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Martin J Stone - One of the best experts on this subject based on the ideXlab platform.
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structural basis of Receptor sulfotyrosine recognition by a cc Chemokine the n terminal region of CCR3 bound to ccl11 eotaxin 1
Structure, 2014Co-Authors: Christopher J Millard, Justin P Ludeman, Meritxell Canals, Jessica L Bridgford, Mark G Hinds, Daniel John Clayton, Arthur Christopoulos, Richard J Payne, Martin J StoneAbstract:Summary Trafficking of leukocytes in immune surveillance and inflammatory responses is activated by Chemokines engaging their Receptors. Sulfation of tyrosine residues in peptides derived from the eosinophil Chemokine Receptor CCR3 dramatically enhances binding to cognate Chemokines. We report the structural basis of this recognition and affinity enhancement. We describe the structure of a CC Chemokine (CCL11/eotaxin-1) bound to a fragment of a Chemokine Receptor: residues 8–23 of CCR3, including two sulfotyrosine residues. We also show that intact CCR3 is sulfated and sulfation enhances Receptor activity. The CCR3 sulfotyrosine residues form hydrophobic, salt bridge and cation-π interactions with residues that are highly conserved in CC Chemokines. However, the orientation of the Chemokine relative to the Receptor N terminus differs substantially from those observed for two CXC Chemokines, suggesting that initial binding of the Receptor sulfotyrosine residues guides subsequent steps in Receptor activation, thereby influencing the Receptor conformational changes and signaling.
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Structural Basis of Receptor Sulfotyrosine Recognition by a CC Chemokine: The N-Terminal Region of CCR3 Bound to CCL11/Eotaxin-1
Structure (London England : 1993), 2014Co-Authors: Christopher J Millard, Justin P Ludeman, Meritxell Canals, Jessica L Bridgford, Mark G Hinds, Daniel John Clayton, Arthur Christopoulos, Richard J Payne, Martin J StoneAbstract:Trafficking of leukocytes in immune surveillance and inflammatory responses is activated by Chemokines engaging their Receptors. Sulfation of tyrosine residues in peptides derived from the eosinophil Chemokine Receptor CCR3 dramatically enhances binding to cognate Chemokines. We report the structural basis of this recognition and affinity enhancement. We describe the structure of a CC Chemokine (CCL11/eotaxin-1) bound to a fragment of a Chemokine Receptor: residues 8–23 of CCR3, including two sulfotyrosine residues. We also show that intact CCR3 is sulfated and sulfation enhances Receptor activity. The CCR3 sulfotyrosine residues form hydrophobic, salt bridge and cation-p interactions with residues that are highly conserved in CC Chemokines. However, the orientation of the Chemokine relative to the Receptor N terminus differs substantially from those observed for two CXC Chemokines, suggesting that initial binding of the Receptor sulfotyrosine residues guides subsequent steps in Receptor activation, thereby influencing the Receptor conformational changes and signaling.
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Tyrosine sulfation influences the Chemokine binding selectivity of peptides derived from Chemokine Receptor CCR3.
Biochemistry, 2011Co-Authors: John Z. Zhu, Justin P Ludeman, Christopher J Millard, Richard J Payne, Levi S. Simpson, Daniel Clayton, Theodore S. Widlanski, Martin J StoneAbstract:The interactions of Chemokines with their G protein-coupled Receptors play critical roles in the control of leukocyte trafficking in normal homeostasis and in inflammatory responses. Tyrosine sulfation is a common post-translational modification in the amino-terminal regions of Chemokine Receptors. However, tyrosine sulfation of Chemokine Receptors is commonly incomplete or heterogeneous. To investigate the possibility that differential sulfation of two adjacent tyrosine residues could bias the responses of Chemokine Receptor CCR3 to different Chemokines, we have studied the binding of three Chemokines (eotaxin-1/CCL11, eotaxin-2/CCL24, and eotaxin-3/CCL26) to an N-terminal CCR3-derived peptide in each of its four possible sulfation states. Whereas the nonsulfated peptide binds to the three Chemokines with approximately equal affinity, sulfation of Tyr-16 gives rise to 9-16-fold selectivity for eotaxin-1 over the other two Chemokines. Subsequent sulfation of Tyr-17 contributes additively to the affinity for eotaxin-1 and eotaxin-2 but cooperatively to the affinity for eotaxin-3. The doubly sulfated peptide selectively binds to both eotaxin-1 and eotaxin-3 approximately 10-fold more tightly than to eotaxin-2. Nuclear magnetic resonance chemical shift mapping indicates that these variations in affinity probably result from only subtle differences in the Chemokine surfaces interacting with these Receptor peptides. These data support the proposal that variations in sulfation states or levels may regulate the responsiveness of Chemokine Receptors to their cognate Chemokines.
