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Membrane and Immunity
Responsable: Franck Fieschi (Professor, UJF) Group:
IntroductionMany steps of cell/cell, host/pathogen interactions and cellular activation are required to access to the immune response. Those events require a broad range of membrane proteins that are often specific of a peculiar cell type. Those proteins of primary relevance to human health (pathogen receptors, chemokine receptors, adhesion molecules) constitute challenging projects for academic research but also for pharmaceutical applications. Despite the difficulties linked to that type of project, considering the methodological pitfalls in eukaryotic membrane protein research, we have initiated three scientific projects in this context:
NADPH oxidase complex: a membrane multi component enzymatic complex
During a bacterial infection, the neutrophil NADPH oxidase initiates the destruction of the phagocyted microorganisms at the infection site by the production of large amounts of superoxide ions and derived products (H2O2, HOCl). The toxicity of the produced species requires a tight regulation of this system by the cell. This macromolecular complex comprises a membrane component the b558 flavocytochrome and several cytosolic proteins. Consecutively to an activation signal the complex associates at the membrane and initiates the electron transfer from NADPH to O2. We investigate with a structural and functional approach the various components of the system as well as some of the steps leading to the structural modifications involved in the complex assembly at the membrane and to the activation of the enzymatic reaction. Collaborations:
Host-pathogen interactions
During HIV infection, the virus is first captured by the muquous epithelia and then is transported to the lymphoid organs where it reaches its major cellular targets: the T lymphocytes. At the level of the epithelia, the virus is captured via the interaction of its surface glycoproteins with a membrane lectin expressed on the dendritic cell surface: DC-SIGN. Once attached, the intact virus is driven by these dendritic cells to the T lymphocytes. The virus can then infect the target cell through the interaction with several cell receptors including CXCR4 or CCR5. These various proteins are involved in the virus capture, transport and infection and constitute the subject of either methodological developments or of structural and functional studies. A significant effort has been placed in the development of an overexpression strategy for these G protein coupled receptors, CXCR4 and CCR5. As for DC-SIGN, we characterize the biological and structural properties of this oligomeric receptor and investigate the properties of its interaction with its targets. Collaborations involving several groups from the Institut Pasteur, Paris, on the DC-SIGN project allowed the study of other pathologies in which DC-SIGN has been shown to be involved, including the Dengue, virus C hepatitis or the Cytomegalovirus. Finally, a collaboration with a saccharide chemist group in Spain enables the development of multivalent molecules (glycodendrimers) specific to DC-SIGN in order to inhibit the virus/DC-SIGN interaction or to develop functionalised vectors to target the dendritic cells. Other lectins from dendritic cells present interaction properties with pathogens. Among these lectins, we work also on Langerine. This specific lectin of Langerhans cells is also able to recognize HIV. Moreover, it is involved in the formation of an original organite, the Birbeck granule. We are studying the physiologic ligands of this lectin, its structural properties as well as its role in Birbeck granule formation. Collaborations:
