What is an immunological synapse?

Microbes and Infection 12 (2010) 438e445 www.elsevier.com/locate/micinf

Review

What is an immunological synapse? Jose´ Luis Rodrı´guez-Ferna´ndez*, Lorena Riol-Blanco1, Cristina Delgado-Martı´n1 Centro de Investigaciones Biolo´gicas, Consejo Superior de Investigaciones Cientı´ficas, 28040 Madrid, Spain Received 5 February 2010; accepted 1 March 2010 Available online 12 March 2010

Abstract Immunological synapses (IS) are emerging as highly organized 3D structures -formed by surface and cytoplasmic signalling and cytoskeletal molecules e that assemble at the zone of contact between a T cell and an antigen presenting cell (APC). The IS control functions that allow APC and T cells modulate the immune response. Ó 2010 Elsevier Masson SAS. All rights reserved. Keywords: Immunological synapse; Dendritic cell; Antigen presenting cells; Lymphocyte; Cellecell contact

1. Introduction Immunological synapses (IS) have been defined as ‘‘stable T celleantigen presenting cell junctions’’ [1,2]. Although there are several types of IS, with different types of APC and T cells involved, in this review, we focus largely on the IS formed between dendritic cells (DCs) and CD4 T cells. Hereafter, we name IS-DC and IS-T cell, to the structures formed at the DC and CD4 side of the IS, respectively. It would be even more informative to denote these IS as ISDC(op CD4) and IS-CD4(op DC), respectively, with the subscripts indicating the cell opposite to the one whose IS is under analysis. This nomenclature has the additional advantage of allowing a better definition of most cellecell interactions described so far in the immune system, including previously described interactions of CD4 T cells with plasmacytoid dendritic cells, B cells with different APCs, NK and cytotoxic CD8 T cells with target cells. On the other hand, this terminology allows a better description of novel IS that may be formed between T cells and other recently described APCs (gd T cells, basophils and mast cells). Most studies on the IS have focused on the IS-T cell and, consequently, this is the region on which most information is * Corresponding author. Tel.: þ34 91 837 3112; fax: þ34 91 536 0432. E-mail address: [email protected] (J.L. Rodrı´guez-Ferna´ndez). 1 Both authors contributed equally to this review. 1286-4579/$ - see front matter Ó 2010 Elsevier Masson SAS. All rights reserved. doi:10.1016/j.micinf.2010.03.003

available [1e3]. This is likely one of the reasons why the concept of IS is often used as a synonymous of IS-T cell, although this is an oversimplification and it is more appropriate to indicate clearly the cells involved in IS formation. In this review, we summarize data on the structure and function of the IS-CD4 T cell and compare it with recent information on the structure and function of the IS-DC. From this analysis it becomes apparent that, despite differences in the molecular composition between these two types of leukocytes, a common pattern is shared by these two regions. In this regard, the IS emerge as highly dynamic and complex 3D structures formed in both, the T cell and the APC, at the contact zone between these cells. In these regions takes place an intense signalling which is most likely required to maintain the organization and to control the functions of these superstructures. 2. Stability is a key feature of the IS Early in vitro studies demonstrated the high duration of the IS formed between APC and T cells [4]. It was also observed that several hours of interaction between DCs and T cells were necessary before T cell activation could be observed [4]. Biophysical analysis confirms that the IS reflects the strong interactions that keep APC and T cells bound together [5]. Two-photon microscopy studies have also shown that DCs and T cells maintain, consistent with IS formation, long interactions in the lymph nodes. Thus, durability is an important

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feature of the IS observed both in vitro and in vivo (see [4] and references therein). The long duration of the IS suggests sophisticated molecular mechanisms controlling the stability of the cytoskeletal networks that maintain these structures. As discussed below, receptors located at the IS-T cell and also at the IS-DC, must relay signals that lead to the activation of adhesion receptors in these areas and to the organization of a stable network of cytoskeletal components necessary to confer stability to these regions. With respect to cytoskeletal remodeling that maintains the stability of the IS-T cell and IS-DC, it has been suggested that the stabilization of the IS requires of the preactivation of T cells, which results in the up-regulation of CD40L in these cells. This ligand binds to CD40 in mature DCs, leading to the reorganization of the actin cytoskeleton in the DC, sustaining DC-T cell binding, which allows T cell activation and proliferation [6]. Therefore, as discussed below, surface and signalling molecules in the APC and the T cells are responsible of organizing the cytoskeleton and conferring stability at these structures. Finally, and also related to the concept of the durability of the IS between T and DCs, it has also been shown that longlasting IS, observed with mature DCs, correlate with the development of immunity, while more shorter interactions, observed when T cells interact with immature DCs, correlate with tolerance [7]. Recently, interactions of mature DCs and T cells that last for several hours, although still for shorter periods of time than those which induce immunity, were also observed when interactions between DCs and T cells lead to tolerance [8]. The latter observation is equally interestingly in the context of the different functions of the IS because it suggests the possibility of the existence of IS specialized in tolerance. These observations are also providing some hint on why it is necessary a stable IS. The structural stability and, consequently, the durability of IS, are probably required to allow the IS-APC and IS-T cell to carry out highly specialized functions involved in immunity, including, among other tasks, the control of the transcription of genes involved in T cell activation that require hours [9]. 3. Deconstructing the IS-T cell Three basic sets of molecules are found at the IS-T cell, namely, surface proteins, on the membrane, and cytoskeletal and signalling molecules, in the cytoplasmic regions [1]. Below we will briefly analyse some basic molecular components and discuss the signalling that takes place at the IS-T cell, emphasizing on recent new data on this structure. 3.1. Surface proteins at the IS (T cell) These molecules in most cases relay, directly or indirectly, intracellular signals that control the organization and functions of this region [1]. Among the surface proteins are included receptors and ligands that support adhesive interactions between T cells and APCs and, in addition, play a key role in initiating the formation and organization of the IS-T cell. The

