Analyse intégrative de l’activation des lymphocytes T

Publications de l’équipe

Année de publication : 2016

Claire Hivroz, Michael Saitakis (2016 Feb 26)

Biophysical Aspects of T Lymphocyte Activation at the Immune Synapse.

Frontiers in immunology : 46 : DOI : 10.3389/fimmu.2016.00046 En savoir plus

T lymphocyte activation is a pivotal step of the adaptive immune response. It requires the recognition by T-cell receptors (TCR) of peptides presented in the context of major histocompatibility complex molecules (pMHC) present at the surface of antigen-presenting cells (APCs). T lymphocyte activation also involves engagement of costimulatory receptors and adhesion molecules recognizing ligands on the APC. Integration of these different signals requires the formation of a specialized dynamic structure: the immune synapse. While the biochemical and molecular aspects of this cell-cell communication have been extensively studied, its mechanical features have only recently been addressed. Yet, the immune synapse is also the place of exchange of mechanical signals. Receptors engaged on the T lymphocyte surface are submitted to many tensile and traction forces. These forces are generated by various phenomena: membrane undulation/protrusion/retraction, cell mobility or spreading, and dynamic remodeling of the actomyosin cytoskeleton inside the T lymphocyte. Moreover, the TCR can both induce force development, following triggering, and sense and convert forces into biochemical signals, as a bona fide mechanotransducer. Other costimulatory molecules, such as LFA-1, engaged during immune synapse formation, also display these features. Moreover, T lymphocytes themselves are mechanosensitive, since substrate stiffness can modulate their response. In this review, we will summarize recent studies from a biophysical perspective to explain how mechanical cues can affect T lymphocyte activation. We will particularly discuss how forces are generated during immune synapse formation; how these forces affect various aspects of T lymphocyte biology; and what are the key features of T lymphocyte response to stiffness.


Année de publication : 2015

Omar I Vivar, Giulia Masi, Jean-Marie Carpier, Joao G Magalhaes, Donatella Galgano, Gregory J Pazour, Sebastian Amigorena, Claire Hivroz, Cosima T Baldari (2015 Dec 31)

IFT20 controls LAT recruitment to the immune synapse and T-cell activation in vivo.

Proceedings of the National Academy of Sciences of the United States of America : 386-91 : DOI : 10.1073/pnas.1513601113 En savoir plus

Biogenesis of the immune synapse at the interface between antigen-presenting cells and T cells assembles and organizes a large number of membrane proteins required for effective signaling through the T-cell receptor. We showed previously that the intraflagellar transport protein 20 (IFT20), a component of the intraflagellar transport system, controls polarized traffic during immune synapse assembly. To investigate the role of IFT20 in primary CD4(+) T cells in vitro and in vivo, we generated mice bearing a conditional defect of IFT20 expression in T cells. We show that in the absence of IFT20, although cell spreading and the polarization of the centrosome were unaffected, T-cell receptor (TCR)-mediated signaling and recruitment of the signaling adaptor LAT (linker for activation of T cells) at the immune synapse were reduced. As a consequence, CD4(+) T-cell activation and proliferation were also defective. In vivo, conditional IFT20-deficient mice failed to mount effective antigen-specific T-cell responses, and their T cells failed to induce colitis after adoptive transfer to Rag(-/-) mice. IFT20 is therefore required for the delivery of the intracellular pool of LAT to the immune synapse in naive primary T lymphocytes and for effective T-cell responses in vivo.

Nathalie Bufi, Michael Saitakis, Stéphanie Dogniaux, Oscar Buschinger, Armelle Bohineust, Alain Richert, Mathieu Maurin, Claire Hivroz, Atef Asnacios (2015 May 9)

Human Primary Immune Cells Exhibit Distinct Mechanical Properties that Are Modified by Inflammation.

Biophysical journal : 2181-90 : DOI : 10.1016/j.bpj.2015.03.047 En savoir plus

T lymphocytes are key modulators of the immune response. Their activation requires cell-cell interaction with different myeloid cell populations of the immune system called antigen-presenting cells (APCs). Although T lymphocytes have recently been shown to respond to mechanical cues, in particular to the stiffness of their environment, little is known about the rigidity of APCs. In this study, single-cell microplate assays were performed to measure the viscoelastic moduli of different human myeloid primary APCs, i.e., monocytes (Ms, storage modulus of 520 +90/-80 Pa), dendritic cells (DCs, 440 +110/-90 Pa), and macrophages (MPHs, 900 +110/-100 Pa). Inflammatory conditions modulated these properties, with storage moduli ranging from 190 Pa to 1450 Pa. The effect of inflammation on the mechanical properties was independent of the induction of expression of commonly used APC maturation markers, making myeloid APC rigidity an additional feature of inflammation. In addition, the rigidity of human T lymphocytes was lower than that of all myeloid cells tested and among the lowest reported (Young’s modulus of 85 ± 5 Pa). Finally, the viscoelastic properties of myeloid cells were dependent on both their filamentous actin content and myosin IIA activity, although the relative contribution of these parameters varied within cell types. These results indicate that T lymphocytes face different cell rigidities when interacting with myeloid APCs in vivo and that this mechanical landscape changes under inflammation.


