Imagerie spatio-temporelle de la dynamique des organelles et des endomembranes

Publications de l’équipe

Année de publication : 2008

Jean Salamero (2008 Nov 29)

[The < green-revolution > is underway].

Médecine sciences : M/S : 987-8 : DOI : 10.1051/medsci/20082411987 En savoir plus
Résumé

Replier
Sylvie Tourne, Blandine Maitre, Anthony Collmann, Emilie Layre, Sabrina Mariotti, François Signorino-Gelo, Caroline Loch, Jean Salamero, Martine Gilleron, Catherine Angénieux, Jean-Pierre Cazenave, Lucia Mori, Daniel Hanau, Germain Puzo, Gennaro De Libero, Henri de la Salle (2008 Mar 8)

Cutting edge: a naturally occurring mutation in CD1e impairs lipid antigen presentation.

Journal of immunology (Baltimore, Md. : 1950) : 3642-6 En savoir plus
Résumé

The human CD1a-d proteins are plasma membrane molecules involved in the presentation of lipid Ags to T cells. In contrast, CD1e is an intracellular protein present in a soluble form in late endosomes or lysosomes and is essential for the processing of complex glycolipid Ags such as hexamannosylated phosphatidyl-myo-inositol, PIM(6). CD1e is formed by the association of beta(2)-microglobulin with an alpha-chain encoded by a polymorphic gene. We report here that one variant of CD1e with a proline at position 194, encoded by allele 4, does not assist PIM(6) presentation to CD1b-restricted specific T cells. The immunological incompetence of this CD1e variant is mainly due to inefficient assembly and poor transport of this molecule to late endosomal compartments. Although the allele 4 of CD1E is not frequent in the population, our findings suggest that homozygous individuals might display an altered immune response to complex glycolipid Ags.

Replier

Année de publication : 2007

Stéphanie Miserey-Lenkei, François Waharte, Annick Boulet, Marie-Hélène Cuif, Danielle Tenza, Amed El Marjou, Graça Raposo, Jean Salamero, Laurent Héliot, Bruno Goud, Solange Monier (2007 Aug 30)

Rab6-interacting protein 1 links Rab6 and Rab11 function.

Traffic (Copenhagen, Denmark) : 1385-403 En savoir plus
Résumé

Rab11 and Rab6 guanosine triphosphatases are associated with membranes of the recycling endosomes (REs) and Golgi complex, respectively. Evidence indicates that they sequentially regulate a retrograde transport pathway between these two compartments, suggesting the existence of proteins that must co-ordinate their functions. Here, we report the characterization of two isoforms of a protein, Rab6-interacting protein 1 (R6IP1), originally identified as a Rab6-binding protein. R6IP1 also binds to Rab11A in its GTP-bound conformation. In interphase cells, R6IP1 is targeted to the Golgi in a Rab6-dependent manner but can associate with Rab11-positive compartments when the level of Rab11A is increased within the cells. Fluorescence resonance energy transfer analysis using fluorescence lifetime imaging shows that the overexpression of R6IP1 promotes an interaction between Rab11A and Rab6 in living cells. Accordingly, the REs marked by Rab11 and transferrin receptor are depleted from the cell periphery and accumulate in the pericentriolar area. However, endosomal and Golgi membranes do not appear to fuse with each other. We also show that R6IP1 function is required during metaphase and cytokinesis, two mitotic steps in which a role of Rab6 and Rab11 has been previously documented. We propose that R6IP1 may couple Rab6 and Rab11 function throughout the cell cycle.

Replier
Stéphanie Uzan-Gafsou, Huguette Bausinger, Fabienne Proamer, Solange Monier, Dan Lipsker, Jean-Pierre Cazenave, Bruno Goud, Henri de la Salle, Daniel Hanau, Jean Salamero (2007 Jun 1)

Rab11A controls the biogenesis of Birbeck granules by regulating Langerin recycling and stability.

