Self determinant selection and acquisition of the autoimmune T cell repertoire

Autologous proteins are continuously processed and presented in the form of peptides associated with self major histocompatibility (MHC) molecules at the surface of antigen-presenting cells for interaction with autoreactive T cells. During thymic selection, the presentation of self peptides is an essential element in the establishment of the T cell repertoire. Developing T cells which recognize self peptide/self MHC complexes with sufficient affinity are clonally deleted. However, we and others have recently demonstrated that a variety of self peptides, despite their high binding affinity to MHC molecules, never reach the threshold of presentation to ensure negative selection (cryptic self peptides). This mechanism may have been selected to avoid excessive purging of T cell repertoire during ontogeny. However, T cells directed to cryptic self determinants represent a continuous threat for the initiation of autoimmunity in adults. Supporting this view, recent studies have documented the involvement of cryptic self peptide presentation in different autoimmune diseases. In this article, we examine the factors that govern the selection of self peptides for presentation to autoreactive T cells in vivo and discuss their contribution to both the induction and the maintenance of self tolerance. In addition, we analyze the mechanisms by which the hierarchy of determinants on a self protein can be disrupted, thereby leading to the presentation of previously cryptic self peptides and the induction of an autoimmune T-cell-mediated process.     

Understanding crypticity is the key to revealing the pathogenesis of autoimmunity

In this opinion, we propose that the hierarchy of antigenic determinants within self-antigens is the major influence in molding the potentially autoreactive T-cell repertoire. The well processed and presented determinants constitute a 'dominant self', whereas the poorly processed and/or presented determinants will be invisible to T cells and comprise a 'cryptic self', which we consider a fundamental cornerstone of a theory of autoimmunity. It accounts for the large repertoire of self-reactive clones because a similar hierarchy is established in the thymus and controls positive and negative selection. Furthermore, this residual T-cell repertoire, largely directed against cryptic determinants, will contain some T cells with sufficient affinity for MHC and antigen that enables them to respond under inflammatory conditions, thus facilitating presentation of previously cryptic determinants.     

Intramolecular and intermolecular spreading during the course of organ allograft rejection

Summary: There are two distinct pathways by which T cells may MHC alloantigens. The direct pathway involves T-cell recognition of intact MHC molecules expressed by donor antigen-presenting cells (APCs). The second, or indirect, pathway describes T-cell recognition of peptides derived from the processing and presentation of allogeneic MHC molecules on self APCs. Recent data demonstrates that indirect recognition plays a central role in both acute and chronic rejection of human organ allografts. Our studies have shown that, at the onset of primary acute rejection, recipient T-cell responses lo donor HLA-DR alloantigens are limited to a single dominant determinant present on erne of the disparate alloantigens and restricted by one of the responder's HLA-DR molecules. In allograft recipients with recurring episodes of rejection, and/or at the onset of chronic rejection, recipient T-cell reactivity may spread lo other epitopes within the allogeneic MHC molecule as well as to other alloantigens expressed by graft tissue. Both quantitative and qualitative alterations in T-cell allopeptide reactivity are associated with increased risk of cellular and/or humoral rejection. These studies provide a basis for the design of new therapeutic strategies and for immunologic monitoring of transplant recipients.     

MHC Class II-Dependent Peptide Antigen Versus Superantigen Presentation to T Cells

T lymphocytes expressing the CD4 coreceptor can be activated by two classes of major histocompatibility complex (MHC) class II-bound ligands. The elaboration of a conventional T-cell mediated immune response involves recognition of an antigenic peptide bound to the MHC class II molecules by a T-cell receptor (TCR) specific to that particular antigen. Conversely, superantigens (SAgs) also bind to MHC class II molecules and activate T cells, leading to a completely different functional outcome; indeed, SAg-responsive T cells die through apoptosis following stimulation. Superantigens are proteins that are secreted by various bacteria. They interact with the TCR using molecular determinants that are distinct from the residues involved in the recognition of nominal antigenic peptides. Despite the similarities between the recognition of the two classes of ligands by the TCR, considerable structural difference is observed. Here, we discuss the current knowledge on the presentation of SAgs to T cells and compare the different aspects of the SAg response with the recognition of antigenic peptide/MHC complexes.     