Année de publication : 2014

Marc Clement, Giulia Fornasa, Stéphane Loyau, Marion Morvan, Francesco Andreata, Kevin Guedj, Jamila Khallou-Laschet, Paola Larghi, Delphine Le Roux, Georges Bismuth, Gilles Chiocchia, Claire Hivroz, Debra K Newman, Antonino Nicoletti, Giuseppina Caligiuri (2014 Oct 4)

Upholding the T cell immune-regulatory function of CD31 inhibits the formation of T/B immunological synapses in vitro and attenuates the development of experimental autoimmune arthritis in vivo.

Journal of autoimmunity : 23-33 : DOI : 10.1016/j.jaut.2014.09.002 En savoir plus

CD31, a trans-homophilic inhibitory receptor expressed on both T- and B-lymphocytes, drives the mutual detachment of interacting leukocytes. Intriguingly, T cell CD31 molecules relocate to the immunological synapse (IS), where the T and B cells establish a stable interaction. Here, we show that intact CD31 molecules, which are able to drive an inhibitory signal, are concentrated at the periphery of the IS but are excluded from the center of the IS. At this site, were the cells establish the closest contact, the CD31 molecules are cleaved, and most of the extracellular portion of the protein, including the trans-homophilic binding sites, is shed from the cell surface. T cells lacking CD31 trans-homophilic binding sites easily establish stable interactions with B cells; at the opposite, CD31 signaling agonists inhibit T/B IS formation as well as the ensuing helper T cell activation and function. Confocal microscopy and flow cytometry analysis of experimental T/B IS shows that the T cell inhibitory effects of CD31 agonists depend on SHP-2 signaling, which reduces the phosphorylation of ZAP70. The analysis of synovial tissue biopsies from patients affected by rheumatoid arthritis showed that T cell CD31 molecules are excluded from the center of the T/B cell synapses in vivo. Interestingly, the administration of CD31 agonists in vivo significantly attenuated the development of the clinical signs of collagen-induced arthritis in DBA1/J mice. Altogether, our data indicate that the T cell co-inhibitory receptor CD31 prevents the formation of functional T/B immunological synapses and that therapeutic strategies aimed at sustaining CD31 signaling will attenuate the development of autoimmune responses in vivo.


Année de publication : 2013

Lionel Le Bourhis, Mathilde Dusseaux, Armelle Bohineust, Stéphanie Bessoles, Emmanuel Martin, Virginie Premel, Maxime Coré, David Sleurs, Nacer-Eddine Serriari, Emmanuel Treiner, Claire Hivroz, Philippe Sansonetti, Marie-Lise Gougeon, Claire Soudais, Olivier Lantz (2013 Oct 10)

MAIT cells detect and efficiently lyse bacterially-infected epithelial cells.

PLoS pathogens : e1003681 : DOI : 10.1371/journal.ppat.1003681 En savoir plus

Mucosal associated invariant T cells (MAIT) are innate T lymphocytes that detect a large variety of bacteria and yeasts. This recognition depends on the detection of microbial compounds presented by the evolutionarily conserved major-histocompatibility-complex (MHC) class I molecule, MR1. Here we show that MAIT cells display cytotoxic activity towards MR1 overexpressing non-hematopoietic cells cocultured with bacteria. The NK receptor, CD161, highly expressed by MAIT cells, modulated the cytokine but not the cytotoxic response triggered by bacteria infected cells. MAIT cells are also activated by and kill epithelial cells expressing endogenous levels of MRI after infection with the invasive bacteria Shigella flexneri. In contrast, MAIT cells were not activated by epithelial cells infected by Salmonella enterica Typhimurium. Finally, MAIT cells are activated in human volunteers receiving an attenuated strain of Shigella dysenteriae-1 tested as a potential vaccine. Thus, in humans, MAIT cells are the most abundant T cell subset able to detect and kill bacteria infected cells.