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integrins VLA4 (a4b1) (ligand VCAM-1 on the APC) and LFA-1 (aLb2) (ligands Intercellular adhesion molecule-1 and -3, ICAM-1, -3, on the APC) play a key role during these processes. The absence of ICAM-1, either on T cells or DCs, prevents IS formation [4]. Interestingly, VLA4 is observed at the IS-T cell even when the APC is not expressing its ligand VCAM-1 [10]. Moreover, integrins at the IS-T cell are able to induce a variety of intracellular signalling that may be involved not only in the regulation of adhesion to APC, but also in other functions of T cells. In this regard, T cells deficient in LFA-1 display a much reduced number of IS-T cell with DCs when the latter cells present low doses of antigen; although the percentage of IS-T cell increases significantly when the dose of antigen displayed by the DCs is high, suggesting that LFA-1 relay signals that lower the dose of antigen required for IS formation [11]. The T cell receptor (TCR) is a central surface receptor component of the IS-T cell that plays a key role in T cell activation [12]. Engagements of TCR by cognate antigens presented by APCs are believed to be important for IS-T cell formation. In this regard, signals generated from the TCR in T cells lead to the activation of LFA-1, engagement of the ligand ICAM-1 on APC and, eventually, to IS-T cell formation. IS-T cell co-stimulatory molecules CD2 and CD28 are also required for T cell activation. Deficiencies of either CD2 or CD28 co-stimulatory molecules do not greatly affect IS-T cell formation in T cells [13]. However, T cells that lack both molecules present a reduction of almost 60% in the number of IS when compared to wild type control T cells, suggesting that these receptors cooperate to induce IS formation [13]. Finally, it has been shown that stimulation of TCR and CD28 can relay signals that result in the polymerization of actin [12], suggesting that these molecules can contribute to IS-T cell formation by regulating the structure of these regions (see below). Emphasizing on the complexity of this region, new surface molecules are continuously identified at the IS-T cell. In this regard, in experiments where DCs were used as APCs, STIM1 and Orai1, two calcium release-activated calcium (CRAC) channels, were observed to cluster at the IS-T cell [14]. Upon TCR stimulation, the Caþ2-activated Kþ channel KCa3.1 also compartmentalize at the IS-CD4 T cells [15]. Since upon engagement of TCR by antigens displayed on the APCs, a rapid increase in intracellular Caþ2 takes place in T cells and sustained intracellular Caþ2 concentrations is necessary for T cell gene expression and activation, it is possible that these channels contribute to keep high intracellular Caþ2 concentration [14]. The cytokine receptor interferon-g-receptor (IFg-R), clusters at the IS-T cell, where it can contribute to T cell helper differentiation [16]. Chemokine receptors CCR5 and CXCR4 locate to the IS-T cell where they may behave as costimulatory molecules [17]. The accumulation of these receptors at the IS-T cell suggests that this region may contribute to target selectively T cells in response to cytokines and chemokines (see below). Interestingly, the cellular prion protein (PrPC), which is a GPI-anchored cell surface protein,

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is also found at the IS-T cell. However, in contrast to DCs (see below), the absence of this protein does not affect the activation of T cells [18]. Among the set of surface proteins that form the IS-T cell have also been identified proteins previously observed at other cellecell junctions, including the neuromuscular junction [19]. Agrin, a proteoglycan and aggregating factor at neuromuscular junctions has also been observed in the IS-T cells, where probably performs similar aggregating functions [20]. Another neuronal protein, the semaphoring receptor neuropilin, has also been observed at the IS-T cell, where it may also contribute to IS-T cell formation [21]. Another feature of the IS-T cell is the high degree of organization of the surface molecules at this region [22,23]. Initial studies performed with T cells that either formed IS with B cells or that were allowed to adhere to lipid bilayers, which were used as surrogate APCs, provided a relatively simple ‘‘bull-eyed’’ pattern of the IS-T cell, with the TCR at the center of the IS-T cells and LFA-1 in an outer ring; however, recent analysis where DCs have been used as APC, show that the IS-T cell may display a more complex organization with multiple foci of TCR receptors, instead of a single central cluster [24]. Receptor topology appears to be an important factor to determine the location of the different surface components and the functionality of IS-T cell. In this regard, similar size pair surface proteins, including TCR, and co-stimulatory molecules CD4, CD8, CD28 and CD2, which extend from the surface of the cell about 7e8 nm tend to aggregate at the central parts of the IS-T cell. Whereas larger proteins like CD43 and CD45, which protrude some 45 nm over the cell, remain at the outer parts of the IS-T cell. The importance of surface protein topology is dramatically shown in experiments where it was observed that the activation of T cell by APC was inhibited when the extracellular domain of the CD2 ligand CD48 was increased in length by adding an additional domain [19]. Finally, to the organization surface proteins at the IS-T cell are believed to contribute the interactions that these proteins establish with the cytoskeleton (see below) and their inclusion in different domains, including tetraspanin and rafts domains [25,26]. 3.2. Cytoskeletal molecules at the IS-T cell Actin, tubulin and cytoskeletal-associated proteins are important to maintain the organization of the IS-T cell [27]. Apart from their structural roles, cytoskeletal proteins can also contribute to control the signalling in this region by providing scaffold that can be bound by different signalling molecules [1]. Concomitant with IS-T cell formation, a variety of cytoskeletal proteins are recruited to this region, which is also known to be a site of active actin polymerization [28]. Moreover, disruption of actin organization by using pharmacological inhibitors or interfering with the activity of actinremodeling regulators, like cofilin, disrupts IS formation and block T cell activation [29]. It has also been shown that upon stimulation of LFA-1, F-actin in the IS-T cell is organized in