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Regulation of Chemokine recognition by site-specific tyrosine sulfation of Receptor peptides.
Chemistry & biology, 2009Co-Authors: Levi S. Simpson, John Z. Zhu, Theodore S. Widlanski, Martin J StoneAbstract:Summary Sulfation of tyrosine is a common posttranslational modification of secreted proteins that influences numerous physiological and pathological processes. Studies of tyrosine sulfation have been hindered by the difficulty of introducing sulfate groups at specific positions of peptides and proteins. Here we report a general strategy for synthesis of peptides containing sulfotyrosine at one or more specific position(s). The approach provides a substantial improvement in both yield and convenience over existing methods. Using synthetic sulfopeptides derived from the Chemokine Receptor CCR3, we demonstrate that sulfation enhances affinity for the Chemokine eotaxin by ∼7-fold or more than 28-fold, depending on which of two adjacent tyrosine residues is sulfated. The synthetic methodology will substantially enhance efforts to understand the functional and structural consequences of protein tyrosine sulfation.
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NMR solution structure and backbone dynamics of the CC Chemokine eotaxin-3.
Biochemistry, 2001Co-Authors: Kristen L. Mayer, Michael R. Mayer, Martin J StoneAbstract:Eotaxin-3 is one of three related Chemokines that specifically activate Chemokine Receptor CCR3. We report the 3D structure and backbone dynamics of eotaxin-3 determined by NMR spectroscopy. Eotaxin-3 is monomeric under the conditions in this study and consists of an unstructured N-terminus before the first two conserved cysteine residues, an irregularly structured N-loop following the second conserved cysteine, a single turn of 3(10)-helix, a three-stranded antiparallel beta-sheet, an alpha-helix, and an unstructured C-terminal tail. As in other Chemokines, the alpha-helix packs against one face of the beta-sheet. The average backbone and heavy atom rmsd values of the 20 structures (residues 9-65) are 0.44 and 1.01 A, respectively. A comparison between the structures of eotaxin-3 and related Chemokines suggests that the electrostatic potential in the vicinity of a surface groove and the structure of the beta2-beta3 turn may be important for maintaining Receptor specificity. The backbone dynamics of eotaxin-3 were determined from 15N NMR relaxation data using the extended model free dynamics formalism. Large amplitude motions on the picosecond to nanosecond time scale were observed in both termini and in some residues in the N-loop, the beta1-beta2 turn, and the beta3 strand; the location of these residues suggests a possible role for dynamics in Receptor binding and activation. In contrast to eotaxin, eotaxin-3 exhibits no substantial mobility on the microsecond to millisecond time scale.
Amato De Paulis - One of the best experts on this subject based on the ideXlab platform.
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Dysregulation of the IgE/FcϵRI network in HIV-1 infection
The Journal of allergy and clinical immunology, 2001Co-Authors: Gianni Marone, Giovanni Florio, Angelica Petraroli, Amato De PaulisAbstract:Abstract Serum IgE levels are increased in adults and children with HIV-1 infection and could be a marker of poor prognosis. Allergic reactions and adverse reactions to drugs are also increased in HIV-1–infected individuals. An imbalance between a T H 1-like and a T H 2-like cytokine profile has been documented in HIV-1 infection. We have found that HIV-1 gp120 from different clades is a potent stimulus for histamine and cytokine (IL-4 and IL-13) release from basophils. Gp120 acts as a viral superantigen, interacting with the V H 3 region of IgE to induce mediator release from human FcϵRI + cells. Human basophils and mast cells express the Chemokine Receptor CCR3, which binds the Chemokines eotaxin and RANTES. By interacting with the CCR3 Receptor on FcϵRI + cells, HIV-1 Tat protein is a potent chemoattractant for human basophils and lung mast cells. Tat protein also induced IL-4 and IL-13 release from basophils. Preincubation of basophils with Tat protein upregulated the surface expression of the CCR3 Receptor. Extracellular Tat can influence the directional migration of human FcϵRI + cells, the expression of Chemokine Receptor CCR3, and the release of T H 2 cytokines. Because Tat protein is actively released by HIV-1–infected cells, our results indicate a novel mechanism by which FcϵRI + cells are rendered more susceptible to infection with CCR3-tropic HIV-1 isolates; that is, two HIV-1 proteins, gp120 and Tat, trigger the release of cytokines critical for T H 2 polarization from FcϵRI + cells, and Tat upregulates β-Chemokine Receptor CCR3 on these cells. (J Allergy Clin Immunol 2001;107:22-30.)
