- Email: [email protected]
Initiation of TCR signalling revisited
Opinion
TRENDS in Immunology
Vol.24 No.8 August 2003
425
Initiation of TCR signalling revisited Alain Trautmann and Clotilde Randriamampita Institut COCHIN, INSERM – CNRS, 22 rue Me´chain, 75014 Paris, France
T-cell receptor (TCR) multimerization has long been considered as an absolute prerequisite for T-cell activation. This view has been challenged by several recent reports showing that monomeric peptide–MHC (pMHC) complexes in solution could be stimulatory for T cells and that even a single pMHC at the immunological synapse was sufficient to trigger a T-cell Ca21 response. Two incompatible models have been proposed to explain these new findings: the pseudodimer and the heterodimerization models. We consider that the heterodimerization model applied to adhesion-primed T cells provides an explanation for a larger set of experimental observations. The mechanisms by which T-cell receptor (TCR) signalling is initiated on antigen recognition still remain a mystery. A wide variety of models have been proposed and discussed in detail [1–5]. This Opinion will focus on the issue of valency of the TCR ligand. For years, immunologists have agreed on the idea that TCR signalling could only be initiated by multimerized TCR ligands and not by monovalent ones. This view was based on the fact that antibody-induced multimerization of TCR–CD3 complexes was indeed stimulatory and that in most reports, Ca2þ responses could only be triggered in T cells by multimerized peptide–MHC (pMHC) complexes and not by monomeric pMHC. This was true for CD4þ [6–8] as well as for CD8þ T cells [9]. However, some data did not fit with the multimerization model, in particular the observation that a single pMHC per target cell was sufficient to trigger cytolytic activity in CD8þ T cells [10]. Admittedly, the estimate of one ligand per target cell in this paper was a statistical one and the actual ligand number could easily be two or three on the relevant target cells. However, the probability that these two or three pMHC would be located next to each other at the cell surface was low. For the same reasons, the fact that a few pMHC at the immunological synapse between T cells and dendritic cells (DCs) were sufficient to trigger a Ca2þ response in the T cell argues against the multimerization model [11]. T-cell Ca21 responses can be elicited by monomeric pMHC complexes The multimerization model was further challenged by the observation that monomeric pMHC in solution could elicit Ca2þ responses in CD8þ T cells, provided CD8 was not truncated, that is, was able to associate with Lck [12]. The interpretation of this finding has been hotly debated since (see later). Consistent with this initial finding, Irvine et al. recently showed that a single pMHC could trigger a T-cell Ca2þ response [13]. This demonstration resulted from an
impressive technological achievement, in which highresolution fluorescence microscopy enabled visualization at immunological synapses of individual MHC class II complexes labelled with phycoerythrin. Care was taken to ascertain that single pMHC were not confused with small aggregates of pMHC. A key point in this paper is the assumption that self-pMHC displayed by antigen-presenting cells (APCs) might help recognizing foreign pMHC. The fundamental role of the coreceptor was underlined in the different sets of data because the triggering of Ca2þ responses by soluble monomeric pMHC or by single pMHC at the immunological synapse strictly depended on the presence of either CD8 [12] or CD4 [13]. Heterodimerization and pseudodimer models Thus, two distinct sets of findings led to the identical conclusion that monomeric pMHC could elicit Ca2þ responses in T cells. Two main models, the heterodimerization and the pseudodimer models have been proposed to interpret these findings (schemes illustrating them are shown in Ref. [14]). A third model, called kineticsegregation, compatible with the possibility that monomeric pMHC are activatory, will be discussed later. The heterodimerization model was initially proposed for both CD4þ and CD8þ T cells [15 –17] but major experimental support in favour of this model was obtained with CD8þ T cells [12]. This model postulates that TCR signalling can be initiated in CD8þ T cells by the heterodimerization of the TCR and the coreceptor CD8 being associated with Lck, enabling Lck to phosphorylate CD3 chains and to initiate TCR signalling. Such a model fits well with previous observations that a potent stimulation results from the cross-linking of CD3 and the coreceptor, CD8 or CD4 (see Ref. [18]). An intriguing feature of this trimeric CD8–MHC–(TCR–CD3) interaction is that the TCR–MHC class I interaction is markedly enhanced by CD8 [19,20], despite the fact that the affinity of soluble MHC class I for CD8 is low, maybe .100 times lower than that of TCR–MHC class I [21]. This presumably reveals that the cohesion of the complex is considerably reinforced by additional direct or indirect interactions, presumably between CD3 and CD8.
