-04-01

    How to design a ligand to bind multiple targets, rather than to a single target, is the focus of this review. Rational polypharmacology draws on knowledge that is both broad ranging and hierarchical. Computer-aided multitarget ligand design methods are described according to their nested knowledge level. Ligand-only and then receptor-ligand strategies are first described; followed by the metabolic network viewpoint. Subsequently strategies that view infectious diseases as multigenomic targets are discussed, and finally the disease level interpretation of medicinal therapy is considered. As yet there is no consensus on how best to proceed in designing a multitarget ligand. The current methodologies are bought together in an attempt to give a practical overview of how polypharmacology design might be best initiated.

  13. Development of chiral sulfoxide ligands for asymmetric catalysis.

    Science.gov (United States)

    Trost, Barry M; Rao, Meera

    2015-04-20

    Nitrogen-, phosphorus-, and oxygen-based ligands with chiral backbones have been the historic workhorses of asymmetric transition-metal-catalyzed reactions. On the contrary, sulfoxides containing chirality at the sulfur atom have mainly been used as chiral auxiliaries for diastereoselective reactions. Despite several distinct advantages over traditional ligand scaffolds, such as the proximity of the chiral information to the metal center and the ability to switch between S and O coordination, these compounds have only recently emerged as a versatile class of chiral ligands. In this Review, we detail the history of the development of chiral sulfoxide ligands for asymmetric catalysis. We also provide brief descriptions of metal-sulfoxide bonding and strategies for the synthesis of enantiopure sulfoxides. Finally, insights into the future development of this underutilized ligand class are discussed.

  14. Galectin-3 is a new MerTK-specific eat-me signal

    OpenAIRE

    Caberoy, Nora B.; Alvarado, Gabriela; Bigcas, Jo-Lawrence; Li, Wei

    2012-01-01

    Phagocytosis of apoptotic cells and cellular debris is a critical process of maintaining tissue and immune homeostasis. Defects in the phagocytosis process cause autoimmunity and degenerative diseases. Phagocytosis ligands or “eat-me” signals control the initiation of the process by linking apoptotic cells to receptors on phagocyte surface and triggering signaling cascades for cargo engulfment. Eat-me signals are traditionally identified on a case-by-case basis with challenges, and the identi...

  15. Non-Canonical Hh Signaling in Cancer—Current Understanding and Future Directions

    OpenAIRE

    Dongsheng Gu; Jingwu Xie

    2015-01-01

    As a major regulatory pathway for embryonic development and tissue patterning, hedgehog signaling is not active in most adult tissues, but is reactivated in a number of human cancer types. A major milestone in hedgehog signaling in cancer is the Food and Drug Administration (FDA) approval of a smoothened inhibitor Vismodegib for treatment of basal cell carcinomas. Vismodegib can block ligand-mediated hedgehog signaling, but numerous additional clinical trials have failed to show significant i...

  16. HAMP domain conformers that propagate opposite signals in bacterial chemoreceptors.

    Directory of Open Access Journals (Sweden)

    Michael V Airola

    Full Text Available HAMP domains are signal relay modules in >26,000 receptors of bacteria, eukaryotes, and archaea that mediate processes involved in chemotaxis, pathogenesis, and biofilm formation. We identify two HAMP conformations distinguished by a four- to two-helix packing transition at the C-termini that send opposing signals in bacterial chemoreceptors. Crystal structures of signal-locked mutants establish the observed structure-to-function relationships. Pulsed dipolar electron spin resonance spectroscopy of spin-labeled soluble receptors active in cells verify that the crystallographically defined HAMP conformers are maintained in the receptors and influence the structure and activity of downstream domains accordingly. Mutation of HR2, a key residue for setting the HAMP conformation and generating an inhibitory signal, shifts HAMP structure and receptor output to an activating state. Another HR2 variant displays an inverted response with respect to ligand and demonstrates the fine energetic balance between "on" and "off" conformers. A DExG motif found in membrane proximal HAMP domains is shown to be critical for responses to extracellular ligand. Our findings directly correlate in vivo signaling with HAMP structure, stability, and dynamics to establish a comprehensive model for HAMP-mediated signal relay that consolidates existing views on how conformational signals propagate in receptors. Moreover, we have developed a rational means to manipulate HAMP structure and function that may prove useful in the engineering of bacterial taxis responses.

  17. HAMP domain conformers that propagate opposite signals in bacterial chemoreceptors.

    Science.gov (United States)

    Airola, Michael V; Sukomon, Nattakan; Samanta, Dipanjan; Borbat, Peter P; Freed, Jack H; Watts, Kylie J; Crane, Brian R

    2013-01-01

    HAMP domains are signal relay modules in >26,000 receptors of bacteria, eukaryotes, and archaea that mediate processes involved in chemotaxis, pathogenesis, and biofilm formation. We identify two HAMP conformations distinguished by a four- to two-helix packing transition at the C-termini that send opposing signals in bacterial chemoreceptors. Crystal structures of signal-locked mutants establish the observed structure-to-function relationships. Pulsed dipolar electron spin resonance spectroscopy of spin-labeled soluble receptors active in cells verify that the crystallographically defined HAMP conformers are maintained in the receptors and influence the structure and activity of downstream domains accordingly. Mutation of HR2, a key residue for setting the HAMP conformation and generating an inhibitory signal, shifts HAMP structure and receptor output to an activating state. Another HR2 variant displays an inverted response with respect to ligand and demonstrates the fine energetic balance between "on" and "off" conformers. A DExG motif found in membrane proximal HAMP domains is shown to be critical for responses to extracellular ligand. Our findings directly correlate in vivo signaling with HAMP structure, stability, and dynamics to establish a comprehensive model for HAMP-mediated signal relay that consolidates existing views on how conformational signals propagate in receptors. Moreover, we have developed a rational means to manipulate HAMP structure and function that may prove useful in the engineering of bacterial taxis responses.

  18. Strategies for exploring TGF-β signaling in Drosophila.

    Science.gov (United States)

    Peterson, Aidan J; O'Connor, Michael B

    2014-06-15

    The TGF-β pathway is an evolutionarily conserved signal transduction module that mediates diverse biological processes in animals. In Drosophila, both the BMP and Activin branches are required for viability. Studies rooted in classical and molecular genetic approaches continue to uncover new developmental roles for TGF-β signaling. We present an overview of the secreted ligands, transmembrane receptors and cellular Smad transducer proteins that compose the core pathway in Drosophila. An assortment of tools have been developed to conduct tissue-specific loss- and gain-of-function experiments for these pathway components. We discuss the deployment of these reagents, with an emphasis on appropriate usage and limitations of the available tools. Throughout, we note reagents that are in need of further improvement or development, and signaling features requiring further study. A general theme is that comparison of phenotypes for ligands, receptors, and Smads can be used to map tissue interactions, and to separate canonical and non-canonical signaling activities. Core TGF-β signaling components are subject to multiple layers of regulation, and are coupled to context-specific inputs and outputs. In addition to fleshing out how TGF-β signaling serves the fruit fly, we anticipate that future studies will uncover new regulatory nodes and modes and will continue to advance paradigms for how TGF-β signaling regulates general developmental processes.

  19. Ligand-based receptor tyrosine kinase partial agonists: New paradigm for cancer drug discovery?

    Science.gov (United States)

    Riese, David J.

    2010-01-01

    Introduction Receptor tyrosine kinases (RTKs) are validated targets for oncology drug discovery and several RTK antagonists have been approved for the treatment of human malignancies. Nonetheless, the discovery and development of RTK antagonists has lagged behind the discovery and development of agents that target G-protein coupled receptors. In part, this is because it has been difficult to discover analogs of naturally-occurring RTK agonists that function as antagonists. Areas covered Here we describe ligands of ErbB receptors that function as partial agonists for these receptors, thereby enabling these ligands to antagonize the activity of full agonists for these receptors. We provide insights into the mechanisms by which these ligands function as antagonists. We discuss how information concerning these mechanisms can be translated into screens for novel small molecule- and antibody-based antagonists of ErbB receptors and how such antagonists hold great potential as targeted cancer chemotherapeutics. Expert opinion While there have been a number of important key findings into this field, the identification of the structural basis of ligand functional specificity is still of the greatest importance. While it is true that, with some notable exceptions, peptide hormones and growth factors have not proven to be good platforms for oncology drug discovery; addressing the fundamental issues of antagonistic partial agonists for receptor tyrosine kinases has the potential to steer oncology drug discovery in new directions. Mechanism based approaches are now emerging to enable the discovery of RTK partial agonists that may antagonize both agonist-dependent and –independent RTK signaling and may hold tremendous promise as targeted cancer chemotherapeutics. PMID:21532939

  20. Arrest functions of the MIF ligand/receptor axes in atherogenesis

    Directory of Open Access Journals (Sweden)

    Sabine eTillmann

    2013-05-01

    Full Text Available Macrophage migration inhibitory factor (MIF has been defined as an important chemokine-like function (CLF chemokine with an essential role in monocyte recruitment and arrest. Adhesion of monocytes to the vessel wall and their transendothelial migration are critical in atherogenesis and many other inflammatory diseases. Chemokines carefully control all steps of the monocyte recruitment process. Those chemokines specialized in controlling arrest are typically immobilized on the endothelial surface, mediating the arrest of rolling monocytes by chemokine receptor-triggered pathways. The chemokine receptor CXCR2 functions as an important arrest receptor on monocytes. An arrest function has been revealed for the bona fide CXCR2 ligands CXCL1 and CXCL8, but genetic studies also suggested that additional arrest chemokines are likely to be involved in atherogenic leukocyte recruitment. While CXCR2 is known to interact with numerous CXC chemokine ligands, the CLF-chemokine MIF, which structurally does not belong to the CXC chemokine sub-family, was surprisingly identified as a non-cognate ligand of CXCR2, responsible for critical arrest functions during the atherogenic process. MIF was originally identified as macrophage migration inhibitory factor, but is now known as a potent inflammatory cytokine with chemokine-like functions including chemotaxis and leukocyte arrest. This review will cover the mechanisms underlying these functions, including MIF’s effects on LFA1 integrin activity and signal transduction, and will discuss the structural similarities between MIF and the bona fide CXCR2 ligand CXCL8 while emphasizing the structural differences. As MIF also interacts with CXCR4, a chemokine receptor implicated in CXCL12-elicited lymphocyte arrest, the arrest potential of the MIF/CXCR4 axis will also be scrutinized as well as the recently identified role of pericyte MIF in attracting leukocytes exiting through venules as part of the pericyte 'motility

  1. Dopamine D3 receptor ligands for drug addiction treatment: update on recent findings.

    Science.gov (United States)

    Le Foll, Bernard; Collo, Ginetta; Rabiner, Eugenii A; Boileau, Isabelle; Merlo Pich, Emilio; Sokoloff, Pierre

    2014-01-01

    The dopamine D3 receptor is located in the limbic area and apparently mediates selective effects on motivation to take drugs and drug-seeking behaviors, so that there has been considerable interest on the possible use of D3 receptor ligands to treat drug addiction. However, only recently selective tools allowing studying this receptor have been developed. This chapter presents an overview of findings that were presented at a symposium on the conference Dopamine 2013 in Sardinia in May 2013. Novel neurobiological findings indicate that drugs of abuse can lead to significant structural plasticity in rodent brain and that this is dependent on the availability of functional dopamine D3 autoreceptor, whose activation increased phosphorylation in the ERK pathway and in the Akt/mTORC1 pathway indicating the parallel engagement of a series of intracellular signaling pathways all involved in cell growth and survival. Preclinical findings using animal models of drug-seeking behaviors confirm that D3 antagonists have a promising profile to treat drug addiction across drugs of abuse type. Imaging the D3 is now feasible in human subjects. Notably, the development of (+)-4-propyl-9-hydroxynaphthoxazine ligand used in positron emission tomography (PET) studies in humans allows to measure D3 and D2 receptors based on the area of the brain under study. This PET ligand has been used to confirm up-regulation of D3 sites in psychostimulant users and to reveal that tobacco smoking produces elevation of dopamine at the level of D3 sites. There are now novel antagonists being developed, but also old drugs such as buspirone, that are available to test the D3 hypothesis in humans. The first results of clinical investigations are now being provided. Overall, those recent findings support further exploration of D3 ligands to treat drug addiction. PMID:24968784

  2. Regulation mechanisms of the FLT3-ligand after irradiation

    International Nuclear Information System (INIS)

    The hematopoietic compartment is one of the most severely damaged after chemotherapy, radiotherapy or accidental irradiations. Whatever its origin, the resulting damage to the bone marrow remains difficult to evaluate. Thus, it would be of great interest to get a biological indicator of residual hematopoiesis in order to adapt the treatment to each clinical situation. Recent results indicated that the plasma Flt3 ligand concentration was increased in patients suffering from either acquired or induced aplasia, suggesting that Flt3 ligand might be useful as a biological indicator of bone marrow status. We thus followed in a mouse model as well as in several clinical situations the variations in plasma Flt3 ligand concentration, after either homogeneous or heterogeneous irradiations. These variations were correlated to the number of hematopoietic progenitors and to other parameters such as duration and depth of pancytopenia. The results indicated that the concentration of Flt3 ligand in the blood reflects the bone marrow status, and that the follow-up of plasma Flt3 ligand concentration could give predictive information about the bone marrow function and the duration and severity of pancytopenia and thrombocytopenia. Nevertheless, the clinical use of Flt3 ligand as a biological indicator of bone marrow damage require the knowledge of the mechanisms regulating the variations in plasma Flt3 ligand concentration. We thus developed a study in the mouse model. The results indicated that the variations in plasma Flt3 ligand variations were not solely due to a balance between its production by lymphoid cells and its consumption by hematopoietic cells. Moreover, we showed that T lymphocytes are not the main regulator of plasma Flt3 ligand concentration as previously suggested, and that other cell types, possibly including bone marrow stromal cells, might be strongly implicated. These results also suggest that the Flt3 ligand is a main systemic regulator of hematopoiesis

  3. Metrical oxidation states of 2-amidophenoxide and catecholate ligands: structural signatures of metal-ligand π bonding in potentially noninnocent ligands.

    Science.gov (United States)

    Brown, Seth N

    2012-02-01

    Catecholates and 2-amidophenoxides are prototypical "noninnocent" ligands which can form metal complexes where the ligands are best described as being in the monoanionic (imino)semiquinone or neutral (imino)quinone oxidation state instead of their closed-shell dianionic form. Through a comprehensive analysis of structural data available for compounds with these ligands in unambiguous oxidation states (109 amidophenolates, 259 catecholates), the well-known structural changes in the ligands with oxidation state can be quantified. Using these correlations, an empirical "metrical oxidation state" (MOS) which gives a continuous measure of the apparent oxidation state of the ligand can be determined based on least-squares fitting of its C-C, C-O, and C-N bond lengths to this single parameter (a simple procedure for doing so is provided via a spreadsheet in the Supporting Information). High-valent d(0) metal complexes, particularly those of vanadium(V) and molybdenum(VI), have ligands with unexpectedly positive, and generally nonintegral, MOS values. The structural effects in these complexes are attributed not to electron transfer, but rather to amidophenoxide- or catecholate-to-metal π bonding, an interpretation supported by the systematic variation of the MOS values as a function of the degree of competition with the other π-donating groups in the structures. PMID:22260321

  4. SCFHOS ubiquitin ligase mediates the ligand-induced down-regulation of the interferon-α receptor

    OpenAIRE

    Suresh Kumar, K.G.; Tang, Weigang; Ravindranath, Abhilash K.; Clark, William A.; Croze, Ed; Fuchs, Serge Y.

    2003-01-01

    Down-regulation of activated signaling receptors in response to their ligands plays a key role in restricting the extent and duration of the signaling. Mechanisms underlying down-regulation of the type I interferon receptor consisting of IFNAR1 and IFNAR2 subunits remain largely unknown. Here we show that IFNAR1 interacts with the Homolog of Slimb (HOS) F-box protein in a phosphorylation-dependent manner, and that this interaction is promoted by interferon α (IFNα). IFNAR1 is ubiquitinated by...

  5. Ligand-size and ligand-chain hydrophilicity effects on the relaxometric properties of ultrasmall Gd2O3 nanoparticles

    Science.gov (United States)

    Tegafaw, Tirusew; Xu, Wenlong; Lee, Sang Hyup; Chae, Kwon Seok; Cha, Hyunsil; Chang, Yongmin; Lee, Gang Ho

    2016-06-01

    The relaxometric properties of ultrasmall Gd2O3 nanoparticles coated with various ligands were investigated. These ligands include small diacids with hydrophobic chains, namely, succinic acid (Mw = 118.09 amu), glutaric acid (Mw = 132.12 amu), and terephthalic acid (Mw = 166.13 amu), and large polyethylenimines (PEIs) with hydrophilic chains, namely, PEI-1300 ( M ¯ n = 1300 ) and PEI-10000 ( M ¯ n = 10000 ). Ligand-size and ligand-chain hydrophilicity effects were observed. The longitudinal (r1) and transverse (r2) water proton relaxivities generally decreased with increasing ligand-size (the ligand-size effect). The ligand-size effect was weaker for PEI because its hydrophilic chains allow water molecules to access the nanoparticle (the ligand-chain hydrophilicity effect). This result was explained on the basis of the magnetic dipole interaction between the dipoles of the nanoparticle and water proton. In addition, all samples were found to be non-toxic in cellular cytotoxicity tests.

  6. The ligand-receptor-G-protein ternary complex as a GTP-synthase. steady-state proton pumping and dose-response relationships for beta -adrenoceptors.

    Science.gov (United States)

    Broadley, K J; Nederkoorn, P H; Timmerman, H; Timms, D; Davies, R H

    2000-07-21

    Steady-state solutions are developed for the rate of G alpha.GTP production in a synthase model of the ligand-receptor-G-protein ternary complex activated by a ligand-receptor proton pumping mechanism. The effective rate, k(31), defining the proton transfer, phosphorylation and G alpha.GTP release is a controlling rate of the synthase in the presence of a ligand with an efficient mode of signal activation, the ligand-receptor interaction taking place under effectively equilibrium conditions. The composite rate, however, becomes an amplifying factor in any dose-response relationship. The amplification is a triple product of the rate, k(31), the equilibrium constant associated with the activation of the proton signal, K(act)and the fraction of agonist conformer transmitting the signal, f(*). Where the rate of activation of the proton signal becomes critically inefficient, the rate of activation, k(act 1)replaces k(31)K(act). A correlation between beta(1)-adrenergic receptor-stimulated GDP release and adenylate cyclase activation shows that this correlation is not unique to an exchange reaction. Within the initiating Tyr-Arg-Tyr receptor proton shuttle mechanism, the position of Arg(r156) paralleldictates the high-(R(p)) and low-(R(u)) ligand-binding affinities. These states are close to R(*)and R(0)of the equilibrium model (De Lean et al., 1980, J. Biol. Chem.255, 7108-7117). An increased rate of hydrogen ion diffusion into a receptor mutant can give rise to constitutive activity while increased rates of G-protein release and changes in receptor state balance can contribute to the resultant level of action. Constitutive action will arise from a faster rate of G-protein release alone if proton diffusion in the wild-type receptor contributes to a basal level of G-protein activation. Competitive ligand-receptor occupancy for constitutive mutants shows that, where the rate of G-protein activation from the proportion of ligand-occupied receptors is less than the

  7. Signal transmission through the CXC chemokine receptor 4 (CXCR4) transmembrane helices.

    Science.gov (United States)

    Wescott, Melanie P; Kufareva, Irina; Paes, Cheryl; Goodman, Jason R; Thaker, Yana; Puffer, Bridget A; Berdougo, Eli; Rucker, Joseph B; Handel, Tracy M; Doranz, Benjamin J

    2016-08-30

    The atomic-level mechanisms by which G protein-coupled receptors (GPCRs) transmit extracellular ligand binding events through their transmembrane helices to activate intracellular G proteins remain unclear. Using a comprehensive library of mutations covering all 352 residues of the GPCR CXC chemokine receptor 4 (CXCR4), we identified 41 amino acids that are required for signaling induced by the chemokine ligand CXCL12 (stromal cell-derived factor 1). CXCR4 variants with each of these mutations do not signal properly but remain folded, based on receptor surface trafficking, reactivity to conformationally sensitive monoclonal antibodies, and ligand binding. When visualized on the structure of CXCR4, the majority of these residues form a continuous intramolecular signaling chain through the transmembrane helices; this chain connects chemokine binding residues on the extracellular side of CXCR4 to G protein-coupling residues on its intracellular side. Integrated into a cohesive model of signal transmission, these CXCR4 residues cluster into five functional groups that mediate (i) chemokine engagement, (ii) signal initiation, (iii) signal propagation, (iv) microswitch activation, and (v) G protein coupling. Propagation of the signal passes through a "hydrophobic bridge" on helix VI that coordinates with nearly every known GPCR signaling motif. Our results agree with known conserved mechanisms of GPCR activation and significantly expand on understanding the structural principles of CXCR4 signaling. PMID:27543332

  8. PPARγ ligand production is tightly linked to clonal expansion during initiation of adipocyte differentiation

    DEFF Research Database (Denmark)

    Hallenborg, Philip; Petersen, Rasmus Koefoed; Feddersen, Søren;

    2014-01-01

    Adipocyte differentiation is orchestrated by the ligand-activated nuclear receptor PPAR. Endogenous ligands comprise oxidized derivatives of arachidonic acid and structurally similar PUFAs. Although expression of PPAR peaks in mature adipocytes, ligands are produced primarily at the onset...

  9. Gas adsorption and gas mixture separations using mixed-ligand MOF material

    Science.gov (United States)

    Hupp, Joseph T.; Mulfort, Karen L.; Snurr, Randall Q.; Bae, Youn-Sang

    2011-01-04

    A method of separating a mixture of carbon dioxiode and hydrocarbon gas using a mixed-ligand, metal-organic framework (MOF) material having metal ions coordinated to carboxylate ligands and pyridyl ligands.