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Expression of the Chemokine Receptor CCR3 on human mast cells.
International archives of allergy and immunology, 2001Co-Authors: Amato De Paulis, Gianni Marone, Francesco Annunziato, Luisa Di Gioia, Sergio Romagnani, Maria Carfora, Chiara Beltrame, Paola RomagnaniAbstract:Background: The aim of this study was to investigate whether human mast cells express functional active CCR3 Receptors, which are activated by CC Chemokines. These ligands include t
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Role of human FcεRI+ cells in HIV-1 infection
Immunological reviews, 2001Co-Authors: Gianni Marone, Giovanni Florio, Angelica Petraroli, Massimo Triggiani, Amato De PaulisAbstract:Summary: Enhanced serum IgE levels in adults and children with HIV-1 infection could be a marker of poor prognosis. HIV-1 infection is believed to involve a switch toward a “TH2-like” cytokine pattern. HIV-1 gp120 from different clades is a potent stimulus for histamine release from human basophils and mast cells. Gp120 also induces IL-4 and IL-13 synthesis from basophils. It functions as a viral superantigen by interacting with the VH3 region of IgE to induce mediator release from human FceRI+ cells. The Chemokine Receptor CCR3, which binds the Chemokines eotaxin and RANTES, is expressed by basophils and lung mast cells. By interacting with the CCR3 Receptor on FceRI+ cells, HIV-1 Tat protein is a potent chemoattractant for basophils and lung mast cells. Tat protein also induces IL-4 and IL-13 release from basophils. Incubation of basophils with Tat protein upregulates the surface expression of the CCR3 Receptor, a co-Receptor of HIV-1 infection. Extracellular Tat affects the directional migration of human FceRI+ cells, CCR3 expression and TH2 cytokines release. We have shown that HIV-1 proteins gp120 and Tat trigger the release of cytokines critical for TH2 polarization from FceRI+ cells through two distinct mechanisms. In addition, Tat upregulates the β-Chemokine Receptor CCR3, making FceRI+ cells more susceptible to infection with CCR3 tropic HIV-1 isolates. This paper is dedicated to Rita Levi-Montalcini who first suggested an involvement of FceRI+ cells in HIV-1 infection. This work was supported by a grant from the Istituto Superiore Sanita (AIDS project 40B.64 and 40A.67), CNR (Target project Biotechnology No. 99.00216.PF31 and No. 99.00401. PF49) and MURST (Rome, Italy).
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Tat protein is an HIV-1-encoded beta-Chemokine homolog that promotes migration and up-regulates CCR3 expression on human Fc epsilon RI+ cells.
Journal of immunology (Baltimore Md. : 1950), 2000Co-Authors: Amato De Paulis, Maria Carfora, R. De Palma, L. Di Gioia, Nella Prevete, Giovanna Tosi, Roberto S. Accolla, Gianni MaroneAbstract:Human basophils and mast cells express the Chemokine Receptor CCR3, which binds the Chemokines eotaxin and RANTES. HIV-1 Tat protein is a potent chemoattractant for basophils and lung mast cells obtained from healthy individuals seronegative for Abs to HIV-1 and HIV-2. Tat protein induced a rapid and transient Ca 2+ influx in basophils and mast cells, analogous to β-Chemokines. Tat protein neither induced histamine release from human basophils and mast cells nor increased IL-3-stimulated histamine secretion from basophils. The chemotactic activity of Tat protein was blocked by preincubation of FceRI + cells with anti-CCR3 Ab. Preincubation of Tat with a mAb anti-Tat (aa 1–86) blocked the migration induced by Tat. In contrast, a mAb specific for the basic region (aa 46–60) did not inhibit the chemotactic effect of Tat protein. Tat protein or eotaxin desensitized basophils to a subsequent challenge with the autologous or the heterologous stimulus. Preincubation of basophils with Tat protein up-regulated the level of CCR3 mRNA and the surface expression of the CCR3 Receptor. Tat protein is the first identified HIV-1-encoded β-Chemokine homologue that influences the directional migration of human FceRI + cells and the expression of surface Receptor CCR3 on these cells.