The heterodimerization model … postulates that TCR signalling can be initiated in CD81 T cells by [TCR] heterodimerization … and … CD8 being associated with Lck …
Corresponding author: Alain Trautmann ([email protected]). http://treimm.trends.com 1471-4906/03/$ - see front matter q 2003 Elsevier Ltd. All rights reserved. doi:10.1016/S1471-4906(03)00182-0
426
Opinion
TRENDS in Immunology
The pseudodimer model was proposed to explain data obtained with CD4 T cells. It is postulated that a foreign antigen-engaged TCR can be crosslinked by one CD4 molecule to a different, self-ligand-binding TCR [13]. This model is consistent with the ability of MHC class II molecules to form dimers (or ‘dimers of dimers’, by taking into account the fact that each MHC class II molecule is a ab heterodimer). The existence of such ‘dimers of dimers’ has been proposed, based on crystallographic data [22,23]. In addition, a substantial loss of stimulatory capacity has been measured for APCs expressing mutated MHC class II molecules, presenting an altered dimerization interface [24]. The pseudodimer model consituted a clever alternative to the heterodimerization model, which was discarded for two reasons, one theoretical and one experimental. The theoretical argument was that structural evidence shows that CD4 engages MHC class II molecules at an angle that appears to preclude simultaneous interaction with the TCR bound to that MHC molecule [25]. However, as recently discussed [14], if one considers an interaction between the coreceptor and CD3, possibly CD3d [26], rather than with the TCR, the angle of the coreceptor – MHC interaction remains perfectly compatible with the heterodimerization model. The experimental reason is that the heterodimerization model predicts that monomeric pMHC in solution should constitute an efficient stimulus. In view of their own previous results [7], and of the ‘peptidetransfer objection’ discussed later, Irvine et al. considered that this prediction was not fulfilled.
The pseudodimer model … postulate[s] that a foreign antigenengaged TCR can be crosslinked by one CD4 molecule to a different, selfligand-binding TCR. ‘Peptide-transfer objection’ is not valid The heterodimerization model has recently been criticized by two groups who established that under some conditions, monomeric pMHC can indeed trigger various responses by CD8 T cells, such as CD69 expression or TCR downregulation [27,28]. However, this phenomenon was apparently a result of the transfer of peptide from soluble pMHC to MHC molecules at the T-cell surface, followed by T-cell– T-cell interactions. As a result, the effective stimulus was not soluble pMHC but the T-cell MHC molecules loaded by peptide transfer. Indeed, monomeric pMHC was unable to activate T cells that were deficient in class I MHC. With class I positive T cells, T-cell activation, as a result of peptide transfer, could be readily mimicked by adding free peptide. The ‘peptide-transfer objection’ appeared to definitely rule out that monomeric pMHC in solution could trigger TCR signalling, and therefore invalidates the heterodimerization model. http://treimm.trends.com
Vol.24 No.8 August 2003
… monomeric pMHC was unable to activate T cells that were deficient in class I MHC. However, it should be stressed that the peptide-transfer hypothesis had been tested after T cells and exogenous pMHC had been in contact for several hours. When the assay was performed in the seconds or minutes following the addition of pMHC, there was absolutely no sign of peptide transfer. Moreover, these short-term stimulations by monomeric pMHC did lead to small but clearly detectable responses that could not be mimicked by mere peptide addition [28]. This was consistent with a previous observation that monomeric pMHC could bind to a significant fraction of the surface TCRs, suggesting that monomeric binding might trigger partial signals not sufficient to initiate activation processes unless coupled with other stimuli [8]. The initial report describing Ca2þ responses induced by monomeric pMHC [12] was based on a short-term assay that should not have been affected by peptide transfer. Most importantly, the control performed with soluble peptide alone (which has been shown to mimic peptide transfer [28]) was performed, and was clearly negative [12]. This argues strongly against the peptide-transfer hypothesis. Finally, the assay was performed with T cells adhering on the glass coverslip to permit Ca2þ imaging. The cells were therefore immobile and could not touch each other. Thus, the ‘peptide-transfer objection’ seems considerably weakened, and the stimulatory capacity of monomeric pMHC cannot be disregarded. Adhesion-induced T-cell priming Two recent papers provide new data that help to clarify the situation. They both show that adherent T cells do respond to monomeric pMHC complexes although the same T cells, once in suspension, fail to respond [29,30]. This effect was observed when T cells adhered to different substrates, such as fibronectin [30], glass or immobilized antibodies directed against MHC class I, CD11 or CD18, and also on adhesion to DCs [29]. This important phenomenon, which we call ‘adhesion-induced T-cell priming’, fits well with the following facts: the initial report on the stimulatory capacity of monomeric pMHC complexes was based on data obtained with adherent CD8þ T cells [12]; a single pMHC is stimulatory when a cytotoxic T cell adheres to its target [10]; at an immunological synapse, a single pMHC could trigger a T-cell Ca2þ response [13]; when T cells are in suspension they are unresponsive to monomeric pMHC [7– 9].
… adherent T cells do respond to monomeric pMHC complexes although the same T cells, once in suspension, fail to respond.