Contemplating immunopeptidomes to better predict them

The identification of T-cell epitopes is key for a complete molecular understanding of immune recognition mechanisms in infectious diseases, autoimmunity and cancer. T-cell epitopes further provide targets for personalized vaccines and T-cell therapy, with several therapeutic applications in cancer immunotherapy and elsewhere. T-cell epitopes consist of short peptides displayed on Major Histocompatibility Complex (MHC) molecules. The recent advances in mass spectrometry (MS) based technologies to profile the ensemble of peptides displayed on MHC molecules – the so-called immunopeptidome – had a major impact on our understanding of antigen presentation and MHC ligands. On the one hand, these techniques enabled researchers to directly identify hundreds of thousands of peptides presented on MHC molecules, including some that elicited T-cell recognition. On the other hand, the data collected in these experiments revealed fundamental properties of antigen presentation pathways and significantly improved our ability to predict naturally presented MHC ligands and T-cell epitopes across the wide spectrum of MHC alleles found in human and other organisms. Here we review recent computational developments to analyze experimentally determined immunopeptidomes and harness these data to improve our understanding of antigen presentation and MHC binding specificities, as well as our ability to predict MHC ligands. We further discuss the strengths and limitations of the latest approaches to move beyond predictions of antigen presentation and tackle the challenges of predicting TCR recognition and immunogenicity.     

T cells from multiple sclerosis patients recognize multiple epitopes on Self-IgG

The highly diversified variable regions of immunoglobulin (Ig) molecules contain immunogenic determinants denoted idiotopes. We have previously reported that T cells from multiple sclerosis (MS) patients recognize IgG from autologous cerebrospinal fluid (CSF), and mapped a T-cell epitope to an IgG idiotope. To test the ability of CSF IgG molecules to elicit a broad polyclonal T-cell response in MS, we have analysed T-cell responses in the blood and CSF against idiotope peptides spanning complementarity determining region (CDR) 3 and somatic mutations within the variable regions of monoclonal CSF IgG. Consistent with a diversified idiotope-specific T-cell repertoire, CD4(+) T cells from both patients recognized several idiotope peptides presented by HLA-DR molecules. Mutations were critical for T-cell recognition, as T cells specific for a mutated CDR1 peptide did not recognize corresponding germline-encoded peptides. One T-cell clone recognized both an idiotope peptide and the B-cell clone expressing this idiotope, compatible with endogenous processing and presentation of this idiotope by B cells. These results suggest that mutated CSF IgG from MS patients carry several T-cell epitopes, which could mediate intrathecal IgG production and inflammation in MS through idiotope-driven T-B-cell collaboration.     

PA28 and the proteasome immunosubunits play a central and independent role in the production of MHC class I-binding peptides in vivo

Proteasomes play a fundamental role in the processing of intracellular antigens into peptides that bind to MHC class I molecules for the presentation of CD8(+) T cells. Three IFN-γ-inducible catalytic proteasome (immuno)subunits as well as the IFN-γ-inducible proteasome activator PA28 dramatically accelerate the generation of a subset of MHC class I-presented antigenic peptides. To determine whether these IFN-γ-inducible proteasome components play a compounded role in antigen processing, we generated mice lacking both PA28 and immunosubunits β5i/LMP7 and β2i/MECL-1. Analyses of MHC class I cell-surface levels ex vivo demonstrated that PA28 deficiency reduced the production of MHC class I-binding peptides both in cells with and without immunosubunits, in the latter cells further decreasing an already diminished production of MHC ligands in the absence of immunoproteasomes. In contrast, the immunosubunits but not PA28 appeared to be of critical importance for the induction of CD8(+) T-cell responses to multiple dominant Influenza and Listeria-derived epitopes. Taken together, our data demonstrate that PA28 and the proteasome immunosubunits use fundamentally different mechanisms to enhance the supply of MHC class I-binding peptides; however, only the immunosubunit-imposed effects on proteolytic epitope processing appear to have substantial influence on the specificity of pathogen-specific CD8(+) T-cell responses.     

