Crosstalk between T lymphocytes and dendritic cells

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Critical Reviews™ in Immunology, 32(2):139-155 (2012)

11040-8401/12/$35.00 © 2012 by Begell House, Inc.

Crosstalk between T Lymphocytes and Dendritic Cells

Claire Hivroz1,2,* , Karine Chemin1,2,3, Marie Tourret1,2, & Armelle Bohineust1,2

1Institut Curie, Centre de Recherche, Pavillon Pasteur, 26 Rue d’ULM, Paris, F-75248; 2 INSERM, Unité 932, Immunité et Cancer, Paris, F-75248, France; 3 Center for Molecular Medicine (CMM), Rheumatology Unit, Department of Medicine, Karolinska Institutet, SE-17176 Stockholm, Sweden

* Address all correspondence to: Claire Hivroz, PhD, Institut Curie, 26 rue d’Ulm Paris F-75248 France; Tel.:(33) 1 56 24 68 35; Fax: (33)1 44 07 07 85; claire.

[email protected]

ABSTRACT: 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 cyto-kines, 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.

KEY WORDS: Immunological synapse, cytoskeleton, dendritic cell, T lymphocyte.

ABBREVIATIONS: DC: dendritic cell; IS: immunological synapse; MTOC: microtubule organizing center; SMAC: supramolecular activating complex.

I. INTRODUCTION

Dendritic cells (DCs) are key cells in the regula-tion of T-cell responses. They orchestrate immune responses by inducing both tolerance to self-antigens and resistance to infectious pathogens.1,2 The ability of DCs to induce tolerance or immu-nity depends on their state of activation, which regulates DCs ability to process antigen and to express costimulatory molecules and cytokines.3 Activation of DCs also controls DC migration to the appropriate sites, in tissues or lymphoid organs, wherein they prime T cells.4,5 Different pathways initiate maturation of DCs. One of these pathways

involves microbial products, which act through toll-like receptors (TLR) expressed on immature DCs.6 Another pathway of activation is induced by signals from cells of the innate immune system such as natural killer (NK) cells, NK T cells, and gd T cells (reviewed by Reis e Sousa7).

Finally, CD4+ T-helper lymphocytes are also potent activators of DCs. The CD40 ligand (CD40L) expressed on activated and memory T cells plays a key role in this process both in murine T cells8–11 and in human T cells.12,13 Cytokines pro-duced by T lymphocytes, such as IL-4 and IFN-g, have also been shown to play a crucial role in the activation of DCs.14–16 This CD4+ T-cell–depen-

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dent activation of DCs has been called “licensing”17 or “education.”18 This activation is probably crucial to amplify DC activation by pathogenic infections that do not provide sufficient inflammatory media-tors to induce a functional maturation of DCs. CD4+ T-cell–dependent activation might also be required when the DC, which has encountered the antigen in the periphery, is not the same as the DC that presents the Ag in the lymphoid organs. In such cases, resident DCs, which have an immature phenotype, have not been in contact with the infectious agents or infected cells, and maturation by T-cell–dependent activating signals are thus crucial for their maturation.

Licensing or education of DCs by CD4+ T cells has been shown to require cognate interactions between CD4+ T lymphocytes and DCs.16,18–20 T-cell education of DCs is carefully targeted. DCs receive their activation signals from a T cell through cell-surface molecules and/or soluble factors that are delivered across a specialized interaction zone called the immunological synapse, which orga-nizes the crosstalk between T lymphocytes and DCs. Herein we review the specificities of the immunological synapses (ISs) formed between T lymphocytes and DCs and the role these structures have on DC and T lymphocyte activation.

II. ROLE OF IS IN CROSSTALK BETWEEN T CELLS AND DCS

The adaptive immune response is initiated by spe-cific interactions between antigen-loaded, mature DCs and naïve CD4+ T cells. The activation of a T cell by a DC is controlled by the formation of a structured area at the DC–T-cell interface that has been called the immunological synapse (IS). This contact area is organized in time and space and involved the segregation and clustering of membrane proteins and cytosolic molecules at the interface between a T cell and a DC as well as a remodeling of both the actin and the microtubule cytoskeletons on both sides of the contact, i.e., the T-cell and DC sides of the IS. In this section we

review and discuss the formation and the role of the IS on the DC side and the T-cell side.