  10. Identification of Target Ligands of CORYNE in Arabidopsis by Phage Display Library

    Institute of Scientific and Technical Information of China (English)

    Heng Zhao; Shuzhen Li; Jiping Sheng; Lin Shen; Yuhui Yang; Bin Yao

    2011-01-01

    CORYNE (CRN) plays important roles in stem cell division and differentiation of shoot apical meristem (SAM) in Arabidopsis thaliana. The cytoplasmic kinase domain of CRN has been cloned and expressed in Escherichia coil, and further purified by two consecutive steps of affinity chromatography. By using this purified CRN as a ligand, a 12-mer random-peptide library was used to determine the specific amino acid sequences binding with the recombinant CRN molecule. After four rounds of biopanning, positive phage clones were isolated and sequenced, and further tested by enzyme linked immunosorbent assay for their binding ability and specificity. Two positive clones that specifically bind to the intracellular protein kinase domain of CRN have been identified. Alignment of these peptides and the kinase-associated protein phosphatase (KAPP) shows high similarity, indicating that KAPP might interact with the cytoplasmic kinase domain of CRN and negatively regulate the CLV signal. Our current study would be helpful to better understand the CLV3 signal pathway.

  11. Teratogenic effects of triphenyltin on embryos of amphibian (Xenopus tropicalis): a phenotypic comparison with the retinoid X and retinoic acid receptor ligands.

    Science.gov (United States)

    Yu, Lin; Zhang, Xiaoli; Yuan, Jing; Cao, Qinzhen; Liu, Junqi; Zhu, Pan; Shi, Huahong

    2011-09-15

    Triphenyltin (TPT) has high binding affinity with the retinoid X receptor (RXR) in animals. The natural ligand of RXR, 9-cis-retinoic acid (RA), is known to induce featured malformations in vertebrate embryos by disrupting RA signal. Limited information is available on the TPT effects on amphibians. We exposed embryos of amphibian (Xenopus tropicalis) to TPT, 9-cis-RA, all-trans-RA (ligand of retinoic acid receptor, RAR), and LGD1069 (a selective ligand of RXR). The 72h LC50 of TPT was 5.25 μg Sn/L, and 72h EC50 was 0.96 μg Sn/L. TPT induced multiple malformations including enlarged proctodaeum and narrow fins. TPT at 5 μg Sn/L inhibited the differentiation of skins and muscles. The reduced brain, loss of external eyes and bent axis were observed in RXR and RAR ligands treatments. TPT and tributyltin (TBT) inhibited the mRNA expression of RXRα and increased that of TRβ. The phenotypes of malformations induced by TPT were similar to those by TBT and were much different from those by the RXR and RAR ligands. These results indicated that TPT was acute toxic and had high teratogenicity to amphibian embryos, and that TPT induced phenotypes of malformations. TPT and TBT might have a similar teratogenic mechanism, which seems not to be mainly mediated through RA signal. PMID:21820800

  12. Electronic spectra and photophysics of platinum(II) complexes with alpha-diimine ligands - Solid-state effects. I - Monomers and ligand pi dimers

    Science.gov (United States)

    Miskowski, Vincent M.; Houlding, Virginia H.

    1989-01-01

    Two types of emission behavior for Pt(II) complexes containing alpha-diimine ligands have been observed in dilute solution. If the complex also has weak field ligands such as chloride, ligand field (d-d) excited states become the lowest energy excited states. If only strong field ligands are present, a diimine 3(pi-pi/asterisk/) state becomes the lowest. In none of the cases studied did metal-to-ligand charge transfer excited state lie lowest.

  13. OSBP-Related Proteins: Liganding by Glycerophospholipids Opens New Insight into Their Function

    Directory of Open Access Journals (Sweden)

    Vesa M. Olkkonen

    2013-11-01

    Full Text Available Oxysterol-binding protein (OSBP and its homologs designated OSBP-related (ORP or OSBP-like (OSBPL proteins constitute a conserved family of lipid binding/transfer proteins (LTP in eukaryotes. The mechanisms of ORP function have remained incompletely understood, but they have been implicated as intracellular sterol sensors or transporters. A number of studies have provided evidence for the roles of ORPs at membrane contact sites (MCS, where endoplasmic reticulum is closely apposed with other organelle limiting membranes. ORPs are postulated to either transport sterols over MCSs or control the activity of enzymatic effectors or assembly of protein complexes with functions in signaling and lipid metabolism. Studies of yeast Saccharomyces cerevisiae ORPs Osh4p, Osh3p, Osh6p and Osh7p have revealed that ORPs do not exclusively bind sterols within their OSBP-related ligand-binding domain (ORD: The Osh4p ORD accommodates either sterols or phosphatidylinositol-4-phosphate (PI4P, and the Osh3p ORD was shown to specifically bind PI4P, the binding cavity being too narrow for a sterol to fit in. Most recently, Osh6p and Osh7p were demonstrated to show specific affinity for phosphatidylserine (PS, and to play a role in the intracellular transport of this glycerophospholipid; Additionally, two mammalian ORPs were shown to bind PS. Thus, the term frequently used for ORPs/OSBPLs, oxysterol-binding proteins, is a misnomer. While a number of ORPs bind oxysterols or cholesterol, other family members appear to interact with phospholipid ligands to regulate lipid fluxes, organelle lipid compositions and cell signaling. As a conclusion, ORPs are LTPs with a wide ligand spectrum and marked functional heterogeneity.

  14. OSBP-related proteins: liganding by glycerophospholipids opens new insight into their function.

    Science.gov (United States)

    Olkkonen, Vesa M

    2013-01-01

    Oxysterol-binding protein (OSBP) and its homologs designated OSBP-related (ORP) or OSBP-like (OSBPL) proteins constitute a conserved family of lipid binding/transfer proteins (LTP) in eukaryotes. The mechanisms of ORP function have remained incompletely understood, but they have been implicated as intracellular sterol sensors or transporters. A number of studies have provided evidence for the roles of ORPs at membrane contact sites (MCS), where endoplasmic reticulum is closely apposed with other organelle limiting membranes. ORPs are postulated to either transport sterols over MCSs or control the activity of enzymatic effectors or assembly of protein complexes with functions in signaling and lipid metabolism. Studies of yeast Saccharomyces cerevisiae ORPs Osh4p, Osh3p, Osh6p and Osh7p have revealed that ORPs do not exclusively bind sterols within their OSBP-related ligand-binding domain (ORD): The Osh4p ORD accommodates either sterols or phosphatidylinositol-4-phosphate (PI4P), and the Osh3p ORD was shown to specifically bind PI4P, the binding cavity being too narrow for a sterol to fit in. Most recently, Osh6p and Osh7p were demonstrated to show specific affinity for phosphatidylserine (PS), and to play a role in the intracellular transport of this glycerophospholipid; Additionally, two mammalian ORPs were shown to bind PS. Thus, the term frequently used for ORPs/OSBPLs, oxysterol-binding proteins, is a misnomer. While a number of ORPs bind oxysterols or cholesterol, other family members appear to interact with phospholipid ligands to regulate lipid fluxes, organelle lipid compositions and cell signaling. As a conclusion, ORPs are LTPs with a wide ligand spectrum and marked functional heterogeneity. PMID:24196413

  15. Regulation mechanisms of the FLT3-ligand after irradiation; Mecanismes de regulation du FLT3-ligand apres irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Prat-Lepesant, M

    2005-06-15

    The hematopoietic compartment is one of the most severely damaged after chemotherapy, radiotherapy or accidental irradiations. Whatever its origin, the resulting damage to the bone marrow remains difficult to evaluate. Thus, it would be of great interest to get a biological indicator of residual hematopoiesis in order to adapt the treatment to each clinical situation. Recent results indicated that the plasma Flt3 ligand concentration was increased in patients suffering from either acquired or induced aplasia, suggesting that Flt3 ligand might be useful as a biological indicator of bone marrow status. We thus followed in a mouse model as well as in several clinical situations the variations in plasma Flt3 ligand concentration, after either homogeneous or heterogeneous irradiations. These variations were correlated to the number of hematopoietic progenitors and to other parameters such as duration and depth of pancytopenia. The results indicated that the concentration of Flt3 ligand in the blood reflects the bone marrow status, and that the follow-up of plasma Flt3 ligand concentration could give predictive information about the bone marrow function and the duration and severity of pancytopenia and thrombocytopenia. Nevertheless, the clinical use of Flt3 ligand as a biological indicator of bone marrow damage require the knowledge of the mechanisms regulating the variations in plasma Flt3 ligand concentration. We thus developed a study in the mouse model. The results indicated that the variations in plasma Flt3 ligand variations were not solely due to a balance between its production by lymphoid cells and its consumption by hematopoietic cells. Moreover, we showed that T lymphocytes are not the main regulator of plasma Flt3 ligand concentration as previously suggested, and that other cell types, possibly including bone marrow stromal cells, might be strongly implicated. These results also suggest that the Flt3 ligand is a main systemic regulator of hematopoiesis

  16. Functional characterization of a chimeric soluble Fas ligand polymer with in vivo anti-tumor activity.

    Science.gov (United States)

    Daburon, Sophie; Devaud, Christel; Costet, Pierre; Morello, Aurore; Garrigue-Antar, Laure; Maillasson, Mike; Hargous, Nathalie; Lapaillerie, Delphine; Bonneu, Marc; Dechanet-Merville, Julie; Legembre, Patrick; Capone, Myriam; Moreau, Jean-François; Taupin, Jean-Luc

    2013-01-01

    Binding of ligand FasL to its receptor Fas triggers apoptosis via the caspase cascade. FasL itself is homotrimeric, and a productive apoptotic signal requires that FasL be oligomerized beyond the homotrimeric state. We generated a series of FasL chimeras by fusing FasL to domains of the Leukemia Inhibitory Factor receptor gp190 which confer homotypic oligomerization, and analyzed the capacity of these soluble chimeras to trigger cell death. We observed that the most efficient FasL chimera, called pFasL, was also the most polymeric, as it reached the size of a dodecamer. Using a cellular model, we investigated the structure-function relationships of the FasL/Fas interactions for our chimeras, and we demonstrated that the Fas-mediated apoptotic signal did not solely rely on ligand-mediated receptor aggregation, but also required a conformational adaptation of the Fas receptor. When injected into mice, pFasL did not trigger liver injury at a dose which displayed anti-tumor activity in a model of human tumor transplanted to immunodeficient animals, suggesting a potential therapeutic use. Therefore, the optimization of the FasL conformation has to be considered for the development of efficient FasL-derived anti-cancer drugs targeting Fas. PMID:23326557

  17. Functional characterization of a chimeric soluble Fas ligand polymer with in vivo anti-tumor activity.

    Directory of Open Access Journals (Sweden)

    Sophie Daburon

    Full Text Available Binding of ligand FasL to its receptor Fas triggers apoptosis via the caspase cascade. FasL itself is homotrimeric, and a productive apoptotic signal requires that FasL be oligomerized beyond the homotrimeric state. We generated a series of FasL chimeras by fusing FasL to domains of the Leukemia Inhibitory Factor receptor gp190 which confer homotypic oligomerization, and analyzed the capacity of these soluble chimeras to trigger cell death. We observed that the most efficient FasL chimera, called pFasL, was also the most polymeric, as it reached the size of a dodecamer. Using a cellular model, we investigated the structure-function relationships of the FasL/Fas interactions for our chimeras, and we demonstrated that the Fas-mediated apoptotic signal did not solely rely on ligand-mediated receptor aggregation, but also required a conformational adaptation of the Fas receptor. When injected into mice, pFasL did not trigger liver injury at a dose which displayed anti-tumor activity in a model of human tumor transplanted to immunodeficient animals, suggesting a potential therapeutic use. Therefore, the optimization of the FasL conformation has to be considered for the development of efficient FasL-derived anti-cancer drugs targeting Fas.

  18. Notch ligand Delta-like 1 promotes the metastasis of melanoma by enhancing tumor adhesion

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, J.P. [Department of Orthopedic Surgery, Tangdu Hospital, The Fourth Military Medical University, Xi' an (China); Li, N. [Department of Oncology, Tangdu Hospital, The Fourth Military Medical University, Xi' an (China); Bai, W.Z.; Qiu, X.C.; Ma, B.A.; Zhou, Y.; Fan, Q.Y.; Shan, L.Q. [Department of Orthopedic Surgery, Tangdu Hospital, The Fourth Military Medical University, Xi' an (China)

    2014-03-28

    Notch signaling plays a vital role in tumorigenicity and tumor progression by regulating proliferation, invasion, and the tumor microenvironment. Previous research by our group indicated that Notch ligand Delta-like 1 (Dll1) is involved in angiogenesis in melanoma, and we noticed that it took a longer time to trypsinize Dll1-expressing B16 melanoma cells than the control cells. In this article, we extended our study to investigate the effects of Dll1 on tumor cell adhesion and metastasis. Dll1 overexpression activated Notch signaling in B16 tumor cells and significantly enhanced the adhering capacity of B16 tumor cells both in vitro and in vivo. B16-Dll1 cells also had a higher metastatic potential than their counterpart in the mouse model of lung metastasis. Along with increased Dll1 expression, N-cadherin, but not E-cadherin, was upregulated in B16-Dll1 cells. These data suggested that Notch ligand Dll1 may enhance the adhesion and metastasis of melanoma cells by upregulation of N-cadherin.

  19. Signal Processing of Random Physiological Signals

    CERN Document Server

    Lessard, Charles

    2006-01-01

    Signal Processing of Random Physiological Signals presents the most widely used techniques in signal and system analysis. Specifically, the book is concerned with methods of characterizing signals and systems. Author Charles Lessard provides students and researchers an understanding of the time and frequency domain processes which may be used to evaluate random physiological signals such as brainwave, sleep, respiratory sounds, heart valve sounds, electromyograms, and electro-oculograms.Another aim of the book is to have the students evaluate actual mammalian data without spending most or all

  20. L-DOPA is an endogenous ligand for OA1.

    Directory of Open Access Journals (Sweden)

    Vanessa M Lopez

    2008-09-01

    Full Text Available Albinism is a genetic defect characterized by a loss of pigmentation. The neurosensory retina, which is not pigmented, exhibits pathologic changes secondary to the loss of pigmentation in the retina pigment epithelium (RPE. How the loss of pigmentation in the RPE causes developmental defects in the adjacent neurosensory retina has not been determined, but offers a unique opportunity to investigate the interactions between these two important tissues. One of the genes that causes albinism encodes for an orphan GPCR (OA1 expressed only in pigmented cells, including the RPE. We investigated the function and signaling of OA1 in RPE and transfected cell lines. Our results indicate that OA1 is a selective L-DOPA receptor, with no measurable second messenger activity from two closely related compounds, tyrosine and dopamine. Radiolabeled ligand binding confirmed that OA1 exhibited a single, saturable binding site for L-DOPA. Dopamine competed with L-DOPA for the single OA1 binding site, suggesting it could function as an OA1 antagonist. OA1 response to L-DOPA was defined by several common measures of G-protein coupled receptor (GPCR activation, including influx of intracellular calcium and recruitment of beta-arrestin. Further, inhibition of tyrosinase, the enzyme that makes L-DOPA, resulted in decreased PEDF secretion by RPE. Further, stimulation of OA1 in RPE with L-DOPA resulted in increased PEDF secretion. Taken together, our results illustrate an autocrine loop between OA1 and tyrosinase linked through L-DOPA, and this loop includes the secretion of at least one very potent retinal neurotrophic factor. OA1 is a selective L-DOPA receptor whose downstream effects govern spatial patterning of the developing retina. Our results suggest that the retinal consequences of albinism caused by changes in melanin synthetic machinery may be treated by L-DOPA supplementation.

  1. Characterization of methacrylate chromatographic monoliths bearing affinity ligands.

    Science.gov (United States)

    Černigoj, Urh; Vidic, Urška; Nemec, Blaž; Gašperšič, Jernej; Vidič, Jana; Lendero Krajnc, Nika; Štrancar, Aleš; Podgornik, Aleš

    2016-09-16

    We investigated effect of immobilization procedure and monolith structure on chromatographic performance of methacrylate monoliths bearing affinity ligands. Monoliths of different pore size and various affinity ligands were prepared and characterized using physical and chromatographic methods. When testing protein A monoliths with different protein A ligand densities, a significant nonlinear effect of ligand density on dynamic binding capacity (DBC) for IgG was obtained and accurately described by Langmuir isotherm curve enabling estimation of protein A utilization as a function of ligand density. Maximal IgG binding capacity was found to be at least 12mg/mL exceeding theoretical monolayer adsorption value of 7.8mg/mL assuming hexagonal packing and IgG hydrodynamic diameter of 11nm. Observed discrepancy was explained by shrinkage of IgG during adsorption on protein A experimentally determined through calculated adsorbed IgG layer thickness of 5.4nm from pressure drop data. For monoliths with different pore size maximal immobilized densities of protein A as well as IgG dynamic capacity linearly correlates with monolith surface area indicating constant ligand utilization. Finally, IgGs toward different plasma proteins were immobilized via the hydrazide coupling chemistry to provide oriented immobilization. DBC was found to be flow independent and was increasing with the size of bound protein. Despite DBC was lower than IgG capacity to immobilized protein A, ligand utilization was higher. PMID:27554023

  2. Peptide signalling in plant development: functional analysis of CLE ligands in Arabidopsis

    NARCIS (Netherlands)

    Fiers, M.A.

    2007-01-01

    In multicellular organisms, cell to cell communication is essential for coordinating growth and differentiation. In plants most known intercellular communication is mediated by phytohormones such as auxin, cytokinin, GA, ABA, ethylene and brassinosteroids. However, in recent years, several putative

  3. Sequential Notch signalling at the boundary of fringe expressing and non-expressing cells.

    Directory of Open Access Journals (Sweden)

    Tobias Troost

    Full Text Available Wing development in Drosophila requires the activation of Wingless (Wg in a small stripe along the boundary of Fringe (Fng expressing and non-expressing cells (FB, which coincides with the dorso-ventral (D/V boundary of the wing imaginal disc. The expression of Wg is induced by interactions between dorsal and ventral cells mediated by the Notch signalling pathway. It appears that mutual signalling from dorsal to ventral and ventral to dorsal cells by the Notch ligands Serrate (Ser and Delta (Dl respectively establishes a symmetric domain of Wg that straddles the D/V boundary. The directional signalling of these ligands requires the modification of Notch in dorsal cells by the glycosyltransferase Fng and is based on the restricted expression of the ligands with Ser expression to the dorsal and that of Dl to the ventral side of the wing anlage. In order to further investigate the mechanism of Notch signalling at the FB, we analysed the function of Fng, Ser and Dl during wing development at an ectopic FB and at the D/V boundary. We find that Notch signalling is initiated in an asymmetric fashion on only one side of the FB. During this initial asymmetric phase, only one ligand is required, with Ser initiating Notch-signalling at the D/V and Dl at the ectopic FB. Furthermore, our analysis suggests that Fng has also a positive effect on Ser signalling. Because of these additional properties, differential expression of the ligands, which has been a prerequisite to restrict Notch activation to the FB in the current model, is not required to restrict Notch signalling to the FB.

  4. Basic digital signal processing

    CERN Document Server

    Lockhart, Gordon B

    1985-01-01

    Basic Digital Signal Processing describes the principles of digital signal processing and experiments with BASIC programs involving the fast Fourier theorem (FFT). The book reviews the fundamentals of the BASIC program, continuous and discrete time signals including analog signals, Fourier analysis, discrete Fourier transform, signal energy, power. The text also explains digital signal processing involving digital filters, linear time-variant systems, discrete time unit impulse, discrete-time convolution, and the alternative structure for second order infinite impulse response (IIR) sections.

  5. Database of ligand-induced domain movements in enzymes

    Directory of Open Access Journals (Sweden)

    Hayward Steven

    2009-03-01

    Full Text Available Abstract Background Conformational change induced by the binding of a substrate or coenzyme is a poorly understood stage in the process of enzyme catalysed reactions. For enzymes that exhibit a domain movement, the conformational change can be clearly characterized and therefore the opportunity exists to gain an understanding of the mechanisms involved. The development of the non-redundant database of protein domain movements contains examples of ligand-induced domain movements in enzymes, but this valuable data has remained unexploited. Description The domain movements in the non-redundant database of protein domain movements are those found by applying the DynDom program to pairs of crystallographic structures contained in Protein Data Bank files. For each pair of structures cross-checking ligands in their Protein Data Bank files with the KEGG-LIGAND database and using methods that search for ligands that contact the enzyme in one conformation but not the other, the non-redundant database of protein domain movements was refined down to a set of 203 enzymes where a domain movement is apparently triggered by the binding of a functional ligand. For these cases, ligand binding information, including hydrogen bonds and salt-bridges between the ligand and specific residues on the enzyme is presented in the context of dynamical information such as the regions that form the dynamic domains, the hinge bending residues, and the hinge axes. Conclusion The presentation at a single website of data on interactions between a ligand and specific residues on the enzyme alongside data on the movement that these interactions induce, should lead to new insights into the mechanisms of these enzymes in particular, and help in trying to understand the general process of ligand-induced domain closure in enzymes. The website can be found at: http://www.cmp.uea.ac.uk/dyndom/enzymeList.do

  6. A nuclear magnetic resonance-based structural rationale for contrasting stoichiometry and ligand binding site(s) in fatty acid-binding proteins.