Opinion
TRENDS in Immunology
Could the response of adherent cells be as a result of monomeric ligand adsorbed on the glass coverslip? Such a phenomenon could indeed affect long-term responses. However, we consider it highly improbable that a significant ligand immobilization can occur on the coverslip, at the lower face of adherent cells, in the few seconds separating ligand addition and the beginning of the Ca2þ response. The mechanism of ‘adhesion-induced T-cell priming’ is likely to be related to several phenomena triggered by T-cell adhesion. Thus, T-cell adhesion causes a marked increase in the Ca2þ content of intracellular stores; it also causes a significant increase in the amount of phosphatidylinositol 4,5-bisphosphate in the T-cell plasma membrane [29]. Both phenomena should converge to amplify the subsequent TCR- and inositol 1,4,5-triphosphatedependent Ca2þ influx. T-cell adhesion also leads to the phosphorylation of three tyrosine kinases, PYK-2 (prolinerich tyrosine kinase-2), Lck and Fyn and of a cytoskeletalassociated protein, paxillin [30]. Adhesion through integrins also activates the MAPK (mitogen-activated protein kinase) pathway and it has recently been reported that MAPK has a positive role in effective TCR signalling [31]. Of all these events, which are the ones that are key for ‘adhesion-induced T-cell priming’ remains to be determined. Whatever the explanation, the detection threshold for TCR signalling is significantly lowered in a T cell primed by adhesion, so that they become responsive to monomeric pMHC. T-cell priming by self-peptides In addition to ‘adhesion-induced T-cell priming’, a distinct and important type of T-cell priming should be considered, which could be called ‘T-cell priming by self-peptides’. Stefanova et al. have recently demonstrated that in vivo, antigen-independent T-cell interactions with APCs transiently facilitate the antigen reactivity of mature T cells [32]. As a result, freshly isolated CD4þ T cells are significantly more responsive to antigenic stimulation than T cells that have been kept in culture, even for as short a time as 30 min. This difference is only visible under conditions in which self-MHC class II recognition in vivo can take place. This is a priming effect, that is, the interaction of a T cell with an APC bearing self-peptides would be important just before the antigen-specific stimulus is delivered. Which model better explains all available data? A positive aspect of the pseudodimer model is that it attributes a possible function to ‘dimers of dimers’ of MHC class II molecules, even though there is no direct evidence in favour of a hetero-pentameric structure including two TCR – MHC couples bridged by one CD4 molecule. However, this model fails to predict the activation of T cells by soluble monomeric pMHC and the differential behaviour of adherent and suspended T cells. The heterodimerization model applied to adhesion-primed T cells is consistent with all the results recently published on the question of the valency of efficient TCR ligands. It postulates the existence of a simpler heterotrimeric structure (MHC, TCR –CD3, coreceptor) likely to exist [19,20], which is only efficient in http://treimm.trends.com
Vol.24 No.8 August 2003
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T cells that have been primed by adhesion. This model should be further tested by examining if adherent CD8þ T cells lacking the relevant MHC class I molecule do respond to soluble monomeric pMHC class I, and if adherent CD4þ T cells can be stimulated by soluble monomeric pMHC class II. The latter experiment would be important in view of the fact that the heterodimerization model was discussed in depth for CD8 and not for CD4 T cells, whereas the reverse was true for the pseudodimer model. Finally, it should be stressed that in both models, Lck recruitment is expected to be due to its association with a coreceptor, CD4 or CD8, in accordance with the coreceptor dependence of T-cell activation generally observed. However, in a minority of cases, T-cell activation is not affected by antibodies preventing the MHC – coreceptor interaction, that is, the activation of these T cells is coreceptorindependent. In those specific cases, Lck recruitement and/or activation must be due to an undetermined phenomenon, predicted neither by the heterodimerization nor by the pseudodimer model but which is compatible with a third model, the kinetic – segregation model [5,33]. This model postulates that TCR signalling can be initiated in regions of close APC – T-cell membrane apposition, leaving a cleft too narrow to accommodate the largesized phosphatase CD45. CD45 exclusion would result in an enhanced kinase:phosphatase ratio that might lead to an increased CD3 tyrosine phosphorylation if a concomitant increase in local concentration of TCR– CD3 takes place, and this might be the case on specific pMHC binding. Several features of this model still await an experimental basis. This model makes no prediction on the minimum number of TCRs that need to bind specific pMHC but it is compatible with a single pMHC –TCR interaction being activatory. However, this model does not readily explain how soluble monomeric pMHC can be stimulatory. Compared to the other models mentioned so far, a weak point is that it does not take into account the major importance of the coreceptors in TCR signalling for some T cells. But this becomes an advantage when considering the case of coreceptor-independent T-cell signalling. In conclusion, the phenomenon of adhesion-induced T-cell priming sheds new light on the question of the minimum valency of efficient TCR ligands. The previous conclusion that TCR signalling could only be triggered by multimerized ligands remains true but only under nonphysiological conditions when T cells are in suspension, which is not the normal condition for antigen recognition in vivo. It would make sense that the TCR signalling machinery can exist under two states, a low sensitivity state when a travelling T cell is in the lymph or blood flow and a high sensitivity state when the T cell has just become adherent to various cell types of the spleen or lymph nodes, including DCs. Both ‘T-cell priming by self-peptides’ and ‘adhesion-induced T-cell priming’ could converge for the induction of this high sensitivity state for TCR signalling. Acknowledgements This work has been supported by grants from Centre National de la Recherche Scientifique, Institut National de la Sante´ et de la Recherche Me´dicale, and Ligue Nationale contre le Cancer.