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Mechanisms of IgE elevation in HIV-1 infection.
Critical reviews in immunology, 2000Co-Authors: Gianni Marone, Giovanni Florio, Angelica Petraroli, Massimo Triggiani, Amato De PaulisAbstract:Serum IgE levels are high in adults and children with HIV-1 infection and could be a marker of poor prognosis. Allergic reactions and adverse reactions to drugs also tend to increase in HIV-1-infected individuals. An imbalance between a "T(H)1-like" and a "T(H)2-like" cytokine profile has been documented in HIV-1 infection. We have demonstrated that HIV-1 gp 120 from different clades is a stimulus for histamine and cytokine (IL-4 and IL-13) release from basophils. Gp 120 acts as a viral superantigen, interacting with the V(H)3 region of IgE to induce mediator release from human Fc epsilonRI+ cells. Human basophils and mast cells express the Chemokine Receptor CCR3, which binds the Chemokines eotaxin and RANTES. By interacting with the CCR3 Receptor on Fc epsilonRI+ cells, HIV-I Tat protein is a potent chemoattractant for human basophils and lung mast cells. Preincubation of basophils with Tat protein upregulates mRNA CCR3 and the surface expression of this Chemokine Receptor. Tat also induces IL-4 and IL-13 release from basophils. Extracellular Tat can influence the directional migration of human Fc epsilonRI+ cells, the expression of Chemokine Receptor CCR3, and the release of T(H)2 cytokines. Our results indicate two novel mechanisms by which two HIV-1 proteins, gp120 and Tat, trigger the release of cytokines critical for T(H)2 polarization from human Fc epsilonRI+ cells.
Ulf Forssmann - One of the best experts on this subject based on the ideXlab platform.
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Chemokine Receptor antagonists: a novel therapeutic approach
2004Co-Authors: Jörn Elsner, Ulf Forssmann, S.e. Escher, Jörn Elsner, S.e. Escher, Jçrn ElsnerAbstract:The aim of this review is to give an overview of the role of Chemokines, particularly ligands of the CC Chemokine Receptor CCR3, in allergic diseases and to show the new concept in the treatment of allergies using Chemokine Receptor antagonists. Allergic diseases such as allergic asthma, allergic rhinitis and atopic dermatitis are characterized by a complex interaction of different cell types and mediators. Among this, Th2 cells, mast cells, basophils and eosinophils are found in the inflamed tissue due to the attraction of Chemokines. Of all the known Chemokine Receptors, the Chemokine Receptor CCR3 seems to play the major role in allergic diseases which is supported by the detection of this Receptor on the cell types mentioned above. Therefore, academic and industrial research focus on compounds to block this Receptor. To date, certain Chemokine Receptor antagonists derived from peptides and small molecules exist to block the Chemokine Receptor CCR3. However, the in vivo data about these compounds and the mechanisms of Receptor interaction are poorly understood, as yet. For the development of additional Chemokine Receptor antagonists, more details about the interaction between the ligands and their Receptors are required. Therefore, additional studies will lead to the identification of novel CCR3 Chemokine Receptor antagonists, which can be therapeutically used in allergic asthma, allergic rhinitis, and atopic dermatitis.
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Chemokine Receptor antagonists: a novel therapeutic approach in allergic diseases.
Allergy, 2004Co-Authors: Jörn Elsner, S.e. Escher, Jörn Elsner, S.e. Escher, Ulf ForssmannAbstract:The aim of this review is to give an overview of the role of Chemokines, particularly ligands of the CC Chemokine Receptor CCR3, in allergic diseases and to show the new concept in the treatment of allergies using Chemokine Receptor antagonists. Allergic diseases such as allergic asthma, allergic rhinitis and atopic dermatitis are characterized by a complex interaction of different cell types and mediators. Among this, Th2 cells, mast cells, basophils and eosinophils are found in the inflamed tissue due to the attraction of Chemokines. Of all the known Chemokine Receptors, the Chemokine Receptor CCR3 seems to play the major role in allergic diseases which is supported by the detection of this Receptor on the cell types mentioned above. Therefore, academic and industrial research focus on compounds to block this Receptor. To date, certain Chemokine Receptor antagonists derived from peptides and small molecules exist to block the Chemokine Receptor CCR3. However, the in vivo data about these compounds and the mechanisms of Receptor interaction are poorly understood, as yet. For the development of additional Chemokine Receptor antagonists, more details about the interaction between the ligands and their Receptors are required. Therefore, additional studies will lead to the identification of novel CCR3 Chemokine Receptor antagonists, which can be therapeutically used in allergic asthma, allergic rhinitis, and atopic dermatitis.