    Science.gov (United States)

    He, Yan; Estephan, Rima; Yang, Xiaomin; Vela, Adriana; Wang, Hsin; Bernard, Cédric; Stark, Ruth E

    2011-03-01

    Liver fatty acid-binding protein (LFABP) is a 14 kDa cytosolic polypeptide, differing from other family members in the number of ligand binding sites, the diversity of bound ligands, and the transfer of fatty acid(s) to membranes primarily via aqueous diffusion rather than direct collisional interactions. Distinct two-dimensional (1)H-(15)N nuclear magnetic resonance (NMR) signals indicative of slowly exchanging LFABP assemblies formed during stepwise ligand titration were exploited, without determining the protein-ligand complex structures, to yield the stoichiometries for the bound ligands, their locations within the protein binding cavity, the sequence of ligand occupation, and the corresponding protein structural accommodations. Chemical shifts were monitored for wild-type LFABP and an R122L/S124A mutant in which electrostatic interactions viewed as being essential to fatty acid binding were removed. For wild-type LFABP, the results compared favorably with the data for previous tertiary structures of oleate-bound wild-type LFABP in crystals and in solution: there are two oleates, one U-shaped ligand that positions the long hydrophobic chain deep within the cavity and another extended structure with the hydrophobic chain facing the cavity and the carboxylate group lying close to the protein surface. The NMR titration validated a prior hypothesis that the first oleate to enter the cavity occupies the internal protein site. In contrast, (1)H and (15)N chemical shift changes supported only one liganded oleate for R122L/S124A LFABP, at an intermediate location within the protein cavity. A rationale based on protein sequence and electrostatics was developed to explain the stoichiometry and binding site trends for LFABPs and to put these findings into context within the larger protein family. PMID:21226535

  7. Extracellular matrix component signaling in cancer

    DEFF Research Database (Denmark)

    Multhaupt, Hinke A. B.; Leitinger, Birgit; Gullberg, Donald;

    2016-01-01

    Cell responses to the extracellular matrix depend on specific signaling events. These are important from early development, through differentiation and tissue homeostasis, immune surveillance, and disease pathogenesis. Signaling not only regulates cell adhesion cytoskeletal organization and...... motility but also provides survival and proliferation cues. The major classes of cell surface receptors for matrix macromols. are the integrins, discoidin domain receptors, and transmembrane proteoglycans such as syndecans and CD44. Cells respond not only to specific ligands, such as collagen, fibronectin......, or basement membrane glycoproteins, but also in terms of matrix rigidity. This can regulate the release and subsequent biol. activity of matrix-bound growth factors, for example, transforming growth factor-β. In the environment of tumors, there may be changes in cell populations and their receptor...

  8. Ligand-based identification of environmental estrogens

    Energy Technology Data Exchange (ETDEWEB)

    Waller, C.L. [Environmental Protection Agency, Research Triangle Park, NC (United States); Oprea, T.I. [Los Alamos National Lab., NM (United States); Chae, K. [National Institute of Environmental Health Sciences, Research Triangle Park, NC (United States)] [and others

    1996-12-01

    Comparative molecular field analysis (CoMFA), a three-dimensional quantitative structure-activity relationship (3D-QSAR) paradigm, was used to examine the estrogen receptor (ER) binding affinities of a series of structurally diverse natural, synthetic, and environmental chemicals of interest. The CoMFA/3D-QSAR model is statistically robust and internally consistent, and successfully illustrates that the overall steric and electrostatic properties of structurally diverse ligands for the estrogen receptor are both necessary and sufficient to describe the binding affinity. The ability of the model to accurately predict the ER binding affinity of an external test set of molecules suggests that structure-based 3D-QSAR models may be used to supplement the process of endocrine disrupter identification through prioritization of novel compounds for bioassay. The general application of this 3D-QSAR model within a toxicological framework is, at present, limited only by the quantity and quality of biological data for relevant biomarkers of toxicity and hormonal responsiveness. 28 refs., 12 figs., 9 tabs.

  9. Niobium tetrahalide complexes with neutral diphosphine ligands.

    Science.gov (United States)

    Benjamin, Sophie L; Chang, Yao-Pang; Hector, Andrew L; Jura, Marek; Levason, William; Reid, Gillian; Stenning, Gavin

    2016-05-10

    The reactions of NbCl4 with diphosphine ligands o-C6H4(PMe2)2, Me2PCH2CH2PMe2 or Et2PCH2CH2PEt2 in a 1 : 2 molar ratio in MeCN solution produced eight-coordinate [NbCl4(diphosphine)2]. [NbBr4(diphosphine)2] (diphosphine = o-C6H4(PMe2)2 or Me2PCH2CH2PMe2) were made similarly from NbBr4. X-ray crystal structures show that [NbCl4{o-C6H4(PMe2)2}2] has a dodecahedral geometry, but the complexes with dimethylene-backboned diphosphines are distorted square antiprisms. The Nb-P distances and niobium tetrabromide, conveniently made from NbCl4 and BBr3, is a chain polymer with edge-linked NbBr6 octahedra and alternating long and short Nb-Nb distances, the latter ascribed to Nb-Nb bonds. PMID:27094082

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2007-11-09

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

  11. N-Acetylglucosamine Functions in Cell Signaling

    Directory of Open Access Journals (Sweden)

    James B. Konopka

    2012-01-01

    Full Text Available The amino sugar N-acetylglucosamine (GlcNAc is well known for the important structural roles that it plays at the cell surface. It is a key component of bacterial cell wall peptidoglycan, fungal cell wall chitin, and the extracellular matrix of animal cells. Interestingly, recent studies have also identified new roles for GlcNAc in cell signaling. For example, GlcNAc stimulates the human fungal pathogen Candida albicans to undergo changes in morphogenesis and expression of virulence genes. Pathogenic E. coli responds to GlcNAc by altering the expression of fimbriae and CURLI fibers that promote biofilm formation and GlcNAc stimulates soil bacteria to undergo changes in morphogenesis and production of antibiotics. Studies with animal cells have revealed that GlcNAc influences cell signaling through the posttranslational modification of proteins by glycosylation. O-linked attachment of GlcNAc to Ser and Thr residues regulates a variety of intracellular proteins, including transcription factors such as NFκB, c-myc, and p53. In addition, the specificity of Notch family receptors for different ligands is altered by GlcNAc attachment to fucose residues in the extracellular domain. GlcNAc also impacts signal transduction by altering the degree of branching of N-linked glycans, which influences cell surface signaling proteins. These emerging roles of GlcNAc as an activator and mediator of cellular signaling in fungi, animals, and bacteria will be the focus of this paper.

  12. Signaling via the NFκB system.

    Science.gov (United States)

    Mitchell, Simon; Vargas, Jesse; Hoffmann, Alexander

    2016-05-01

    The nuclear factor kappa B (NFκB) family of transcription factors is a key regulator of immune development, immune responses, inflammation, and cancer. The NFκB signaling system (defined by the interactions between NFκB dimers, IκB regulators, and IKK complexes) is responsive to a number of stimuli, and upon ligand-receptor engagement, distinct cellular outcomes, appropriate to the specific signal received, are set into motion. After almost three decades of study, many signaling mechanisms are well understood, rendering them amenable to mathematical modeling, which can reveal deeper insights about the regulatory design principles. While other reviews have focused on upstream, receptor proximal signaling (Hayden MS, Ghosh S. Signaling to NF-κB. Genes Dev 2004, 18:2195-2224; Verstrepen L, Bekaert T, Chau TL, Tavernier J, Chariot A, Beyaert R. TLR-4, IL-1R and TNF-R signaling to NF-κB: variations on a common theme. Cell Mol Life Sci 2008, 65:2964-2978), and advances through computational modeling (Basak S, Behar M, Hoffmann A. Lessons from mathematically modeling the NF-κB pathway. Immunol Rev 2012, 246:221-238; Williams R, Timmis J, Qwarnstrom E. Computational models of the NF-KB signalling pathway. Computation 2014, 2:131), in this review we aim to summarize the current understanding of the NFκB signaling system itself, the molecular mechanisms, and systems properties that are key to its diverse biological functions, and we discuss remaining questions in the field. WIREs Syst Biol Med 2016, 8:227-241. doi: 10.1002/wsbm.1331 For further resources related to this article, please visit the WIREs website. PMID:26990581

  13. Calcium signaling in pluripotent stem cells.

    Science.gov (United States)

    Apáti, Ágota; Pászty, Katalin; Erdei, Zsuzsa; Szebényi, Kornélia; Homolya, László; Sarkadi, Balázs

    2012-04-28

    Pluripotent stem cells represent a new source of biological material allowing the exploration of signaling phenomena during normal cell development and differentiation. Still, the calcium signaling pathways and intracellular calcium responses to various ligands or stress conditions have not been sufficiently explored as yet in embryonic or induced pluripotent stem cells and in their differentiated offspring. This is partly due to the special culturing conditions of these cell types, the rapid morphological and functional changes in heterogeneous cell populations during early differentiation, and methodological problems in cellular calcium measurements. In this paper, we review the currently available data in the literature on calcium signaling in pluripotent stem cells and discuss the potential shortcomings of these studies. Various assay methods are surveyed for obtaining reliable data both in undifferentiated embryonic stem cells and in specific, stem cell-derived human tissues. In this paper, we present the modulation of calcium signaling in human embryonic stem cells (hESC) and in their derivates; mesenchymal stem cell like (MSCl) cells and cardiac tissues using the fluorescent calcium indicator Fluo-4 and confocal microscopy. LPA, trypsin and angiotensin II were effective in inducing calcium signals both in HUES9 and MSCl cells. Histamine and thrombin induced calcium signal exclusively in the MSCl cells, while ATP was effective only in HUES9 cells. There was no calcium signal evoked by GABA, even at relatively high concentrations. In stem cell-derived cardiomyocytes a rapid increase in the beating rate and an increase of the calcium signal peaks could be observed after the addition of adrenaline, while verapamil led to a strong decrease in cellular calcium and stopped spontaneous contractions in a relaxed state.

  14. Protein carbonylation as a novel mechanism in redox signaling.

    Science.gov (United States)

    Wong, Chi Ming; Cheema, Amrita K; Zhang, Lihua; Suzuki, Yuichiro J

    2008-02-15

    Reactive oxygen species serve as second messengers for signal transduction; however, molecular targets of oxidant signaling have not been defined. Here, we show that ligand-receptor-mediated signaling promotes reactive oxygen species-dependent protein carbonylation. Treatment of pulmonary artery smooth muscle cells with endothelin-1 increased protein carbonyls. Carbonylation of the majority of proteins occurred transiently, suggesting that there is also a mechanism for decarbonylation induced by endothelin-1. Decarbonylation was suppressed by inhibition of thioredoxin reductase, and cellular thioredoxin was upregulated during the decarbonylation phase. These results indicate that endothelin-1 promotes oxidant signaling as well as thioredoxin-mediated reductive signaling to regulate carbonylation and decarbonylation mechanisms. In cells treated with endothelin receptor antagonists, hydrogen peroxide scavengers, or an iron chelator, we identified, via mass spectrometry, proteins that are carbonylated in a receptor- and Fenton reaction-dependent manner, including annexin A1, which promotes apoptosis and suppresses cell growth. Carbonylation of annexin A1 by endothelin-1 was followed by proteasome-dependent degradation of this protein. We propose that carbonylation and subsequent degradation of annexin A1 may play a role in endothelin-mediated cell growth and survival, important events in pulmonary vascular remodeling. Protein carbonylation in response to ligand-receptor interactions represents a novel mechanism in redox signaling.

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

    Science.gov (United States)

    Gutierrez-Mazariegos, Juliana; Schubert, Michael; Laudet, Vincent

    2014-01-01

    Retinoic acid (RA) is a vitamin A-derived morphogen controlling important developmental processes in vertebrates, and more generally in chordates, including axial patterning and tissue formation and differentiation. In the embryo, endogenous RA levels are controlled by RA synthesizing and degrading enzymes and the RA signal is transduced by two retinoid receptors: the retinoic acid receptor (RAR) and the retinoid X receptor (RXR). Both RAR and RXR are members of the nuclear receptor superfamily of ligand-activated transcription factors and mainly act as heterodimers to activate the transcription of target genes in the presence of their ligand, all-trans RA. This signaling pathway was long thought to be a chordate innovation, however, recent findings of gene homologs involved in RA signaling in the genomes of a wide variety of non-chordate animals, including ambulacrarians (sea urchins and acorn worms) and lophotrochozoans (annelids and mollusks), challenged this traditional view and suggested that the RA signaling pathway might have a more ancient evolutionary origin than previously thought. In this chapter, we discuss the evolutionary history of the RA signaling pathway, and more particularly of the RARs, which might have experienced independent gene losses and duplications in different animal lineages. In sum, the available data reveal novel insights into the origin of the RA signaling pathway as well as into the evolutionary history of the RARs. PMID:24962881

  16. Effect of size and conformation of the ligand on asialoglycoprotein receptor-mediated ligand internalization and degradation in rat hepatocytes

    International Nuclear Information System (INIS)

    The rates of internalization and degradation of 125-I-labeled desialylated cyanogen bromide fragment I of orosomucoid (AS-CNBr-I) and its reduced and carboxymethylated derivative (AS-RC-CNBr-I) were compared with those of 125I-labeled asialoorosomucoid (ASOR) in rat hepatocytes. At 30 nM the rates of internalization and degradation of 125I-AS-CNBr-I were greater than those of 125I-ASOR. 125I-AS-RC-CNBr-I also had a lower rate of internalization and degradation. In contrast to 125I-ASOR, when degradation was inhibited by 5 μM colchicine there was a significant intracellular accumulation of the smaller ligands. At 40C the hepatocytes were found to bind the fragmented ligands more than 125I-ASOR. Incubation of the cells with bound ligand at 370 indicated that diacytosis of 125I-ASOR was greater than the smaller ligands. Colchincine markedly enhanced diacytosis of 125I-ASOR. On the other hand, there were marked accumulation of the smaller ligands by colchicine. These results suggest that the rates of internalization, degradation and diacytosis of the ligand are affected by the size and conformation of the ligand through different rates of receptor binding and intracellular transport

  17. HybridDock: A Hybrid Protein-Ligand Docking Protocol Integrating Protein- and Ligand-Based Approaches.

    Science.gov (United States)

    Huang, Sheng-You; Li, Min; Wang, Jianxin; Pan, Yi

    2016-06-27

    Structure-based molecular docking and ligand-based similarity search are two commonly used computational methods in computer-aided drug design. Structure-based docking tries to utilize the structural information on a drug target like protein, and ligand-based screening takes advantage of the information on known ligands for a target. Given their different advantages, it would be desirable to use both protein- and ligand-based approaches in drug discovery when information for both the protein and known ligands is available. Here, we have presented a general hybrid docking protocol, referred to as HybridDock, to utilize both the protein structures and known ligands by combining the molecular docking program MDock and the ligand-based similarity search method SHAFTS, and evaluated our hybrid docking protocol on the CSAR 2013 and 2014 exercises. The results showed that overall our hybrid docking protocol significantly improved the performance in both binding affinity and binding mode predictions, compared to the sole MDock program. The efficacy of the hybrid docking protocol was further confirmed using the combination of DOCK and SHAFTS, suggesting an alternative docking approach for modern drug design/discovery. PMID:26317502

  18. Biomimetic macroporous hydrogels: protein ligand distribution and cell response to the ligand architecture in the scaffold.

    Science.gov (United States)

    Savina, Irina N; Dainiak, Maria; Jungvid, Hans; Mikhalovsky, Sergey V; Galaev, Igor Yu

    2009-01-01

    Macroporous hydrogels (MHs), cryogels, are a new type of biomaterials for tissue engineering that can be produced from any natural or synthetic polymer that forms a gel. Synthetic MHs are rendered bioactive by surface or bulk modifications with extracellular matrix components. In this study, cell response to the architecture of protein ligands, bovine type-I collagen (CG) and human fibrinogen (Fg), immobilised using different methods on poly(2-hydroxyethyl methacrylate) (pHEMA) macroporous hydrogels (MHs) was analysed. Bulk modification was performed by cross-linking cryo-co-polymerisation of HEMA and poly(ethylene glycol)diacrylate (PEGA) in the presence of proteins (CG/pHEMA and Fg/pHEMA MHs). The polymer surface was modified by covalent immobilisation of the proteins to the active epoxy (ep) groups present on pHEMA after hydrogel fabrication (CG-epHEMA and Fg-epHEMA MHs). The concentration of proteins in protein/pHEMA and protein-epHEMA MHs was 80-85 and 130-140 mug/ml hydrogel, respectively. It was demonstrated by immunostaining and confocal laser scanning microscopy that bulk modification resulted in spreading of CG in the polymer matrix and spot-like distribution of Fg. On the contrary, surface modification resulted in spot-like distribution of CG and uniform spreading of Fg, which evenly coated the surface. Proliferation rate of fibroblasts was higher on MHs with even distribution of the ligands, i.e., on Fg-epHEMA and CG/pHEMA. After 30 days of growth, fibroblasts formed several monolayers and deposited extracellular matrix filling the pores of these MHs. The best result in terms of cell proliferation was obtained on Fg-epHEMA. The ligands displayed on surface of these scaffolds were in native conformation, while in bulk-modified CG/pHEMA MHs most of the proteins were buried inside the polymer matrix and were less accessible for interactions with specific antibodies and cells. The method used for MH modification with bioligands strongly affects spatial

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

    OpenAIRE

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

    2008-01-01

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

  20. Hedgehog signaling regulates segment formation in the annelid Platynereis

    OpenAIRE

    Dray, Nicolas; Tessmar-Raible, Kristin; Le Gouar, Martine; Vibert, Laura; Christodoulou, Foteini; Schipany, Katharina; Guillou, Aurélien; Zantke, Juliane; Snyman, Heidi; Béhague, Julien; Vervoort, Michel; Arendt, Detlev; Balavoine, Guillaume

    2010-01-01

    Annelids and arthropods share a similar segmented organization of the body whose evolutionary origin remains unclear. The Hedgehog signaling pathway, prominent in arthropod embryonic segment patterning has not been shown to have a similar function outside arthropods. We show that the ligand Hedgehog, the receptor Patched, and the transcription factor Gli are all expressed in striped patterns prior to morphological segment appearance in the annelid Platynereis dumerilii. Treatments with small ...

  1. Ligand-based reactivity of a platinum bisdithiolene: double diene addition yields a new C2-chiral chelate ligand.

    Science.gov (United States)

    Kerr, Mitchell J; Harrison, Daniel J; Lough, Alan J; Fekl, Ulrich

    2009-10-01

    The reaction of Pt(tfd)(2) [tfd = S(2)C(2)(CF(3))(2)] with excess 2,3-dimethyl-1,3-butadiene initially yields the expected 1:1 adduct, in which the diene has added across two sulfur atoms on separate tfd ligands. However, within 1 day at 50 degrees C, this kinetic product quantitatively converts into a thermodynamic product where two dienes have added to one tfd ligand via unprecedented addition across the dithiolene CS bonds. The new reaction is highly selective for the C(2)-symmetric diastereomer. A new chiral bisthioether chelate ligand has formed in the product, which has been characterized crystallographically. PMID:19634863

  2. Modification of diphenylamine-linked bis(oxazoline)ligands:Tuning of electronic effect and rigidity of ligand skeleton

    Institute of Scientific and Technical Information of China (English)

    LIU Han; LI Wei; DU DaMing

    2009-01-01

    The electronic effect of diphenylamine-linked bis(oxazoline) ligands was tuned through introduction of electron-withdrawing bromo and nitro substituents onto the 4 and 4' position.The variation of the NH bond acidity was determined by the different chemical shifts of NH.The catalytic activity and enantioselectivity of the modified ligands were tested in the asymmetric FriedeI-Crafts alkylation of indole with β-nitrostyrene.The effect of iigand skeleton rigidity was also investigated through the synthesis of iminodibenzyl-linked bis(oxazoline) ligands and evaluation of their catalytic activity in Friedel-Crafts alkylation.

  3. Capacity of Diffusion-based Molecular Communication with Ligand Receptors

    CERN Document Server

    Einolghozati, Arash; Fekri, Faramarz

    2012-01-01

    A diffusion-based molecular communication system has two major components: the diffusion in the medium, and the ligand-reception. Information bits, encoded in the time variations of the concentration of molecules, are conveyed to the receiver front through the molecular diffusion in the medium. The receiver, in turn, measures the concentration of the molecules in its vicinity in order to retrieve the information. This is done via ligand-reception process. In this paper, we develop models to study the constraints imposed by the concentration sensing at the receiver side and derive the maximum rate by which a ligand-receiver can receive information. Therefore, the overall capacity of the diffusion channel with the ligand receptors can be obtained by combining the results presented in this paper with our previous work on the achievable information rate of molecular communication over the diffusion channel.

  4. Structural basis for ligand recognition of incretin receptors

    DEFF Research Database (Denmark)

    Underwood, Christina Rye; Parthier, Christoph; Reedtz-Runge, Steffen

    2010-01-01

    The glucose-dependent insulinotropic polypeptide (GIP) receptor and the glucagon-like peptide-1 (GLP-1) receptor are homologous G-protein-coupled receptors (GPCRs). Incretin receptor agonists stimulate the synthesis and secretion of insulin from pancreatic β-cells and are therefore promising agents...... for the treatment of type 2 diabetes. It is well established that the N-terminal extracellular domain (ECD) of incretin receptors is important for ligand binding and ligand specificity, whereas the transmembrane domain is involved in receptor activation. Structures of the ligand-bound ECD of incretin receptors have...... appear to be the main driving force for ligand binding to the ECD of incretin receptors. Obviously, the-still missing-structures of full-length incretin receptors are required to construct a complete picture of receptor function at the molecular level. However, the progress made recently in structural...

  5. Unique advantages of organometallic supporting ligands for uranium complexes

    Energy Technology Data Exchange (ETDEWEB)

    Diaconescu, Paula L. [Univ. of California, Los Angeles, CA (United States); Garcia, Evan [Univ. of California, Los Angeles, CA (United States)

    2014-05-31

    The objective of our research project was to study the reactivity of uranium complexes supported by ferrocene-based ligands. In addition, this research provides training of graduate students as the next generation of actinide scientists.

  6. Synthesis and Catalytic Activity of Two New Cyclic Tetraaza Ligands

    Directory of Open Access Journals (Sweden)

    Burkhard König

    2003-05-01

    Full Text Available Two new chiral cyclic tetraaza ligands were synthesized and characterized. Their catalytic activity was tested in the asymmetric addition of diethylzinc to benzaldehyde. The expected secondary alcohol was obtained in moderate yields, but with very low enantioselectivity.

  7. Steered molecular dynamics simulations of protein-ligand interactions

    Institute of Scientific and Technical Information of China (English)

    XU; Yechun; SHEN; Jianhua; LUO; Xiaomin; SHEN; Xu; CHEN; Ka

    2004-01-01

    Studies of protein-ligand interactions are helpful to elucidating the mechanisms of ligands, providing clues for rational drug design. The currently developed steered molecular dynamics (SMD) is a complementary approach to experimental techniques in investigating the biochemical processes occurring at microsecond or second time scale, thus SMD may provide dynamical and kinetic processes of ligand-receptor binding and unbinding, which cannot be accessed by the experimental methods. In this article, the methodology of SMD is described, and the applications of SMD simulations for obtaining dynamic insights into protein-ligand interactions are illustrated through two of our own examples. One is associated with the simulations of binding and unbinding processes between huperzine A and acetylcholinesterase, and the other is concerned with the unbinding process of α-APA from HIV-1 reverse transcriptase.