428
Opinion
TRENDS in Immunology
Vol.24 No.8 August 2003
References 1 Germain, R.N. (1997) T cell signaling: the importance of receptor clustering. Curr. Biol. 7, R640 – R644 2 McKeithan, T.W. (1995) Kinetic proofreading in T-cell receptor signal transduction. Proc. Natl. Acad. Sci. U. S. A. 92, 5042 – 5046 3 Lanzavecchia, A. et al. (1999) From TCR engagement to T cell activation: a kinetic view of T cell behavior. Cell 96, 1 – 4 4 Germain, R.N. and Stefanova, I. (1999) The dynamics of T cell receptor signaling: complex orchestration and the key roles of tempo and cooperation. Annu. Rev. Immunol. 17, 467 – 522 5 van der Merwe, P.A. (2001) The TCR triggering puzzle. Immunity 14, 665 – 668 6 Symer, D.E. et al. (1992) Inhibition or activation of human T cell receptor transfectants is controlled by defined, soluble antigen arrays. J. Exp. Med. 176, 1421– 1430 7 Boniface, J.J. et al. (1998) Initiation of signal transduction through the T cell receptor requires the multivalent engagement of peptide/MHC ligands. Immunity 9, 459– 466 8 Cochran, J.R. et al. (2000) The relationship of MHC-peptide binding and T cell activation probed using chemically defined MHC class II oligomers. Immunity 12, 241 – 250 9 Daniels, M.A. and Jameson, S.C. (2000) Critical role for CD8 in T cell receptor binding and activation by peptide/major histocompatibility complex multimers. J. Exp. Med. 191, 335– 346 10 Sykulev, Y. et al. (1996) Evidence that a single peptide-MHC complex on a target cell can elicit a cytolytic T cell response. Immunity 4, 565 – 571 11 Delon, J. et al. (1998) Antigen-dependent and antigen-independent Ca2þ responses triggered in T cells by dendritic cells compared to B cells. J. Exp. Med. 188, 1473– 1484 12 Delon, J. et al. (1998) CD8 expression allows T cell signaling by monomeric peptide– MHC complexes. Immunity 9, 467 – 473 13 Irvine, D.J. et al. (2002) Direct observation of ligand recognition by T cells. Nature 419, 845 – 849 14 van der Merwe, P.A. (2002) Do T cell receptors do it alone? Nat. Immunol. 3, 1122 – 1123 15 Salter, R.D. et al. (1990) A binding site for the T-cell co-receptor CD8 on the a3 domain of HLA-A2. Nature 345, 41 – 46 16 Jameson, S.C. and Bevan, M.J. (1995) T cell receptor antagonists and partial agonists. Immunity 2, 1 – 11 17 Madrenas, J. and Germain, R.N. (1996) Variant TCR ligands: new insights into the molecular basis of antigen- dependent signal transduction and T-cell activation. Semin. Immunol. 8, 83 – 101 18 Anderson, P. et al. (1987) Cross-linking of T3 (CD3) with T4 (CD4)
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21 22 23 24
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26 27
28
29 30
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enhances the proliferation of resting T lymphocytes. J. Immunol. 139, 678– 682 Luescher, I.F. et al. (1995) CD8 modulation of T-cell antigen receptorligand interactions on living cytotoxic T lymphocytes. Nature 373, 353– 356 Denkberg, G. et al. (2001) Critical role for CD8 in binding of MHC tetramers to TCR: CD8 antibodies block specific binding of human tumor-specific MHC-peptide tetramers to TCR. J. Immunol. 167, 270– 276 Davis, S.J. et al. (2003) The nature of molecular recognition by T cells. Nat. Immunol. 4, 217 – 224 Brown, J.H. et al. (1993) Three-dimensional structure of human class II histocompatibility antigen HLA-DR1. Nature 364, 33 – 39 Fremont, D.H. et al. (1996) Structures of an MHC class II molecule with covalently bound single peptides. Science 272, 1001 – 1004 Lindstedt, R. et al. (2001) Amino acid substitutions in the putative MHC class II ‘dimer of dimers’ interface inhibit CD4þ T cell activation. J. Immunol. 166, 800– 808 Wang, J.H. et al. (2001) Crystal structure of the human CD4 N-terminal two-domain fragment complexed to a class II MHC molecule. Proc. Natl. Acad. Sci. U. S. A. 98, 10799 – 10804 Doucey, M.A. et al. (2003) CD3d establishes a functional link between the T cell receptor and CD8. J. Biol. Chem. 278, 3257 – 3264 Ge, Q. et al. (2002) Soluble peptide-MHC monomers cause activation of CD8þ T cells through transfer of the peptide to T cell MHC molecules. Proc. Natl. Acad. Sci. U. S. A. 99, 13729 – 13734 Schott, E. et al. (2002) Class I negative CD8 T cells reveal the confounding role of peptide-transfer onto CD8 T cells stimulated with soluble H2-Kb molecules. Proc. Natl. Acad. Sci. U. S. A. 99, 13735 – 13740 Randriamampita, C. et al. (2003) T cell adhesion lowers the threshold for antigen detection. Eur. J. Immunol. 33, 1215– 1223 Doucey, M.A. et al. Convergence of b1/b3 integrin- and TCR/CD8mediated signals promotes activation of CTL mediated cytotoxicity. J. Biol. Chem. 10.1074/jbc.302709200 (http://intl.jbc.org/) Stefanova, I. et al. (2003) TCR ligand discrimination is enforced by competing ERK positive and SHP-1 negative feedback pathways. Nat. Immunol. 4, 248 – 254 Stefanova, I.I. et al. (2002) Self-recognition promotes the foreign antigen sensitivity of naı¨ve T lymphocytes. Nature 420, 429– 434 Davis, S.J. and van der Merwe, P.A. (1996) The structure and ligand interactions of CD2: implications for T-cell function. Immunol. Today 17, 177 – 187