A. Distribution of Membrane and Cytosolic Proteins at the IS between T Lymphocytes and DCs

1. Organization of the IS on the T-Cell Side

The first analyses of ISs were performed with B cells or artificial planar lipid bilayers containing key molecules involved in T-cell activation.21,22 These pioneering studies showed that T-cell receptors (TCRs) progressively accumulate at the central part of the IS, called the central supramolecular activat-ing complex cSMAC, whereas integrins segregate from TCRs to form the so-called peripheral SMAC (pSMAC), 21,22 and bulky membrane proteins, such as CD43, are excluded from the central zone and gather at the distal SMAC (dSMAC).23–25 Pattern-ing of the membrane receptors is accompanied by a spatial distribution of cytosolic signaling molecules (reviewed by Smith-Garvin et al.26) [e.g., Protein Kinase C-theta (PKC-t),21 Linker of Activated T cells (LAT)27,28] that are found in the cSMAC, whereas others, such as talin, are found at the pSMAC.21 This concentric pattern of proteins has been called the bull’s eye pattern or, more recently, the Kupfer’s synapse in reference to the first description of this peculiar structure by Kupfer et al.21 This concentric pattern is not the only way to structure the contact area (reviewed byTrautmann and Valitutti29). Indeed, several groups (including ours) have shown that the structure of the contact zone between DCs and T lymphocytes is different from that observed in T-cell–B-cell interfaces or in T lymphocytes in contact with artificial lipid bilayers containing pMHC and costimulatory molecules. These differences probably illustrate the unique properties of the DCs.

DCs have been shown to induce structured contacts with T lymphocytes in the absence of specific antigens.30–32 Indeed, the accumulation of

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the TCR–CD3 complexes, segregation of LFA-1 from TCR,31 exclusion of CD43,30–32 and enrich-ment of PKC-q33 have been observed in the absence of specific antigenic stimulation. An elegant study by Brossard et al., who compared T-cell–B-cell ISs with T-cell–DC ISs using both electron and light microscopy, revealed the formation of mul-tifocal synapses in T-cell–DC ISs, characterized by submicronic contact spots, with no large-scale segregation of CD3 and LFA-1.31 Another layer of complexity stems from the fact that DCs at different stages of maturation form different ISs. Indeed, mature DCs form stable conjugates with naïve T cells and induce the formation of Kupfer’s synapses, whereas immature DCs form few stable conjugates with no organized ISs.32,34

Moreover, mature DCs can form long-lasting interactions with naïve T cells, even in the absence of Ag, whereas immature DCs establish only short intermittent contacts, suggesting that these short contacts prevent the formation of organized ISs and full T-cell activation by immature DCs. This has been observed in murine bone marrow DCs,34 murine plasmacytoid DCs,35 and human monocyte-derived DCs (Figure 1).32 These in vitro observations correlate with results obtained in vivo using biphotonic microscopy, which show that the interactions of both CD4+- and CD8+-naïve T cells with mature priming-inducing DCs are more stable than the contacts with resting tolerance-inducing DCs.36–38 Although in vivo studies contribute valuable data on interaction kinetics, little infor-mation is available on the molecular dynamics at the T-cell–DC interface. Indeed, live imaging of molecules involved in the formation of the IS in intact tissues has been technically challenging. A pioneer study by Germain et al. used confocal imaging to show explanted lymph nodes in which the contact zone between T cells and DCs excluded CD43.39 In a more recent study, the distribution of LAT-EGFP (LAT-Enhanced Green Fluorescent Protein) molecules during antigen recognition by activated CD4+ T cells was tracked using bipho-tonic microscopy in lymph nodes.40 This study showed that most of the interactions of recently

activated CD4+ T cells with DCs were dynamic, with no persistent large-scale accumulation of LAT in the T-cell–DC contact zone.40 Yet this study did not look at the accumulation of LAT in resting T cells interacting with DCs at differ-ent stages of maturation. Finally, TCR dynamics in naïve T cells interacting with DCs in lymph nodes was followed in a transgenic mouse express-ing a TCR–GFP fusion.41 This study showed that interaction of naïve T cells with antigen-bearing DCs induced a rapid and brief clustering of TCR in the synaptic zone that was rapidly followed by internalization. Moreover, the study revealed that T cells are able to actively aggregate TCRs, to flux calcium, and to internalize TCR clusters, even in a motile synapse.41 These results suggest that the formation of a stable cSMAC is not indispensable for full T-cell activation by DCs, and they confirm in vitro studies, showing that T cells can sum up signals resulting from multiple interactions with macrophages and then induce the NFAT-dependent GFP transcription.42 The “serial encounter” model43,44 postulates that not only long-lasting contacts but also sequential short-lived interactions with DCs (5–20 min each) can lead to T-cell activation. However, as revealed by our study, multiple short-lived contacts might induce some T-cell activation events but not oth-ers.32 Innovation in imaging techniques will bring valuable information on the molecular nature and dynamic of the contact between T lymphocytes and DCs.