Molecular biology of the cell : 3169-79 En savoir plus
Résumé

The extent to which Rab GTPases, Rab-interacting proteins, and cargo molecules cooperate in the dynamic organization of membrane architecture remains to be clarified. Langerin, a recycling protein accumulating in the Rab11-positive compartments of Langerhans cells, induces the formation of Birbeck granules (BGs), which are membrane subdomains of the endosomal recycling network. We investigated the role of Rab11A and two members of the Rab11 family of interacting proteins, Rip11 and RCP, in Langerin traffic and the biogenesis of BGs. The overexpression of a dominant-negative Rab11A mutant or Rab11A depletion strongly influenced Langerin traffic and stability and the formation of BGs, whereas modulation of other Rab proteins involved in dynamic regulation of the endocytic-recycling pathway had no effect. Impairment of Rab11A function led to a missorting of Langerin to lysosomal compartments, but inhibition of Langerin degradation by chloroquine did not restore the formation of BGs. Loss of RCP, but not of Rip11, also had a modest, but reproducible effect on Langerin stability and BG biogenesis, pointing to a role for Rab11A-RCP complexes in these events. Our results show that Rab11A and Langerin are required for BG biogenesis, and they illustrate the role played by a Rab GTPase in the formation of a specialized subcompartment within the endocytic-recycling system.

Replier
Victor Racine, Martin Sachse, Jean Salamero, Vincent Fraisier, Alain Trubuil, Jean-Baptiste Sibarita (2007 Mar 21)

Visualization and quantification of vesicle trafficking on a three-dimensional cytoskeleton network in living cells.

Journal of microscopy : 214-28 En savoir plus
Résumé

Recent progress in biology and microscopy has made it possible to acquire multidimensional data on rapid cellular activities. Unfortunately, the data analysis needed to describe the observed biological process still remains a major bottleneck. We here present a novel method of studying membrane trafficking by monitoring vesicular structures moving along a three-dimensional cytoskeleton network. It allows the dynamics of such structures to be qualitatively and quantitatively investigated. Our tracking method uses kymogram analysis to extract the consistent part of the temporal information and to allow the meaningful representation of vesicle dynamics. A fully automatic extension of this method, together with a statistical tool for dynamic comparisons, allows the precise analysis and comparison of overall speed distributions and directions. It can handle typical complex situations, such as a dense set of vesicles moving at various velocities, fusing and dissociating with each other or with other cell compartments. The overall approach has been characterized and validated on synthetic data. We chose the Rab6A protein, a GTPase involved in the regulation of intracellular membrane trafficking, as a molecular model. The application of our method to GFP-Rab6A stable cells acquired using fast four-dimensional deconvolution video-microscopy gives considerable cellular dynamic information unreachable using other techniques.

Replier

Année de publication : 2005

Henri de la Salle, Sabrina Mariotti, Catherine Angenieux, Martine Gilleron, Luis-Fernando Garcia-Alles, Dag Malm, Thomas Berg, Samantha Paoletti, Blandine Maître, Lionel Mourey, Jean Salamero, Jean Pierre Cazenave, Daniel Hanau, Lucia Mori, Germain Puzo, Gennaro De Libero (2005 Nov 29)

Assistance of microbial glycolipid antigen processing by CD1e.

Science (New York, N.Y.) : 1321-4 En savoir plus
Résumé

Complexes between CD1 molecules and self or microbial glycolipids represent important immunogenic ligands for specific subsets of T cells. However, the function of one of the CD1 family members, CD1e, has yet to be determined. Here, we show that the mycobacterial antigens hexamannosylated phosphatidyl-myo-inositols (PIM6) stimulate CD1b-restricted T cells only after partial digestion of the oligomannose moiety by lysosomal alpha-mannosidase and that soluble CD1e is required for this processing. Furthermore, recombinant CD1e was able to bind glycolipids and assist in the digestion of PIM6. We propose that, through this form of glycolipid editing, CD1e helps expand the repertoire of glycolipidic T cell antigens to optimize antimicrobial immune responses.

Replier

Année de publication : 2004

Ray McDermott, Huguette Bausinger, Dominique Fricker, Danièle Spehner, Fabienne Proamer, Dan Lipsker, Jean-Pierre Cazenave, Bruno Goud, Henri De La Salle, Jean Salamero, Daniel Hanau (2004 Jun 12)

Reproduction of Langerin/CD207 traffic and Birbeck granule formation in a human cell line model.