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TRENDS in Immunology
Vol.24 No.8 August 2003
425
Initiation of TCR signalling revisited Alain Trautmann and Clotilde Randriamampita Institut COCHIN, INSERM – CNRS, 22 rue Me´chain, 75014 Paris, France
T-cell receptor (TCR) multimerization has long been considered as an absolute prerequisite for T-cell activation. This view has been challenged by several recent reports showing that monomeric peptide–MHC (pMHC) complexes in solution could be stimulatory for T cells and that even a single pMHC at the immunological synapse was sufficient to trigger a T-cell Ca21 response. Two incompatible models have been proposed to explain these new findings: the pseudodimer and the heterodimerization models. We consider that the heterodimerization model applied to adhesion-primed T cells provides an explanation for a larger set of experimental observations. The mechanisms by which T-cell receptor (TCR) signalling is initiated on antigen recognition still remain a mystery. A wide variety of models have been proposed and discussed in detail [1–5]. This Opinion will focus on the issue of valency of the TCR ligand. For years, immunologists have agreed on the idea that TCR signalling could only be initiated by multimerized TCR ligands and not by monovalent ones. This view was based on the fact that antibody-induced multimerization of TCR–CD3 complexes was indeed stimulatory and that in most reports, Ca2þ responses could only be triggered in T cells by multimerized peptide–MHC (pMHC) complexes and not by monomeric pMHC. This was true for CD4þ [6–8] as well as for CD8þ T cells [9]. However, some data did not fit with the multimerization model, in particular the observation that a single pMHC per target cell was sufficient to trigger cytolytic activity in CD8þ T cells [10]. Admittedly, the estimate of one ligand per target cell in this paper was a statistical one and the actual ligand number could easily be two or three on the relevant target cells. However, the probability that these two or three pMHC would be located next to each other at the cell surface was low. For the same reasons, the fact that a few pMHC at the immunological synapse between T cells and dendritic cells (DCs) were sufficient to trigger a Ca2þ response in the T cell argues against the multimerization model [11]. T-cell Ca21 responses can be elicited by monomeric pMHC complexes The multimerization model was further challenged by the observation that monomeric pMHC in solution could elicit Ca2þ responses in CD8þ T cells, provided CD8 was not truncated, that is, was able to associate with Lck [12]. The interpretation of this finding has been hotly debated since (see later). Consistent with this initial finding, Irvine et al. recently showed that a single pMHC could trigger a T-cell Ca2þ response [13]. This demonstration resulted from an
impressive technological achievement, in which highresolution fluorescence microscopy enabled visualization at immunological synapses of individual MHC class II complexes labelled with phycoerythrin. Care was taken to ascertain that single pMHC were not confused with small aggregates of pMHC. A key point in this paper is the assumption that self-pMHC displayed by antigen-presenting cells (APCs) might help recognizing foreign pMHC. The fundamental role of the coreceptor was underlined in the different sets of data because the triggering of Ca2þ responses by soluble monomeric pMHC or by single pMHC at the immunological synapse strictly depended on the presence of either CD8 [12] or CD4 [13]. Heterodimerization and pseudodimer models Thus, two distinct sets of findings led to the identical conclusion that monomeric pMHC could elicit Ca2þ responses in T cells. Two main models, the heterodimerization and the pseudodimer models have been proposed to interpret these findings (schemes illustrating them are shown in Ref. [14]). A third model, called kineticsegregation, compatible with the possibility that monomeric pMHC are activatory, will be discussed later. The heterodimerization model was initially proposed for both CD4þ and CD8þ T cells [15 –17] but major experimental support in favour of this model was obtained with CD8þ T cells [12]. This model postulates that TCR signalling can be initiated in CD8þ T cells by the heterodimerization of the TCR and the coreceptor CD8 being associated with Lck, enabling Lck to phosphorylate CD3 chains and to initiate TCR signalling. Such a model fits well with previous observations that a potent stimulation results from the cross-linking of CD3 and the coreceptor, CD8 or CD4 (see Ref. [18]). An intriguing feature of this trimeric CD8–MHC–(TCR–CD3) interaction is that the TCR–MHC class I interaction is markedly enhanced by CD8 [19,20], despite the fact that the affinity of soluble MHC class I for CD8 is low, maybe .100 times lower than that of TCR–MHC class I [21]. This presumably reveals that the cohesion of the complex is considerably reinforced by additional direct or indirect interactions, presumably between CD3 and CD8.