2. Organization of the IS on the DC Side

The organization of the synaptic zone on the T-cell side has been the object of intense scrutiny. In contrast, little is known on the organization of membrane and cytosolic proteins on the antigen-presenting cell (APC) side, probably because APCs were thought to only play a passive role because they could be replaced by beads coated with activat-ing antibodies or artificial lipid bilayers engrafted with activating molecules. However, several studies


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have shown that surface and cytoplasmic molecules expressed by DCs presented a specific spatial distri-bution at the IS. For example, DC counter receptors for T cell membrane molecules redistribute at the IS. Indeed, clustering of major histocompatibility molecules (MHC) class II molecules on the surface of the TCR molecules have been shown to take place in DCs and to play a role in T-cell activa-tion.45,46 Costimulatory molecules such as CD80, CD86, and adhesion molecules are also recruited at the IS between DCs and T lymphocytes.46,47 Another costimulatory molecule, CD70, is also delivered to the IS.48 This molecule binds the CD27 molecule on T cells and controls CD8+

T-cell effective immunity.49 CD70 intracellular trafficking is controlled, like the MHC class II, by the invariant chain (Ii), suggesting a coordinate arrival at the IS of these two key activators of T cells (Figure 1).50 Other surface receptors present on DCs are also recruited in the T-cell–DC syn-aptic zone. The semaphorin receptor plexin-A1, which is expressed in DCs and strongly induced by the MHC class II transactivator (CIITA),51 is enriched at the IS with T cells.52 One of its counter receptors, neuropilin-1, is also recruited at the IS on the T-cell side.53 Plexin-A1 expression by DCs and neuropilin expression by T lympho-cytes, respectively, are required for T-cell activa-

FIGURE 1: Formation of synapses between CD4+ T cells and immature or mature DCs. On the T cell side, T cells establish short intermittent contact with immature DCs and long contacts with mature DCs.32,34,35 CD43 is not excluded from the immunological synapse formed with immature DC, whereas it is excluded when DC are mature.31 TCR/CD3, PKCq and LAT are enriched in immunological synapses formed with mature DCs. This enrichment is not observed with immature DCs.32,34,35 Microscopy images show contact between human CD4+ T cells and autologous immature or mature DCs and were performed as in Blanchard et al.32 On the DC side, interaction of T cells with antigen-pulsed DCs induce the recruitment of MHC class II,45,46 CD80, CD86, 46,47 and CD70 48 at the immunological synapse as well as dissolution of podosomes.132



tion because silencing of plexin-A151,52 or use of neuropilin neutralizing antibodies 53 inhibits T-cell activation. Notch1 receptor and Notch ligands expressed by both T cells and DCs also show a specific clustering at the IS. Indeed, Notch1 recep-tors on TH cells and Notch ligands (Delta-like1, Jagged1) on DCs cluster in their apposed central SMACs, whereas Notch1 receptor on DCs and Notch ligands on TH cluster in their “interphac-ing” peripheral SMACs on the T-cell–DC IS.54 DCs can also present chemokines at their plasma membranes; these tethered chemokines influence T-cell motility and subsequent T-cell reactivity.55

Finally, similar to those in T lymphocytes, cytosolic signaling molecules are also recruited at the IS on the DC side. Indeed, the Akt kinase has been shown to be recruited at the IS on the DC side,56 where it might play a role in the survival of DCs induced by T cells.57 In DCs also, like in T cells, adaptor proteins involved in signal-ing cascade are recruited at the IS. The cytosolic adaptor molecule SKAP-HOM, which is similar to the T-cell–specific homologue SKAP55 and is expressed by DCs, is recruited at the IS.58 Another scaffolding protein, spinophilin, which is present in neuronal dendritic spines and regulates synaptic transmission, has been shown to be dynamically recruited to the IS in an antigen-dependent man-ner and to be required for optimal T-cell activa-tion.59 The SCIMP protein, a new palmitoylated transmembrane adaptor protein, is also localized in the IS and seems to be involved in MHC class II signaling.60