the form of a ‘‘cloud’’ that lowers the threshold required for subsequent T cell activation [30]. The actin cytoskeleton plays also an important role in the organization of surface proteins at the IS-T cell. In this regard, myosin motors transport surface protein towards the IS-T cell [31]. In addition, ERM (ezrine radixinemoesin) family of proteins, which connect the actin cytosleketon to different surface proteins, contribute to restrict proteins like CD43 to the outer ring of the IS-T cell [32]. Perturbation of ERM proteins blunt IS-T cell formation and T cell activation, suggesting the key role of ERM proteins in these processes [28,33]. During IS-T cell formation, the Microtubule-Organizing Center (MTOC) and the Golgi apparatus get oriented towards the IS-T cell [27]. Reorientation of these cytosolic organelles is believed to be required for the delivery of signalling and cytoskeletal molecules to the IS-T cell. 3.3. Signalling molecules at the IS-T cell The clustering of signalling molecules at the cytoplasmic regions of the IS-T cell suggests a flurry of signalling processes that, predictably, are involved in the regulation of the structure and function(s) of this region [1,34]; although the specific contribution of individual signalling molecules to the function(s) and/or organization of these regions is just starting to be clarified. A detailed description of all the signalling that takes place at the IS-T cell is beyond the scope of this short review. We briefly provide an overview of the main signalling molecules observed in this area to emphasize on the one hand on the complexity and, on the other hand, on the organized nature of signalling that takes place the IS-T cell. Among the signalling molecules observed in this region are found kinases, including tyrosine kinases like, Lck, ZAP-70 and Fyn [1]. These kinases are important for the transmission of signals from the TCR, a receptor that is devoid of enzymatic activity. Lck initiates the signalling from the TCR; ZAP-70, is responsible of phosphorylating the adaptor Linker for activation of T cells (LAT). Phosphorylated-LAT recruits a variety of enzymes including phospholipase C-g (PLC-g), the Grb2/SOS complex, PI3K and the GADS/SLP-76/Nck/Vav complex, that connect LAT to the small GTP-ases Rac and to the recruitment of PKCq [35]. The kinase p21-activated kinase 1 (PAK1), also translocates and becomes activated at the IS-T cell [36]. Activation of PI3K at the IS-T cells, results in accumulation of phosphatidylinositol (3,4,5) triphosphate (PIP3) at the cytoplasmic leaflet of the plasma membrane [37]. The increase in PIP3 preceded the increase in intracellular calcium following stimulation of the TCR. Apparently, the increase in PIP3 is not related to IS formation, because neither the number of IS-T cell formed nor the signalling that lead to phosphotyrosine accumulation was altered when PI3K was inhibited [38]. PIP3 controls the localization and the activation of PH-domaincontaining proteins, including the serineethreonine kinase Akt, which is also activated at the IS-T cell [39]. Active Akt phosphorylates and inactivates FOXO1, resulting in enhanced proliferation of T cells [39]. The kinase transforming Growth Factor beta (TGF-b)-activated kinase (TAK) is also recruited

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to the IS-T cell from where it can modulates NFkB-dependent gene expression by phosphorylating the NFkB regulator IkB kinase (IKK) [40]. PAK1, forming a complex with PAK interacting exchange factor (PIX), and G protein coupled receptor kinase interactor (GIT), is recruited and activated at the IS-T cell, from where this kinase regulates T cell transcriptional responses [36]. Adaptors are molecules with no enzymatic activity that serve to connect signalling and cytoskeletal molecules. The adaptor CD2AP was earlier appreciated as an important regulator of the organization of the IS-T cell [1]. Subsequently, a variety of adaptors has been described in the IS-T cell, including Uncoordinated 119 (Unc119) [41], Linker for activation of T cells (LAT) [42], Adhesion and degranulation promoting adaptor protein (ADAP) [43], SH2 domaincontaining leukocyte phosphoprotein of 76 kDa (SLP-76) [43] and Discs large homolog 1 (Dlgh1) [44]. Unc119, which controls the activation of the GTPase rat brain 11 (Rab11), a family protein involved in intracellular transport, regulates the recruitment of the actin motor myosin 5B and the organization of a multiprotein complex on endosomes that transport the kinase Lck to the IS-T cell [45], a process that is important for the formation of the IS-T cell and for T cell activation. Endosomes also transport to the IS-T cell other signalling proteins, like the adaptor LAT [42], and surface receptors, like the TCR [46]. In this regard, the intracellular transport regulator Rab35 can controls IS-T cell formation by controlling TCR enrichment in this region [47]. Intraflagellar transport 20 (IFT20), a component of the molecular complex that promotes cilia assembly, also contributes to the trafficking of TCR-CD3 complex to the IS-T cell [48]. LAT which, as indicated above, is phosphorylated by the kinase ZAP-70, recruits a variety of signalling molecules involved in T activation, including Phospholipase Cg1, the adaptor Grb2 and the guanine nucleotide exchange factor Son of sevenless (SOS), GADS and SLP-76, all involved in T cell activation [1]. ADAP, which interacts with dynein, a motor protein that transports cargo on microtubules, has been involved in the regulation of MTOC polarization at the IS-T cell [49]. Dglh1, an adaptor observed at neuromuscular junctions, also localizes to the IS-T cell, where it forms a complex with Lck-ZAP-70and WiskotteAldrich syndrome protein (WASp). Dlgh1 orchestrate antigen-induced actin polymerization leading to T cell synapse assembly and also regulates the Ca2þ dependent transcription factor family nuclear factor of activated T cell (NFAT), cytokine production and effector function of T cells [44,50,51]. A variety of actin regulators accumulate in the IS-T cell, including small GTPases (Rac1, Rho and Rap), the actin nucleator actin-related protein 2/3 (Arp2/3), different guanine nucleotide exchange factors (GEFs) (e.g. Vav) and other actin regulators like WASP, WAVE, and HS1 [28]. At the IS-T cells accumulate also deacetylases (e.g. HDAC6), which regulate the tubulin cytoskeleton [1,26]. Interestingly, HDAC6, which also binds F-actin, is involved in clearing misfolded proteins from the cell. Finally, further testifying on the complexity of this structure, at the IS-T cell are also found clustered

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mitochondria which are believed to be an important component of these regions required for T cell activation upon IS formation [52]. 4. Functions of the IS-T cell The complexity of the signalling at the IS-T cell clearly suggests that this is a functionally important region of T cells [53e56]. Several hypotheses, not mutually exclusive, have been put forward to explain the mechanisms whereby the IS-T cell may control the functions of T cells. Next we discuss very briefly these hypotheses [53e56]. IS formation could be a molecular mechanism used to arrest lymphocytes and allow the subsequent activation of these cells. As discussed above, adhesion receptor at the IS-T cell regulate the adhesion between APC and T cells, the stopping in the lymph nodes of the highly motile lymphocytes and the organization of the cytoskeleton in the T cell. The stability of the IS and the immobilization of the T cells permits an adequate activation of these cells. The IS-T cell can be a regulator of TCR signalling and T cell activation. Initial studies on the IS-T cells posited the concept that this region controls the activity of the T cell receptor (TCR) [22,57]. However, since the signalling from the TCR was maximal well before the IS-T cells was completely formed, it was suggested that TCR signalling does not require previous IS formation [58]. As endocytosis was observed at the central region of the IS-T cell, it was proposed that the IS-T cell could regulate the removal of the TCR once the signalling from this receptors was terminated [55,59]. More recently, a combination of in silico and in vitro analysis of the IS-T cell has led to a proposal that incorporates these prior views on the IS. The new studies suggest that the IS-T cell is an adaptive system that boosts low and attenuates high TCR signals. In this regard, strong antigenic stimuli lead to the rapid degradation of the TCR and weak antigenic stimuli abate the rate of degradation of the TCR [60]. Thus, the IS-T cell may be a molecular platform that maintains the signalling in the presence of antigens with different strengths [59]. The intercellular space at the IS could be a location where cytokines and chemokines could be secreted and consequently directed to regulate T cells. At the IS-T cell cluster CCR5 and CXCR4 and interferon-gamma receptors (IFNg-R) implying that these receptors can be targeted selectively at the IS region. Furthermore the co-clustering of IFNg-R and TCR was prevented in the presence of IL4 [16]. In T cells, interleukin 2 (IL-2) and interferon-g (IF-g) are secreted exclusively in intercellular space of the IS, while other cytokines like TNFa and CCL3 are released multidirectionally [61]. Earlier studies appreciated that the IS-T cell may control the orientation of the MTOC of the T cell and also the secretion apparatus of these cells. The segregation of IFNg-R at the IS-T cell has led to the suggestion that the commitment of na€ıve CD4 T cells into Th1 T cells can be controlled from this structure [16]. The IS-T cell could regulate asymmetric cell division. It has also been suggested that the polarization of the molecules at the IS-T cell is responsible for the asymmetric division of T cells, which results in the