The heterodimerization model … postulates that TCR signalling can be initiated in CD81 T cells by [TCR] heterodimerization … and … CD8 being associated with Lck …
Corresponding author: Alain Trautmann ([email protected]). http://treimm.trends.com 1471-4906/03/$ - see front matter q 2003 Elsevier Ltd. All rights reserved. doi:10.1016/S1471-4906(03)00182-0
426
Opinion
TRENDS in Immunology
The pseudodimer model was proposed to explain data obtained with CD4 T cells. It is postulated that a foreign antigen-engaged TCR can be crosslinked by one CD4 molecule to a different, self-ligand-binding TCR [13]. This model is consistent with the ability of MHC class II molecules to form dimers (or ‘dimers of dimers’, by taking into account the fact that each MHC class II molecule is a ab heterodimer). The existence of such ‘dimers of dimers’ has been proposed, based on crystallographic data [22,23]. In addition, a substantial loss of stimulatory capacity has been measured for APCs expressing mutated MHC class II molecules, presenting an altered dimerization interface [24]. The pseudodimer model consituted a clever alternative to the heterodimerization model, which was discarded for two reasons, one theoretical and one experimental. The theoretical argument was that structural evidence shows that CD4 engages MHC class II molecules at an angle that appears to preclude simultaneous interaction with the TCR bound to that MHC molecule [25]. However, as recently discussed [14], if one considers an interaction between the coreceptor and CD3, possibly CD3d [26], rather than with the TCR, the angle of the coreceptor – MHC interaction remains perfectly compatible with the heterodimerization model. The experimental reason is that the heterodimerization model predicts that monomeric pMHC in solution should constitute an efficient stimulus. In view of their own previous results [7], and of the ‘peptidetransfer objection’ discussed later, Irvine et al. considered that this prediction was not fulfilled.
The pseudodimer model … postulate[s] that a foreign antigenengaged TCR can be crosslinked by one CD4 molecule to a different, selfligand-binding TCR. ‘Peptide-transfer objection’ is not valid The heterodimerization model has recently been criticized by two groups who established that under some conditions, monomeric pMHC can indeed trigger various responses by CD8 T cells, such as CD69 expression or TCR downregulation [27,28]. However, this phenomenon was apparently a result of the transfer of peptide from soluble pMHC to MHC molecules at the T-cell surface, followed by T-cell– T-cell interactions. As a result, the effective stimulus was not soluble pMHC but the T-cell MHC molecules loaded by peptide transfer. Indeed, monomeric pMHC was unable to activate T cells that were deficient in class I MHC. With class I positive T cells, T-cell activation, as a result of peptide transfer, could be readily mimicked by adding free peptide. The ‘peptide-transfer objection’ appeared to definitely rule out that monomeric pMHC in solution could trigger TCR signalling, and therefore invalidates the heterodimerization model. http://treimm.trends.com
Vol.24 No.8 August 2003
… monomeric pMHC was unable to activate T cells that were deficient in class I MHC. However, it should be stressed that the peptide-transfer hypothesis had been tested after T cells and exogenous pMHC had been in contact for several hours. When the assay was performed in the seconds or minutes following the addition of pMHC, there was absolutely no sign of peptide transfer. Moreover, these short-term stimulations by monomeric pMHC did lead to small but clearly detectable responses that could not be mimicked by mere peptide addition [28]. This was consistent with a previous observation that monomeric pMHC could bind to a significant fraction of the surface TCRs, suggesting that monomeric binding might trigger partial signals not sufficient to initiate activation processes unless coupled with other stimuli [8]. The initial report describing Ca2þ responses induced by monomeric pMHC [12] was based on a short-term assay that should not have been affected by peptide transfer. Most importantly, the control performed with soluble peptide alone (which has been shown to mimic peptide transfer [28]) was performed, and was clearly negative [12]. This argues strongly against the peptide-transfer hypothesis. Finally, the assay was performed with T cells adhering on the glass coverslip to permit Ca2þ imaging. The cells were therefore immobile and could not touch each other. Thus, the ‘peptide-transfer objection’ seems considerably weakened, and the stimulatory capacity of monomeric pMHC cannot be disregarded. Adhesion-induced T-cell priming Two recent papers provide new data that help to clarify the situation. They both show that adherent T cells do respond to monomeric pMHC complexes although the same T cells, once in suspension, fail to respond [29,30]. This effect was observed when T cells adhered to different substrates, such as fibronectin [30], glass or immobilized antibodies directed against MHC class I, CD11 or CD18, and also on adhesion to DCs [29]. This important phenomenon, which we call ‘adhesion-induced T-cell priming’, fits well with the following facts: the initial report on the stimulatory capacity of monomeric pMHC complexes was based on data obtained with adherent CD8þ T cells [12]; a single pMHC is stimulatory when a cytotoxic T cell adheres to its target [10]; at an immunological synapse, a single pMHC could trigger a T-cell Ca2þ response [13]; when T cells are in suspension they are unresponsive to monomeric pMHC [7– 9].
… adherent T cells do respond to monomeric pMHC complexes although the same T cells, once in suspension, fail to respond.