B. Remodeling of the Cytoskeleton at the Immunological Synapse

1. On the T-Cell Side

One of the hallmarks of the formation of the IS is the remodeling of both the actin and micro-tubule T-cell cytoskeletons in the contact zone. After recognition of the pMHC complexes at the surface of an APC, T lymphocytes polarize their

polymerized actin in the contact zone (reviewed by Burkhardt et al.61). Signals from the TCR initi-ate this program of actin cytoskeletal rearrange-ments, but actin remodeling also regulates TCR signaling as revealed by the inhibitory effects of drugs that inhibit dynamic actin polymerization on T-cell activation.62,63 T-cell–APC conjugation results in morphological changes, as the stimulated T cell rounds up and accumulates filamentous actin (F-actin) at the stimulatory interface. Rapid inactivation of the ezrin-radixin-moesin proteins after antigen recognition also contributes to T-cell morphological changes by regulating the cortical rigidity of T cells64 and the collapse of microvilli.65,66 Indeed, it results in disanchoring of the cortical actin cytoskeleton from the plasma membrane decreasing cellular rigidity 64 that contributes to more efficient T cell-antigen-presenting cell con-jugate formation. Accumulation of F-actin at the T-cell–APC interface is the result of TCR-induced localized activation of multiple actin regulatory and polymerizing pathways, among which are formins and Arp2/3,67 the Wiskott-Aldrich syn-drome protein (WASp),68–70 small Rho GTPases such as Cdc42 71–73 and Rac.67,74 Activation of the T cell in response to an APC also results in the polarization of the microtubule organizing center (MTOC) toward the T-cell–APC contact site.75 Mature DCs induce this MTOC polarity more efficiently than immature DCs.34,35 T-cell MTOC polarity also requires TCR signaling, and several signaling molecules have been shown to be involved by others and us in this polarization. Among oth-ers, the p56Lck 76 and ZAP70 tyrosine kinases,77 the LAT and SLP76 adaptors,28,78 PKCs,79–81 and the casein kinase 1d82 have been shown to control T-cell MTOC polarity at the IS. Yet, the mechanisms by which MTOC polarity toward the APC is controlled are far from being understood. In particular, how TCR signaling might control motors such as dynein, which has been shown to be involved in MTOC polarity,83–85 remains unknown. Moreover, the link between actin remodeling and MTOC polarity at the synapse are not yet entirely clear. However, a recent study by Alcover et al.


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showed that ezrin that binds to actin also controls MTOC polarity.86

Although the T-cell MTOC polarity at the IS is well established, its role is still a matter of debate. MTOC polarity has been involved in TCR signaling 85 and enrichment of the TCR at the IS.87 Yet, inhibition of MTOC polarity in T cells does not always correlate with inhibition of T-cell activation. Indeed, it seems that microtubule remodeling at the IS might rather control down-regulation of TCR signaling, 86 and inhibitors of MTOC polarity do not inhibit cytokine production by T lymphocytes.88 Although the role of MTOC polarization in T cells is not entirely unraveled, the polarized delivery of effectors may be required to establish a high local concentration of effectors at the IS and/or to ensure the antigen-specificity of the immune responses by avoiding bystander effects. Indeed, MTOC polarization at the IS induces a reorientation of the Golgi and secretory apparatus to the cell–cell interface, which controls the polarized secretion of lymphokines89–91 (Figure 2) and lytic granules92,93 toward the interacting APC or target cell. This polarity has been suggested to control the directional delivery of cytokines or harmful cyto-toxic granules, thus maintaining the Ag specificity and avoiding effects on neighboring cells.