Paola Larghi, David J Williamson, Jean-Marie Carpier, Stéphanie Dogniaux, Karine Chemin, Armelle Bohineust, Lydia Danglot, Katharina Gaus, Thierry Galli, Claire Hivroz (2013 Feb 15)

VAMP7 controls T cell activation by regulating the recruitment and phosphorylation of vesicular Lat at TCR-activation sites.

Nature immunology : 723-31 : DOI : 10.1038/ni.2609 En savoir plus

The mechanisms by which Lat (a key adaptor in the T cell antigen receptor (TCR) signaling pathway) and the TCR come together after TCR triggering are not well understood. We investigate here the role of SNARE proteins, which are part of protein complexes involved in the docking, priming and fusion of vesicles with opposing membranes, in this process. Here we found, by silencing approaches and genetically modified mice, that the vesicular SNARE VAMP7 was required for the recruitment of Lat-containing vesicles to TCR-activation sites. Our results indicated that this did not involve fusion of Lat-containing vesicles with the plasma membrane. VAMP7, which localized together with Lat on the subsynaptic vesicles, controlled the phosphorylation of Lat, formation of the TCR-Lat-signaling complex and, ultimately, activation of T cells. Our findings suggest that the transport and docking of Lat-containing vesicles with target membranes containing TCRs regulates TCR-induced signaling.


Année de publication : 2012

Claire Hivroz, Karine Chemin, Marie Tourret, Armelle Bohineust (2012 Dec 11)

Crosstalk between T lymphocytes and dendritic cells.

Critical reviews in immunology : 139-55 En savoir plus

Dendritic cells (DCs) are professional antigen-presenting cells (APCs) with the unique property of inducing priming and differentiation of naïve CD4+ and CD8+ T cells into helper and cytotoxic effectors. Their efficiency is due to their unique ability to process antigen, express costimulatory molecules, secrete cytokines, and migrate to tissues or lymphoid organs to prime T cells. DCs also play an important role in T-cell peripheral tolerance. There is ample evidence that the DC ability to present antigens is regulated by CD4+ helper T cells. Indeed, interactions between surface receptors and ligands expressed respectively by T cells and DCs, as well as T-cell-derived cytokines modify DC functions. This T-cell-induced modification of DCs has been called « education » or « licensing. » This intimate crosstalk between DCs and T lymphocytes is key in establishing appropriate adaptive immune responses. It requires cognate interactions between T lymphocytes and DCs, which are organized in time and space by structures called immunological synapses. Here we discuss the particular aspects of immunological synapses formed between T cells and DCs and the role these organized interactions have in T-cell-DC crosstalk.

Fabian Hauck, Clotilde Randriamampita, Emmanuel Martin, Stéphane Gerart, Nathalie Lambert, Annick Lim, Jean Soulier, Zosia Maciorowski, Fabien Touzot, Despina Moshous, Pierre Quartier, Sébastien Heritier, Stéphane Blanche, Fréderic Rieux-Laucat, Nicole Brousse, Isabelle Callebaut, André Veillette, Claire Hivroz, Alain Fischer, Sylvain Latour, Capucine Picard (2012 Sep 19)

Primary T-cell immunodeficiency with immunodysregulation caused by autosomal recessive LCK deficiency.

The Journal of allergy and clinical immunology : 1144-1152.e11 : DOI : 10.1016/j.jaci.2012.07.029 En savoir plus

Signals emanating from the antigen T-cell receptor (TCR) are required for T-cell development and function. The T lymphocyte-specific protein tyrosine kinase (Lck) is a key component of the TCR signaling machinery. On the basis of its function, we considered LCK a candidate gene in patients with combined immunodeficiency.

Karine Chemin, Armelle Bohineust, Stéphanie Dogniaux, Marie Tourret, Sarah Guégan, Francesc Miro, Claire Hivroz (2012 Jul 20)

Cytokine secretion by CD4+ T cells at the immunological synapse requires Cdc42-dependent local actin remodeling but not microtubule organizing center polarity.

Journal of immunology (Baltimore, Md. : 1950) : 2159-68 : DOI : 10.4049/jimmunol.1200156 En savoir plus

Cytokine secretion by T lymphocytes plays a central role in mounting adaptive immune responses. However, little is known about how newly synthesized cytokines, once produced, are routed within T cells and about the mechanisms involved in regulating their secretions. In this study, we investigated the role of cytoskeleton remodeling at the immunological synapse (IS) in cytokine secretion. We show that a key regulator of cytoskeleton remodeling, the Rho GTPase Cdc42, controls IFN-γ secretion by primary human CD4+ T lymphocytes. Surprisingly, microtubule organizing center polarity at the IS, which does not depend on Cdc42, is not required for cytokine secretion by T lymphocytes, whereas microtubule polymerization is required. In contrast, actin remodeling at the IS, which depends on Cdc42, controls the formation of the polymerized actin ring at the IS, the dynamic concentration of IFN-γ-containing vesicles inside this ring, and the secretion of these vesicles. These results reveal a previously unidentified role of Cdc42-dependent actin remodeling in cytokine exocytosis at the IS.