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eotaxin 2 a novel cc Chemokine that is selective for the Chemokine Receptor CCR3 and acts like eotaxin on human eosinophil and basophil leukocytes
Journal of Experimental Medicine, 1997Co-Authors: Ulf Forssmann, Mariagrazia Uguccioni, Pius Loetscher, Clemens A Dahinden, Hanno Langen, Marcus ThelenAbstract:A novel human CC Chemokine consisting of 78 amino acids and having a molecular mass of 8,778.3 daltons (VVIPSPCCMF FVSKRIPENR VVSYQLSSRS TCLKAGVIFT TKKGQQ SCGD PKQEWVQRYM KNLDAKQKKA SPRARAVA) was isolated together with three minor COOH-terminally truncated variants with 73, 75, and 76 residues. The new Chemokine was termed eotaxin-2 because it is functionally very similar to eotaxin. In terms of structure, however, eotaxin and eotaxin-2 are rather distant, they share only 39% identical amino acids and differ almost completely in the NH2-terminal region. Eotaxin-2 induced chemotaxis of eosinophils as well as basophils, with a typically bimodal concentration dependence, and the release of histamine and leukotriene C4 from basophils that had been primed with IL-3. In all assays, eotaxin-2 had the same efficacy as eotaxin, but was somewhat less potent. The migration and the release responses were abrogated in the presence of a monoclonal antibody that selectively blocks the eotaxin Receptor, CCR3, indicating that eotaxin-2, like eotaxin, acts exclusively via CCR3. Receptor usage was also studied in desensitization experiments by measuring [Ca2+]i changes in eosinophils. Complete cross-desensitization was observed between eotaxin-2, eotaxin and MCP-4 confirming activation via CCR3. No Ca2+ mobilization was obtained in neutrophils, monocytes and lymphocytes, in agreement with the lack of chemotactic responsiveness. Intradermal injection of eotaxin-2 in a rhesus monkey (100 or 1,000 pmol per site) induced a marked local infiltration of eosinophils, which was most pronounced in the vicinity of postcapillary venules and was comparable to the effect of eotaxin.
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eotaxin 2 a novel cc Chemokine that is selective for the Chemokine Receptor CCR3 and acts like eotaxin on human eosinophil and basophil leukocytes
Journal of Experimental Medicine, 1997Co-Authors: Ulf Forssmann, Mariagrazia Uguccioni, Pius Loetscher, Clemens A Dahinden, Hanno Langen, Marcus Thelen, Marco BaggioliniAbstract:A novel human CC Chemokine consisting of 78 amino acids and having a molecular mass of 8,778.3 daltons (VVIPSPCCMF FVSKRIPENR VVSYQLSSRS TCLKAGVIFT TKKGQQ SCGD PKQEWVQRYM KNLDAKQKKA SPRARAVA) was isolated together with three minor COOH-terminally truncated variants with 73, 75, and 76 residues. The new Chemokine was termed eotaxin-2 because it is functionally very similar to eotaxin. In terms of structure, however, eotaxin and eotaxin-2 are rather distant, they share only 39% identical amino acids and differ almost completely in the NH2-terminal region. Eotaxin-2 induced chemotaxis of eosinophils as well as basophils, with a typically bimodal concentration dependence, and the release of histamine and leukotriene C4 from basophils that had been primed with IL-3. In all assays, eotaxin-2 had the same efficacy as eotaxin, but was somewhat less potent. The migration and the release responses were abrogated in the presence of a monoclonal antibody that selectively blocks the eotaxin Receptor, CCR3, indicating that eotaxin-2, like eotaxin, acts exclusively via CCR3. Receptor usage was also studied in desensitization experiments by measuring [Ca2+]i changes in eosinophils. Complete cross-desensitization was observed between eotaxin-2, eotaxin and MCP-4 confirming activation via CCR3. No Ca2+ mobilization was obtained in neutrophils, monocytes and lymphocytes, in agreement with the lack of chemotactic responsiveness. Intradermal injection of eotaxin-2 in a rhesus monkey (100 or 1,000 pmol per site) induced a marked local infiltration of eosinophils, which was most pronounced in the vicinity of postcapillary venules and was comparable to the effect of eotaxin.
Jörn Elsner - One of the best experts on this subject based on the ideXlab platform.