Polarized secretion does concern not only soluble proteins, such as cytokines, but also trans-membrane proteins, such as FasL94 and CTLA-4.95,96 Together with Valitutti et al., we recently showed that CD154 (CD40L) is recruited and concentrated at the IS formed between human primary CD4+ T cells and autologous DCs, and furthermore, this recruitment requires T-cell polar-ity at the IS.88,97 Our study shows that CD154 is rapidly recruited at the IS. The dynamic analysis of the GFP-CD154 recruitment at the IS suggests that two pools of CD154 are recruited: one is at the plasma membrane and one is intracellular. Because CD154 surface expression is low even in activated T cells, recruitment of the endocytic pool is probably crucial to provide a “bolus” of CD154 signaling, which is crucial for CD154/CD40 signaling and IL-12 production by DCs

(Figure 2).97 Interestingly, our results demonstrate that some T-cell-dependent activation of DCs, such as expression of costimulatory molecules and IL-6 and IL-8 productions, do not need T-cell MTOC polarity, whereas IL-12 production does. These results have important consequences on T-cell–DC crosstalk because they suggest that in poor polarizing conditions CD4+ T cells are unable to induce DC licensing or to induce Th1 polarization, which depends on IL-12 production (reviewed by Trinchieri98). Since then, CD154-CD40 interactions have also been shown to provide anti-apoptotic signals to DCs,56,99,100 to control cross-presentation of antigen by DCs,17 and to overcome peripheral T-cell tolerance.101 T-cell polarity and CD154 recruitment at the IS might also play a role in these events. It remains to be determined whether other molecules of the TNFR family involved in the crosstalk between T lymphocytes and DCs, such as OX40, CD134 (reviewed by Croft102), and LIGHT (reviewed by Schneider et al.103) also require T-cell polarity to signal in DCs.

One recent study by Davis et al. showed inter-esting features of the ISs formed between CD4+ T cells and B cells or DCs.104 In this study, research-ers used electron microscopy and 3D tomography to characterize the synapses of antigen-specific CD4+ T cells recognizing B cells and DCs at different time points. They showed that there are at least four distinct stages in synapse formation, proceeding over several hours, including an initial stage involving invasive T-cell pseudopodia that penetrate deeply into the antigen-presenting cell.104 These results suggest that considerable forces are developed by CD4+ T lymphocytes that may be important to search for ligands at the surface of the APC, induce the TCR signaling pathways, and exclude bulky proteins from the contact zone. These forces might also be involved in a mechanism called trogocytosis by which T cells acquire MHC class I and II molecules as well as costimulatory molecules and membrane patches from APC, including DCs.105–107 Acquisition by T cells of molecules from the APC might participate



in ending the immune responses by decreasing the Ag presentation by APC or by converting T cells in APC allowing fratricide lysis of T cells by cytotoxic T lymphocytes.108

Our recent data show that T cells do indeed develop actin-dependent forces on APC and that these forces adapt to the rigidity of the APC they encounter.109 Thus T cells might use their cyto-skeleton to probe the biomechanical environment of the APC with which they interact, gathering information that might contribute to the induction

of their functional program. These forces applied on DCs by T cells might also contribute to the modification of DCs by T lymphocytes.

2. The Special Case of the IS Formed between Regulatory T Cells and DCs

Regulatory T cells (Treg cells or Tregs) play impor-tant roles in the prevention of autoimmunity,110 maternal–fetal tolerance,111 and other immune

FIGURE 2: T-cell polarization toward the DC is required to induce CD40L/CD40 signaling in DCs. Upon recognition by T cells of the peptide-MHC class II presented by DC, the T cell polarized its MTOC toward the cognate DC. This is accompanied by polarization of the Golgi apparatus and of vesicles containing cytokines such as IFN-g. This polarity leads to an enrichment of CD40L in the center of the interaction zone88,97 that is required to induce CD40-dependent signaling in DCs 97 and production of IL-1288,97 by DCs. Microscopy images were performed as in Miro et al.16 It remains to be determined whether this enrichment of CD40L in the synaptic zone is also required for other CD40-dependent functions such as DC survival,56,99,100 antigen cross-presentation,17 and overcoming T-cell tolerance.101 Human CD4+ T lymphocytes interacting with autologous monocyte-derived DCs. Labelling for IFN-g, TNF-R, IL-2, and CD40L are shown (white arrows). Microscopy images show contact between human CD4+ T cells and autologous immature or mature DCs and were performed as in Miro et al.16