The Journal of investigative dermatology : 72-7 En savoir plus
Résumé

Birbeck granules (BG) are organelles specific to Langerhans cells (LCs), which form where the C-type lectin Langerin accumulates. Their function remains obscure due to morphologic and dynamic alterations induced by maturation of isolated LC. In this study, we attempted to reconstitute Langerin traffic and BG formation in the endosomal pathway of a human melanoma cell line. In the selected Langerin-transfected cell line, M10-22E, Langerin is distributed between the early recycling endosomal compartment and the plasma membrane, as in LC. Whereas mainly concentrated in membranes related to the Rab11(+) endosomal recycling compartment at the steady state, Langerin also recycles in M10-22E cells and drives BG biogenesis in the endosomal recycling compartment. Interruption of endocytosis or recycling induces redistribution of intracellular Langerin with an associated alteration in BG location and morphology. We have, therefore, generated a stable, Langerin-transfected cell line in which Langerin traffic and distribution and BG morphology replicate that seen in freshly isolated LC. This practical model can now be used to further delineate the nature and function of BG.

Replier
Laure A Perrin-Cocon, Christian L Villiers, Jean Salamero, Françoise Gabert, Patrice N Marche (2004 Mar 9)

B cell receptors and complement receptors target the antigen to distinct intracellular compartments.

Journal of immunology (Baltimore, Md. : 1950) : 3564-72 En savoir plus
Résumé

The processing of exogenous Ags is an essential step for the generation of immunogenic peptides that will be presented to T cells. This processing relies on the efficient intracellular targeting of Ags, because it depends on the content of the compartments in which Ags are delivered in APCs. Opsonization of Ags by the complement component C3 strongly enhances their presentation by B cells and increases their immunogenicity in vivo. To investigate the role of C3 in the targeting of Ags, we compared the intracellular traffic of proteins internalized by complement receptor (CR) and B cell receptor (BCR) in B lymphocytes. Whereas both receptors are able to induce efficient Ag presentation, their intracellular pathways are different. CR ligand is delivered to compartments containing MHC class II molecules (MHC-II) but devoid of transferrin receptor and Lamp-2, whereas BCR rapidly targets its ligand toward Lamp-2-positive, late endosomal MHC-II-enriched compartments through intracellular vesicles containing transferrin receptor. CR and BCR are delivered to distinct endocytic pathways, and the kinetic evolution of the protein content of these pathways is very different. Both types of compartments contain MHC-II, but CR-targeted compartments receive less neosynthesized MHC-II than do BCR-targeted compartments. The targeting induced by CR toward compartments that are distinct from BCR-targeted compartments probably participates in C3 modulation of Ag presentation.

Replier

Année de publication : 2003

Marion de Toledo, Francesca Senic-Matuglia, Jean Salamero, Gilles Uze, Franck Comunale, Philippe Fort, Anne Blangy (2003 Sep 10)

The GTP/GDP cycling of rho GTPase TCL is an essential regulator of the early endocytic pathway.

Molecular biology of the cell : 4846-56 En savoir plus
Résumé

Rho GTPases are key regulators of actin dynamics. We report that the Rho GTPase TCL, which is closely related to Cdc42 and TC10, localizes to the plasma membrane and the early/sorting endosomes in HeLa cells, suggesting a role in the early endocytic pathway. Receptor-dependent internalization of transferrin (Tf) is unaffected by suppression of endogenous TCL by small interfering RNA treatment. However, Tf accumulates in Rab5-positive uncoated endocytic vesicles and fails to reach the early endosome antigen-1-positive early endosomal compartments and the pericentriolar recycling endosomes. Moreover, Tf release upon TCL knockdown is significantly slower. Conversely, in the presence of dominant active TCL, internalized Tf accumulates in early endosome antigen-1-positive early/sorting endosomes and not in perinuclear recycling endosomes. Tf recycles directly from the early/sorting endosomes and it is normally released by the cells. The same phenotype is generated by replacing the C terminus of dominant active Cdc42 and TC10 with that of TCL, indicating that all three proteins share downstream effector proteins. Thus, TCL is essential for clathrin-dependent endocytosed receptors to enter the early/sorting endosomes. Furthermore, the active GTPase favors direct recycling from early/sorting endosomes without accumulating in the perinuclear recycling endosomes.