Nadia Bourouina, Julien Husson, Claire Hivroz, Nelly Henry (2012 Mar 15)

Biomimetic droplets for artificial engagement of living cell surface receptors: the specific case of the T-cell.

Langmuir : the ACS journal of surfaces and colloids : 6106-13 : DOI : 10.1021/la300398a En savoir plus

Liquid colloids, in the form of droplets grafted with specific biomolecules, are emerging as potential biomimetic systems. Here we show for the first time the possibility of forming hybrid conjugates between an advanced living cell model, the T-cell of the Jurkat cell line, and a specifically grafted droplet. Using T-cells expressing a fluorescent chimeric protein associated with the TCR/CD3 complex and fluorescent ligand-grafted droplets, we demonstrate formation of an interfacial contact concentrated in linking molecules, the morphology and dynamics of which strongly depend on the targeted receptor. The sequence of events ranges from the initial concentration of molecules following an unbound molecule gradient to active actin-driven spreading and fragmentation of the contact, ending with droplet internalization. We observed synchronized colocalization of receptors and ligands driven by cell dynamics and closely mirrored by the droplet interface. Using intracellular calcium probe Fura-2, we also showed that the cell/droplet interaction can trigger the T-cell signaling cascade. By examining molecular dynamics using FRAP measurements, we observed a nearly frozen cell droplet joining interface. Taken together, our results point to liquid colloids as promising new tools both for probing cell surface interactions and receptor dynamics and for manipulating biological cell functions.


Année de publication : 2011

Julien Husson, Karine Chemin, Armelle Bohineust, Claire Hivroz, Nelly Henry (2011 May 17)

Force generation upon T cell receptor engagement.

PloS one : e19680 : DOI : 10.1371/journal.pone.0019680 En savoir plus

T cells are major players of adaptive immune response in mammals. Recognition of an antigenic peptide in association with the major histocompatibility complex at the surface of an antigen presenting cell (APC) is a specific and sensitive process whose mechanism is not fully understood. The potential contribution of mechanical forces in the T cell activation process is increasingly debated, although these forces are scarcely defined and hold only limited experimental evidence. In this work, we have implemented a biomembrane force probe (BFP) setup and a model APC to explore the nature and the characteristics of the mechanical forces potentially generated upon engagement of the T cell receptor (TCR) and/or lymphocyte function-associated antigen-1 (LFA-1). We show that upon contact with a model APC coated with antibodies towards TCR-CD3, after a short latency, the T cell developed a timed sequence of pushing and pulling forces against its target. These processes were defined by their initial constant growth velocity and loading rate (force increase per unit of time). LFA-1 engagement together with TCR-CD3 reduced the growing speed during the pushing phase without triggering the same mechanical behavior when engaged alone. Intracellular Ca(2+) concentration ([Ca(2+)](i)) was monitored simultaneously to verify the cell commitment in the activation process. [Ca(2+)](i) increased a few tens of seconds after the beginning of the pushing phase although no strong correlation appeared between the two events. The pushing phase was driven by actin polymerization. Tuning the BFP mechanical properties, we could show that the loading rate during the pulling phase increased with the target stiffness. This indicated that a mechanosensing mechanism is implemented in the early steps of the activation process. We provide here the first quantified description of force generation sequence upon local bidimensional engagement of TCR-CD3 and discuss its potential role in a T cell mechanically-regulated activation process.


Année de publication : 2010

Marie Tourret, Sarah Guégan, Karine Chemin, Stéphanie Dogniaux, Francesc Miro, Armelle Bohineust, Claire Hivroz (2010 Oct 27)

T cell polarity at the immunological synapse is required for CD154-dependent IL-12 secretion by dendritic cells.