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responses that can be detrimental to the organism (reviewed by Sakaguchi and Sakaguchi112). Interac-tions of Treg cells with DCs are required for their activation under physiological conditions,113 and they do form ISs with DCs.114,115 ISs between Tregs and DCs are different from synapses formed between T-helper cells and DCs. Neuropilin-1 (Nrp-1), which is expressed by most Treg cells but not naïve T-helper cells, has been shown to pro-mote prolonged interactions with immature DCs, thus resulting in higher sensitivity of Tregs to low concentrations of antigen.116 Tregs were also shown to be more mobile than naïve T cells in vitro, out-competing the latter by aggregating around DCs. 115 These aggregations of Tregs around DCs were shown to specifically down-regulate the expression of CD80/86 on DCs, suggesting that Treg might prevent antigen-reactive naïve T cells from being activated by antigen-presenting DCs by competing for their access to DCs and decreasing the ability of the DCs to present antigens.115 Some of these effects might be due at least in part to the difference of localization of PKCq at the synapse between TH cells and Treg cells (reviewed by Zanin-Zhorov et al.117). Indeed, PKCq, which is recruited at the IS in TH cells,21 is sequestered away from the IS in Treg, and this seems to enhance the function of Treg.118 Moreover, PKCq recruitment at the IS has been shown to destabilize synapse formation in naïve T cells.119 Thus, Tregs that do not polarize PKCq at the synapse form more stable conjugates with DCs and thus compete with naïve T cells for their inter-actions with DCs. In the “ménage à trois” between TH, Treg and DC, Treg cells have been shown to significantly inhibit the recruitment of protein kinase Cq (PKCq) to the IS of the TH cell as long as both T cells were of the same Ag specificity and were contacting the same DC.114 In vitro Tregs have also been shown to inhibit the polarization of the secre-tory machinery of TH toward the APC, including DC, through a TGF-b–dependent mechanism.120 Thus Tregs can block the targeted delivery of the helper signal to DCs.

It remains to be determined whether these effects require Treg-dependent modifications of

DCs or if the DC serves as a “platform” to allow cognate interactions between Treg and TH cells. It would surely be interesting to look at the potential modifications on the DC side of DCs interacting with Treg cells.

3. On the DC SideAgain, analysis of cytoskeleton remodeling at the IS on the DC side has not been given com-parable attention in the research field. Analyzes are sparse, yet results obtained on DCs often mirror some of the aspects first revealed in T lymphocytes. One of the specificities of DCs is that unlike B cells they are much more “active” when contacting T cells. Indeed, in vitro and in vivo analyses have revealed that they project dendrites that are likely to facilitate interactions with T cells.121,122 Studies have shown that DC actin cytoskeleton focally polarizes during the formation of conjugates with CD4+ T cells and that inhibitors of DC actin polymerization reduce the formation of conjugates as well as T-cell activation.123 This type of membrane activity is generally controlled by the actin cytoskeleton, which is in turn regulated by small guanosine triphosphatases (GTPases) of the Rho family.124 Indeed, the Rho family of GTPases, including Rac1 and Rac2, has been shown to control the formation of dendrites in mature DCs control-ling T/DC interactions.125 Moreover, Rac1 and Cdc42, in which activities are increased by matu-ration, have also been involved in the transport of essential immunostimulatory molecules (i.e., MHC class I and CD86) to the DC surface.126 Other proteins involved in actin remodeling have been shown to play a role in activation of T cells by DC. The Wiskott-Aldrich syndrome protein (WASp), key regulator of actin polymerization in hematopoietic cells, has been shown to control both DC migration to the lymph nodes, stabi-lization of T-DC conjugates, IS formation in T cells, and activation of naïve T cells127–129 (and reviewed by Monypenny et al.130). Notably, actin cytoskeleton remodeling of DCs at the IS formed between NKs and DCs was recently shown to also



control NK activity. Indeed, WASp-dependent actin reorganization in the DCs was reported to allow for MHC-I clustering at the immuno-logical NK–DC synapse, thereby preventing the formation of a lytic synapse in the NK cells and preventing DC killing.131

All of these studies stress the fact that forma-tion of an effective contact between T cells and DCs is dependent on active cytoskeletal rearrange-ments in DCs.