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Donor to donor variation of the Chemokine Receptor CCR1 on human eosinophils and its prolonged internalization by RANTES
The Journal of Allergy and Clinical Immunology, 2004Co-Authors: Yasmin Dulkys, S.e. Escher, Jörn ElsnerAbstract:Abstract Rationale Whereas recent studies underlies the fundamental importance of the CC Chemokine Receptor CCR3 for the recruitment of eosinophils in allergic diseases controversial data about the expression of CCR1 on eosinophils exist. Therefore, the purpose of this study was to investigate the expression and regulation of CCR1 on eosinophils. Methods Flow cytometric analysis of whole blood eosinophils or CD16-negative selected freshly isolated eosinophils from atopic and non-atopic donors using an anti-CCR1 mAb. Results We found that 32% of the donors (n=24) were positive and 68% (n=49) were negative to express CCR1. Pre-incubation of isolated CCR1-positive eosinophils for 24 h with IL-3 (p=0.021), RANTES (p Conclusions A subpopulation of human donors are able to express CCR1 on the surface of eosinophils which helps to explain the different response of eosinophils to MIP-1α in previous studies. It is tempting to speculate whether the higher positive charge of RANTES in comparison to its overall negative charge of MIP-1α is responsible for the prolonged CCR1 internalization in response to RANTES.
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Chemokine Receptor antagonists: a novel therapeutic approach
2004Co-Authors: Jörn Elsner, Ulf Forssmann, S.e. Escher, Jörn Elsner, S.e. Escher, Jçrn ElsnerAbstract:The aim of this review is to give an overview of the role of Chemokines, particularly ligands of the CC Chemokine Receptor CCR3, in allergic diseases and to show the new concept in the treatment of allergies using Chemokine Receptor antagonists. Allergic diseases such as allergic asthma, allergic rhinitis and atopic dermatitis are characterized by a complex interaction of different cell types and mediators. Among this, Th2 cells, mast cells, basophils and eosinophils are found in the inflamed tissue due to the attraction of Chemokines. Of all the known Chemokine Receptors, the Chemokine Receptor CCR3 seems to play the major role in allergic diseases which is supported by the detection of this Receptor on the cell types mentioned above. Therefore, academic and industrial research focus on compounds to block this Receptor. To date, certain Chemokine Receptor antagonists derived from peptides and small molecules exist to block the Chemokine Receptor CCR3. However, the in vivo data about these compounds and the mechanisms of Receptor interaction are poorly understood, as yet. For the development of additional Chemokine Receptor antagonists, more details about the interaction between the ligands and their Receptors are required. Therefore, additional studies will lead to the identification of novel CCR3 Chemokine Receptor antagonists, which can be therapeutically used in allergic asthma, allergic rhinitis, and atopic dermatitis.
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Chemokine Receptor antagonists: a novel therapeutic approach in allergic diseases.
Allergy, 2004Co-Authors: Jörn Elsner, S.e. Escher, Jörn Elsner, S.e. Escher, Ulf ForssmannAbstract:The aim of this review is to give an overview of the role of Chemokines, particularly ligands of the CC Chemokine Receptor CCR3, in allergic diseases and to show the new concept in the treatment of allergies using Chemokine Receptor antagonists. Allergic diseases such as allergic asthma, allergic rhinitis and atopic dermatitis are characterized by a complex interaction of different cell types and mediators. Among this, Th2 cells, mast cells, basophils and eosinophils are found in the inflamed tissue due to the attraction of Chemokines. Of all the known Chemokine Receptors, the Chemokine Receptor CCR3 seems to play the major role in allergic diseases which is supported by the detection of this Receptor on the cell types mentioned above. Therefore, academic and industrial research focus on compounds to block this Receptor. To date, certain Chemokine Receptor antagonists derived from peptides and small molecules exist to block the Chemokine Receptor CCR3. However, the in vivo data about these compounds and the mechanisms of Receptor interaction are poorly understood, as yet. For the development of additional Chemokine Receptor antagonists, more details about the interaction between the ligands and their Receptors are required. Therefore, additional studies will lead to the identification of novel CCR3 Chemokine Receptor antagonists, which can be therapeutically used in allergic asthma, allergic rhinitis, and atopic dermatitis.
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The CC Chemokine Receptor 3 CCR3 is functionally expressed on eosinophils but not on neutrophils.