Opinion
TRENDS in Immunology
Could the response of adherent cells be as a result of monomeric ligand adsorbed on the glass coverslip? Such a phenomenon could indeed affect long-term responses. However, we consider it highly improbable that a significant ligand immobilization can occur on the coverslip, at the lower face of adherent cells, in the few seconds separating ligand addition and the beginning of the Ca2þ response. The mechanism of ‘adhesion-induced T-cell priming’ is likely to be related to several phenomena triggered by T-cell adhesion. Thus, T-cell adhesion causes a marked increase in the Ca2þ content of intracellular stores; it also causes a significant increase in the amount of phosphatidylinositol 4,5-bisphosphate in the T-cell plasma membrane [29]. Both phenomena should converge to amplify the subsequent TCR- and inositol 1,4,5-triphosphatedependent Ca2þ influx. T-cell adhesion also leads to the phosphorylation of three tyrosine kinases, PYK-2 (prolinerich tyrosine kinase-2), Lck and Fyn and of a cytoskeletalassociated protein, paxillin [30]. Adhesion through integrins also activates the MAPK (mitogen-activated protein kinase) pathway and it has recently been reported that MAPK has a positive role in effective TCR signalling [31]. Of all these events, which are the ones that are key for ‘adhesion-induced T-cell priming’ remains to be determined. Whatever the explanation, the detection threshold for TCR signalling is significantly lowered in a T cell primed by adhesion, so that they become responsive to monomeric pMHC. T-cell priming by self-peptides In addition to ‘adhesion-induced T-cell priming’, a distinct and important type of T-cell priming should be considered, which could be called ‘T-cell priming by self-peptides’. Stefanova et al. have recently demonstrated that in vivo, antigen-independent T-cell interactions with APCs transiently facilitate the antigen reactivity of mature T cells [32]. As a result, freshly isolated CD4þ T cells are significantly more responsive to antigenic stimulation than T cells that have been kept in culture, even for as short a time as 30 min. This difference is only visible under conditions in which self-MHC class II recognition in vivo can take place. This is a priming effect, that is, the interaction of a T cell with an APC bearing self-peptides would be important just before the antigen-specific stimulus is delivered. Which model better explains all available data? A positive aspect of the pseudodimer model is that it attributes a possible function to ‘dimers of dimers’ of MHC class II molecules, even though there is no direct evidence in favour of a hetero-pentameric structure including two TCR – MHC couples bridged by one CD4 molecule. However, this model fails to predict the activation of T cells by soluble monomeric pMHC and the differential behaviour of adherent and suspended T cells. The heterodimerization model applied to adhesion-primed T cells is consistent with all the results recently published on the question of the valency of efficient TCR ligands. It postulates the existence of a simpler heterotrimeric structure (MHC, TCR –CD3, coreceptor) likely to exist [19,20], which is only efficient in http://treimm.trends.com
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T cells that have been primed by adhesion. This model should be further tested by examining if adherent CD8þ T cells lacking the relevant MHC class I molecule do respond to soluble monomeric pMHC class I, and if adherent CD4þ T cells can be stimulated by soluble monomeric pMHC class II. The latter experiment would be important in view of the fact that the heterodimerization model was discussed in depth for CD8 and not for CD4 T cells, whereas the reverse was true for the pseudodimer model. Finally, it should be stressed that in both models, Lck recruitment is expected to be due to its association with a coreceptor, CD4 or CD8, in accordance with the coreceptor dependence of T-cell activation generally observed. However, in a minority of cases, T-cell activation is not affected by antibodies preventing the MHC – coreceptor interaction, that is, the activation of these T cells is coreceptorindependent. In those specific cases, Lck recruitement and/or activation must be due to an undetermined phenomenon, predicted neither by the heterodimerization nor by the pseudodimer model but which is compatible with a third model, the kinetic – segregation model [5,33]. This model postulates that TCR signalling can be initiated in regions of close APC – T-cell membrane apposition, leaving a cleft too narrow to accommodate the largesized phosphatase CD45. CD45 exclusion would result in an enhanced kinase:phosphatase ratio that might lead to an increased CD3 tyrosine phosphorylation if a concomitant increase in local concentration of TCR– CD3 takes place, and this might be the case on specific pMHC binding. Several features of this model still await an experimental basis. This model makes no prediction on the minimum number of TCRs that need to bind specific pMHC but it is compatible with a single pMHC –TCR interaction being activatory. However, this model does not readily explain how soluble monomeric pMHC can be stimulatory. Compared to the other models mentioned so far, a weak point is that it does not take into account the major importance of the coreceptors in TCR signalling for some T cells. But this becomes an advantage when considering the case of coreceptor-independent T-cell signalling. In conclusion, the phenomenon of adhesion-induced T-cell priming sheds new light on the question of the minimum valency of efficient TCR ligands. The previous conclusion that TCR signalling could only be triggered by multimerized ligands remains true but only under nonphysiological conditions when T cells are in suspension, which is not the normal condition for antigen recognition in vivo. It would make sense that the TCR signalling machinery can exist under two states, a low sensitivity state when a travelling T cell is in the lymph or blood flow and a high sensitivity state when the T cell has just become adherent to various cell types of the spleen or lymph nodes, including DCs. Both ‘T-cell priming by self-peptides’ and ‘adhesion-induced T-cell priming’ could converge for the induction of this high sensitivity state for TCR signalling. Acknowledgements This work has been supported by grants from Centre National de la Recherche Scientifique, Institut National de la Sante´ et de la Recherche Me´dicale, and Ligue Nationale contre le Cancer.