Replier
Catherine Angénieux, Jean Salamero, Dominique Fricker, Jean-Marie Wurtz, Blandine Maître, Jean-Pierre Cazenave, Daniel Hanau, Henri de la Salle (2003 Apr 3)

Common characteristics of the human and rhesus macaque CD1e molecules: conservation of biochemical and biological properties during primate evolution.

Immunogenetics : 842-9 En savoir plus
Résumé

In humans, a family of five genes encodes the CD1 molecules. Four of these proteins, CD1a, b, c, and d, are expressed on the plasma membrane and traffic between the cell surface and endocytic compartments, where they are loaded with antigenic glycolipids. The existence of human CD1e was demonstrated recently. This molecule surprisingly remains inside the cell, accumulating mainly in the Golgi compartments of immature dendritic cells and in the late endosomes of mature dendritic cells. In the latter compartments, CD1e is cleaved and becomes soluble. To determine whether these properties were specific to human CD1e, we investigated the presence and characteristics of CD1e in the rhesus macaque, an evolutionarily distant species of the primate lineage. Our results show that the cellular and biochemical properties of the human and simian CD1e molecules are similar, suggesting that the particular intracellular distribution of CD1e is important for its physiological and/or immunological function.

Replier

Année de publication : 2002

Ray Mc Dermott, Umit Ziylan, Danièle Spehner, Huguette Bausinger, Dan Lipsker, Mieke Mommaas, Jean-Pierre Cazenave, Graça Raposo, Bruno Goud, Henri de la Salle, Jean Salamero, Daniel Hanau (2002 Jan 26)

Birbeck granules are subdomains of endosomal recycling compartment in human epidermal Langerhans cells, which form where Langerin accumulates.

Molecular biology of the cell : 317-35 En savoir plus
Résumé

Birbeck granules are unusual rod-shaped structures specific to epidermal Langerhans cells, whose origin and function remain undetermined. We investigated the intracellular location and fate of Langerin, a protein implicated in Birbeck granule biogenesis, in human epidermal Langerhans cells. In the steady state, Langerin is predominantly found in the endosomal recycling compartment and in Birbeck granules. Langerin internalizes by classical receptor-mediated endocytosis and the first Birbeck granules accessible to endocytosed Langerin are those connected to recycling endosomes in the pericentriolar area, where Langerin accumulates. Drug-induced inhibition of endocytosis results in the appearance of abundant open-ended Birbeck granule-like structures appended to the plasma membrane, whereas inhibition of recycling induces Birbeck granules to merge with a tubular endosomal network. In mature Langerhans cells, Langerin traffic is abolished and the loss of internal Langerin is associated with a concomitant depletion of Birbeck granules. Our results demonstrate an exchange of Langerin between early endosomal compartments and the plasma membrane, with dynamic retention in the endosomal recycling compartment. They show that Birbeck granules are not endocytotic structures, rather they are subdomains of the endosomal recycling compartment that form where Langerin accumulates. Finally, our results implicate ADP-ribosylation factor proteins in Langerin trafficking and the exchange between Birbeck granules and other endosomal membranes.

Replier
Dan Lipsker, Umit Ziylan, Danièle Spehner, Fabienne Proamer, Huguette Bausinger, Pascale Jeannin, Jean Salamero, Alain Bohbot, Jean-Pierre Cazenave, Robert Drillien, Yves Delneste, Daniel Hanau, Henri de la Salle (2002 Jan 25)

Heat shock proteins 70 and 60 share common receptors which are expressed on human monocyte-derived but not epidermal dendritic cells.