The structural basis of T-cell allorecognition

Foreign allogeneic major histocompatibility complex (MHC) class I and class II molecules elicit an exceptionally vigorous T-cell response. A small component of the alloresponse comprises CD4+ T cells that recognize allogeneic MHC indirectly after processing into peptide fragments that are bound and presented by self-MHC class II. The majority of alloreactive T cells directly recognize intact allogeneic MHC molecules expressed on foreign cells. Some alloreactive T-cell interactions with allogeneic MHC molecules are indifferent to the bound peptide, but evidence suggests that most show specificity to peptide. The vigor and diversity of the direct alloreactive T-cell response can therefore be explained by summation of numerous responses to each of the peptides in the novel set bound by allogeneic MHC molecules. Structural studies definitively show that the overall mechanism of T-cell receptor (TCR) recognition of self-MHC and allogeneic MHC molecules is similar. Many alloreactive T cells recognize several different combinations of MHC and bound peptide that do not necessarily possess structural homology. Flexibility within the TCR structure allows adaptation to the different contact surfaces. Crossreactivity seems to be an intrinsic property of the TCR required, because a single TCR must possess the ability to interact with both self-MHC during positive selection and at least one combination of foreign antigenic peptide presented by self-MHC. Recognition of allogeneic MHC molecules is an inadvertent consequence of the need for TCR crossreactivity.     

The existence of two types of proteasome, the constitutive proteasome and the immunoproteasome, may serve as another layer of protection against autoimmunity

Negative selection of CD8 single positive thymocytes is based on the presentation through the major histocompatibility complex (MHC) class I pathway of peptides derived from degradation of self-proteins by the constitutive proteasome and the immunoproteasome in the thymus. Then naïve CD8+ T-cells can be primed by mature dendritic cells. In mature dendritic cells peptides presented by MHC class I molecules are derived from degradation of endogenous self-proteins or through the process of cross-presentation from degradation of exogenous proteins by the immunoproteasome. In the absence of infection, peripheral cells display peptides on MHC class I molecules derived from degradation of endogenous self-proteins by the constitutive proteasome. The pool of peptides derived from protein degradation by the constitutive proteasome differs from the pool peptides derived from protein degradation by the immunoproteasome. Thus the probability of an autoreactive naïve CD8+ T-cell that escaped negative selection, and converted by a mature dendritic cell to autoreactive cytolytic T-cell, to kill a normal cell in the periphery, is reduced.     

Efficient MHC class I-peptide binding is required but does not ensure MHC class I-restricted immunogenicity

Cytotoxic T lymphocyte (CTL) epitopes are naturally processed peptides bound and presented by major histocompatibility (MHC) class I molecules. Since they are expressed at the cell surface in sufficient amounts to be recognized by CTL, it is generally believed, and in some cases demonstrated, that they bind efficiently to MHC class I molecules in vivo. Based on this knowledge, candidate CTL epitopes are now searched for by identifying peptides that efficiently bind to MHC class I molecules in vitro. We analysed whether this approach is valid by systematically investigating the relationship between MHC class I-peptide binding and peptide immunogenicity. Fifteen peptides that represent known CTL epitopes were tested for their MHC class I binding ability. In a comparative study with 83 peptides that bear the appropriate MHC class I allele-specific motifs but do not contain known CTL epitopes, the CTL epitope-bearing peptides showed the highest binding affinity for MHC class I. This was true for two MHC class I alleles in two different assay systems that monitor peptide-MHC class I binding. Furthermore, selected motif-bearing K^b binding peptides were used to induce peptide-specific CTL responses in mice. Only a subset of the high affinity K^b binding peptides induced reproducible peptide-specific CTL responses, whereas none of the low affinity K^b binding peptides induced a response. Taken together, these results indicate that efficient peptide-MHC class I binding is required for immunogenicity. Vice versa, immunogenicity is not guaranteed by efficient peptide-MHC class I binding, implying that additional factors are involved. Nevertheless, selection of candidate CTL epitopes on the basis of MHC class I binding seems valid. Our data indicate that, although an excess of peptides might be selected, the chance of missing immunogenic peptides is minimal.     

Major histocompatibility complex class I-restricted presentation of protein antigens without prior intracellular processing

Proteins in their native form are incapable of stimulating antigen (Ag)-specific T cells, which can only recognize major histocompatibility complex (MHC)-bound peptides that have been generated by intracellular processing within antigen-presenting cells (APCs). Here, we show that APCs can trigger MHC class I-restricted T-cell responses after presenting proteins without conventional intracellular processing, provided the immunostimulatory MHC class I-binding peptide sequence is incorporated at the carboxy-terminal position. Such MHC-bound proteins do not stimulate T cells directly, because the contact between MHC/peptide complex and its cognate ligand is sterically hindered by the amino-terminal bulk of the protein. Removal of the latter via an extracellular Ag proteolysis by the T-cell- and/or APC-derived enzymes is required for effective T-cell stimulation. Our data challenge the established concept that only small peptides can bind to the MHC class I molecules.     