Journal of immunology (Baltimore, Md. : 1950) : 6809-18 : DOI : 10.4049/jimmunol.1001501 En savoir plus

Ag-specific interaction between T lymphocytes and dendritic cells (DCs) leads to both T cell and DC activation. CD154 (CD40 ligand)/CD40 interactions have been shown to play a major, although not exclusive, role in this functional cross-talk. Interactions between T cells and DCs are structured by an immunological synapse (IS), characterized by polarization of the T cell microtubule cytoskeleton toward the interacting DCs. Yet the role T cell polarization may play in T cell-induced DC activation is mostly unknown. In this study, we address the role of T cell polarity in CD154-dependent activation of DCs in a human model, using two different tools to block T cell polarity (i.e., a microtubule depolymerizing drug and an inhibitor of atypical protein kinase C). We show that CD154 is recruited and concentrated at the IS formed between human primary T cells and autologous DCs and that this recruitment requires T cell polarity at the IS. Moreover, we show that T cell polarization at the IS controls T cell-dependent CD154-CD40 signaling in DCs as well as CD154-dependent IL-12 secretion by DCs. This study shows that T cell polarity at the IS plays a key role in CD154/CD40-dependent cross-talk between CD4(+) T cells and DCs.

Nelly Henry, Claire Hivroz (2010 Jun 2)

Early T-cell activation biophysics.

HFSP journal : 401-11 : DOI : 10.2976/1.3254098 En savoir plus

The T-cell is one of the main players in the mammalian immune response. It ensures antigen recognition at the surface of antigen-presenting cells in a complex and highly sensitive and specific process, in which the encounter of the T-cell receptor with the agonist peptide associated with the major histocompatibility complex triggers T-cell activation. While signaling pathways have been elucidated in increasing detail, the mechanism of TCR triggering remains highly controversial despite active research published in the past 10 years. In this paper, we present a short overview of pending questions on critical initial events associated with T-cell triggering. In particular, we examine biophysical approaches already in use, as well as future directions. We suggest that the most recent advances in fluorescence super-resolution imaging, coupled with the new classes of genetic fluorescent probes, will play an important role in elucidation of the T-cell triggering mechanism. Beyond this aspect, we predict that exploration of mechanical cues in the triggering process will provide new clues leading to clarification of the entire mechanism.


Année de publication : 2009

Cinzia Nobile, Dominika Rudnicka, Milena Hasan, Nathalie Aulner, Françoise Porrot, Christophe Machu, Olivier Renaud, Marie-Christine Prévost, Claire Hivroz, Olivier Schwartz, Nathalie Sol-Foulon (2009 Dec 18)

HIV-1 Nef inhibits ruffles, induces filopodia, and modulates migration of infected lymphocytes.

Journal of virology : 2282-93 : DOI : 10.1128/JVI.02230-09 En savoir plus

The HIV-1 Nef protein is a pathogenic factor modulating the behavior of infected cells. Nef induces actin cytoskeleton changes and impairs cell migration toward chemokines. We further characterized the morphology, cytoskeleton dynamics, and motility of HIV-1-infected lymphocytes. By using scanning electron microscopy, confocal immunofluorescence microscopy, and ImageStream technology, which combines flow cytometry and automated imaging, we report that HIV-1 induces a characteristic remodeling of the actin cytoskeleton. In infected lymphocytes, ruffle formation is inhibited, whereas long, thin filopodium-like protrusions are induced. Cells infected with HIV with nef deleted display a normal phenotype, and Nef expression alone, in the absence of other viral proteins, induces morphological changes. We also used an innovative imaging system to immobilize and visualize living individual cells in suspension. When combined with confocal « axial tomography, » this technique greatly enhances three-dimensional optical resolution. With this technique, we confirmed the induction of long filopodium-like structures in unfixed Nef-expressing lymphocytes. The cytoskeleton reorganization induced by Nef is associated with an important impairment of cell movements. The adhesion and spreading of infected cells to fibronectin, their spontaneous motility, and their migration toward chemokines (CXCL12, CCL3, and CCL19) were all significantly decreased. Therefore, Nef induces complex effects on the lymphocyte actin cytoskeleton and cellular morphology, which likely impacts the capacity of infected cells to circulate and to encounter and communicate with bystander cells.

Christie-Ann McCarl, Capucine Picard, Sara Khalil, Takumi Kawasaki, Jens Röther, Alexander Papolos, Jeffery Kutok, Claire Hivroz, Francoise Ledeist, Katrin Plogmann, Stephan Ehl, Gundula Notheis, Michael H Albert, Bernd H Belohradsky, Janbernd Kirschner, Anjana Rao, Alain Fischer, Stefan Feske (2009 Dec 17)

ORAI1 deficiency and lack of store-operated Ca2+ entry cause immunodeficiency, myopathy, and ectodermal dysplasia.

The Journal of allergy and clinical immunology : 1311-1318.e7 : DOI : 10.1016/j.jaci.2009.10.007 En savoir plus

Defects in the development or activation of T cells result in immunodeficiency associated with severe infections early in life. T-cell activation requires Ca2+ influx through Ca2+-release activated Ca2+ (CRAC) channels encoded by the gene ORAI1.