  8. Increased CD40 ligand in patients with acute anterior uveitis

    DEFF Research Database (Denmark)

    Øgard, Carsten; Sørensen, Torben Lykke; Krogh, Erik

    2005-01-01

    The inflammatory response in acute anterior uveitis (AU) is believed to be primarily mediated by autoreactive T-cells. We wanted to evaluate whether the T-cell activation marker CD40 ligand is involved in the AU immunopathogenesis.......The inflammatory response in acute anterior uveitis (AU) is believed to be primarily mediated by autoreactive T-cells. We wanted to evaluate whether the T-cell activation marker CD40 ligand is involved in the AU immunopathogenesis....

  9. Reversible Size Control of Silver Nanoclusters via Ligand-exchange

    KAUST Repository

    Bootharaju, Megalamane Siddaramappa

    2015-05-21

    The properties of atomically monodisperse noble metal nanoclusters (NCs) are intricately intertwined with their precise molecular formula. The vast majority of size-specific NC syntheses start from the reduction of the metal salt and thiol ligand mixture. Only in gold was it recently shown that ligand-exchange could induce the growth of NCs from one atomically precise species to another; a process of yet unknown reversibility. Here, we present a process for the ligand-exchange-induced growth of atomically precise silver NCs, in a biphasic liquid-liquid system, which is particularly of interest because of its complete reversibility and ability to occur at room temperature. We explore this phenomenon in-depth using Ag35(SG)18 [SG= glutathionate] and Ag44(4-FTP)30 [4-FTP= 4-fluorothiophenol] as model systems. We show that the ligand-exchange conversion of Ag35(SG)18 into Ag44(4-FTP)30 is rapid (< 5 min) and direct, while the reverse process proceeds slowly through intermediate cluster sizes. We adapt a recently developed theory of reverse Ostwald ripening to model the NCs’ interconvertibility. The model’s predictions are in good agreement with the experimental observations, and they highlight the importance of small changes in the ligand-metal binding energy in determining the final equilibrium NC size. Based on the insight provided by this model, we demonstrated experimentally that by varying the choice of ligands, ligand-exchange can be used to obtain different sized NCs. The findings in this work establish ligand-exchange as a versatile tool for tuning cluster sizes.

  10. Coordination chemistry of poly(thioether)borate ligands

    OpenAIRE

    Riordan, Charles G.

    2010-01-01

    This review traces the development and application of the tris(thioether)borate ligands, tripodal ligands with highly polarizable thioether donors. Areas of emphasis include the basic coordination chemistry of the mid-to-late first row transition metals (Fe, Ni, Co, Cu), and the role of the thioether substituent in directing complex formation, the modeling of zinc thiolate protein active sites, high-spin organo-iron and organo-cobalt chemistry, the preparation of monovalent complexes of Fe, C...

  11. Metallogel formation in aqueous DMSO by perfluoroalkyl decorated terpyridine ligands.

    Science.gov (United States)

    Tatikonda, Rajendhraprasad; Bhowmik, Sandip; Rissanen, Kari; Haukka, Matti; Cametti, Massimo

    2016-08-01

    Terpyridine based ligands 1 and 2, decorated with a C8F17 perfluorinated tag, are able to form stable thermoreversible gels in the presence of several d-block metal chloride salts. The gel systems obtained have been characterized by NMR, X-ray diffraction, electron microscopies and Tgel experiments in order to gain insights into the observed different behaviour of the two similar ligands, also in terms of the effect of additional common anionic species. PMID:27460754

  12. Synthesis and evaluation of potential ligands for nuclear waste processing

    OpenAIRE

    Iqbal, M.

    2012-01-01

    The research presented in this thesis deals with the synthesis and evaluation of new potential ligands for the complexation of actinide and lanthanide ions either for their extraction from bulk radioactive waste or their stripping from an extracted organic phase for final processing of the waste. In particular, the aim of the work described here is the development of new ligands with improved separation and extraction efficiency. Separation of actinides (An) and lanthanides (Ln) is a challeng...

  13. Heterobifunctional PEG Ligands for Bioconjugation Reactions on Iron Oxide Nanoparticles

    OpenAIRE

    Maarten Bloemen; Thomas Van Stappen; Pieter Willot; Jeroen Lammertyn; Guy Koeckelberghs; Nick Geukens; Ann Gils; Thierry Verbiest

    2014-01-01

    Ever since iron oxide nanoparticles have been recognized as promising scaffolds for biomedical applications, their surface functionalization has become even more important. We report the synthesis of a novel polyethylene glycol-based ligand that combines multiple advantageous properties for these applications. The ligand is covalently bound to the surface via a siloxane group, while its polyethylene glycol backbone significantly improves the colloidal stability of the particle in complex envi...

  14. Analysis of cell locomotion on ligand gradient substrates.

    Science.gov (United States)

    Sarvestani, Alireza S; Jabbari, Esmaiel

    2009-06-01

    Directional cell motility plays a key role in many biological processes like morphogenesis, inflammation, wound repair, angiogenesis, immune response, and tumor metastasis. Cells respond to the gradient in surface ligand density by directed locomotion towards the direction of higher ligand density. Theoretical models which address the physical basis underlying the regulatory effect of ligand gradient on cell motility are highly desirable. Predictive models not only contribute to a better understanding of biological processes, but they also provide a quantitative interconnection between cell motility and biophysical properties of the extracellular matrix (ECM) for rational design of biomaterials as scaffolds in tissue engineering. In this work, we consider a one-dimensional (1D) continuum viscoelastic model to predict the cell velocity in response to linearly increasing density of surface ligands on a substrate. The cell is considered as a 1D linear viscoelastic object with position dependent elasticity due to the variation in actin network density. The cell-substrate interaction is characterized by a frictional force, controlled by the density of ligand-receptor pairs. The generation of contractile stresses is described in terms of kinetic equations for the reactions between actins, myosins, and guanine nucleotide regulatory proteins. The model predictions show a reasonable agreement with experimentally measured cell speeds, considering biologically relevant values for the model parameters. The model predicts a biphasic relationship between cell speed and slope of gradient as well as a maximum limiting speed after a finite migration time. For a given slope of ligand gradient, the onset of the limiting speed appears at longer times for substrates with lower ligand gradients. The model can be applied to the design of biomaterials as scaffolds for guided tissue regeneration as it predicts an optimum range for the slope of ligand gradient. PMID:19205048

  15. Delivering carbide ligands to sulfide-rich clusters.

    Science.gov (United States)

    Reinholdt, Anders; Herbst, Konrad; Bendix, Jesper

    2016-02-01

    The propensity of the terminal ruthenium carbide Ru(C)Cl2(PCy3)2 (RuC) to form carbide bridges to electron-rich transition metals enables synthetic routes to metal clusters with coexisting carbide and sulfide ligands. Electrochemical experiments show the Ru≡C ligand to exert a relatively large electron-withdrawing effect compared with PPh3, effectively shifting redox potentials.

  16. Tetrapyrroles as Endogenous TSPO Ligands in Eukaryotes and Prokaryotes: Comparisons with Synthetic Ligands

    Science.gov (United States)

    Veenman, Leo; Vainshtein, Alex; Yasin, Nasra; Azrad, Maya; Gavish, Moshe

    2016-01-01

    The 18 kDa translocator protein (TSPO) is highly 0conserved in eukaryotes and prokaryotes. Since its discovery in 1977, numerous studies established the TSPO’s importance for life essential functions. For these studies, synthetic TSPO ligands typically are applied. Tetrapyrroles present endogenous ligands for the TSPO. Tetrapyrroles are also evolutionarily conserved and regulate multiple functions. TSPO and tetrapyrroles regulate each other. In animals TSPO-tetrapyrrole interactions range from effects on embryonic development to metabolism, programmed cell death, response to stress, injury and disease, and even to life span extension. In animals TSPOs are primarily located in mitochondria. In plants TSPOs are also present in plastids, the nuclear fraction, the endoplasmic reticulum, and Golgi stacks. This may contribute to translocation of tetrapyrrole intermediates across organelles’ membranes. As in animals, plant TSPO binds heme and protoporphyrin IX. TSPO-tetrapyrrole interactions in plants appear to relate to development as well as stress conditions, including salt tolerance, abscisic acid-induced stress, reactive oxygen species homeostasis, and finally cell death regulation. In bacteria, TSPO is important for switching from aerobic to anaerobic metabolism, including the regulation of photosynthesis. As in mitochondria, in bacteria TSPO is located in the outer membrane. TSPO-tetrapyrrole interactions may be part of the establishment of the bacterial-eukaryote relationships, i.e., mitochondrial-eukaryote and plastid-plant endosymbiotic relationships. PMID:27271616

  17. Tetrapyrroles as Endogenous TSPO Ligands in Eukaryotes and Prokaryotes: Comparisons with Synthetic Ligands

    Directory of Open Access Journals (Sweden)

    Leo Veenman

    2016-06-01

    Full Text Available The 18 kDa translocator protein (TSPO is highly 0conserved in eukaryotes and prokaryotes. Since its discovery in 1977, numerous studies established the TSPO’s importance for life essential functions. For these studies, synthetic TSPO ligands typically are applied. Tetrapyrroles present endogenous ligands for the TSPO. Tetrapyrroles are also evolutionarily conserved and regulate multiple functions. TSPO and tetrapyrroles regulate each other. In animals TSPO-tetrapyrrole interactions range from effects on embryonic development to metabolism, programmed cell death, response to stress, injury and disease, and even to life span extension. In animals TSPOs are primarily located in mitochondria. In plants TSPOs are also present in plastids, the nuclear fraction, the endoplasmic reticulum, and Golgi stacks. This may contribute to translocation of tetrapyrrole intermediates across organelles’ membranes. As in animals, plant TSPO binds heme and protoporphyrin IX. TSPO-tetrapyrrole interactions in plants appear to relate to development as well as stress conditions, including salt tolerance, abscisic acid-induced stress, reactive oxygen species homeostasis, and finally cell death regulation. In bacteria, TSPO is important for switching from aerobic to anaerobic metabolism, including the regulation of photosynthesis. As in mitochondria, in bacteria TSPO is located in the outer membrane. TSPO-tetrapyrrole interactions may be part of the establishment of the bacterial-eukaryote relationships, i.e., mitochondrial-eukaryote and plastid-plant endosymbiotic relationships.

  18. Tetrapyrroles as Endogenous TSPO Ligands in Eukaryotes and Prokaryotes: Comparisons with Synthetic Ligands.

    Science.gov (United States)

    Veenman, Leo; Vainshtein, Alex; Yasin, Nasra; Azrad, Maya; Gavish, Moshe

    2016-01-01

    The 18 kDa translocator protein (TSPO) is highly 0conserved in eukaryotes and prokaryotes. Since its discovery in 1977, numerous studies established the TSPO's importance for life essential functions. For these studies, synthetic TSPO ligands typically are applied. Tetrapyrroles present endogenous ligands for the TSPO. Tetrapyrroles are also evolutionarily conserved and regulate multiple functions. TSPO and tetrapyrroles regulate each other. In animals TSPO-tetrapyrrole interactions range from effects on embryonic development to metabolism, programmed cell death, response to stress, injury and disease, and even to life span extension. In animals TSPOs are primarily located in mitochondria. In plants TSPOs are also present in plastids, the nuclear fraction, the endoplasmic reticulum, and Golgi stacks. This may contribute to translocation of tetrapyrrole intermediates across organelles' membranes. As in animals, plant TSPO binds heme and protoporphyrin IX. TSPO-tetrapyrrole interactions in plants appear to relate to development as well as stress conditions, including salt tolerance, abscisic acid-induced stress, reactive oxygen species homeostasis, and finally cell death regulation. In bacteria, TSPO is important for switching from aerobic to anaerobic metabolism, including the regulation of photosynthesis. As in mitochondria, in bacteria TSPO is located in the outer membrane. TSPO-tetrapyrrole interactions may be part of the establishment of the bacterial-eukaryote relationships, i.e., mitochondrial-eukaryote and plastid-plant endosymbiotic relationships.

  19. Tetrapyrroles as Endogenous TSPO Ligands in Eukaryotes and Prokaryotes: Comparisons with Synthetic Ligands.

    Science.gov (United States)

    Veenman, Leo; Vainshtein, Alex; Yasin, Nasra; Azrad, Maya; Gavish, Moshe

    2016-01-01

    The 18 kDa translocator protein (TSPO) is highly 0conserved in eukaryotes and prokaryotes. Since its discovery in 1977, numerous studies established the TSPO's importance for life essential functions. For these studies, synthetic TSPO ligands typically are applied. Tetrapyrroles present endogenous ligands for the TSPO. Tetrapyrroles are also evolutionarily conserved and regulate multiple functions. TSPO and tetrapyrroles regulate each other. In animals TSPO-tetrapyrrole interactions range from effects on embryonic development to metabolism, programmed cell death, response to stress, injury and disease, and even to life span extension. In animals TSPOs are primarily located in mitochondria. In plants TSPOs are also present in plastids, the nuclear fraction, the endoplasmic reticulum, and Golgi stacks. This may contribute to translocation of tetrapyrrole intermediates across organelles' membranes. As in animals, plant TSPO binds heme and protoporphyrin IX. TSPO-tetrapyrrole interactions in plants appear to relate to development as well as stress conditions, including salt tolerance, abscisic acid-induced stress, reactive oxygen species homeostasis, and finally cell death regulation. In bacteria, TSPO is important for switching from aerobic to anaerobic metabolism, including the regulation of photosynthesis. As in mitochondria, in bacteria TSPO is located in the outer membrane. TSPO-tetrapyrrole interactions may be part of the establishment of the bacterial-eukaryote relationships, i.e., mitochondrial-eukaryote and plastid-plant endosymbiotic relationships. PMID:27271616

  20. Digital signal processing laboratory

    CERN Document Server

    Kumar, B Preetham

    2011-01-01

    INTRODUCTION TO DIGITAL SIGNAL PROCESSING Brief Theory of DSP ConceptsProblem SolvingComputer Laboratory: Introduction to MATLAB®/SIMULINK®Hardware Laboratory: Working with Oscilloscopes, Spectrum Analyzers, Signal SourcesDigital Signal Processors (DSPs)ReferencesDISCRETE-TIME LTI SIGNALS AND SYSTEMS Brief Theory of Discrete-Time Signals and SystemsProblem SolvingComputer Laboratory: Simulation of Continuous Time and Discrete-Time Signals and Systems ReferencesTIME AND FREQUENCY ANALYSIS OF COMMUNICATION SIGNALS Brief Theory of Discrete-Time Fourier Transform (DTFT), Discrete Fourier Transform

  1. Multiplexing oscillatory biochemical signals.

    Science.gov (United States)

    de Ronde, Wiet; ten Wolde, Pieter Rein

    2014-04-01

    In recent years it has been increasingly recognized that biochemical signals are not necessarily constant in time and that the temporal dynamics of a signal can be the information carrier. Moreover, it is now well established that the protein signaling network of living cells has a bow-tie structure and that components are often shared between different signaling pathways. Here we show by mathematical modeling that living cells can multiplex a constant and an oscillatory signal: they can transmit these two signals simultaneously through a common signaling pathway, and yet respond to them specifically and reliably. We find that information transmission is reduced not only by noise arising from the intrinsic stochasticity of biochemical reactions, but also by crosstalk between the different channels. Yet, under biologically relevant conditions more than 2 bits of information can be transmitted per channel, even when the two signals are transmitted simultaneously. These observations suggest that oscillatory signals are ideal for multiplexing signals. PMID:24685537

  2. NMU signaling promotes endometrial cancer cell progression by modulating adhesion signaling.

    Science.gov (United States)

    Lin, Ting-Yu; Wu, Fang-Ju; Chang, Chia-Lin; Li, Zhongyou; Luo, Ching-Wei

    2016-03-01

    Neuromedin U (NMU) was originally named based on its strong uterine contractile activity, but little is known regarding its signaling/functions in utero. We identified that NMU and one of its receptors, NMUR2, are not only present in normal uterine endometrium but also co-expressed in endometrial cancer tissues, where the NMU level is correlated with the malignant grades and survival of patients. Cell-based assays further confirmed that NMU signaling can promote cell motility and proliferation of endometrial cancer cells derived from grade II tumors. Activation of NMU pathway in these endometrial cancer cells is required in order to sustain expression of various adhesion molecules, such as CD44 and integrin alpha1, as well as production of their corresponding extracellular matrix ligands, hyaluronan and collagen IV; it also increased the activity of SRC and its downstream proteins RHOA and RAC1. Thus, it is concluded that NMU pathway positively controls the adhesion signaling-SRC-Rho GTPase axis in the tested endometrial cancer cells and that changes in cell motility and proliferation can occur when there is manipulation of NMU signaling in these cells either in vitro or in vivo. Intriguingly, this novel mechanism also explains how NMU signaling promotes the EGFR-driven and TGFβ receptor-driven mesenchymal transitions. Through the above axis, NMU signaling not only can promote malignancy of the tested endometrial cancer cells directly, but also helps these cells to become more sensitive to niche growth factors in their microenvironment. PMID:26849234

  3. Serum albumin ligand binding volumes using high pressure denaturation

    International Nuclear Information System (INIS)

    Highlights: ► We use pressure shift assay to study the thermodynamics of decanoate and dodecanoate ligand binding to human serum albumin. ► Pressure shift assay provides information on ligand binding volumes. ► The ligands stabilized human serum albumin against both pressure and temperature denaturation. ► ANS is a strong human serum albumin stabilizer and competes with lipids for the same binding sites. - Abstract: The pressure shift assay (PSA, also termed either PressureFluor or differential pressure fluorimetry) was used to study the thermodynamics of decanoate and dodecanoate lipid binding to human serum albumin (HSA) in the temperature range from 25 °C to 80 °C and the pressure range from 0.1 MPa to 400 MPa. The ligands stabilized HSA against both pressure and temperature denaturation. The P–T phase diagram for HSA bound to saturated fatty acids is shown. Pressure induced HSA denaturation reversibility is demonstrated via either intrinsic tryptophan or extrinsic probe 1,8-anilinonaphthalene sulfonate (ANS) fluorescence. The effect of guanidinium in a PSA was studied. PSA provides information on ligand binding volumes. The volume changes from protein–ligand binding are thermodynamically important and could be used in designing compounds with specific volumetric binding properties.

  4. Predicting Efficient Antenna Ligands for Tb(III) Emission

    Energy Technology Data Exchange (ETDEWEB)

    Samuel, Amanda P.S.; Xu, Jide; Raymond, Kenneth

    2008-10-06

    A series of highly luminescent Tb(III) complexes of para-substituted 2-hydroxyisophthalamide ligands (5LI-IAM-X) has been prepared (X = H, CH{sub 3}, (C=O)NHCH{sub 3}, SO{sub 3}{sup -}, NO{sub 2}, OCH{sub 3}, F, Cl, Br) to probe the effect of substituting the isophthalamide ring on ligand and Tb(III) emission in order to establish a method for predicting the effects of chromophore modification on Tb(III) luminescence. The energies of the ligand singlet and triplet excited states are found to increase linearly with the {pi}-withdrawing ability of the substituent. The experimental results are supported by time-dependent density functional theory (TD-DFT) calculations performed on model systems, which predict ligand singlet and triplet energies within {approx}5% of the experimental values. The quantum yield ({Phi}) values of the Tb(III) complex increases with the triplet energy of the ligand, which is in part due to the decreased non-radiative deactivation caused by thermal repopulation of the triplet. Together, the experimental and theoretical results serve as a predictive tool that can be used to guide the synthesis of ligands used to sensitize lanthanide luminescence.

  5. Porphyrin-based design of bioinspired multitarget quadruplex ligands.

    Science.gov (United States)

    Laguerre, Aurélien; Desbois, Nicolas; Stefan, Loic; Richard, Philippe; Gros, Claude P; Monchaud, David

    2014-09-01

    Secondary nucleic acid structures, such as DNA and RNA quadruplexes, are potential targets for cancer therapies. Ligands that interact with these targets could thus find application as anticancer agents. Synthetic G-quartets have recently found numerous applications, including use as bioinspired G-quadruplex ligands. Herein, the design, synthesis and preliminary biophysical evaluation of a new prototype multitarget G-quadruplex ligand, (PNA)PorphySQ, are reported, where peptidic nucleic acid guanine ((PNA)G) was incorporated in the porphyrin-templated synthetic G-quartet (PorphySQ). Using fluorescence resonance energy transfer (FRET)-melting experiments, PorphySQ was shown to possess enhanced quadruplex-interacting properties thanks to the presence of four positively charged (PNA)G residues that improve its electrostatic interactions with the binding site of both DNA and RNA quadruplexes (i.e., their negatively charged and accessible G-quartets), thereby making (PNA)PorphySQ an interesting prototype of a multitarget ligand. Both the chemical stability and water solubility of (PNA)PorphySQ are improved over the non-PNA derivative (PorphySQ), which are desirable properties for drug development, and while improvements remain to be made, this ligand is a promising lead for the further development of multitarget G-quadruplex ligands. PMID:24678052

  6. Mitochondrial DNA from hepatocytes as a ligand for TLR9: Drivers of nonalcoholic steatohepatitis?

    Science.gov (United States)

    Handa, Priya; Vemulakonda, Akhila; Kowdley, Kris V; Uribe, Misael; Méndez-Sánchez, Nahum

    2016-08-21

    Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide, affecting approximately one third of the Western world. It consists of a wide spectrum of liver disorders, ranging from fatty liver to nonalcoholic steatohepatitis (NASH), which consists of steatosis, ballooning injury and inflammation. Despite an alarming growth in the statistics surrounding NAFLD, there are as yet no effective therapies for its treatment. Innate immune signaling has been thought to play a significant role in initiating and augmenting hepatic inflammation, contributing to the transition from nonalcoholic fatty liver to NASH. An immune response is triggered by countless signals called damage-associated molecular patterns (DAMPs) elicited by lipid-laden and damaged hepatocytes, which are recognized by pattern recognition receptors (PRRs) on hepatic immune cells to initiate inflammatory signaling. In this editorial, in addition to summarizing innate immune signaling in NAFLD and discussing potential therapies that target innate immune pathways, we have described a recent study that demonstrated that mitochondrial DNA serves as a DAMP activating a hepatic PRR, TLR9, in mice and in the plasma of NASH patients. In addition to identifying a new ligand for TLR9 during NASH progression, the study shows that blocking TLR9 reverses NASH, paving the way for the development of future NASH therapy. PMID:27610009

  7. Structural Basis of Ligand Binding by a C-di-GMP Riboswitch

    Energy Technology Data Exchange (ETDEWEB)

    Smith, K.; Lipchock, S; Ames, T; Wang, J; Breaker, R; Strobel, S

    2009-01-01

    The second messenger signaling molecule bis-(3{prime}-5{prime})-cyclic dimeric guanosine monophosphate (c-di-GMP) regulates many processes in bacteria, including motility, pathogenesis and biofilm formation. c-di-GMP-binding riboswitches are important downstream targets in this signaling pathway. Here we report the crystal structure, at 2.7 {angstrom} resolution, of a c-di-GMP riboswitch aptamer from Vibrio cholerae bound to c-di-GMP, showing that the ligand binds within a three-helix junction that involves base-pairing and extensive base-stacking. The symmetric c-di-GMP is recognized asymmetrically with respect to both the bases and the backbone. A mutant aptamer was engineered that preferentially binds the candidate signaling molecule c-di-AMP over c-di-GMP. Kinetic and structural data suggest that genetic regulation by the c-di-GMP riboswitch is kinetically controlled and that gene expression is modulated through the stabilization of a previously unidentified P1 helix, illustrating a direct mechanism for c-di-GMP signaling.