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simultaneous generation of memory and effector T cells [62]. Taken together, the results discussed above show that the IS-T cell may control multiple functions in T cells. 5. Deconstructing the IS-DC When compared to the knowledge garnered on the IS-T cell, the information available on the IS-DC is much more limited. However, despite this paucity of information, the slowly emerging data suggest that the IS-DC presents, similar to the IS-T cell, a design that includes surface proteins and, in the cytoplasmic regions, structural and signalling components. 5.1. Surface proteins at the IS-DC On DCs, Major histocompatibility complex class II (MHC II) molecules cluster opposite to CD3 aggregates on T cells [26]. DCs are able to form IS with T cells even in the absence of MHC I or MHC II molecules, which suggests that MHC proteins are not required for IS-DC formation [63]. The integrin ligands ICAM-1 and ICAM-3 form a ring in the outer part of the IS (DC), opposite to the integrin LFA-1 on the IS-T cell [26]. As indicated above, LFA-1-ICAM binding is necessary to allow the interactions between DCs and T cells during IS formation [4]. CD40 molecules are found at the central regions of the IS-DCs [64]. At the IS-DCs are also found B7-1 and B7-2. Although the latter molecules can be both ligands of the T cell co-stimulatory receptors CD28 and CTLA-4; however, B7-1 binds preferentially to CTLA-4 and B7-2 to CD28. B7-1 may enhance the number of IS formed between APC and T cells and the antigen-dependent proliferation of the latter cells. Interestingly, even in the absence of B7-1 and B7-2, CD28 localize, although at a slightly reduced level, at the IS-T cell. However, CTLA-4 requires either of these two receptors to localize to the IS-T cell [65]. CD70, a TNF-related transmembrane protein, also localizes to the IS-DCs. The interaction of CD70 on the IS-DC with CD27 on the IS-T cell is essential for the priming of T cells [66]. Moreover, the vesicular trafficking of CD70 and MHC class II is regulated by the microtubule-associated dynein motor complex [66]. The semaphorin receptor Plexin-A1 also localized at the IS-DCs [67]. Reduction of Plexin-A1 in DCs by using interference RNA, blunts the T cell stimulatory ability of peptide-pulse DCs and the actin clustered at the IS-DC [66,68]. Plexin co-localizes at the IS-DCs with its co-receptor Neuropilin [21]. Interference with Neuropilin using neutralizing antibodies, inhibits T cell-DC clustering and DC-induced proliferation of T cells [21]. The Notch receptor and Notch ligands (Delta like 1 and Jagged 1) have also been found in the IS-DCs, with the ligands locating at the central part of this IS and the receptors on an outer peripheral ring [69]. After IS-DC formation, processed Notch 1 receptors intracellular domains accumulate in the nucleus of DCs, where they can regulate DC gene expression [69]. The cellular prion protein (PrPC) which, as indicated above, locates at the IS-T cell, also accumulates at the IS-DC [18]. Moreover, although PrPC deficiency does not affect the maturation of DCs, DCs

lacking PrPC are not able to induce a full activation of T cells [18]. A thoroughly study on the organization of surface proteins at the IS-DC has not been performed so far. However, resembling the situation at the IS-T cell, surface molecules of the IS-DC, e.g. MHC II molecules, localize also in rafts and in domains that include the tetraspanin CD81, suggesting that surface proteins at the IS-DC may be also organized [26,70]. 5.2. Cytoskeletal molecules at the IS-DC In the cytoplasmic regions of the IS-DC cluster F-actin and the actin bundling protein fascin [71,72]. Disruption of F-actin in DCs by treating the cells with cytochalasin D, blocks IS formation and T cell activation [71,72]. F-actin is probably necessary to maintain the structure and the signalling from the IS-DC. Microtubules also play a role in the organization of the IS-DC. Long microtubule ‘‘railroads’’, which originate in the MTOC and that get oriented towards the IS-DC, bind motor proteins that are instrumental in transporting as cargo, along the microtubules and towards the IS-DC, vesicules called endolysosomal tubules that contain MHC II, and other molecules like, probably, CD70 [66,73,74]. Thus, microtubule ‘‘railroads’’ serve as efficient systems to deliver specific components to the IS-DC. 5.3. Signaling molecules at the IS-DC At the IS-DC are observed clusters of tyrosine phosphorylated proteins indicating that these are active signalling regions [72]. A variety of actin regulators are also found in the IS-DC, including phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) [75] and vasodilatador-stimulated phosphoprotein (VASP) [76]. The small GTPase Rho is activated in response to the stimulation of the Plexin-A1 receptor [67]. IS-DC deficient in WiscotteAldrich syndrome protein (WASP), an actin nucleating factor that is activated by Rho and (PI(4,5)P2), do not form stable IS with T cells, which results in impaired T cell priming [77,78]. The kinase Akt that participates in the regulation of DC survival (see below) also localizes to the IS-DC [72]. Adaptors, which as indicated above, are key transducers of information in different signalling regions, including IS-T cells have also been localized to these regions. Spinophilin, a PDZ-containing adaptor described at the neural synapses, clusters at the IS-DC [79]. DCs deficient in spinophilin display have a reduced ability to present antigen [79]. DCs lacking the DC adaptor SKAP-HOM show delays in IS-DC formation, suggesting that this molecule may also locate at the IS-DC [80]. 6. Function(s) of the IS-DC Since DCs are APC it seems reasonable that one of the functions of the IS-DC could be to facilitate an adequate activation of T cells. In this regard, opposite to the CD3 on T cells, cluster MHC molecules on the DCs. The clustering of MHC at the IS-DC correlates with the ability to induce T cell activation [26]. Surface molecules at the IS-DC can contribute