We demonstrated that CD4+ T-cell-depen-dent signals induced cytoskeleton remodeling in DCs. This remodeling was characterized by striking elongation of immature DCs that change from a roundish shape of 10–15 mm in diameter to an elongated “neuron-like” shape that is 50–80 mm in length.132 These elongated DCs established contact with both CD4+ T cells and other DCs and tended to form a network of interdigitat-ing DCs after 4–6 h of antigen-specific contact with T cells. These changes were accompanied by modifications of the actin cytoskeleton that were dependent on antigen-specific interaction between T cells and DCs. In our study, DCs showed actin filaments along the plasma mem-brane, as well as polymerized actin-rich “cups” in the contact zones with T cells. Moreover, CD4+ T cells induced dissolution of actin-rich dynamic structures, called podosomes, found on several cells from the myeloid lineage133 (Figure 1). This dissolution of podosomes induced by T cells induced a stronger mobility of the DCs that were then able to migrate. We were able to show that these modifications were induced by chemokines produced by T-cell–DC Ag-specific contacts and that they contribute to a better activation of T cells.132 Recent studies have also shown that regulatory T cells (Treg) modify the chemoattractant properties of DCs by inhibiting their production of chemokines.134,135 This could result in less recruitment of T cells in the lymph nodes and could explain in part the inhibitory effects of Tregs on T-cell immune responses. It remains unclear whether direct contact between Tregs and DCs is important for these effects, and

it will be interesting to observe the morphologi-cal modifications Tregs can induce in DCs after cognate interactions.

Altogether, these studies show that Ag-specific interactions between T cells and DCs induce actin remodeling both on the T-cell side and on the DC side, and that these modifications are key in regulating T-cell and DC functions.

Studies have also shown a role for DC micro-tubule dynamic and MTOC polarity on T-cell activation by DCs. DCs activated by Toll-like receptor (TLR) agonists have been shown to reori-ent their MTOC toward the interacting T cell dur-ing antigen-specific synapse formation through a Cdc42-dependent mechanism. IL-12 was enriched around the MTOC at early time points after TLR ligation and was polarized toward the DC–T-cell interface in antigen-specific synapses.136 The same reciprocal reorientation of MTOC in T-cell–B-cell conjugates has also been recently shown.137 These results suggest that MTOC polarization in DCs, as in T cells, might increase the local concentra-tion of proinflammatory mediators at the IS, thus controlling T-cell priming.

III. CONCLUSION AND PERSPECTIVES

Ample evidence shows that the organization of the contact zone between T lymphocytes and DCs plays a crucial role in both T-cell and DC activation. Much research has been done to elu-cidate the movement of receptors and remodeling of the cytoskeleton on the T-cell side of the IS. However, because studying the IS on the T-cell APC has been replaced by artificial lipid bilayers or beads presenting ligands of the TCR and of co-receptors, the APC has long been thought to be passive in IS formation. It is now clear that clustering of membrane and signaling molecules and remodeling of the cytoskeleton on the APC side, especially in DCs, plays an important role in T-cell activation. It would now be interesting to study the remodeling of the contact zone in different DC subpopulations, which have been


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described both in murine and human models to have distinct functions. Such studies should bring valuable information on the correlation of IS formation on the DC side and functions such as cross-presentation, cytokine secretion, and TH polarization. Moreover, recent studies have shown that T lymphocytes secrete microvesicles that contain microRNAs,138,139 which can act on cognate APCs. APCs including DCs have also been shown to secrete these microvesicles.140 The IS may thus be an ideal place for secretion of these microvesicles by both cellular partners. The precise role this secretion might have on both T lymphocytes and DCs remains to be determined.

Finally, current state-of-the-art approaches almost exclusively focus on the role of the spa-tial distribution of protein receptors, signaling molecules, and cytoskeleton remodeling in the communication between cells at the IS. However, it is highly likely that other factors, such as physi-cal parameters (e.g., forces and mechanics and/or membrane stiffness) contribute to the formation of the IS and the activation of both cellular part-ners. Indeed, remodeling of the cytoskeleton on both the T-cell and DC sides of the IS strongly suggest that forces are exerted that might be sensed by the cells and modify their fate. The biomechanical properties of the different APCs, including DCs, should thus be investigated as well as the role these mechanical properties have on IS formation and T lymphocyte activation. These investigations would open up a new area of research in immunology.

ACKNOWLEDGMENTS

This study was supported by the Institut Curie, the Institut National de Santé et de Recherche Médicale (INSERM). AB and MT were funded by la Ligue contre le Cancer and KC by la Fondation pour la Recherche Médicale. The authors are not aware of any affiliations, memberships, funding, or financial holdings that might be perceived as affecting the objectivity of this review.

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Crosstalk between T lymphocytes and dendritic cells

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