  8. Structural basis of ligand binding by a c-di-GMP riboswitch.

    Science.gov (United States)

    Smith, Kathryn D; Lipchock, Sarah V; Ames, Tyler D; Wang, Jimin; Breaker, Ronald R; Strobel, Scott A

    2009-12-01

    The second messenger signaling molecule bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP) regulates many processes in bacteria, including motility, pathogenesis and biofilm formation. c-di-GMP-binding riboswitches are important downstream targets in this signaling pathway. Here we report the crystal structure, at 2.7 A resolution, of a c-di-GMP riboswitch aptamer from Vibrio cholerae bound to c-di-GMP, showing that the ligand binds within a three-helix junction that involves base-pairing and extensive base-stacking. The symmetric c-di-GMP is recognized asymmetrically with respect to both the bases and the backbone. A mutant aptamer was engineered that preferentially binds the candidate signaling molecule c-di-AMP over c-di-GMP. Kinetic and structural data suggest that genetic regulation by the c-di-GMP riboswitch is kinetically controlled and that gene expression is modulated through the stabilization of a previously unidentified P1 helix, illustrating a direct mechanism for c-di-GMP signaling. PMID:19898477

  9. TLR2 ligands induce NF-κB activation from endosomal compartments of human monocytes.

    Directory of Open Access Journals (Sweden)

    Karim J Brandt

    Full Text Available Localization of Toll-like receptors (TLR in subcellular organelles is a major strategy to regulate innate immune responses. While TLR4, a cell-surface receptor, signals from both the plasma membrane and endosomal compartments, less is known about the functional role of endosomal trafficking upon TLR2 signaling. Here we show that the bacterial TLR2 ligands Pam3CSK4 and LTA activate NF-κB-dependent signaling from endosomal compartments in human monocytes and in a NF-κB sensitive reporter cell line, despite the expression of TLR2 at the cell surface. Further analyses indicate that TLR2-induced NF-κB activation is controlled by a clathrin/dynamin-dependent endocytosis mechanism, in which CD14 serves as an important upstream regulator. These findings establish that internalization of cell-surface TLR2 into endosomal compartments is required for NF-κB activation. These observations further demonstrate the need of endocytosis in the activation and regulation of TLR2-dependent signaling pathways.

  10. Robust detection of peak signals for lateral flow immunoassays

    Science.gov (United States)

    Kim, Jongwon; Kim, Jong Dae; Nahm, Kie Bong; Choi, Eui Yul; Lee, Geumyoung

    2011-02-01

    Template matching method is presented to identify the peaks from the scanned signals of lateral flow immunoassay strips. The template is composed of two pulses separated by the distance of the control and the target ligand line in the assay, and is convolved with the scanned signal to deliver the maximum at the center of the two peaks. The peak regions were identified with the predefined distances from the center. Glycosylated haemoglobin immunoassay strips and fluorescent strip readers from Boditechmed Inc. were tested to estimate the lot and reader variations of the concentration measurands. The results showed the robustness of the propose method.

  11. Ligands turning around in the midst of protein conformers: the origin of ligand-protein mating. A NMR view.

    Science.gov (United States)

    Pertinhez, T A; Spisni, A

    2011-01-01

    Protein-ligand binding is a puzzling process. Many theories have been devised since the pioneering key-and-lock hypothesis based on the idea that both the protein and the ligand have a rigid single conformation. Indeed, molecular motion is the essence of the universe. Consequently, not only proteins are characterized by an extraordinary conformational freedom, but ligands too can fluctuate in a rather vast conformational space. In this scenario, the quest to understand how do they match is fascinating. Recognizing that the inherent dynamics of molecules is the key factor controlling the success of the binding and, subsequently, their chemical/biological function, here we present a view of this process from the NMR stand point. A description of the most relevant NMR parameters that can provide insights, at atomic level, on the mechanisms of protein-ligand binding is provided in the final section. PMID:20939791

  12. A Ferrocene-Based Catecholamide Ligand: the Consequences of Ligand Swivel for Directed Supramolecular Self-Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Mugridge, Jeffrey; Fiedler, Dorothea; Raymond, Kenneth

    2010-02-04

    A ferrocene-based biscatecholamide ligand was prepared and investigated for the formation of metal-ligand supramolecular assemblies with different metals. Reaction with Ge(IV) resulted in the formation of a variety of Ge{sub n}L{sub m} coordination complexes, including [Ge{sub 2}L{sub 3}]{sup 4-} and [Ge{sub 2}L{sub 2}({mu}-OMe){sub 2}]{sup 2-}. The ligand's ability to swivel about the ferrocenyl linker and adopt different conformations accounts for formation of many different Ge{sub n}L{sub m} species. This study demonstrates why conformational ligand rigidity is essential in the rational design and directed self-assembly of supramolecular complexes.

  13. The Notch ligand JAG1 is required for sensory progenitor development in the mammalian inner ear.

    Directory of Open Access Journals (Sweden)

    Amy E Kiernan

    2006-01-01

    Full Text Available In mammals, six separate sensory regions in the inner ear are essential for hearing and balance function. Each sensory region is made up of hair cells, which are the sensory cells, and their associated supporting cells, both arising from a common progenitor. Little is known about the molecular mechanisms that govern the development of these sensory organs. Notch signaling plays a pivotal role in the differentiation of hair cells and supporting cells by mediating lateral inhibition via the ligands Delta-like 1 and Jagged (JAG 2. However, another Notch ligand, JAG1, is expressed early in the sensory patches prior to cell differentiation, indicating that there may be an earlier role for Notch signaling in sensory development in the ear. Here, using conditional gene targeting, we show that the Jag1 gene is required for the normal development of all six sensory organs within the inner ear. Cristae are completely lacking in Jag1-conditional knockout (cko mutant inner ears, whereas the cochlea and utricle show partial sensory development. The saccular macula is present but malformed. Using SOX2 and p27kip1 as molecular markers of the prosensory domain, we show that JAG1 is initially expressed in all the prosensory regions of the ear, but becomes down-regulated in the nascent organ of Corti by embryonic day 14.5, when the cells exit the cell cycle and differentiate. We also show that both SOX2 and p27kip1 are down-regulated in Jag1-cko inner ears. Taken together, these data demonstrate that JAG1 is expressed early in the prosensory domains of both the cochlear and vestibular regions, and is required to maintain the normal expression levels of both SOX2 and p27kip1. These data demonstrate that JAG1-mediated Notch signaling is essential during early development for establishing the prosensory regions of the inner ear.

  14. O-glycans direct selectin ligands to lipid rafts on leukocytes.

    Science.gov (United States)

    Shao, Bojing; Yago, Tadayuki; Setiadi, Hendra; Wang, Ying; Mehta-D'souza, Padmaja; Fu, Jianxin; Crocker, Paul R; Rodgers, William; Xia, Lijun; McEver, Rodger P

    2015-07-14

    Palmitoylated cysteines typically target transmembrane proteins to domains enriched in cholesterol and sphingolipids (lipid rafts). P-selectin glycoprotein ligand-1 (PSGL-1), CD43, and CD44 are O-glycosylated proteins on leukocytes that associate with lipid rafts. During inflammation, they transduce signals by engaging selectins as leukocytes roll in venules, and they move to the raft-enriched uropods of polarized cells upon chemokine stimulation. It is not known how these glycoproteins associate with lipid rafts or whether this association is required for signaling or for translocation to uropods. Here, we found that loss of core 1-derived O-glycans in murine C1galt1(-/-) neutrophils blocked raft targeting of PSGL-1, CD43, and CD44, but not of other glycosylated proteins, as measured by resistance to solubilization in nonionic detergent and by copatching with a raft-resident sphingolipid on intact cells. Neuraminidase removal of sialic acids from wild-type neutrophils also blocked raft targeting. C1galt1(-/-) neutrophils or neuraminidase-treated neutrophils failed to activate tyrosine kinases when plated on immobilized anti-PSGL-1 or anti-CD44 F(ab')2. Furthermore, C1galt1(-/-) neutrophils incubated with anti-PSGL-1 F(ab')2 did not generate microparticles. In marked contrast, PSGL-1, CD43, and CD44 moved normally to the uropods of chemokine-stimulated C1galt1(-/-) neutrophils. These data define a role for core 1-derived O-glycans and terminal sialic acids in targeting glycoprotein ligands for selectins to lipid rafts of leukocytes. Preassociation of these glycoproteins with rafts is required for signaling but not for movement to uropods.

  15. Ligand-specific sequential regulation of transcription factors for differentiation of MCF-7 cells

    Directory of Open Access Journals (Sweden)

    Toyoda Tetsuro

    2009-11-01

    Full Text Available Abstract Background Sharing a common ErbB/HER receptor signaling pathway, heregulin (HRG induces differentiation of MCF-7 human breast cancer cells while epidermal growth factor (EGF elicits proliferation. Although cell fates resulting from action of the aforementioned ligands completely different, the respective gene expression profiles in early transcription are qualitatively similar, suggesting that gene expression during late transcription, but not early transcription, may reflect ligand specificity. In this study, based on both the data from time-course quantitative real-time PCR on over 2,000 human transcription factors and microarray of all human genes, we identified a series of transcription factors which may control HRG-specific late transcription in MCF-7 cells. Results We predicted that four transcription factors including EGR4, FRA-1, FHL2, and DIPA should have responsibility of regulation in MCF-7 cell differentiation. Validation analysis suggested that one member of the activator protein 1 (AP-1 family, FOSL-1 (FRA-1 gene, appeared immediately following c-FOS expression, might be responsible for expression of transcription factor FHL2 through activation of the AP-1 complex. Furthermore, RNAi gene silencing of FOSL-1 and FHL2 resulted in increase of extracellular signal-regulated kinase (ERK phosphorylation of which duration was sustained by HRG stimulation. Conclusion Our analysis indicated that a time-dependent transcriptional regulatory network including c-FOS, FRA-1, and FHL2 is vital in controlling the ERK signaling pathway through a negative feedback loop for MCF-7 cell differentiation.

  16. Functional Selectivity and Antidepressant Activity of Serotonin 1A Receptor Ligands

    Directory of Open Access Journals (Sweden)

    Zdzisław Chilmonczyk

    2015-08-01

    Full Text Available Serotonin (5-HT is a monoamine neurotransmitter that plays an important role in physiological functions. 5-HT has been implicated in sleep, feeding, sexual behavior, temperature regulation, pain, and cognition as well as in pathological states including disorders connected to mood, anxiety, psychosis and pain. 5-HT1A receptors have for a long time been considered as an interesting target for the action of antidepressant drugs. It was postulated that postsynaptic 5-HT1A agonists could form a new class of antidepressant drugs, and mixed 5-HT1A receptor ligands/serotonin transporter (SERT inhibitors seem to possess an interesting pharmacological profile. It should, however, be noted that 5-HT1A receptors can activate several different biochemical pathways and signal through both G protein-dependent and G protein-independent pathways. The variables that affect the multiplicity of 5-HT1A receptor signaling pathways would thus result from the summation of effects specific to the host cell milieu. Moreover, receptor trafficking appears different at pre- and postsynaptic sites. It should also be noted that the 5-HT1A receptor cooperates with other signal transduction systems (like the 5-HT1B or 5-HT2A/2B/2C receptors, the GABAergic and the glutaminergic systems, which also contribute to its antidepressant and/or anxiolytic activity. Thus identifying brain specific molecular targets for 5-HT1A receptor ligands may result in a better targeting, raising a hope for more effective medicines for various pathologies.

  17. Correlating RANK ligand/RANK binding kinetics with osteoclast formation and function.

    Science.gov (United States)

    Warren, Julia T; Zou, Wei; Decker, Corinne E; Rohatgi, Nidhi; Nelson, Christopher A; Fremont, Daved H; Teitelbaum, Steven L

    2015-11-01

    The interaction between Receptor Activator of NF-κB Ligand (RANKL) and its receptor RANK is essential for the differentiation and bone resorbing capacity of the osteoclast. Osteoprotegerin (OPG), a soluble homodimer, acts as a decoy receptor for RANKL and thus inhibits osteoclastogenesis. An imbalance in the RANKL/RANK/OPG axis, with decreased OPG and/or increased RANKL, is associated with diseases that favor bone loss, including osteoporosis. Recently, we established a yeast surface display system and screened libraries of randomly mutated RANKL proteins to identify mutations that abolish binding to OPG while preserving recognition of RANK. These efforts yielded several RANKL variants possessing substantially higher affinity for RANK compared to their wild-type (WT) counterpart. Using recombinant RANKL mutant proteins, we find those with increased affinity for RANK produce more robust signaling in osteoclast lineage cells and have greater osteoclastogenic potential. Our results are the first to document gain of function RANKL mutations. They indicate that the physiological RANKL/RANK interaction is not optimized for maximal signaling and function, perhaps reflecting the need to maintain receptor specificity within the tumor necrosis factor superfamily (TNFSF). Instead, we find, a biphasic relationship exists between RANKL/RANK affinity and osteoclastogenic capacity. In our panel of RANKL variants, this relationship is driven entirely by manipulation of the kinetic off-rate. Our structure-based and yeast surface display-derived insights into manipulating this critical signaling axis may aid in the design of novel anti-resorptive therapies as well as provide a paradigm for design of other receptor-specific TNF superfamily ligand variants. PMID:25864714

  18. electronic Ligand Builder and Optimisation Workbench (eLBOW): A tool for ligand coordinate and restraint generation

    Energy Technology Data Exchange (ETDEWEB)

    Moriarty, Nigel; Grosse-Kunstleve, Ralf; Adams, Paul

    2009-07-01

    The electronic Ligand Builder and Optimisation Workbench (eLBOW) is a program module of the PHENIX suite of computational crystallographic software. It's designed to be a flexible procedure using simple and fast quantum chemical techniques to provide chemically accurate information for novel and known ligands alike. A variety of input formats and options allow for the attainment of a number of diverse goals including geometry optimisation and generation of restraints.

  19. Integration of screening and identifying ligand(s) from medicinal plant extracts based on target recognition by using NMR spectroscopy

    OpenAIRE

    sprotocols

    2015-01-01

    Authors: Yalin Tang, Qian Shang, Junfeng Xiang, Qianfan Yang, Qiuju Zhou, Lin Li, Hong Zhang, Qian Li, Hongxia Sun, Aijiao Guan, Wei Jiang & Wei Gai ### Abstract This protocol presents the screening of ligand(s) from medicinal plant extracts based on target recognition by using NMR spectroscopy. A detailed description of sample preparation and analysis process is provided. NMR spectroscopies described here are 1H NMR, diffusion-ordered spectroscopy (DOSY), relaxation-edited NMR, ...

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2007-10-15

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

  1. Synthesis, characterization and catalytic activity of copper(II) complexes containing a redox-active benzoxazole iminosemiquinone ligand.

    Science.gov (United States)

    Balaghi, S Esmael; Safaei, Elham; Chiang, Linus; Wong, Edwin W Y; Savard, Didier; Clarke, Ryan M; Storr, Tim

    2013-05-21

    A tridentate benzoxazole-containing aminophenol ligand HL(BAP) was synthesized and complexed with Cu(II). The resulting Cu(II) complexes were characterized by X-ray, IR, UV-vis-NIR spectroscopies, and magnetic susceptibility studies, demonstrating that the ligand is oxidized to the o-iminosemiquinone form [L(BIS)](-) in the isolated complexes. L(BIS)Cu(II)Cl exhibits a distorted tetrahedral geometry, while L(BIS)Cu(II)OAc is square pyramidal. In both solid state structures the ligand is coordinated to Cu(II)via the benzoxazole, as well as the nitrogen and oxygen atoms from the o-iminosemiquinone moiety. The chloride, or acetate group occupies the fourth and/or fifth positions in L(BIS)Cu(II)Cl and L(BIS)Cu(II)OAc, respectively. Magnetic susceptibility measurements indicate that both complexes are diamagnetic due to antiferromagnetic coupling between the d(9) Cu(II) centre and iminosemiquinone ligand radical. Electrochemical studies of the complexes demonstrate both a quasi-reversible reduction and oxidation process for the Cu complexes. While L(BIS)Cu(II)X (X = Cl) is EPR-silent, chemical oxidation affords a species with an EPR signal consistent with ligand oxidation to form a d(9) Cu(II) iminoquinone species. In addition, chemical reduction results in a Cu(II) centre most likely bound to an amidophenoxide. Mild and efficient oxidation of alcohol substrates to the corresponding aldehydes was achieved with molecular oxygen as the oxidant and L(BIS)Cu(II)X-Cs2CO3 as the catalyst.

  2. Anti-CD20 antibody promotes cancer escape via enrichment of tumor-evoked regulatory B cells expressing low levels of CD20 and CD137L.

    Science.gov (United States)

    Bodogai, Monica; Lee Chang, Catalina; Wejksza, Katarzyna; Lai, Jinping; Merino, Maria; Wersto, Robert P; Gress, Ronald E; Chan, Andrew C; Hesdorffer, Charles; Biragyn, Arya

    2013-04-01

    The possible therapeutic benefits of B-cell depletion in combating tumoral immune escape have been debated. In support of this concept, metastasis of highly aggressive 4T1 breast cancer cells in mice can be abrogated by inactivation of tumor-evoked regulatory B cells (tBreg). Here, we report the unexpected finding that B-cell depletion by CD20 antibody will greatly enhance cancer progression and metastasis. Both murine and human tBregs express low levels of CD20 and, as such, anti-CD20 mostly enriches for these cells. In the 4T1 model of murine breast cancer, this effect of enriching for tBregs suggests that B-cell depletion by anti-CD20 may not be beneficial at all in some cancers. In contrast, we show that in vivo-targeted stimulation of B cells with CXCL13-coupled CpG oligonucleotides (CpG-ODN) can block cancer metastasis by inhibiting CD20(Low) tBregs. Mechanistic investigations suggested that CpG-ODN upregulates low surface levels of 4-1BBL on tBregs to elicit granzyme B-expressing cytolytic CD8(+) T cells, offering some explanative power for the effect. These findings underscore the immunotherapeutic importance of tBreg inactivation as a strategy to enhance cancer therapy by targeting both the regulatory and activating arms of the immune system in vivo. PMID:23365136

  3. A thermal responsive affinity ligand for precipitation of sialylated proteins

    Directory of Open Access Journals (Sweden)

    Lindsay Arnold

    2016-01-01

    Full Text Available We report here the development of a thermal responsive affinity ligand specific to sialic acid, sialic acid containing oligosaccharides, glycoproteins, and other sialylated glycoconjugates. The ligand is a fusion protein of 40 repeats of pentapeptide of an elastin like polymer (ELP and the 21 kD sialic acid binding domain of a Vibrio cholera neuraminidase (VCNA. For cost-effective synthesis, the fusion protein was targeted to the periplasmic space of an E. coli lpp deletion mutant, resulting in its secretion to the growth medium. A pre-induction heat-shock step at 42 ˚C for 20 minutes was necessary to achieve high level expression of the ligand. Under optimized induction condition (18 ˚C, 0.1 mM IPTG and 48 hours of post-induction cultivation, the ligand was produced to about 100 mg/L. The ligand exhibited a transition temperature of 52 ˚C, which could be depressed to 37 ˚C with the addition of 0.5 M NaCl. Using fetuin as a model sialylated protein, the ligand was applied in an affinity precipitation process to illustrate its potential application in glycoprotein isolation. The ligand captured 100% fetuin from an aqueous solution when the molar ratio of ligand to fetuin was 10 to 1, which was lower than the expected for full titration of sialic acid on the glycoprotein by the lectin. Elution of fetuin from ligand was achieved with PBS buffer containing 2 mM sialic acid. To evaluate how protein and other contaminants influence the recovery of sialylated proteins, CHO medium was spiked into the fetuin solution. The predominant protein species in CHO medium was found to be albumin. Although its removal of over 94% was evident, purified fetuin contained some albumin due to its over-abundance. Additional experiments with albumin contaminant of varying concentrations showed that below 1 mg/L, albumin had no impact on the affinity precipitation, whereas above 10 mg/L, some albumin was co-purified with fetuin. However, even at 50 mg/ml, fetuin

  4. Biotechnological Fluorescent Ligands of the Bradykinin B1 Receptor: Protein Ligands for a Peptide Receptor.

    Directory of Open Access Journals (Sweden)

    Xavier Charest-Morin

    Full Text Available The bradykinin (BK B1 receptor (B1R is a peculiar G protein coupled receptor that is strongly regulated to the point of being inducible in immunopathology. Limited clinical evidence suggests that its expression in peripheral blood mononuclear cells is a biomarker of active inflammatory states. In an effort to develop a novel imaging/diagnostic tool, we report the rational design and testing of a fusion protein that is a ligand of the human B1R but not likely to label peptidases. This ligand is composed of a fluorescent protein (FP (enhanced green FP [EGFP] or mCherry prolonged at its N-terminus by a spacer peptide and a classical peptide agonist or antagonist (des-Arg9-BK, [Leu8]des-Arg9-BK, respectively. The design of the spacer-ligand joint peptide was validated by a competition assay for [3H]Lys-des-Arg9-BK binding to the human B1R applied to 4 synthetic peptides of 18 or 19 residues. The labeling of B1R-expressing cells with EGFP or mCherry fused with 7 of such peptides was performed in parallel (microscopy. Both assays indicated that the best design was FP-(Asn-Glyn-Lys-des-Arg9-BK; n = 15 was superior to n = 5, suggesting benefits from minimizing steric hindrance between the FP and the receptor. Cell labeling concerned mostly plasma membranes and was inhibited by a B1R antagonist. EGFP-(Asn-Gly15-Lys-des-Arg9-BK competed for the binding of [3H]Lys-des-Arg9-BK to human recombinant B1R, being only 10-fold less potent than the unlabeled form of Lys-des-Arg9-BK to do so. The fusion protein did not label HEK 293a cells expressing recombinant human BK B2 receptors or angiotensin converting enzyme. This study identifies a modular C-terminal sequence that can be adapted to protein cargoes, conferring high affinity for the BK B1R, with possible applications in diagnostic cytofluorometry, histology and drug delivery (e.g., in oncology.