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to stabilize the organization of their counterreceptors on the IS-T cell, allowing the correct function of this structure [16,61,62,81,82]. As indicated above, microtubule ‘‘railroads’’, which start at the MTOC, transport MHC II molecules to the IS-DC [73]. Likely that these MHC complexes could be transported to the IS-DC associated to co-stimulatory molecules [83]. It is possible that the IS-DC may orchestrate the transport of different receptors and signalling molecules to this region. Recently, we have also shown that IS-DCs formation protects the DCs from apoptosis both in vitro and also in vivo [72]. Consistent with the pro-survival function of the IS-DC, we observed translocation of the pro-survival kinase Akt1 to the IS-DC [72]. We showed that Akt is important to induce the pro-survival effects of the IS-DC because its pharmacological inhibition reduced the pro-survival effects caused by IS-DC formation [72]. The activation of Akt was also followed by inhibition of transcription factor FOXO, which is proapoptotic in DCs, and by the activation of NFkB, which promotes survival in these cells [72,84]. We also found that stimulation of CD40 induces activation of Akt, suggesting that CD40 can induce anti-apoptotic signalling from the IS-DC [72]. These results are consistent with high durability of the IS [4], implying that during this period from the IS-DC can be relayed to the DC intracellular signals that inhibit apoptosis and allow the activation of T cells. 7. Conclusions So far, the studies on the IS have focused largely on the T cells and the term IS is often used as a synonym of IS-T cell; however, it is clear that equally important is the DC side of the IS and, predictable, any APC involved in stable interactions with T cells. Another drawback of the current studies on the IS is the excessive emphasis on the organization of surface receptors when defining this structure. Although surface receptors probably initiate and possible orchestrate the organization of the whole IS, it is more appropriate to consider the IS-T cell and the IS-DC as organized 3D structures, consisting of surface and cytoplasmic molecules that assemble in the T cell and DC side of the IS. Although there is a considerable degree of variability in the types of APCs and T cells that have been used to analyse IS formation. However, it is very likely that there is a basic set of molecular components of the IS-T cells that could vary in the different types of T cells (e.g. na€ıve or memory CD4 T cells or CD8 T cells) or APCs involved in IS formation. Future studies will have to dissect out how the flurry of signalling that takes place at the IS controls the organization and functions of these structures. Acknowledgements We sincerely apologize to those colleagues whose papers could not be cited due to the brevity of this review. This work was supported by the Ministerio de Educacio´n y Ciencia (grants BFI-2001-0228 and SAF2005-00801) and Red de Investigacio´n en Inflamacio´n y en Enfermedades Reuma´ticas (RIER) (grant RD08/0075 (RETICS Program/Instituto de

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Salud Carlos III)) (J.L.R.-F.), a scholarship associated with grant PI021058, conferred by the Fondo de Investigacio´n Sanitaria (L.R.-B.), and fellowship Formacio´n de Personal Investigador (FPI) conferred by the Ministerio de Educacio´n y Ciencia (C.D.-M.). References [1] M.L. Dustin, The cellular context of T cell signaling. Immunity 30 (2009) 482e492. [2] D.R. Fooksman, S. Vardhana, G. Vasiliver-Shamis, J. Liese, D. Blair, J. Waite, C. Sacrista´n, G. Victoria, A. Zanin-Zhorov, M.L. Dustin, Functional anatomy of T cell activation and synapse formation. Annu. Rev. Immunol. 28 (2010) 1e27. [3] S. Valitutti, L. Dupre´, Plasticity of immunological synapses. Curr. Top. Microbiol. Immunol. 340 (2010) 209e228. [4] D.M. Davis, Mechanisms and functions for the duration of intercellular contacts made by lymphocytes. Nat. Rev. Immunol. 9 (2009) 543e555. [5] B.H. Hosseini, I. Louban, D. Djandji, G.H. Wabnitz, J. Deeg, N. Bulbuc, Y. Samstag, M. Gunzer, J.P. Spatz, G.J. Ha¨mmerling, Immune synapse formation determines interaction forces between T cells and antigenpresenting cells measured by atomic force microscopy. Proc. Natl. Acad. Sci. U. S. A. 106 (2009) 17852e17857. [6] T. Rothoeft, S. Balkow, M. Krummen, S. Beissert, G. Varga, K. Loser, P. Oberbanscheidt, F. van de Boom, S. Grabbe, Structure and duration of contact between dendritic cells and T cells are controlled by T cell activation. Eur. J. Immunol. 36 (2006) 3105e3117. [7] S. Hugues, L. Fetler, L. Bonifaz, J. Helft, F. Amblard, S. Amigorena, Distinct T cell dynamics in lymph nodes during the induction of tolerance and immunity. Nat. Immunol. 5 (2004) 1235e1242. [8] G. Shakhar, R.L. Lindquist, D. Skokos, D. Dudziak, J.H. Huang, M.C. Nussenzweig, M.L. Dustin, Stable T cell-dendritic cell interactions precede the development of both tolerance and immunity in vivo. Nat. Immunol. 6 (2005) 707e714. [9] J.B. Huppa, M. Gleimer, C. Sumen, M.M. Davis, Continuous T-cell receptor signaling required for synapse maintenance and full effector potential. Nat. Immunol. 4 (2003) 749e755. [10] C.C. DeNucci, J.S. Mitchel, Y. Shimizu, Integrin function in T cell homing to lymphoid and non-lymphoid sites: getting there and staying there. Crit. Rev. Immunol. 29 (2009) 87e109. [11] Y. Wang, K. Shibuya, Y. Yamashita, J. Shirakawa, K. Shibata, H. Kai, T. Yokosuka, T. Saito, S.-i. Honda, S. Tahara-Hanaoka, A. Shibuya, LFA-1 decreases the antigen dose for T cell activation in vivo. Int. Immunol 20 (2008) 1119e1127. ´ Shea, J.A. Johnston, J.H. Kehrl, G. Koretzky, L.E. Samelson, Key [12] J.J. O molecules involved in receptor-mediated lymphocyte activation. Curr. Protoc. Immunol. (2001) Chapter 11: Unit 11.9A. [13] J.M. Green, V. Karpitskiy, S.L. Kimzey, A.S. Shaw, Coordinate regulation of T cell activation by CD2 and CD28. J. Immunol. 164 (2000) 3591e3595. [14] M.I. Lioudyno, J.A. Kozak, A. Penna, O. Safrina, S.L. Zhang, D. Sen, J. Roos, K.A. Stauderman, M.D. Cahalan, Orai1 and STIM1 move to the immunological synapse and are up-regulated during T cell activation. Proc. Natl. Acad. Sci. U. S. A. 105 (2008) 2011e2016. [15] S.A. Nicolaou, L. Neumeier, Y. Peng, D.C. Devor, L. Conforti, The Ca2þ activated K(þ) channel KCa3.1 compartmentalizes in the immunological synapse of human T lymphocytes. Am. J. Physiol. Cell Physiol. 292 (2007) C1431eC1439. [16] R.A. Maldonado, M.A. Soriano, C. Perdomo, K. Sigrist, D.J. Irvine, T. Decker, L.H. Glimcher, Control of T helper cell differentiation through cytokine receptor inclusion in the immunological synapse. J. Exp. Med. 4 (2009) 877e892. [17] R.L. Contento, B. Molon, C. Boularan, T. Pozzan, S. Manes, S. Marullo, A. Viola, CXCR4-CCR5: a couple modulating T cell functions. Proc. Natl. Acad. Sci. U. S. A. 105 (2008) 10101e10106.