European journal of immunology : 322-32 En savoir plus
Résumé

Priming of CTL by means of heat shock proteins (hsp) is dependent on antigen-presenting cells (APC), which present the hsp-associated peptides, via their cell surface MHC class I molecules, toCD8(+) T cells. It has not yet been established how human (hu) hsp70 interacts with the major (hu)APC, the dendritic cells (DC). Here we show that (hu)hsp70 is specifically internalized intoCD14(-), Toll-like receptor 4(-) monocyte-derived (hu)DC by receptor-mediated endocytosis. We further demonstrate that (hu)hsp70 and (hu)hsp60 share the same receptors on (hu)monocyte-derived DC. Both molecules as well as MHC class I molecules are spontaneously internalized and reach the MHC class II-enriched compartments. Finally, freshly isolated (hu) epidermal Langerhans cells (LC), the DC of the skin, as well as CD34(+)-derived LC do not bind hsp60 or hsp70. Given the likely importance of the internalization of hsp70 by APC in the induction of the immune responses, the finding that hsp60 and hsp70 are internalized through the same receptor(s) may explain why microbial hsp60 represents a major T cell antigen. This may rationalize the use of microbial hsp60 to prime immune responses against microbes. The lack of hsp60/70 receptors on epidermal LC raises the crucial question as to whether absence of priming of the skin and mucosal immune systems by hsp-polypeptide complexes could account for some tissue-specific diseases. This work also points to a potential advantage of using monocyte-derived DC in human immunotherapeutic applications of hsp60/70.

Replier

Année de publication : 2001

Isabelle Le Blanc, Vincent Blot, Isabelle Bouchaert, Jean Salamero, Bruno Goud, Arielle R Rosenberg, Marie-Christine Dokhélar (2001 Dec 26)

Intracellular distribution of human T-cell leukemia virus type 1 Gag proteins is independent of interaction with intracellular membranes.

Journal of virology : 905-11 En savoir plus
Résumé

Retrovirus Gag proteins are synthesized on free ribosomes, and are sufficient to govern the assembly and release of virus particles. Like type C retroviruses, human T-cell leukemia virus type 1 (HTLV-1) assembles and buds at the plasma membrane. After immunofluorescence staining, HTLV-1 Gag proteins appear as punctuated intracellular clusters, which suggests that they are associated either with intracellular membranes or with the plasma membrane. However, colocalization experiments using a panel of markers demonstrated that Gag proteins were not associated with the membranes involved in the secretory or endocytosis pathway. Small amounts of Gag proteins were detected at the plasma membrane and colocalized with the envelope glycoproteins. Moreover, Gag proteins were excluded from streptolysin-O permeabilized cells and in this respect behaved like cytoplasmic proteins. This suggests that the trafficking of HTLV-1 Gag proteins through the cytoplasm of the host cell is independent of any cell membrane system.

Replier

Année de publication : 2014

Sofia Traikov, Christoph Stange, Thomas Wassmer, Perrine Paul-Gilloteaux, Jean Salamero, Graça Raposo, Bernard Hoflack (1970 Jan 1)

Septin6 and Septin7 GTP binding proteins regulate AP-3- and ESCRT-dependent multivesicular body biogenesis.

PloS one : e109372 : DOI : 10.1371/journal.pone.0109372 En savoir plus
Résumé

Septins (SEPTs) form a family of GTP-binding proteins implicated in cytoskeleton and membrane organization, cell division and host/pathogen interactions. The precise function of many family members remains elusive. We show that SEPT6 and SEPT7 complexes bound to F-actin regulate protein sorting during multivesicular body (MVB) biogenesis. These complexes bind AP-3, an adapter complex sorting cargos destined to remain in outer membranes of maturing endosomes, modulate AP-3 membrane interactions and the motility of AP-3-positive endosomes. These SEPT-AP interactions also influence the membrane interaction of ESCRT (endosomal-sorting complex required for transport)-I, which selects ubiquitinated cargos for degradation inside MVBs. Whereas our findings demonstrate that SEPT6 and SEPT7 function in the spatial, temporal organization of AP-3- and ESCRT-coated membrane domains, they uncover an unsuspected coordination of these sorting machineries during MVB biogenesis. This requires the E3 ubiquitin ligase LRSAM1, an AP-3 interactor regulating ESCRT-I sorting activity and whose mutations are linked with Charcot-Marie-Tooth neuropathies.

Replier