European journal of immunology, 2000Co-Authors: Renate Höchstetter, Yasmin Dulkys, Gustav Dobos, Daniela Kimmig, Alexander Kapp, Jörn ElsnerAbstract:The Chemokine subclasses differ in their biological activity to stimulate different kinds of effector cells via distinct Chemokine Receptors. Controversial results about the expression of the CC Chemokine Receptor CCR3 on the surface of human neutrophils have been described. To find out whether eosinophil contamination might be responsible for these diverse observations, CCR3 expression on highly purified neutrophils and eosinophils was investigated. We enriched neutrophils from a heterogeneous granulocyte population with immunomagnetic beads coated with various anti-CD52 monoclonal antibodies. This procedure was suitable to enrich neutrophils with a purity of up to 99.85%. Reverse transcriptase-PCR revealed that CCR3 mRNA was not expressed by CD52-negative selected neutrophils. In contrast to these cells, CCR3 mRNA could be detected in a heterogeneous granulocyte population and CD16-negative selected eosinophils. In addition, spectrofluorometric measurement of intracellular calcium concentration ([Ca2+]i) demonstrated that CD52-negative selected neutrophils did not show a transient [Ca2+]i increase following stimulation with the CCR3 ligand eotaxin, whereas the heterogeneous granulocyte population as well as eosinophils did respond. Therefore, previous studies demonstrating the expression of CCR3 on human neutrophils have to be re-evaluated because CCR3 mRNA detection on human neutrophils due to contamination by mRNA from eosinophils could not be excluded.
James E. Pease - One of the best experts on this subject based on the ideXlab platform.
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small molecule Receptor agonists and antagonists of CCR3 provide insight into mechanisms of Chemokine Receptor activation
Journal of Biological Chemistry, 2007Co-Authors: Emma L Wise, T J Williams, Cécile E. Duchesnes, Paula C A Da Fonseca, Rodger A Allen, James E. PeaseAbstract:Chemokine Receptor CCR3 is highly expressed by eosinophils and signals in response to binding of the eotaxin family of Chemokines, which are up-regulated in allergic disorders. Consequently, CCR3 blockade is of interest as a possible therapeutic approach for the treatment of allergic disease. We have described previously a bispecific antagonist of CCR1 and CCR3 named UCB35625 that was proposed to interact with the transmembrane residues Tyr-41, Tyr-113, and Glu-287 of CCR1, all of which are conserved in CCR3. Here, we show that cells expressing the CCR3 constructs Y113A and E287Q are insensitive to antagonism by UCB35625 and also exhibit impaired chemotaxis in response to CCL11/eotaxin, suggesting that these residues are important for antagonist binding and also Receptor activation. Furthermore, mutation of the residue Tyr-113 to alanine was found to turn the antagonist UCB35625 into a CCR3 agonist. Screens of small molecule libraries identified a novel specific agonist of CCR3 named CH0076989. This was able to activate eosinophils and transfectants expressing both wild-type CCR3 and a CCR1-CCR3 chimeric Receptor lacking the CCR3 amino terminus, indicating that this region of CCR3 is not required for CH0076989 binding. A direct interaction with the transmembrane helices of CCR3 was supported by mutation of the residues Tyr-41, Tyr-113, and Glu-287 that resulted in complete loss of CH0076989 activity, suggesting that the compound mimics activation by CCL11. We conclude that both agonists and antagonists of CCR3 appear to occupy overlapping sites within the transmembrane helical bundle, suggesting a fine line between agonism and antagonism of Chemokine Receptors.
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Asthma, allergy and Chemokines.
Current drug targets, 2006Co-Authors: James E. PeaseAbstract:In the human, Chemokines represent a structurally related family of more than forty cytokines which act on distinct subsets of leukocytes via specific G protein-coupled Receptors expressed on the cell surface. The induction of select repertoires of Chemokines following exposure to allergen provides a biological basis for the selective leukocyte recruitment, observed both clinically and experimentally. The Chemokine Receptor CCR3 is expressed on the cell surface of eosinophils, Th2 lymphocytes, basophils and mast cells and binds the Eotaxin family of Chemokines (CCL11, CCL24 and CCL26), whose production is upregulated following allergen challenge. Once recruited, eosinophils are a source of growth factors associated with tissue repair and remodelling and also have the ability to induce tissue damage, a capacity that extends from their traditional role in protecting the host against parasitic worms. Thus, impairment of their recruitment by selective blockade of the CCR3:eotaxin axis represents an attractive target for the therapeutic treatment of asthma. In this review, we will examine recent developments in the field and highlight the roles of other Chemokine:Chemokine Receptor axes implicated in leukocyte recruitment during allergic inflammation.