444

J.L. Rodrı´guez-Ferna´ndez et al. / Microbes and Infection 12 (2010) 438e445

[18] C. Ballerini, P. Gourdain, V. Bachy, N. Blanchard, E. Levavasseur, S. Gre´goire, P. Fontes, P. Aucouturier, C. Hivroz, C. Carnaud, Functional implication of cellular prion protein in antigen-driven interactions between T cells and dendritic cells. J. Immunol. 176 (2006) 7254e7262. [19] M.L. Dustin, D.R. Colman, Neural and immunological synaptic relations. Science 298 (2002) 785e789. [20] A.A. Khan, C. Bose, L.S. Yam, M.J. Soloski, F. Rupp, Physiological regulation of the immunological synapse by agrin. Science 292 (2001) 1681e1686. [21] R. Tordjman, Y. Lepelletier, V. Lemarchandel, M. Cambot, P. Gaulard, O. Hermine, P.H. Rome´o, A neuronal receptor, neuropilin-1, is essential for the initiation of the primary immune response. Nat. Immunol. 3 (2002) 477e482. [22] S. Cemerski, J. Das, J. Locasale, P. Arnold, E. Giurisato, M.A. Markiewicz, D. Fremont, P.M. Allen, A.K. Chakraborty, A.S. Shaw, The stimulatory potency of T cell antigens is influenced by the formation of the immunological synapse. Immunity 26 (2007) 345e355. [23] T. Saito, T. Yokosuka, Immunological synapse and microclusters: the site for recognition and activation of T cells. Curr. Opin. Immunol. 18 (2006) 305e313. [24] M.L. Dustin, S.-Y. Tseng, R. Varma, G. Campi, T cell-dendritic cell immunological synapses. Curr. Opin. Immunol. 18 (2006) 512e516. [25] M.C. Miceli, M. Moran, C.D. Chung, V.P. Patel, T. Low, W. Zinnanti, Co-stimulation and counter-stimulation: lipid raft clustering controls TCR signaling and functional outcomes. Semin. Immunol. 13 (2001) 115e128. [26] O. Barreiro, H. De la Fuente, M. Mittelbrun, F. Sanchez-Madrid, Functional insights on the polarized redistribution of leukocytes integrins and their ligands during leukocyte migration and immune interactions. Immunol. Rev. 218 (2007) 147e164. [27] M. Vicente-Manzanares, F. Sanchez-Madrid, Role of the cytoskeleton during leukocyte responses. Nat. Rev. Immunol. 4 (2004) 1e14. [28] D.D. Billadeau, J.C. Nolz, T.S. Go´mez, Regulation of T-cell activation by the cytoskeleton. Nat. Rev. Immunol. 7 (2007) 131e143. [29] S.M. Eibert, K.H. Lee, R. Pipkorn, U. Sester, G.H. Wabnitz, T. Giese, S. C. Meuer, Y. Samstag, Cofilin peptide homologs interfere with immunological synapse formation and T cell activation. Proc. Natl. Acad. Sci. U. S. A. 101 (2004). [30] J. Suzuki, S. Yamasaki, J. Wu, G.A. Koretzky, T. Saito, The actin cloud induced by LFA-1-mediated outside-in signals lowers the threshold for T-cell activation. Blood 109 (2007) 168e175. [31] A.S. Shaw, FERMing up the synapse. Immunity 15 (2001) 683e686. [32] E.J. Allenspach, P. Cullinan, J. Tong, Q. Tang, A.G. Tesciuba, J.L. Cannon, S.M. Takahashi, R. Morgan, J.K. Burkhardt, A.I. Sperling, ERM-dependent movement of CD43 defines a novel protein complex distal to the immunological synapse. Immunity 15 (2001) 739e750. [33] A. Roumier, J.C. Olivo-Marin, M. Arpin, F. Michel, M. Martin, P. Mangeat, O. Acuto, A. Dautry-Varsat, A. Alcover, The membranemicrofilament linker ezrin is involved in the formation of the immunological synapse and in T cell activation. Immunity 15 (2001) 715e728. [34] P. Friedl, A.T. den Boer, M. Gunzer, Tuning immune responses: diversity and adaptation of the immunological synapse. Nat. Rev. Immunol. 5 (2005) 532e545. [35] W. Zhang, R.P. Trible, M. Zhu, S.K. Liu, C.J. McGlade, L.E. Samelson, Association of Grb2, Gads, and phospholipase C-g 1 with phosphorylated LAT tyrosine residues. Effect of LAT tyrosine mutations on T cell antigen receptor-mediated signaling. J. Biol. Chem. 275 (2000) 23355e23361. [36] H. Phee, R.T. Abraham, A. Weiss, Dynamic recruitment of PAK1 to the immunological synapse is mediated by PIX independently of SLP-76 and Vav1. Nat. Immunol. 6 (2005) 608e617. [37] J. Harriague, G. Bismuth, Imaging antigen-induced PI3K activation in T cells. Nat. Immunol. 3 (2002) 1090e1096. [38] P.S. Costello, P.J. Gallagher, D.A. Cantrell, Sustained and dynamic inositol lipid metabolism inside and outside the immunological synapse. Nat. Immunol. 3 (2002) 1082e1089. [39] S. Fabre, V. Lang, J. Harriague, A. Jobart, T.G. Unterman, A. Trautmann, G. Bismuth, Stable activation of phosphatidylinositol 3-kinase in the