Related platformThe MP3 platform (Membrane Protein Purification Platform) corresponds to four protein purification modules of the Akta Xpress type, dedicated to membrane protein purification. Person in charge of the platform : Michel Thépaut. Picture of group
From left to right: EC, IS, MT, LS (back), IH (front), CJB, AP, FF and CV. Publications since 2007Sattin S, Daghetti A, Thépaut M, Berzi A, Sánchez-Navarro M, Tabarani G, Rojo J, Fieschi F, Clerici M, Bernardi A (2010) ACS Chem Biol. 5, 301-312. Inhibition of DC-SIGN-mediated HIV infection by a linear trimannoside mimic in a tetravalent presentation. Arcemisbéhère L, Sen T, Boudier L, Balestre MN, Gaibelet G, Detouillon E, Orcel H, Mendre C, Rahmeh R, Granier S, Vivès C, Fieschi F, Damian M, Durroux T, Banères JL, Mouillac B (2010) J Biol Chem. 285, 6337-6347. Leukotriene BLT2 receptor monomers activate the G(i2) GTP-binding protein more efficiently than dimers. Marcoux J, Thierry E, Vivès C, Signor L, Fieschi F, Forest E (2010) J Am Soc Mass Spectrom 21, 76-79. Investigating alternative acidic proteases for H/D exchange coupled to mass spectrometry: plasmepsin 2 but not plasmepsin 4 is active under quenching conditions. Durand D, Vivès C, Cannella D, Pérez J, Pebay-Peyroula E, Vachette P, Fieschi F (2010) J Struct Biol. 169, 45-53. NADPH oxidase activator p67(phox) behaves in solution as a multidomain protein with semi-flexible linkers. Tabarani G, Thepaut M, Stroebel D, Ebel C, Vives C, Vachette P, Durand D, and Fieschi F. (2009) J. Biol. Chem. 284, 21229-21240. DC-SIGN neck domain is a pH-sensor controlling oligomerization: saxs and hydrodynamic studies of extracellular domain. Thépaut, M, Valladeau, V, Nurisso, A, Kahn, R, Arnou, A, Vivès, C, Saeland, S, Ebel, C, Monnier, C, Dezutter-Dambuyant, C, Imberty, A and Fieschi F. (2009) Biochemistry 48, 2684-2698. Structural studies of langerin and birbeck Granules: a macromolecular organisation model. Marcoux, J, Man, P, Castellan, M, Vivès, C, Forest, E, and Fieschi F (2009) FEBS Letters 583, 835-840. Conformational changes in p47phox upon activation highlighted by mass spectrometry coupled to Hydrogen/Deuterium exchange and limited proteolysis. Angulo, J, Diaz, I, Reina, J, Tabarani, G, Fieschi, F, Rojo, J, and Nieto P (2008). ChemBioChem 9, 2225-7. Saturation transfer difference (STD) NMR spectroscopy characterization of dual binding mode of a mannose dissacharide to DC-SIGN. Timpano, G, Tabarani, G, Anderluh, M, Invernizzi, D, Vasile, F, Potenza, D, Nieto, P, Rojo, J, Fieschi, F and Bernardi, A. (2008) ChemBioChem,9, 1921-30. Synthesis of novel DC-SIGN ligands with an a-fucosylamide anchor. Reina, J, Diaz, I, Nieto, P, Campillo, N, Paez, J, Tabarani, G, Fieschi, F, and Rojo, J (2008) Organic and Biomolecular Chemistry, 6, 2743-54. Docking, Synthesis, and NMR studies of mannosyl trisaccharide ligands for DC-SIGN Lectin. Thepaut, M, Vives, C, Pompidor, G, Kahn, R, and Fieschi, F. (2008) Acta crystallographica. F. 64, 115-8. Overproduction, purification and preliminary crystallographic analysis of the Carbohydrate Recognition Domain of human Langerin. Reina, J, Sattin, S, Invernizzi, D, Mari, S, Martinez-Prats, L, Tabarani, G, Fieschi, F, Delgado, R, Nieto, D, Rojo, J, and Bernardi, A. (2007) ChemMedChem. 2, 1030-1036. 1,2-Mannobioside Mimic: Synthesis, DC-SIGN interaction by NMR and Docking, and Antiviral Activity. Reina, J, Maldonado, OS, Tabarani, G, Fieschi, F, and Rojo J (2007) Bioconjugate Chemistry. 18, 963-9. Mannose glycoconjugates functionalized at positions 1 and 6. Binding analysis to DC-SIGN using Biosensors. Li, XJ, Fieschi, F, Paclet, MH, Grunwald, D, Campion, Y, Morel, F and Stasia, MJ (2007) Journal of Leukocyte Biology 81, 238-249. Leu505 of Nox2 is crucial for optimal p67phox-dependent activation of the flavocytochrome b558 during phagocytic NADPH oxidase assembly. |