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Alanine scanning mutagenesis of the Chemokine Receptor CCR3 reveals distinct extracellular residues involved in recognition of the eotaxin family of Chemokines.
Molecular immunology, 2005Co-Authors: Cécile E. Duchesnes, Timothy J. Williams, Philip M. Murphy, James E. PeaseAbstract:Despite considerable differences in primary structure, the Chemokines eotaxin-1/CCL11, eotaxin-2/CCL24 and eotaxin-3/CCL26 signal via a single Receptor, CCR3, but exhibit different potencies and efficacies. To examine Receptor/ligand interactions in more detail, we performed alanine scanning mutagenesis of 21 charged residues within the extracellular loops (ECLs) of CCR3. Following transient expression in the L1.2 cell line, CCR3 mutants were assessed for their ability to be expressed at the cell surface, bind CCL11 and induce chemotactic responses to CCL11, CCL24 and CCL26. The majority of constructs were well expressed at the cell surface and bound CCL11 with low nanomolar affinity. Exceptions to this rule included the mutants E175A and E176A (ECL2) which were poorly expressed and responded weakly to all three ligands in chemotaxis assays. In contrast, the mutants K26 (amino-terminus) E179 and E180 (ECL2) responded in chemotaxis assays to CCL11 and CCL24, but not to CCL26. Mutation of residues in ECL3 was informative, with the D272A, K277A and D280A mutants exhibiting reduced chemotactic responses to two or more of the three ligands examined, despite being expressed on the cell surface at levels similar to WT CCR3. This suggests a major role for ECL3 in the recognition of all three eotaxins. In summary, distinct acidic and basic residues within CCR3 determine both Receptor expression and activation by the eotaxins. Determining how these Chemokines interact with their Receptor at the molecular level should increase our understanding of the process of Chemokine Receptor activation.
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The carboxyl terminus of the Chemokine Receptor CCR3 contains distinct domains which regulate chemotactic signaling and Receptor down‐regulation in a ligand‐dependent manner
European journal of immunology, 2005Co-Authors: Ian Sabroe, Victoria E L Stubbs, Annelies Jorritsma, Georgina Xanthou, Louise A. Jopling, Paul D. Ponath, Timothy J. Williams, Philip M. Murphy, James E. PeaseAbstract:The Chemokine Receptor CCR3 regulates the chemotaxis of leukocytes implicated in allergic disease, such as eosinophils. Incubation of eosinophils with CCL11, CCL13 or CCL5 resulted in a rapid decrease of cell-surface CCR3 which was replicated using CCR3 transfectants. Progressive truncation of the CCR3 C terminus by 15 amino acids produced three constructs, Δ340, Δ325 and Δ310. Δ340 and Δ325 were able to bind CCL11 with affinities similar to wild-type CCR3. Δ340 transfectants exhibited enhanced migration and reduced Receptor down-regulation in response to CCL11 and CCL13. Δ325 transfectants displayed chemotactic responses to CCL11 and CCL13 similar to wild-type CCR3, and had impaired down-regulation when stimulated with CCL13 but not CCL11. In contrast, neither the Δ325 nor Δ340 truncation affected chemotaxis or Receptor down-regulation induced by CCL5. Δ310 transfectants bound CCL11 poorly and were biologically inactive. Inhibitors of p38 mitogen-activated protein kinase and PI3-kinase antagonized eosinophil shape change responses and chemotaxis of transfectants to CCL11 and CCL13. In contrast, shape change but not chemotaxis was sensitive to inhibition of the extracellular signal-regulated kinase kinase pathway suggesting differential regulation of the two responses. Thus, the CCR3 C terminus contains distinct domains responsible for the regulation of Receptor desensitization and for coupling to chemotactic responses.
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Eotaxin and asthma
Current Opinion in Pharmacology, 2001Co-Authors: James E. Pease, Timothy J. WilliamsAbstract:Abstract Eotaxin is a small protein that is produced in the lungs of asthmatic patients and is a potent chemoattractant for eosinophils. Eotaxin, a CC Chemokine, stimulates the migration of eosinophils from the small blood vessels in the lungs by acting on the CC Chemokine Receptor CCR3, which is located on the leukocyte cell surface. In the past year, three low molecular weight compounds have been developed that can block this Receptor. Such compounds may be developed into orally available drugs aimed at preventing eosinophil recruitment and, hence, the pathogenesis associated with the activation of these cells within the lung tissue.