[40]

[41]

[42]

[43]

[44]

[45]

[46]

[47]

[48]

[49]

[50]

[51]

[52]

[53]

[54] [55] [56] [57]

[58]

[59]

T cell immunological synapse stimulates Akt signaling to FoxO1 nuclear exclusion and cell growth control. J. Immunol. 174 (2005) 4161e4171. P.B. Shambharkar, M. Blonska, B.P. Pappu, H. Li, Y. You, H. Sakurai, B.G. Darnay, H. Hara, J. Penninger, X. Lin, Phosphorylation and ubiquitination of the IkappaB kinase complex by two distinct signaling pathways. EMBO J. 26 (2007) 1794e1805. M.M. Gorska, Q. Liang, Z. Karim, R. Alam, Uncoordinated 119 protein controls trafficking of Lck via the Rab11 endosome and is critical for immunological synapse formation. J. Immunol. 183 (2009) 1675e1684. B. Bonello, N. Blanchard, M.C. Montoya, E. Aguado, C. Langlet, H.-T. He, S. Nunez-Cruz, M. Malissen, F. Sanchez-Madrid, D. Olive, C. Hivroz, Y. Collette, Dynamic recruitment of the adaptor protein LAT: LAT exists in two distinct intracellular pools and controls its own recruitment. J. Cell. Sci. 117 (2004) 1009e1116. H. Wang, F.E. McCann, J.D. Gordan, X. Wu, M. Raab, T.H. Malik, D.M. Davis, C.E. Rudd, ADAP-SLP-76 binding differentially regulates supramolecular activation cluster (SMAC) formation relative to T cellAPC conjugation. J. Exp. Med. 200 (2004) 1063e1074. R. Xavier, S. Rabizadeh, K. Ishiguro, N. Andre, J.B. Ortiz, H. Wachtel, D.G. Morris, M. Lopez-Ilasaca, A.C. Shaw, W. Swat, B. Seed, Discs large (Dlg1) complexes in lymphocyte activation. J. Cell Biol. 166 (2004) 173e178. L.I. Ehrlich, P.J. Ebert, M.F. Krummel, A. Weiss, M.M. Davis, Dynamics of p56lck translocation to the T cell immunological synapse following agonist and antagonist stimulation. Immunity 17 (2002) 809e822. V. Das, B. Nal, A. Dujeancourt, M.I. Thoulouze, T. Galli, P. Roux, A. Dautry-Varsat, A. Alcover, Activation-induced polarized recycling targets T cell antigen receptors to the immunological synapse; involvement of SNARE complexes. Immunity 20 (2004) 577e588. G. Patino-Lopez, X. Dong, K. Ben-Aissa, K.M. Bernot, T. Itoh, M. Fukuda, M.J. Kruhlak, L.E. Samelson, S. Shaw, Rab35 and its GAP EPI64C in T cells regulate receptor recycling and immunological synapse formation. J. Biol. Chem. 283 (2008) 18323e18330. F. Finetti, S.R. Paccani, M.G. Riparbelli, E. Giacomello, G. Perinetti, G.J. Pazour, J.L. Rosenbaum, C.T. Baldari, Intraflagellar transport is required for polarized recycling of the TCR/CD3 complex to the immune synapse. Nat. Cell Biol. 11 (2009) 1332e1339. J. Combs, S.J. Kim, S. Tan, L.A. Ligon, E.L. Holzbaur, J. Kuhn, M. Poenie, Recruitment of dynein to the Jurkat immunological synapse. Proc. Natl. Acad. Sci. U. S. A. 103 (2006) 14883e14888. J.L. Round, T. Tomassian, M. Zhang, V. Patel, S.P. Schoenberger, M.C. Miceli, Dlgh1 coordinates actin polymerization, synaptic T cell receptor and lipid raft aggregation, and effector function in T cells. J. Exp. Med. 201 (2005) 419e430. J.L. Round, L.A. Humphries, T. Tomassian, P. Mittelstadt, M. Zhang, M. C. Miceli, Scaffold protein Dlgh1 coordinates alternative p38 kinase activation, directing T cell receptor signals toward NFAT but not NF-kappaB transcription factors. Nat. Immunol. 8 (2007) 154e161. A. Quintana, C. Schwindling, A.S. Wenning, U. Becherer, J. Rettig, E.C. Schwarz, M. Hoth, T cell activation requires mitochondrial translocation to the immunological synapse. Proc. Natl. Acad. Sci. U. S. A. 104 (2007) 14418e14423. S.J. Davis, P.A. Van der Merwe, The immunological synapse: required for T-cell receptor signaling or directing T-cell effector function? Curr. Biol. 11 (2001) R289eR291. P.A. Van der Merwe, S.J. Davis, The immunological synapsea multitasking system. Science 295 (2002) 1479e1480. P.A. Van der Merwe, Formation and function of the immunological synapse. Curr. Opin. Immunol. 1 (2002) 293e298. S. Cemerski, A. Shaw, Immune synapses in T-cell activation. Curr. Opin. Immunol. 18 (2006) 298e304. A. Grakoui, S.K. Bromley, C. Sumen, M.M. Davis, A.S. Shaw, P.M. Allen, M.L. Dustin, The immunological synapse: a molecular machine controlling T cell activation. Science 285 (1999) 221e227. K.-H. Lee, A.D. Holdorf, M.L. Dustin, A.C. Chan, P.M. Allen, A.S. Shaw, T Cell receptor signaling precedes immunological synapse formation. Science 295 (2002) 1539e1542. K.-H. Lee, A.R. Dinner, C. Tu, G. Campi, S. Raychaudhuri, R. Varma, T.M. Sims, W.R. Burack, H. Wu, J. Wang, O. Kanawaga, M. Markiewicz,

J.L. Rodrı´guez-Ferna´ndez et al. / Microbes and Infection 12 (2010) 438e445

[60]

[61] [62]

[63]

[64]

[65]

[66]

[67]

[68]

[69]

[70]

[71]