  5. MITF Modulates Therapeutic Resistance through EGFR Signaling.

    Science.gov (United States)

    Ji, Zhenyu; Erin Chen, Yiyin; Kumar, Raj; Taylor, Michael; Jenny Njauw, Ching-Ni; Miao, Benchun; Frederick, Dennie T; Wargo, Jennifer A; Flaherty, Keith T; Jönsson, Göran; Tsao, Hensin

    2015-07-01

    Response to targeted therapies varies significantly despite shared oncogenic mutations. Nowhere is this more apparent than in BRAF (V600E)-mutated melanomas where initial drug response can be striking and yet relapse is commonplace. Resistance to BRAF inhibitors have been attributed to the activation of various receptor tyrosine kinases (RTKs), although the underlying mechanisms have been largely uncharacterized. Here, we found that EGFR-induced vemurafenib resistance is ligand dependent. We employed whole-genome expression analysis and discovered that vemurafenib resistance correlated with the loss of microphthalmia-associated transcription factor (MITF), along with its melanocyte lineage program, and with the activation of EGFR signaling. An inverse relationship between MITF, vemurafenib resistance, and EGFR was then observed in patient samples of recurrent melanoma and was conserved across melanoma cell lines and patients' tumor specimens. Functional studies revealed that MITF depletion activated EGFR signaling and consequently recapitulated the resistance phenotype. In contrast, forced expression of MITF in melanoma and colon cancer cells inhibited EGFR and conferred sensitivity to BRAF/MEK inhibitors. These findings indicate that an "autocrine drug resistance loop" is suppressed by melanocyte lineage signal(s), such as MITF. This resistance loop modulates drug response and could explain the unique sensitivity of melanomas to BRAF inhibition. PMID:25789707

  6. Theoretical Study of Molecular Determinants Involved in Signal Binding to the TraR Protein of Agrobacterium tumefaciens

    Directory of Open Access Journals (Sweden)

    N. Kumar

    2005-10-01

    Full Text Available N-acylated homoserine lactone (AHL mediated cell-cell communication in bacteria is dependent on the recognition of the cognate signal by its receptor. This interaction allows the receptor-ligand complex to act as a transcriptional activator, controlling the expression of a range of bacterial phenotypes, including virulence factor expression and biofilm formation. One approach to determine the key features of signal- binding is to model the intermolecular interactions between the receptor and ligand using computational-based modeling software (LigandFit. In this communication, we have modeled the crystal structure of the AHL receptor protein TraR and its AHL signal N-(3- oxooctanoyl-homoserine lactone from Agrobacterium tumefaciens and compared it to the previously reported antagonist behaviour of a number of AHL analogues, in an attempt to determine structural constraints for ligand binding. We conclude that (i a common conformation of the AHL in the hydrophobic and hydrophilic region exists for ligand-binding, (ii a tail chain length threshold of 8 carbons is most favourable for ligand-binding affinity, (iii the positive correlation in the docking studies could be used a virtual screening tool.

  7. Delta-Like Ligand 4 Modulates Liver Damage by Down-Regulating Chemokine Expression.

    Science.gov (United States)

    Shen, Zhe; Liu, Yan; Dewidar, Bedair; Hu, Junhao; Park, Ogyi; Feng, Teng; Xu, Chengfu; Yu, Chaohui; Li, Qi; Meyer, Christoph; Ilkavets, Iryna; Müller, Alexandra; Stump-Guthier, Carolin; Munker, Stefan; Liebe, Roman; Zimmer, Vincent; Lammert, Frank; Mertens, Peter R; Li, Hai; Ten Dijke, Peter; Augustin, Hellmut G; Li, Jun; Gao, Bin; Ebert, Matthias P; Dooley, Steven; Li, Youming; Weng, Hong-Lei

    2016-07-01

    Disrupting Notch signaling ameliorates experimental liver fibrosis. However, the role of individual Notch ligands in liver damage is unknown. We investigated the effects of Delta-like ligand 4 (Dll4) in liver disease. DLL4 expression was measured in 31 human liver tissues by immunohistochemistry. Dll4 function was examined in carbon tetrachloride- and bile duct ligation-challenged mouse models in vivo and evaluated in hepatic stellate cells, hepatocytes, and Kupffer cells in vitro. DLL4 was expressed in patients' Kupffer and liver sinusoidal endothelial cells. Recombinant Dll4 protein (rDll4) ameliorated hepatocyte apoptosis, inflammation, and fibrosis in mice after carbon tetrachloride challenge. In vitro, rDll4 significantly decreased lipopolysaccharide-dependent chemokine expression in both Kupffer and hepatic stellate cells. In bile duct ligation mice, rDll4 induced massive hepatic necrosis, resulting in the death of all animals within 1 week. Inflammatory cell infiltration and chemokine ligand 2 (Ccl2) expression were significantly reduced in rDll4-receiving bile duct ligation mice. Recombinant Ccl2 rescued bile duct ligation mice from rDll4-mediated death. In patients with acute-on-chronic liver failure, DLL4 expression was inversely associated with CCL2 abundance. Mechanistically, Dll4 regulated Ccl2 expression via NF-κB. Taken together, Dll4 modulates liver inflammatory response by down-regulating chemokine expression. rDll4 application results in opposing outcomes in two models of liver damage. Loss of DLL4 may be associated with CCL2-mediated cytokine storm in patients with acute-on-chronic liver failure. PMID:27171900

  8. Ligands for pheromone-sensing neurons are not conformationally activated odorant binding proteins.

    Directory of Open Access Journals (Sweden)

    Carolina Gomez-Diaz

    Full Text Available Pheromones form an essential chemical language of intraspecific communication in many animals. How olfactory systems recognize pheromonal signals with both sensitivity and specificity is not well understood. An important in vivo paradigm for this process is the detection mechanism of the sex pheromone (Z-11-octadecenyl acetate (cis-vaccenyl acetate [cVA] in Drosophila melanogaster. cVA-evoked neuronal activation requires a secreted odorant binding protein, LUSH, the CD36-related transmembrane protein SNMP, and the odorant receptor OR67d. Crystallographic analysis has revealed that cVA-bound LUSH is conformationally distinct from apo (unliganded LUSH. Recombinantly expressed mutant versions of LUSH predicted to enhance or diminish these structural changes produce corresponding alterations in spontaneous and/or cVA-evoked activity when infused into olfactory sensilla, leading to a model in which the ligand for pheromone receptors is not free cVA, but LUSH that is "conformationally activated" upon cVA binding. Here we present evidence that contradicts this model. First, we demonstrate that the same LUSH mutants expressed transgenically affect neither basal nor pheromone-evoked activity. Second, we compare the structures of apo LUSH, cVA/LUSH, and complexes of LUSH with non-pheromonal ligands and find no conformational property of cVA/LUSH that can explain its proposed unique activated state. Finally, we show that high concentrations of cVA can induce neuronal activity in the absence of LUSH, but not SNMP or OR67d. Our findings are not consistent with the model that the cVA/LUSH complex acts as the pheromone ligand, and suggest that pheromone molecules alone directly activate neuronal receptors.

  9. Dinitrosyl iron complexes with thiol-containing ligands as a "working form" of endogenous nitric oxide.

    Science.gov (United States)

    Vanin, Anatoly F

    2016-04-01

    The material presented herein is an overview of the results obtained by our research team during the many years' study of biological activities and occurrence of dinitrosyl iron complexes (DNIC) with thiol-containing ligands in human and animal organisms. With regard to their dose dependence and vast diversity of biological activities, DNIC are similar to the system of endogenous NO, one of the most universal regulators of biological processes. The role of biologically active components in DNIC is played by their iron-dinitrosyl fragments, [Fe(NO)2], endowed with the ability to generate neutral NO molecules and nitrosonium ions (NO(+)). Their release is effected by heme-and thiol-containing proteins, which fulfill the function of biological targets and acceptors of NO and NO(+). Beneficial regulatory effects of DNIC on physiological and metabolic processes are numerous and diverse and include, among other things, lowering of arterial pressure and accelerated healing of skin wounds. In the course of fast decomposition of their Fe(NO)2 fragments (e.g., in the presence of iron chelators), DNIC produce adverse (cytotoxic) effects, which can best be exemplified by their ability to suppress the development of experimental endometriosis in animals. In animal tissues, DNIC with thiol-containing ligands are predominantly represented by the binuclear form, which, contrary to mononuclear DNIC detectable by the 2.03 signal, is EPR-silent. The ample body of evidence on biological activities and occurrence of DNIC gained so far clearly demonstrates that in human and animal organisms DNIC with thiol-containing ligands represent a "working form" of the system of endogenous NO responsible for its accumulation and stabilization in animal tissues as well as its further transfer to its biological targets. PMID:26820635

  10. Expressions of chemokine receptor CXCR4 and its ligand CXCL12 in salivary adenoid cystic carcinoma

    Institute of Scientific and Technical Information of China (English)

    徐晓刚; 吕春堂; 周中华

    2004-01-01

    Objective: To examine expressions of chemokine receptor CXCR4 and its ligand CXCL12 in primary focus and lymphogenous metastasis of salivary adenoid cystic carcinoma (ACC) with lung metastasis. Methods: Using immunohistochemical hypersensitivity catalyzed signal amplification (CSA), expressions of chemokine receptor CXCR4 and ligand CXCL12 were detected in tissue specimens from 20 cases of primary cancer focus and lymphogenous metastasis of salivary adenoid cystic carcinoma, of which 7 cases were associated with lung metastasis and 3 with lympogenons metastasis. Twenty cases of tongue carcinoma (including 10 cases with lymphogenous metastasis) and 15 cases of mucoepidermoid carcinoma (including 5 cases with lymphogenous metastasis) were used as the malignant control group; and salivary mixed tumor ( n =10), tongue leukoceratosis ( n = 10) and cervical lymph node reactive hyperplasia ( n = 10) were used as the benign control group. Results: Expression of CXCR4 in the tissues and lymph metastases of oral and maxillofacial salivary ACC, mucoepidermoid carcinoma and tongue carcinoma was significantly higher than that of the benign control group ( P < 0.05); expression of CXCR4 in the primary focus of ACC was significantly higher than that of the malignant control group; and expression of CXCR4 in the ACC with lung metastasis was 87.1% (6/7), significantly higher than that without lung metastasis( P <0.01 ). There was evident positive expression of CXCL12 in endotheliocytes of microvessels within cancer and paracancer tissues and significantly high expression of CXCL12 in lymphogenous metastasis( P < 0.05). Conclusion: Chemokine receptor CXCR4 and its ligand CXCL12 may be associated with local invasion and lymphogenous metastasis of oral and maxillofacial cancer, especially with lung metastasis of salivary ACC.

  11. Synthesis and enzymatic cleavage of dual-ligand quantum dots

    Energy Technology Data Exchange (ETDEWEB)

    Sewell, Sarah L. [Department of Biomedical Engineering, Vanderbilt University, Nashville, TN (United States); Giorgio, Todd D., E-mail: todd.d.giorgio@vanderbilt.edu [Department of Biomedical Engineering, Vanderbilt University, Nashville, TN (United States); Department of Chemical and Biomolecular Engineering, Vanderbilt University, Nashville, TN (United States)

    2009-05-05

    Site directed therapy promises to minimize treatment-limiting systemic effects associated with cytotoxic agents that have no specificity for pathologic tissues. One general strategy is to target cell surface receptors uniquely presented on particular tissues. Highly specific in vivo targeting of an emerging neoplasm through a single molecular recognition mechanism has not generally been successful. Nonspecific binding and specific binding to non-target cells compromise the therapeutic index of small molecule, ubiquitous cancer targeting ligands. In this work, we have designed and fabricated a nanoparticle (NP) construct that could potentially overcome the current limitations of targeted in vivo delivery. Quantum dots (QDs) were functionalized with a poly(ethylene glycol) (PEG) modified to enable specific cleavage by matrix metalloprotease-7 (MMP-7). The QDs were further functionalized with folic acid, a ligand for a cell surface receptor that is overexpressed in many tumors, but also expressed in some normal tissues. The nanomolecular construct is designed so that the PEG initially conceals the folate ligand and construct binding to cells is inhibited. MMP-7 activated peptide cleavage and subsequent unmasking of the folate ligand occurs only near tumor tissue, resulting in a proximity activated (PA) targeting system. QDs functionalized with both the MMP-7 cleavable substrate and folic acid were successfully synthesized and characterized. The proteolytic capability of the dual ligand QD construct was quantitatively assessed by fluorometric analysis and compared to a QD construct functionalized with only the PA ligand. The dual ligand PA nanoparticles studied here exhibit significant susceptibility to cleavage by MMP-7 at physiologically relevant conditions. The capacity to autonomously convert a biopassivated nanostructure to a tissue-specific targeted delivery agent in vivo represents a paradigm change for site-directed therapies.

  12. Synthesis and enzymatic cleavage of dual-ligand quantum dots

    International Nuclear Information System (INIS)

    Site directed therapy promises to minimize treatment-limiting systemic effects associated with cytotoxic agents that have no specificity for pathologic tissues. One general strategy is to target cell surface receptors uniquely presented on particular tissues. Highly specific in vivo targeting of an emerging neoplasm through a single molecular recognition mechanism has not generally been successful. Nonspecific binding and specific binding to non-target cells compromise the therapeutic index of small molecule, ubiquitous cancer targeting ligands. In this work, we have designed and fabricated a nanoparticle (NP) construct that could potentially overcome the current limitations of targeted in vivo delivery. Quantum dots (QDs) were functionalized with a poly(ethylene glycol) (PEG) modified to enable specific cleavage by matrix metalloprotease-7 (MMP-7). The QDs were further functionalized with folic acid, a ligand for a cell surface receptor that is overexpressed in many tumors, but also expressed in some normal tissues. The nanomolecular construct is designed so that the PEG initially conceals the folate ligand and construct binding to cells is inhibited. MMP-7 activated peptide cleavage and subsequent unmasking of the folate ligand occurs only near tumor tissue, resulting in a proximity activated (PA) targeting system. QDs functionalized with both the MMP-7 cleavable substrate and folic acid were successfully synthesized and characterized. The proteolytic capability of the dual ligand QD construct was quantitatively assessed by fluorometric analysis and compared to a QD construct functionalized with only the PA ligand. The dual ligand PA nanoparticles studied here exhibit significant susceptibility to cleavage by MMP-7 at physiologically relevant conditions. The capacity to autonomously convert a biopassivated nanostructure to a tissue-specific targeted delivery agent in vivo represents a paradigm change for site-directed therapies.

  13. PDGFRaa Signaling Is Regulated through the Primary Cilium in Fibroblasts

    DEFF Research Database (Denmark)

    Schneider, Linda; Clement, Christian Alexandro; Teilmann, S.C.;

    2005-01-01

    Recent findings show that cilia are sensory organelles that display specific receptors and ion channels, which transmit signals from the extracellular environment via the cilium to the cell to control tissue homeostasis and function [ [1] , [2] , [3] , [4] , [5] and [6] ]. Agenesis of primary cilia...... that the primary cilium in fibroblasts [ 11 ] plays a critical role in growth control via platelet-derived growth factor receptor a (PDGFRa), which localizes to the primary cilium during growth arrest in NIH3T3 cells and primary cultures of mouse embryonic fibroblasts. Ligand-dependent activation of PDGFRaa....... Signaling through PDGFRß, which localizes to the plasma membrane, is maintained at comparable levels in wild-type and mutant cells. We propose that ciliary PDGFRaa signaling is linked to tissue homeostasis and to mitogenic signaling pathways....

  14. Slit2/Robo1 signaling in glioma migration and invasion.

    Science.gov (United States)

    Xu, Yun; Li, Wen-Liang; Fu, Li; Gu, Feng; Ma, Yong-Jie

    2010-12-01

    Slit2/Robo1 is a conserved ligand-receptor system, which greatly affects the distribution, migration, axon guidance and branching of neuron cells. Slit2 and its transmembrane receptor Robo1 have different distribution patterns in gliomas. The expression of Slit2 is at very low levels in pilocytic astrocytoma, fibrillary astrocytoma and glioblastoma, while Robo1 is highly expressed in different grades of gliomas at both mRNA and protein levels. Acquisition of insidious invasiveness by malignant glioma cells involves multiple genetic alterations in signaling pathways. Although the specific mechanisms of tumor-suppressive effect of Slit2/Robo1 have not been elucidated, it has been proved that Slit2/Robo1 signaling inhibits glioma cell migration and invasion by inactivation of Cdc42-GTP. With the research development on the molecular mechanisms of Slit2/Robo1 signaling in glioma invasion and migration, Slit2/Robo1 signaling may become a potential target for glioma prevention and treatment.

  15. Microassay for measurement of binding of radiolabelled ligands to cell surface molecules

    International Nuclear Information System (INIS)

    An improved technique for measuring the binding of radiolabelled ligands to cell surface molecules has been developed by modification of a procedure using centrifugation through a water-immiscible oil to separate free and cell-bound ligand. It maximises the percentage of ligand bound since cell-bound and free ligand can be separated easily and reproducibly even when very small reaction volumes are used. This permits low levels of ligand radiolabelling and relatively low numbers of cells to be used

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

    Science.gov (United States)

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

    2016-07-01

    It is becoming increasingly clear that intracellular signaling can be subject to strict spatial control. As the covalent attachment of a signaling ligand to a nanoparticle (NP) impacts ligand-receptor binding, uptake, and trafficking, nanoconjugation provides new opportunities for manipulating intracellular signaling in a controlled fashion. To establish the effect of nanoconjugation on epidermal growth factor (EGF) mediated signaling, we investigate here the intracellular fate of nanoconjugated EGF (NP-EGF) and its bound receptor (EGFR) by quantitative correlated darkfield/fluorescence microscopy and density-based endosomal fractionation. We demonstrate that nanoconjugation prolongs the dwell time of phosphorylated receptors in the early endosomes and that the retention of activated EGFR in the early endosomes is accompanied by an EGF mediated apoptosis at effective concentrations that do not induce apoptosis in the case of free EGF. Overall, these findings indicate nanoconjugation as a rational strategy for modifying signaling that acts by modulating the temporo-spatial distribution of the activated EGF-EGFR ligand-receptor complex.It is becoming increasingly clear that intracellular signaling can be subject to strict spatial control. As the covalent attachment of a signaling ligand to a nanoparticle (NP) impacts ligand-receptor binding, uptake, and trafficking, nanoconjugation provides new opportunities for manipulating intracellular signaling in a controlled fashion. To establish the effect of nanoconjugation on epidermal growth factor (EGF) mediated signaling, we investigate here the intracellular fate of nanoconjugated EGF (NP-EGF) and its bound receptor (EGFR) by quantitative correlated darkfield/fluorescence microscopy and density-based endosomal fractionation. We demonstrate that nanoconjugation prolongs the dwell time of phosphorylated receptors in the early endosomes and that the retention of activated EGFR in the early endosomes is accompanied by an EGF

  17. Synthesis, Spectroscopy, Theoretical, and Electrochemical Studies of Zn(II, Cd(II, and Hg(II Azide and Thiocyanate Complexes of a New Symmetric Schiff-Base Ligand

    Directory of Open Access Journals (Sweden)

    Morteza Montazerozohori

    2013-01-01

    Full Text Available Synthesis of zinc(II/cadmium(II/mercury(II thiocyanate and azide complexes of a new bidentate Schiff-base ligand (L with general formula of MLX2 (M = Zn(II, Cd(II, and Hg(II in ethanol solution at room temperature is reported. The ligand and metal complexes were characterized by using ultraviolet-visible (UV-visible, Fourier transform infrared (FT-IR, 1H- and 13C-NMR spectroscopy and physical characterization, CHN analysis, and molar conductivity. 1H- and 13C-NMR spectra have been studied in DMSO-d6. The reasonable shifts of FT-IR and NMR spectral signals of the complexes with respect to the free ligand confirm well coordination of Schiff-base ligand and anions in an inner sphere coordination space. The conductivity measurements as well as spectral data indicated that the complexes are nonelectrolyte. Theoretical optimization on the structure of ligand and its complexes was performed at the Becke’s three-parameter hybrid functional (B3 with the nonlocal correlation of Lee-Yang-Parr (LYP level of theory with double-zeta valence (LANL2DZ basis set using GAUSSIAN 03 suite of program, and then some theoretical structural parameters such as bond lengths, bond angles, and torsion angles were obtained. Finally, electrochemical behavior of ligand and its complexes was investigated. Cyclic voltammograms of metal complexes showed considerable changes with respect to free ligand.