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References 1 Germain, R.N. (1997) T cell signaling: the importance of receptor clustering. Curr. Biol. 7, R640 – R644 2 McKeithan, T.W. (1995) Kinetic proofreading in T-cell receptor signal transduction. Proc. Natl. Acad. Sci. U. S. A. 92, 5042 – 5046 3 Lanzavecchia, A. et al. (1999) From TCR engagement to T cell activation: a kinetic view of T cell behavior. Cell 96, 1 – 4 4 Germain, R.N. and Stefanova, I. (1999) The dynamics of T cell receptor signaling: complex orchestration and the key roles of tempo and cooperation. Annu. Rev. Immunol. 17, 467 – 522 5 van der Merwe, P.A. (2001) The TCR triggering puzzle. Immunity 14, 665 – 668 6 Symer, D.E. et al. (1992) Inhibition or activation of human T cell receptor transfectants is controlled by defined, soluble antigen arrays. J. Exp. Med. 176, 1421– 1430 7 Boniface, J.J. et al. (1998) Initiation of signal transduction through the T cell receptor requires the multivalent engagement of peptide/MHC ligands. Immunity 9, 459– 466 8 Cochran, J.R. et al. (2000) The relationship of MHC-peptide binding and T cell activation probed using chemically defined MHC class II oligomers. Immunity 12, 241 – 250 9 Daniels, M.A. and Jameson, S.C. (2000) Critical role for CD8 in T cell receptor binding and activation by peptide/major histocompatibility complex multimers. J. Exp. Med. 191, 335– 346 10 Sykulev, Y. et al. (1996) Evidence that a single peptide-MHC complex on a target cell can elicit a cytolytic T cell response. Immunity 4, 565 – 571 11 Delon, J. et al. (1998) Antigen-dependent and antigen-independent Ca2þ responses triggered in T cells by dendritic cells compared to B cells. J. Exp. Med. 188, 1473– 1484 12 Delon, J. et al. (1998) CD8 expression allows T cell signaling by monomeric peptide– MHC complexes. Immunity 9, 467 – 473 13 Irvine, D.J. et al. (2002) Direct observation of ligand recognition by T cells. Nature 419, 845 – 849 14 van der Merwe, P.A. (2002) Do T cell receptors do it alone? Nat. Immunol. 3, 1122 – 1123 15 Salter, R.D. et al. (1990) A binding site for the T-cell co-receptor CD8 on the a3 domain of HLA-A2. Nature 345, 41 – 46 16 Jameson, S.C. and Bevan, M.J. (1995) T cell receptor antagonists and partial agonists. Immunity 2, 1 – 11 17 Madrenas, J. and Germain, R.N. (1996) Variant TCR ligands: new insights into the molecular basis of antigen- dependent signal transduction and T-cell activation. Semin. Immunol. 8, 83 – 101 18 Anderson, P. et al. (1987) Cross-linking of T3 (CD3) with T4 (CD4)
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20
21 22 23 24
25
26 27
28
29 30
31
32 33
enhances the proliferation of resting T lymphocytes. J. Immunol. 139, 678– 682 Luescher, I.F. et al. (1995) CD8 modulation of T-cell antigen receptorligand interactions on living cytotoxic T lymphocytes. Nature 373, 353– 356 Denkberg, G. et al. (2001) Critical role for CD8 in binding of MHC tetramers to TCR: CD8 antibodies block specific binding of human tumor-specific MHC-peptide tetramers to TCR. J. Immunol. 167, 270– 276 Davis, S.J. et al. (2003) The nature of molecular recognition by T cells. Nat. Immunol. 4, 217 – 224 Brown, J.H. et al. (1993) Three-dimensional structure of human class II histocompatibility antigen HLA-DR1. Nature 364, 33 – 39 Fremont, D.H. et al. (1996) Structures of an MHC class II molecule with covalently bound single peptides. Science 272, 1001 – 1004 Lindstedt, R. et al. (2001) Amino acid substitutions in the putative MHC class II ‘dimer of dimers’ interface inhibit CD4þ T cell activation. J. Immunol. 166, 800– 808 Wang, J.H. et al. (2001) Crystal structure of the human CD4 N-terminal two-domain fragment complexed to a class II MHC molecule. Proc. Natl. Acad. Sci. U. S. A. 98, 10799 – 10804 Doucey, M.A. et al. (2003) CD3d establishes a functional link between the T cell receptor and CD8. J. Biol. Chem. 278, 3257 – 3264 Ge, Q. et al. (2002) Soluble peptide-MHC monomers cause activation of CD8þ T cells through transfer of the peptide to T cell MHC molecules. Proc. Natl. Acad. Sci. U. S. A. 99, 13729 – 13734 Schott, E. et al. (2002) Class I negative CD8 T cells reveal the confounding role of peptide-transfer onto CD8 T cells stimulated with soluble H2-Kb molecules. Proc. Natl. Acad. Sci. U. S. A. 99, 13735 – 13740 Randriamampita, C. et al. (2003) T cell adhesion lowers the threshold for antigen detection. Eur. J. Immunol. 33, 1215– 1223 Doucey, M.A. et al. Convergence of b1/b3 integrin- and TCR/CD8mediated signals promotes activation of CTL mediated cytotoxicity. J. Biol. Chem. 10.1074/jbc.302709200 (http://intl.jbc.org/) Stefanova, I. et al. (2003) TCR ligand discrimination is enforced by competing ERK positive and SHP-1 negative feedback pathways. Nat. Immunol. 4, 248 – 254 Stefanova, I.I. et al. (2002) Self-recognition promotes the foreign antigen sensitivity of naı¨ve T lymphocytes. Nature 420, 429– 434 Davis, S.J. and van der Merwe, P.A. (1996) The structure and ligand interactions of CD2: implications for T-cell function. Immunol. Today 17, 177 – 187
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