P.M. Allen, M.L. Dustin, A.K. Chakraborty, A.S. Shaw, The immunological synapse balances T cell receptor signaling and degradation. Science 302 (2003) 1218e1222. S. Cemerski, J. Das, E. Giurisato, M.A. Markiewicz, P.M. Allen, A.K. Chakraborty, A.S. Shaw, The balance between T cell receptor signaling and degradation at the centre of the immunological synapse is determined by antigen quality. Immunity 29 (2008) 414e422. M. Huse, E.J. Quann, M.M. Davis, Shouts, whispers and the kiss of death: directional secretion of T cells. Nat Immunol 9 (2008) 1105e1111. J.T. Chang, V.R. Palanivel, I. Kinjyo, F. Schambach, A.M. Intlekofer, A. Banerjee, S.A. Longworth, K.E. Vinup, P. Mrass, J. Oliaro, N. Killeen, J.S. Orange, S.M. Russell, W. Weninger, S.L. Reiner, Asymmetric T lymphocyte division in the initiation of adaptive immune responses. Science 315 (2007) 1687e1691. P. Revy, M. Sospedra, B. Barbour, A. Trautmann, Functional antigenindependent synapses formed between T cells and dendritic cells. Nat. Immunol. 2 (2001) 925e931. J. Boisvert, S. Edmondson, M.F. Krummel, Immunological synapse formation licences CD40-CD40L accumulation at T-APC contact sites. J. Immunol. 173 (2004) 3647e3652. T. Pentcheva-Hoang, J.G. Egen, K. Wojnoonski, J.P. Allison, B7-1 and B7-2 selectively recruit CTLA-4 and Cd28 to the immunological synapse. Immunity 21 (2004) 401e413. A.M. Keller, T.A. Groothuis, E.A. Veraar, M. Marsman, L. Maillette de Buy Wenniger, H. Janssen, J. Neefjes, J. Borst, Costimulatory ligand CD70 is delivered to the immunological synapse by shared intracellular trafficking with MHC class II molecules. Proc. Natl. Acad. Sci. U. S. A. 104 (2007) 5989e5994. S.Y. Eun, B.P. O’Connor, A.W. Wong, H.W. van Deventer, D.J. Taxman, W. Reed, P. Li, J.S. Blum, K.P. McKinnon, J.P. Ting, Rho activation and actin polarization are dependent on plexin-A1 in dendritic cells. J. Immunol. 177 (2006) 4271e4275. A.W. Wong, W.J. Brickey, D.J. Taxman, H.W. van Deventer, W. Reed, J.X. Gao, P. Zheng, Y. Liu, P. Li, J.S. Blum, K.P. McKinnon, J.P. Ting, CIITA-regulated plexin-A1 affects T-cell-dendritic cell interactions. Nat. Immunol. 4 (2003) 891e898. W.H. Luty, D. Rodeberg, J. Parness, Y. Vyas, Antiparallel segregation of notch components in the immunological synapse directs reciprocal signaling in allogeneic Th:DC conjugates. J. Immunol. 179 (2007) 819e829. A.B. Vogt, S. Spindeldreher, H. Kropshofer, Clustering of MHC-peptide complexes prior to their engagement in the immunological synapse: lipid raft and tetraspan microdomains. Immunol. Rev. 189 (2002) 136e151. M.M. Al-Alwan, G. Rowden, T.D.G. Lee, K.A. West, The dendritic cell cytoskeleton is critical for the formation of the immunological synapse. J. Immunol. 166 (2001) 1452e1456.

445

[72] L. Riol-Blanco, C. Delgado-Martı´n, N. Sa´nchez-Sa´nchez, L.M. AlonsoC, M.D. Gutie´rrez-Lo´pez, G. Martı´nez del Hoyo, J. Navarro, F. SanchezMadrid, C. Caba~nas, P. Sa´nchez-Mateos, J.L. Rodrı´guez-Ferna´ndez, Immunological synapse formation inhibits, via NFkB and FOXO1, the apoptosis of dendritic cells. Nat. Immunol. 10 (2009) 753e760. [73] M. Boes, J. Cerny, R. Massol, M. Op den Brouw, T. Kirchhausen, J. Chen, H.L. Ploegh, T-cell engagement of dendritic cells rapidly rearranges MHC class II transport. Nature 418 (2002) 983e988. [74] J.M. Vyas, Y.-M. Kim, K. Artavanis-Tsakones, J.C. Love, A.G. Van der Veen, H.L. Ploegh, Tubulation of class II MHC compartments ins microtubule dependent and involves multiple endolysosomal membrane proteins in primary dendritic cells. J. Immunol. 178 (2007) 7199e7210. [75] D.R. Fooksman, S.R. Shaikh, S. Boyle, M. Edidin, Phosphatidylinositol 4,5-bisphosphate concentration at the APC side of the immunological synapse is required for effector T cell function. J. Immunol. 182 (2009) 5179e5182. [76] P.J. Fisher, P.A. Bulur, S. Vuk-Pavlovic, F.G. Prendergast, A.B. Dietz, Dendritic cell microvilli: a novel membrane structure associated with the multifocal synapse and T-cell clustering. Blood 112 (2009) 5037e5045. [77] G. Bourna, S. Burns, A.J. Thrasher, Impaired T-cell priming in vivo resulting from dysfunction of WASp-deficient dendritic cells. Blood 110 (2007) 4278e4284. [78] J. Pulecio, E. Tagliani, A. Scholer, F. Prete, L. Fetler, O.R. Burrone, I.F. Benvenut, Expression of WiskotteAldrich syndrome protein in dendritic cells regulates synapse formation and activation of naive CD8 þ T cells. J. Immunol. 181 (2008) 1135e1142. [79] O. Bloom, J.J. Unternaehrer, A. Jiang, J.S. Shin, L. Delamarre, P. Allen, I. Mellman, Spinophilin participates in information transfer at immunological synapses. J. Cell Biol. 181 (2008) 203e211. [80] A. Reinhold, S. Reimann, D. Reinhold, B. Schraven, M. Togni, Expression of SKAP-HOM in DCs is required for an optimal immune response in vivo. J. Leukoc. Biol. 86 (2009) 61e71. [81] J. Jacobelli, P.G. Andres, J. Boisvert, M.F. Krummel, New views of the immunological synapse: variations in assembly and function. Curr. Opin. Immunol. 16 (2004) 345e352. [82] R.A. Maldonado, D.J. Irvine, R. Schreiber, L.H. Glimcher, A role for the immunological synapse in lineage commitment of CD4 lymphocytes. Nature 431 (2004) 527e532. [83] S.J. Turley, K. Inaba, W.S. Garrette, M. Ebersold, J. Unternaehrer, R.M. Steinman, I. Mellman, Transport of peptide-MHC class II complexes in developing dendritic cells. Science 288 (2000) 522e527. [84] C. Escribano, C. Delgado-Martı´n, J. Rodriguez-Fernandez, CCR7dependent stimulation of survival in dendritic cells involves inhibition of GSK3b. J. Immunol. 183 (2009) 6282e6295.