  18. T-cell ligands modulate the cytolytic activity of the CD33/CD3 BiTE antibody construct, AMG 330

    International Nuclear Information System (INIS)

    Preclinical and emerging clinical studies demonstrate that bispecific T-cell engaging (BiTE) antibody constructs can potently lyse targeted tumor cells, but the determinants for their activity remain incompletely understood. Using human acute myeloid leukemia (AML) cell lines engineered to overexpress individual T-cell ligands, we found that expression of the inhibitory ligands, PD-L1 and PD-L2, reduced the cytolytic activity of the BiTE antibody construct targeting CD33, AMG 330; conversely, expression of the activating ligands, CD80 and CD86, augmented the cytotoxic activity of AMG 330. Consistent with these findings, treatment with an activating antibody directed at the co-stimulatory T-cell receptor, CD28, significantly increased AMG 330-induced cytotoxicity in human AML cell lines. Using specimens from 12 patients with newly diagnosed or relapsed/refractory AML, we found that activation of CD28 also increased the activity of AMG 330 in primary human AML cells (P=0.023). Together, our findings indicate that T-cell ligands and co-receptors modulate the anti-tumor activity of the CD33/CD3 BiTE antibody construct, AMG 330. These findings suggest that such ligands/co-receptors could serve as biomarkers of response and that co-treatment strategies with pharmacological modulators of T-cell receptor signaling could be utilized to further enhance the activity of this targeted therapeutic

  19. Coarse-grained molecular dynamics simulations of protein-ligand binding.

    Science.gov (United States)

    Negami, Tatsuki; Shimizu, Kentaro; Terada, Tohru

    2014-09-30

    Coarse-grained molecular dynamics (CGMD) simulations with the MARTINI force field were performed to reproduce the protein-ligand binding processes. We chose two protein-ligand systems, the levansucrase-sugar (glucose or sucrose), and LinB-1,2-dichloroethane systems, as target systems that differ in terms of the size and shape of the ligand-binding pocket and the physicochemical properties of the pocket and the ligand. Spatial distributions of the Coarse-grained (CG) ligand molecules revealed potential ligand-binding sites on the protein surfaces other than the real ligand-binding sites. The ligands bound most strongly to the real ligand-binding sites. The binding and unbinding rate constants obtained from the CGMD simulation of the levansucrase-sucrose system were approximately 10 times greater than the experimental values; this is mainly due to faster diffusion of the CG ligand in the CG water model. We could obtain dissociation constants close to the experimental values for both systems. Analysis of the ligand fluxes demonstrated that the CG ligand molecules entered the ligand-binding pockets through specific pathways. The ligands tended to move through grooves on the protein surface. Thus, the CGMD simulations produced reasonable results for the two different systems overall and are useful for studying the protein-ligand binding processes.

  20. Characterization of a Dchs1 mutant mouse reveals requirements for Dchs1-Fat4 signaling during mammalian development

    OpenAIRE

    Mao, Yaopan; Mulvaney, Joanna; Zakaria, Sana; Yu, Tian; Morgan, Katherine Malanga; Allen, Steve; Basson, M. Albert; Francis-West, Philippa; Irvine, Kenneth D.

    2011-01-01

    The Drosophila Dachsous and Fat proteins function as ligand and receptor, respectively, for an intercellular signaling pathway that regulates Hippo signaling and planar cell polarity. Although gene-targeted mutations in two mammalian Fat genes have been described, whether mammals have a Fat signaling pathway equivalent to that in Drosophila, and what its biological functions might be, have remained unclear. Here, we describe a gene-targeted mutation in a murine Dachsous homolog, Dchs1. Analys...

  1. Aluminum complexes of the redox-active [ONO] pincer ligand.

    Science.gov (United States)

    Szigethy, Géza; Heyduk, Alan F

    2012-07-14

    A series of aluminum complexes containing the tridentate, redox-active ligand bis(3,5-di-tert-butyl-2-phenol)amine ([ONO]H(3)) in three different oxidation states were synthesized. The aluminum halide salts AlCl(3) and AlBr(3) were reacted with the doubly deprotonated form of the ligand to afford five-coordinate [ONHO(cat)]AlX(solv) complexes (1a, X = Cl, solv = OEt(2); 1b, X = Br, solv = THF), each having a trigonal bipyramidal coordination geometry at the aluminum and containing the [ONHO(cat)](2-) ligand with a protonated, sp(3)-hybridized nitrogen donor. The [ONO] ligand platform may also be added to aluminum through the use of the oxidized ligand salt [ONO(q)]K, which was reacted with AlCl(3) in the presence of either diphenylacetylacetonate (acacPh(2)(-)) or 8-oxyquinoline (quinO(-)) to afford [ONO(q)]Al(acacPh(2))Cl (2) or [ONO(q)]Al(quinO)Cl (3), respectively, with well-defined [ONO(q)](-) ligands. Quinonate complexes 2 and 3 were reduced by one electron to afford the corresponding complexes K{[ONO(sq)]Al(acacPh(2))(py)} (4) and K{[ONO(sq)]Al(quinO)(py)} (5), respectively, containing well-defined [ONO(sq)](2-) ligands. The addition of tetrachloro-1,2-quinone to 1a in the presence of pyridine resulted in the expulsion of HCl and the formation of an aluminum complex with two different redox active ligands, [ONO]Al(o-O(2)C(6)Cl(4))(py) (6). Similar results were obtained when 1a was reacted with 9,10-phenanthrenequinone to afford [ONO]Al(o-O(2)C(14)H(8))(py) (7) or with pyrene-4,5-dione to afford [ONO]Al(o-O(2)C(16)H(8))(py) (8). Structural, spectroscopic and preliminary magnetic measurements on 6-8 suggest ligand non-innocent redox behavior in these complexes. PMID:22669327

  2. Ligand Migration and Binding in Myoglobin Mutant L29W

    Science.gov (United States)

    Nienhaus, G. Ulrich; Waschipky, Robert; Nienhaus, Karin; Minkow, Oleksandr; Ostermann, Andreas; Parak, Fritz G.

    2001-09-01

    Myoglobin, a small globular heme protein that binds gaseous ligands such as O2, CO, and NO reversibly at the heme iron, has for many years been a paradigm for studying the effects of structure and dynamics on protein reactions. Time-resolved spectroscopic measurements after photodissociation of the ligand reveal a complex ligand binding reaction with multiple kinetic intermediates, resulting from protein relaxation and movements of the ligand within the protein. To observe structural changes induced by ligand dissociation, we have investigated carbonmonoxy myoglobin (MbCO) mutant L29W using time-resolved infrared spectroscopy in combination with x-ray crystallography. The presence of two distinct infrared stretch bands of the bound CO, AI at 1945 cm-1 and AII at 1955 cm-1, implies that L29W MbCO assumes two different conformations at neutral pH. Low-temperature flash photolysis experiments with monitoring of the absorption changes in the individual CO lines reveal markedly different rebinding properties. While recombination in AII is conceptually simple and well described by a two-state transition involving a distribution of enthalpy barriers, recombination in AI is more complicated: Besides a fast kinetic component, a second, slower kinetic component appears; its population grows with increasing temperature. X-ray crystallography of crystals illuminated below 180 K to photodissociate the CO reveals that the slow component arises from ligands that have migrated from their initial docking site to a remote site within the distal heme pocket. This process occurs in an essentially immobilized, frozen protein. Subsequently, ligands rebind by thermal activation over a barrier that is much higher than the barrier for recombination from the initial docking site. Upon photodissociation above 180 K, ligands escape from the distal pocket, aided by protein fluctuations that transiently open exit channels. The x-ray structure shows a large proportion of ligands in a cavity on

  3. Cell signaling review series

    Institute of Scientific and Technical Information of China (English)

    Aiming Lin; Zhenggang Liu

    2008-01-01

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

  4. Signal sciences workshop. Proceedings

    International Nuclear Information System (INIS)

    This meeting is aimed primarily at signal processing and controls. The technical program for the 1997 Workshop includes a variety of efforts in the Signal Sciences with applications in the Microtechnology Area a new program at LLNL and a future area of application for both Signal/Image Sciences. Special sessions organized by various individuals in Seismic and Optical Signal Processing as well as Micro-Impulse Radar Processing highlight the program, while the speakers at the Signal Processing Applications session discuss various applications of signal processing/control to real world problems. For the more theoretical, a session on Signal Processing Algorithms was organized as well as for the more pragmatic, featuring a session on Real-Time Signal Processing

  5. Retinoid signalling during embryogenesis

    NARCIS (Netherlands)

    Pijnappel, W.W.M.; Hendriks, H.F.J.; Durston, A.J.

    1996-01-01

    Conclusion: Retinoids are suspected to have multiple functions during embryogenesis, which are carried out via various different signal transduction pathways involving active retinoids and nuclear retinoid receptors. Research focuses on the identification of the retinoid signal transduction componen

  6. Elucidating Ligand-Modulated Conformational Landscape of GPCRs Using Cloud-Computing Approaches.

    Science.gov (United States)

    Shukla, Diwakar; Lawrenz, Morgan; Pande, Vijay S

    2015-01-01

    G-protein-coupled receptors (GPCRs) are a versatile family of membrane-bound signaling proteins. Despite the recent successes in obtaining crystal structures of GPCRs, much needs to be learned about the conformational changes associated with their activation. Furthermore, the mechanism by which ligands modulate the activation of GPCRs has remained elusive. Molecular simulations provide a way of obtaining detailed an atomistic description of GPCR activation dynamics. However, simulating GPCR activation is challenging due to the long timescales involved and the associated challenge of gaining insights from the "Big" simulation datasets. Here, we demonstrate how cloud-computing approaches have been used to tackle these challenges and obtain insights into the activation mechanism of GPCRs. In particular, we review the use of Markov state model (MSM)-based sampling algorithms for sampling milliseconds of dynamics of a major drug target, the G-protein-coupled receptor β2-AR. MSMs of agonist and inverse agonist-bound β2-AR reveal multiple activation pathways and how ligands function via modulation of the ensemble of activation pathways. We target this ensemble of conformations with computer-aided drug design approaches, with the goal of designing drugs that interact more closely with diverse receptor states, for overall increased efficacy and specificity. We conclude by discussing how cloud-based approaches present a powerful and broadly available tool for studying the complex biological systems routinely.

  7. The Prelude on Novel Receptor and Ligand Targets Involved in the Treatment of Diabetes Mellitus

    Directory of Open Access Journals (Sweden)

    Venu Gopal Jonnalagadda

    2014-05-01

    Full Text Available Metabolic disorders are a group of disorders, due to the disruption of the normal metabolic process at a cellular level. Diabetes Mellitus and Tyrosinaemia are the majorly reported metabolic disorders. Among them, Diabetes Mellitus is a one of the leading metabolic syndrome, affecting 5 to 7 % of the population worldwide and mainly characterised by elevated levels of glucose and is associated with two types of physiological event disturbances such as impaired insulin secretion and insulin resistance. Up to now, various treatment strategies are like insulin, alphaglucosidase inhibitors, biguanides, incretins were being followed. Concurrently, various novel therapeutic strategies are required to advance the therapy of Diabetes mellitus. For the last few decades, there has been an extensive research in understanding the metabolic pathways involved in Diabetes Mellitus at the cellular level and having the profound knowledge on cell-growth, cell-cycle, and apoptosis at a molecular level provides new targets for the treatment of Diabetes Mellitus. Receptor signalling has been involved in these mechanisms, to translate the information coming from outside. To understand the various receptors involved in these pathways, we must have a sound knowledge on receptors and ligands involved in it. This review mainly summarises the receptors and ligands which are involved the Diabetes Mellitus. Finally, researchers have to develop the alternative chemical moieties that retain their affinity to receptors and efficacy. Diabetes Mellitus being a metabolic disorder due to the glucose surfeit, demands the need for regular exercise along with dietary changes.

  8. NOP Receptor Ligands as Potential Agents for Inflammatory and Autoimmune Diseases

    Directory of Open Access Journals (Sweden)

    Elaine C. Gavioli

    2011-01-01

    Full Text Available Nociceptin/orphanin FQ (N/OFQ is a seventeen-amino acid peptide that is the endogenous ligand of a G-protein-coupled receptor (NOP. Various immune cells express the precursor protein and secrete N/OFQ as well as display binding sites for this peptide. The functional capacity of NOP receptor was demonstrated in vitro and in vivo studies by the ability of N/OFQ to induce chemotaxis of immune cells, to regulate the expression of cytokines and other inflammatory mediators, and to control cellular and humoral immunity. In this context, N/OFQ could modulate the outcome of some inflammatory diseases, such as sepsis and autoimmune pathologies by mechanisms not clearly elucidated yet. In fact, human body fluid revealed increased levels of N/OFQ under sepsis, arthritis, and Parkinson's diagnose. Preclinical studies pointed to the blockade of NOP receptor signaling as successful in treating these experimental conditions. Further preclinical and clinical studies are required to investigate the potential of NOP ligands in treating inflammatory diseases.

  9. Competitive interactions of ligands and macromolecular crowders with maltose binding protein.

    Directory of Open Access Journals (Sweden)

    Andrew C Miklos

    Full Text Available Cellular signaling involves a cascade of recognition events occurring in a complex environment with high concentrations of proteins, polysaccharides, and other macromolecules. The influence of macromolecular crowders on protein binding affinity through hard-core repulsion is well studied, and possible contributions of protein-crowder soft attraction have been implicated recently. Here we present direct evidence for weak association of maltose binding protein (MBP with a polysaccharide crowder Ficoll, and that this association effectively competes with the binding of the natural ligand, maltose. Titration data over wide ranges of maltose and Ficoll concentrations fit well with a three-state competitive binding model. Broadening of MBP (1H-(15N TROSY spectra by the addition of Ficoll indicates weak protein-crowder association, and subsequent recovery of sharp NMR peaks upon addition of maltose indicates that the interactions of the crowder and the ligand with MBP are competitive. We hypothesize that, in the Escherichia coli periplasm, the competitive interactions of polysaccharides and maltose with MBP could allow MBP to shuttle between the peptidoglycan attached to the outer membrane and the ATP-binding cassette transporter in the inner membrane.

  10. Electrostatically induced recruitment of membrane peptides into clusters requires ligand binding at both interfaces.

    Directory of Open Access Journals (Sweden)

    Yuri N Antonenko

    Full Text Available Protein recruitment to specific membrane locations may be governed or facilitated by electrostatic attraction, which originates from a multivalent ligand. Here we explored the energetics of a model system in which this simple electrostatic recruitment mechanism failed. That is, basic poly-L-lysine binding to one leaflet of a planar lipid bilayer did not recruit the triply-charged peptide (O-Pyromellitylgramicidin. Clustering was only observed in cases where PLL was bound to both channel ends. Clustering was indicated (i by the decreased diffusional PLL mobility D(PLL and (ii by an increased lifetime τ(PLL of the clustered channels. In contrast, if PLL was bound to only one leaflet, neither D(PLL nor τ(P changed. Simple calculations suggest that electrostatic repulsion of the unbound ends prevented neighboring OPg dimers from approaching each other. We believe that a similar mechanism may also operate in cell signaling and that it may e.g. contribute to the controversial results obtained for the ligand driven dimerization of G protein-coupled receptors.

  11. Dual ligand/receptor interactions activate urothelial defenses against uropathogenic E. coli.

    Science.gov (United States)

    Liu, Yan; Mémet, Sylvie; Saban, Ricardo; Kong, Xiangpeng; Aprikian, Pavel; Sokurenko, Evgeni; Sun, Tung-Tien; Wu, Xue-Ru

    2015-11-09

    During urinary tract infection (UTI), the second most common bacterial infection, dynamic interactions take place between uropathogenic E. coli (UPEC) and host urothelial cells. While significant strides have been made in the identification of the virulence factors of UPEC, our understanding of how the urothelial cells mobilize innate defenses against the invading UPEC remains rudimentary. Here we show that mouse urothelium responds to the adhesion of type 1-fimbriated UPEC by rapidly activating the canonical NF-κB selectively in terminally differentiated, superficial (umbrella) cells. This activation depends on a dual ligand/receptor system, one between FimH adhesin and uroplakin Ia and another between lipopolysaccharide and Toll-like receptor 4. When activated, all the nuclei (up to 11) of a multinucleated umbrella cell are affected, leading to significant amplification of proinflammatory signals. Intermediate and basal cells of the urothelium undergo NF-κB activation only if the umbrella cells are detached or if the UPEC persistently express type 1-fimbriae. Inhibition of NF-κB prevents the urothelium from clearing the intracellular bacterial communities, leading to prolonged bladder colonization by UPEC. Based on these data, we propose a model of dual ligand/receptor system in innate urothelial defenses against UPEC.

  12. Novel 4-phenylpiperidine-2,6-dione derivatives. Ligands for α1-adrenoceptor subtypes

    KAUST Repository

    Romeo, Giuseppe F.

    2011-07-01

    A number of new 4-phenylpiperidine-2,6-diones bearing at the 1-position an ω-[4-(substituted phenyl)piperazin-1-yl]alkyl moiety were designed and synthesized as ligands for the α1-adrenergic receptor (α1-AR) subtypes. Some synthesized compounds, tested in binding assays for the human cloned α1A-, α1B-, and α1D-AR subtypes, displayed affinities in the nanomolar range. Highest affinity values were found in derivatives having a butyl connecting chain between the 4-phenylpiperidine-2,6-dione and the phenylpiperazinyl moieties. 1-[4-[4-(2-Methoxyphenyl)piperazin-1-yl]butyl]-4-phenylpiperidine-2,6- dione (34) showed the best affinity for the α1A-AR (pK i = 8.74) and 10-fold selectivity compared to the other two α1-AR subtypes. Some representative compounds were also tested in order to evaluate their effects on the signal transduction pathway coupled to α1-AR subtypes. They all blocked norepinephrine-induced stimulation of inositol phospholipid hydrolysis, thus behaving as antagonists. Binding data were used to refine a previously developed pharmacophoric model for α1D-ARs. The revised model shows a highly predictive power and could be useful for the future design of high affinity α1D-AR ligands. © 2011 Elsevier Masson SAS. All rights reserved.

  13. Wingless ligand 5a is a critical regulator of placental growth and survival.

    Science.gov (United States)

    Meinhardt, Gudrun; Saleh, Leila; Otti, Gerlinde R; Haider, Sandra; Velicky, Philipp; Fiala, Christian; Pollheimer, Jürgen; Knöfler, Martin

    2016-01-01

    The maternal uterine environment is likely critical for human placental morphogenesis and development of its different trophoblast subtypes. However, factors controlling growth and differentiation of these cells during early gestation remain poorly elucidated. Herein, we provide evidence that the ligand Wnt5a could be a critical regulator of trophoblast proliferation and survival. Immunofluorescence of tissues and western blot analyses of primary cultures revealed abundant Wnt5a expression and secretion from first trimester decidual and villous stromal cells. The ligand was also detectable in decidual glands, macrophages and NK cells. Wnt5a increased proliferation of villous cytotrophoblasts and cell column trophoblasts, outgrowth on collagen I as well as cyclin A and D1 expression in floating explant cultures, but suppressed camptothecin-induced apoptosis. Similarly, Wnt5a stimulated BrdU incorporation and decreased caspase-cleaved cytokeratin 18 neo-epitope expression in primary cytotrophoblasts. Moreover, Wnt5a promoted activation of the MAPK pathway in the different trophoblast models. Chemical inhibition of p42/44 MAPK abolished cyclin D1 expression and Wnt5a-stimulated proliferation. Compared to controls, MAPK phosphorylation and proliferation of cytotrophoblasts declined upon supplementation of supernatants from Wnt5a gene-silenced decidual or villous stromal cells. In summary, non-canonical Wnt5a signalling could play a role in early human trophoblast development by promoting cell proliferation and survival. PMID:27311852

  14. The Role of TNF Related Apoptosis-Inducing Ligand in Neurodegenerative Diseases

    Institute of Scientific and Technical Information of China (English)

    Y.Huang; N.Erdmann; H.Peng; Y.Zhao; Jialin Zheng

    2005-01-01

    A hallmark of all forms of neurodegenerative diseases is impairment of neuronal functions, and in many cases neuronal cell death. Although the etiology of neurodegenerative diseases may be distinct, different diseases display a similar pathogenesis, for example abnormal immunity within the central nervous system (CNS), activation of macrophage/microglia and the involvement of proinflammatory cytokines. Recent studies show that neurons in a neurodegenerative state undergo a highly regulated programmed cell death, also called apoptosis. TNF-related apoptosis-inducing ligand (TRAIL), a member of the TNF family, has been shown to be involved in apoptosis during many diseases. As one member of a death ligand family, TRAIL was originally thought to target only tumor cells and was not present in CNS. However, recent data showed that TRAIL was unregulated in HIV-1-infected and immune-activated macrophages, a major disease inducing cell during HIV-1-associated dementia (HAD). TRAIL is also induced on neuron by β-amyloid protein, an important pathogen for Alzheimer's disease. In this review, we summarize the possible common aspects that TRAIL involved those neurodegenerative diseases, TRAIL induced apoptosis signaling in the CNS cells, and specific role of TRAIL in individual diseases. Cellular & Molecular Immunology. 2005;2(2):113-122.

  15. The Role of TNF Related Apoptosis-Inducing Ligand in Neurodegenerative Diseases

    Institute of Scientific and Technical Information of China (English)

    Y.Huang; N.Erdmann; H.Peng; Y.Zhao

    2005-01-01

    A hallmark of all forms of neurodegenerative diseases is impairment of neuronal functions, and in many cases neuronal cell death. Although the etiology of neurodegenerative diseases may be distinct, different diseases display a similar pathogenesis, for example abnormal immunity within the central nervous system (CNS), activation of macrophage/microglia and the involvement of proinflammatory cytokines. Recent studies show that neurons in a neurodegenerative state undergo a highly regulated programmed cell death, also called apoptosis. TNF-related apoptosis-inducing ligand (TRAIL), a member of the TNF family, has been shown to be involved in apoptosis during many diseases. As one member of a death ligand family, TRAIL was originally thought to target only tumor cells and was not present in CNS. However, recent data showed that TRAIL was unregulated in HIV-l-infected and immune-activated macrophages, a major disease inducing cell during HIV-l-associated dementia (HAD). TRAIL is also induced on neuron by [$-amyloid protein, an important pathogen for Alzheimer's disease. In this review, we summarize the possible common aspects that TRAIL involved those neurodegenerative diseases, TRAIL induced apoptosis signaling in the CNS cells, and specific role of TRAIL in individual diseases. Cellular & MolecularImmunology. 2005;2(2):113-122.

  16. Cloud computing for protein-ligand binding site comparison.

    Science.gov (United States)

    Hung, Che-Lun; Hua, Guan-Jie

    2013-01-01

    The proteome-wide analysis of protein-ligand binding sites and their interactions with ligands is important in structure-based drug design and in understanding ligand cross reactivity and toxicity. The well-known and commonly used software, SMAP, has been designed for 3D ligand binding site comparison and similarity searching of a structural proteome. SMAP can also predict drug side effects and reassign existing drugs to new indications. However, the computing scale of SMAP is limited. We have developed a high availability, high performance system that expands the comparison scale of SMAP. This cloud computing service, called Cloud-PLBS, combines the SMAP and Hadoop frameworks and is deployed on a virtual cloud computing platform. To handle the vast amount of experimental data on protein-ligand binding site pairs, Cloud-PLBS exploits the MapReduce paradigm as a management and parallelizing tool. Cloud-PLBS provides a web portal and scalability through which biologists can address a wide range of computer-intensive questions in biology and drug discovery.

  17. Kinetics of Receptor-Ligand Interactions in Immune Responses

    Institute of Scientific and Technical Information of China (English)

    Mian Long; Shouqin Lü; Ganyun Sun

    2006-01-01

    Receptor-ligand interactions in blood flow are crucial to initiate the biological processes as inflammatory cascade,platelet thrombosis, as well as tumor metastasis. To mediate cell adhesions, the interacting receptors and ligands must be anchored onto two apposing surfaces of two cells or a cell and a substratum, i.e., the two-dimensional (2D) binding, which is different from the binding of a soluble ligand in fluid phase to a receptor, i.e., three-dimensional (3D) binding. While numerous works have been focused on 3D kinetics of receptor-ligand interactions in immune systems, 2D kinetics and its regulations have less been understood, since no theoretical framework and experimental assays have been established until 1993. Not only does the molecular structure dominate 2D binding kinetics, but the shear force in blood flow also regulates cell adhesions mediated by interacting receptors and ligands. Here we provided the overview of current progresses in 2D bindings and regulations. Relevant issues of theoretical frameworks, experimental measurements, kinetic rates and binding affinities, and force regulations,were discussed.

  18. Cloud computing for protein-ligand binding site comparison.

    Science.gov (United States)

    Hung, Che-Lun; Hua, Guan-Jie

    2013-01-01

    The proteome-wide analysis of protein-ligand binding sites and their interactions with ligands is important in structure-based drug design and in understanding ligand cross reactivity and toxicity. The well-known and commonly used software, SMAP, has been designed for 3D ligand binding site comparison and similarity searching of a structural proteome. SMAP can also predict drug side effects and reassign existing drugs to new indications. However, the computing scale of SMAP is limited. We have developed a high availability, high performance system that expands the comparison scale of SMAP. This cloud computing service, called Cloud-PLBS, combines the SMAP and Hadoop frameworks and is deployed on a virtual cloud computing platform. To handle the vast amount of experimental data on protein-ligand binding site pairs, Cloud-PLBS exploits the MapReduce paradigm as a management and parallelizing tool. Cloud-PLBS provides a web portal and scalability through which biologists can address a wide range of computer-intensive questions in biology and drug discovery. PMID:23762824

  19. Predicting protein ligand binding motions with the conformation explorer

    Directory of Open Access Journals (Sweden)

    Flores Samuel C

    2011-10-01

    Full Text Available Abstract Background Knowledge of the structure of proteins bound to known or potential ligands is crucial for biological understanding and drug design. Often the 3D structure of the protein is available in some conformation, but binding the ligand of interest may involve a large scale conformational change which is difficult to predict with existing methods. Results We describe how to generate ligand binding conformations of proteins that move by hinge bending, the largest class of motions. First, we predict the location of the hinge between domains. Second, we apply an Euler rotation to one of the domains about the hinge point. Third, we compute a short-time dynamical trajectory using Molecular Dynamics to equilibrate the protein and ligand and correct unnatural atomic positions. Fourth, we score the generated structures using a novel fitness function which favors closed or holo structures. By iterating the second through fourth steps we systematically minimize the fitness function, thus predicting the conformational change required for small ligand binding for five well studied proteins. Conclusions We demonstrate that the method in most cases successfully predicts the holo conformation given only an apo structure.

  20. The role of CXCL12-CXCR4 signaling pathway in pancreatic development

    DEFF Research Database (Denmark)

    Katsumoto, Keiichi; Kume, Shoen

    2013-01-01

    Chemokine (C-X-C motif) receptor 4 (CXCR4) is the receptor for chemokine (C-X-C motif) ligand 12 (CXCL12, also known as stromal derived factor-1, Sdf1). CXCR4, a protein consisting 352 amino acids, is known to transduce various signals such as cell differentiation, cell survival, cell proliferation...

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

    DEFF Research Database (Denmark)

    Petersen, Pia Steen; Woldbye, David P D; Madsen, Andreas Nygaard;

    2009-01-01

    The receptor for the orexigenic peptide, ghrelin, is one of the most constitutively active 7TM receptors known, as demonstrated under in vitro conditions. Change in expression of a constitutively active receptor is associated with change in signaling independent of the endogenous ligand. In the f...

  2. Regulation of Notch signaling during T- and B-cell development by O-fucose glycans.

    Science.gov (United States)

    Stanley, Pamela; Guidos, Cynthia J

    2009-07-01

    Notch signaling is required for the development of all T cells and marginal zone (MZ) B cells. Specific roles in T- and B-cell differentiation have been identified for different Notch receptors, the canonical Delta-like (Dll) and Jagged (Jag) Notch ligands, and downstream effectors of Notch signaling. Notch receptors and ligands are post-translationally modified by the addition of glycans to extracellular domain epidermal growth factor-like (EGF) repeats. The O-fucose glycans of Notch cell-autonomously modulate Notch-ligand interactions and the strength of Notch signaling. These glycans are initiated by protein O-fucosyltransferase 1 (Pofut1), and elongated by the transfer of N-acetylglucosamine (GlcNAc) to the fucose by beta1,3GlcNAc-transferases termed lunatic, manic, or radical fringe. This review discusses T- and B-cell development from progenitors deficient in O-fucose glycans. The combined data show that Lfng and Mfng regulate T-cell development by enhancing the interactions of Notch1 in T-cell progenitors with Dll4 on thymic epithelial cells. In the spleen, Lfng and Mfng cooperate to modify Notch2 in MZ B progenitors, enhancing their interaction with Dll1 on endothelial cells and regulating MZ B-cell production. Removal of O-fucose affects Notch signaling in myelopoiesis and lymphopoiesis, and the O-fucose glycan in the Notch1 ligand-binding domain is required for optimal T-cell development.

  3. p116Rip : a new player in RhoA signalling

    NARCIS (Netherlands)

    Mulder, Jacqueline

    2005-01-01

    Cells sense changes in their environment via specific sensors, so-called receptors that are located on the outer surface, at the plasma membrane. Receptors are activated by binding of their cognate ligands, signal molecules such as growth factors, which leads to distinct cellular responses. Stimulat

  4. Crystallization and crystallographic analysis of the ligand-binding domain of the Pseudomonas putida chemoreceptor McpS in complex with malate and succinate

    International Nuclear Information System (INIS)

    The crystallization of the ligand-binding domain of the methyl-accepting chemotaxis protein chemoreceptor McpS (McpS-LBD) is reported. Methyl-accepting chemotaxis proteins (MCPs) are transmembrane proteins that sense changes in environmental signals, generating a chemotactic response and regulating other cellular processes. MCPs are composed of two main domains: a ligand-binding domain (LBD) and a cytosolic signalling domain (CSD). Here, the crystallization of the LBD of the chemoreceptor McpS (McpS-LBD) is reported. McpS-LBD is responsible for sensing most of the TCA-cycle intermediates in the soil bacterium Pseudomonas putida KT2440. McpS-LBD was expressed, purified and crystallized in complex with two of its natural ligands (malate and succinate). Crystals were obtained by both the counter-diffusion and the hanging-drop vapour-diffusion techniques after pre-incubation of McpS-LBD with the ligands. The crystals were isomorphous and belonged to space group C2, with two molecules per asymmetric unit. Diffraction data were collected at the ESRF synchrotron X-ray source to resolutions of 1.8 and 1.9 Å for the malate and succinate complexes, respectively

  5. Danger signals in stroke.

    Science.gov (United States)

    Gelderblom, Mathias; Sobey, Christopher G; Kleinschnitz, Christoph; Magnus, Tim

    2015-11-01

    Danger molecules are the first signals released from dying tissue after stroke. These danger signals bind to receptors on immune cells that will result in their activation and the release of inflammatory and neurotoxic mediators, resulting in amplification of the immune response and subsequent enlargement of the damaged brain volume. The release of danger signals is a central event that leads to a multitude of signals and cascades in the affected and neighbouring tissue, therefore providing a potential target for therapy.

  6. Biomedical signal processing

    CERN Document Server

    Akay, Metin

    1994-01-01

    Sophisticated techniques for signal processing are now available to the biomedical specialist! Written in an easy-to-read, straightforward style, Biomedical Signal Processing presents techniques to eliminate background noise, enhance signal detection, and analyze computer data, making results easy to comprehend and apply. In addition to examining techniques for electrical signal analysis, filtering, and transforms, the author supplies an extensive appendix with several computer programs that demonstrate techniques presented in the text.

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

    OpenAIRE

    Oda, Kanae; Kitano, Hiroaki

    2006-01-01

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

  8. Role of Ret Signaling in the Regulation of the Nigrostriatal Dopaminergic System in Mice

    OpenAIRE

    Mijatovic, Jelena

    2009-01-01

    Glial cell line-derived neurotrophic factor (GDNF) enhances survival of dopamine (DA) neurons in vitro, protects DAergic neurons in animal models of Parkinson s disease (PD), stimulates DA neuron function, regulates postnatal development and supports DA neurons during adulthood. GDNF utilizes a multireceptor complex consisting of a ligand-binding co-receptor GDNF family receptor alpha 1 (GFRα1) and a signal-transducing receptor. The major signaling receptor of GDNF is the receptor tyrosine k...

  9. Intraflagellar Transport, Cilia and Mammalian Hedgehog Signaling: Analysis in Mouse Embryonic Fibroblasts

    OpenAIRE

    Ocbina, Polloneal Jymmiel R.; Anderson, Kathryn V.

    2008-01-01

    Genetic studies in the mouse have shown that Intraflagellar Transport (IFT) is essential for mammalian Hedgehog (Hh) signal transduction. In this study, we take advantage of wild type and IFT mutant mouse embryonic fibroblasts (MEFs) to characterize additional aspects of the relationship between IFT and Hh signaling. Exposure to Sonic hedgehog (Shh) ligand or expression of an activated allele of Smo, SmoA1, activates a Hh reporter in wild-type MEFs, but not in MEFs derived from embryos that l...

  10. Wnt/β-catenin signaling in T cell immunity and cancer immunotherapy

    OpenAIRE

    Gattinoni, Luca; Ji, Yun; Restifo, Nicholas P

    2010-01-01

    Wnt ligands are lipid-modified secreted glycoproteins that regulate embryonic development, cell fate specification and the homeostasis of self-renewing adult tissues. In addition to its well established role in thymocyte development, recent studies have indicated that Wnt/β-catenin signaling is critical for the differentiation, polarization and survival of mature T lymphocytes. Here, we describe our current understanding of Wnt signaling in the biology of post-thymic T cells and discuss how h...

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

    OpenAIRE

    Straßer, Andreas Dominikus

    2014-01-01

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

  12. Tetrapyrrole Signaling in Plants

    Science.gov (United States)

    Larkin, Robert M.

    2016-01-01

    Tetrapyrroles make critical contributions to a number of important processes in diverse organisms. In plants, tetrapyrroles are essential for light signaling, the detoxification of reactive oxygen species, the assimilation of nitrate and sulfate, respiration, photosynthesis, and programed cell death. The misregulation of tetrapyrrole metabolism can produce toxic reactive oxygen species. Thus, it is not surprising that tetrapyrrole metabolism is strictly regulated and that tetrapyrrole metabolism affects signaling mechanisms that regulate gene expression. In plants and algae, tetrapyrroles are synthesized in plastids and were some of the first plastid signals demonstrated to regulate nuclear gene expression. In plants, the mechanism of tetrapyrrole-dependent plastid-to-nucleus signaling remains poorly understood. Additionally, some of experiments that tested ideas for possible signaling mechanisms appeared to produce conflicting data. In some instances, these conflicts are potentially explained by different experimental conditions. Although the biological function of tetrapyrrole signaling is poorly understood, there is compelling evidence that this signaling is significant. Specifically, this signaling appears to affect the accumulation of starch and may promote abiotic stress tolerance. Tetrapyrrole-dependent plastid-to-nucleus signaling interacts with a distinct plastid-to-nucleus signaling mechanism that depends on GENOMES UNCUOPLED1 (GUN1). GUN1 contributes to a variety of processes, such as chloroplast biogenesis, the circadian rhythm, abiotic stress tolerance, and development. Thus, the contribution of tetrapyrrole signaling to plant function is potentially broader than we currently appreciate. In this review, I discuss these aspects of tetrapyrrole signaling.

  13. Environmental Ligands of the Aryl Hydrocarbon Receptor and Their Effects in Models of Adult Liver Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Jan Vondráček

    2016-01-01

    Full Text Available The toxicity of environmental and dietary ligands of the aryl hydrocarbon receptor (AhR in mature liver parenchymal cells is well appreciated, while considerably less attention has been paid to their impact on cell populations exhibiting phenotypic features of liver progenitor cells. Here, we discuss the results suggesting that the consequences of the AhR activation in the cellular models derived from bipotent liver progenitors could markedly differ from those in hepatocytes. In contact-inhibited liver progenitor cells, the AhR agonists induce a range of effects potentially linked with tumor promotion. They can stimulate cell cycle progression/proliferation and deregulate cell-to-cell communication, which is associated with downregulation of proteins forming gap junctions, adherens junctions, and desmosomes (such as connexin 43, E-cadherin, β-catenin, and plakoglobin, as well as with reduced cell adhesion and inhibition of intercellular communication. At the same time, toxic AhR ligands may affect the activity of the signaling pathways contributing to regulation of liver progenitor cell activation and/or differentiation, such as downregulation of Wnt/β-catenin and TGF-β signaling, or upregulation of transcriptional targets of YAP/TAZ, the effectors of Hippo signaling pathway. These data illustrate the need to better understand the potential role of liver progenitors in the AhR-mediated liver carcinogenesis and tumor promotion.

  14. Identification and super-resolution imaging of ligand-activated receptor dimers in live cells

    CERN Document Server

    Winckler, Pascale; Giannone, Gregory; De Giorgi, Francesca; Ichas, François; Sibarita, Jean-Baptiste; Lounis, Brahim; Cognet, Laurent

    2013-01-01

    Molecular interactions are key to many chemical and biological processes like protein function. In many signaling processes they occur in sub-cellular areas displaying nanoscale organizations and involving molecular assemblies. The nanometric dimensions and the dynamic nature of the interactions make their investigations complex in live cells. While super-resolution fluorescence microscopies offer live-cell molecular imaging with sub-wavelength resolutions, they lack specificity for distinguishing interacting molecule populations. Here we combine super-resolution microscopy and single-molecule F\\"orster Resonance Energy Transfer (FRET) to identify dimers of receptors induced by ligand binding and provide super-resolved images of their membrane distribution in live cells. By developing a two-color universal-Point-Accumulation-In-the-Nanoscale-Topography (uPAINT) method, dimers of epidermal growth factor receptors (EGFR) activated by EGF are studied at ultra-high densities, revealing preferential cell-edge sub-...

  15. Characterization of new outer membrane proteins of Pseudomonas aeruginosa using a combinatorial peptide ligand library.

    Science.gov (United States)

    Ben Mlouka, Mohamed Amine; Khemiri, Arbia; Seyer, Damien; Hardouin, Julie; Chan Tchi Song, Philippe; Dé, Emmanuelle; Jouenne, Thierry; Cosette, Pascal

    2015-02-01

    Most often, the use of ProteoMiner beads has been restricted to human serum proteins for the normalization of major proteins, such as albumin. However, there are other situations of interest in which the presence of major proteins would quench the signals of low abundance polypeptides. We propose the use of these beads for investigating the envelope of the gram-negative bacterium Pseudomonas aeruginosa. Initially, we performed comparative 2D electrophoresis to qualitatively evaluate the incidence of the normalization stage. This demonstrated a significant reduction of the major membrane proteins. Thereafter, using shotgun analysis, the same protein extract was targeted by using combinatorial peptide ligand library capture. This treatment yielded 154 additional outer membrane proteins (OMPs) uncovered by the study of the crude sample. PMID:25471289

  16. Is copper(I) really soft? Probing the hardness of Cu(I) with pyridinecarboxaldehyde ligands

    Indian Academy of Sciences (India)

    D Saravanabharathi; M Nethaji; A G Samuelson

    2002-08-01

    Cu(I) complexes of formula Cu(PPh3)2LClO4 [L = 2 or 3 pyridine carboxaldehyde] are synthesised and characterised to explore the coordination of an aldehyde, a hard and neutral oxygen donor to a soft Cu(I) centre. The structural and spectroscopic results illustrate that only in 2-pyridinecarboxaldehyde, the `C=O' group coordinates to soft Cu(I) centres due to a favourable chelate effect, while in 3-pyridinecarboxaldehyde, it remains uncoordinated. Upon chelation via N and O donors, 2-pyridinecarboxaldehyde resembles bipyridine or phenanthroline in terms of its bite angle and spectroscopic features. Such chelation can be easily challenged with coordinating anions like bromide, or more basic pyridines. A drastic change in the MLCT absorption signals the decomplexation of the `C=O’ group. The observed results point out that the Cu(I) centre can readily exchange the hard `O’ donor for softer ligands.

  17. Identification of LasR Ligands through a Virtual Screening Approach

    DEFF Research Database (Denmark)

    Skovstrup, Søren; Le Quement, Sebastian Thordal; Hansen, Thomas;

    2013-01-01

    With the widespread occurrence of bacterial resistance to antibiotics, the development of new strategies beyond conventional treatments is a pursuit taken by public health institutions worldwide. LasR, a transcription factor that controls quorum sensing in Pseudomonas aeruginosa, has emerged...... as an attractive therapeutic target for the next generation of antimicrobial agents. In the present study, a virtual screening workflow combining pharmacophore‐ and structure‐based approaches was used to identify new LasR ligands. Five novel inducers and three inhibitors of LasR activity were validated...... experimentally by use of a cell‐based assay. Interestingly, these compounds are molecularly distinct from the native signal molecule, N‐3‐oxododecanoyl‐L‐homoserine lactone (OHN), and may serve as lead structures for the design of new drugs. The binding modes of these compounds to the OHN binding site in Las...

  18. Analytical developments for screening of lanthanides/ligands interactions

    International Nuclear Information System (INIS)

    This work investigates the potential of hyphenated capillary electrophoresis and inductively coupled mass spectrometry to classify different ligands according to their europium binding affinity in a hydro-organic medium. On the one hand, this method enables to evaluate the affinity of phosphorus-containing ligands in less than two hours and using less than 15 ng of ligand. On the other hand, complexation constants could be determined. The results are in excellent agreement with the values obtained by spectrophotometric titrations.Moreover, a library of copolymers for solid/liquid extraction of europium is investigated. The extraction protocol enables to classify copolymers according to their europium affinity in a hydro-organic medium. This screening requires 60 mg of copolymers. For the most promising recognition properties and selectivity La3+/Eu3+/Lu3+ are evaluated. (author)

  19. Ligand Binding Thermodynamics in Drug Discovery: Still a Hot Tip?

    Science.gov (United States)

    Geschwindner, Stefan; Ulander, Johan; Johansson, Patrik

    2015-08-27

    The use of ligand binding thermodynamics has been proposed as a potential success factor to accelerate drug discovery. However, despite the intuitive appeal of optimizing binding enthalpy, a number of factors complicate routine use of thermodynamic data. On a macroscopic level, a range of experimental parameters including temperature and buffer choice significantly influence the observed thermodynamic signatures. On a microscopic level, solute effects, structural flexibility, and cooperativity lead to nonlinear changes in enthalpy. This multifactorial character hides essential enthalpy contributions of intermolecular contacts, making them experimentally nonobservable. In this perspective, we present three case studies, reflect on some key factors affecting thermodynamic signatures, and investigate their relation to the hydrophobic effect, enthalpy-entropy compensation, lipophilic ligand efficiency, and promiscuity. The studies highlight that enthalpy and entropy cannot be used as direct end points but can together with calculations increase our understanding of ligand binding and identify interesting outliers that do not behave as expected.

  20. Coordination chemistry of N-heterocyclic nitrenium-based ligands.

    Science.gov (United States)

    Tulchinsky, Yuri; Kozuch, Sebastian; Saha, Prasenjit; Mauda, Assaf; Nisnevich, Gennady; Botoshansky, Mark; Shimon, Linda J W; Gandelman, Mark

    2015-05-01

    Comprehensive studies on the coordination properties of tridentate nitrenium-based ligands are presented. N-heterocyclic nitrenium ions demonstrate general and versatile binding abilities to various transition metals, as exemplified by the synthesis and characterization of Rh(I) , Rh(III) , Mo(0) , Ru(0) , Ru(II) , Pd(II) , Pt(II) , Pt(IV) , and Ag(I) complexes based on these unusual ligands. Formation of nitrenium-metal bonds is unambiguously confirmed both in solution by selective (15) N-labeling experiments and in the solid state by X-ray crystallography. The generality of N-heterocyclic nitrenium as a ligand is also validated by a systematic DFT study of its affinity towards all second-row transition and post-transition metals (Y-Cd) in terms of the corresponding bond-dissociation energies.