Superantigens are presented by and activate thymocytes from infants

A high percentage of human fetal and postnatal thymocytes express MHC class II molecules. This raises the possibility that human thymocytes in early life are able to present peptides to other immature T cells and thereby initiate thymic selection of these cells. Here we address this question by exposing newly harvested infant thymocytes to superantigen (Sag) which binds to the T-cell receptor and to MHC class II chains outside the peptide binding groove. The results show that the thymocytes are able to present Sag and to be activated to proliferation as well as apoptosis by Sag presented by other thymocytes. The absence of responses to Sag with mutations in class II binding sites showed that class II molecules were necessary for the responses, and very low expression of class II molecules on CD4-8- cells indicates that the demonstrated T-cell/T-cell interactions are confined to T-cell receptor-positive CD4+8+, CD4+8-, and CD4-8+ cells. These latter subsets were shown to be able to present Sag to each other. These findings suggest that class II+ thymocytes may participate in the selection of self-restricted T cells during a critical period in the shaping of the human immune system. Copyright Copyright 1999 S. Karger AG, Basel     

MHC class II loading of high or low affinity peptides directed by Ii/peptide fusion constructs: implications for T cell activation

CD4(+) T cells recognize peptides presented on the cell surface of antigen presenting cells in the MHC class II context. The biosynthesis and transport of MHC class II molecules depend on the type II transmembrane invariant chain (Ii) and are tightly regulated processes. Ii is known to bind to the MHC class II peptide-binding groove via its class II-associated Ii peptide (CLIP) region early in the biosynthetic pathway to prevent premature peptide binding. In this study we have genetically exchanged CLIP with peptides of either high or low affinity for the class II peptide binding groove and utilized the properties of Ii to manipulate MHC class II loading. An inducible promoter controlled expression of the Ii/peptide fusion constructs, and presentation at different expression levels was studied. Both peptides were excised from Ii and presented on MHC class II molecules as shown by liquid chromatography-tandem mass spectrometry, but the high affinity peptide was presented more efficiently than the low affinity peptide. Both peptides were efficient in eliciting T cell responses at high Ii/peptide concentration independent of the duration of T cell stimulation. The peptides were also able to elicit an IL-2 response at low expression levels; however, the kinetic differed as the T cells required longer duration of T cell contact to reach a significant T cell response. This probably reflects the number of class II/peptide complexes at the cell surface and is discussed.     

Major histocompatibility complex class I-restricted alloreactive CD4+ T cells

Although it is well established that CD4+ T cells generally recognize major histocompatibility complex (MHC) class II molecules, MHC class I-reactive CD4+ T cells have occasionally been reported. Here we describe the isolation and characterization of six MHC class I-reactive CD4+ T-cell lines, obtained by co-culture of CD4+ peripheral blood T cells with the MHC class II-negative, transporter associated with antigen processing (TAP)-negative cell line, T2, transfected with human leucocyte antigen (HLA)-B27. Responses were inhibited by the MHC class I-specific monoclonal antibody (mAb), W6/32, demonstrating the direct recognition of MHC class I molecules. In four cases, the restriction element was positively identified as HLA-A2, as responses by these clones were completely inhibited by MA2.1, an HLA-A2-specific mAb. Interestingly, three of the CD4+ T-cell lines only responded to cells expressing HLA-B27, irrespective of their restricting allele, implicating HLA-B27 as a possible source of peptides presented by the stimulatory MHC class I alleles. In addition, these CD4+ MHC class I alloreactive T-cell lines could recognize TAP-deficient cells and therefore may have particular clinical relevance to situations where the expression of TAP molecules is decreased, such as viral infection and transformation of cells.     

Long‐lasting cross‐presentation of tumor antigen in human DC

DC cross‐present exogenous antigens on MHC class I molecules, a process required for the onset of anti‐tumor immune responses. In order to study the cross‐presentation of tumor antigens by human DC, we compared the pathways of cross‐presentation of long peptides requiring internalization and intracellular processing with the direct presentation of short peptides, which does not require intracellular processing. We found that, after brief incubations with DC, short peptides were presented to CD8⁺ T cells with higher efficiencies than long peptides. After longer times of chase in the absence of peptide, however, the efficiency of presentation of the two types of peptides was reversed. After 2–3 days, DC pulsed with long peptides still activated T cells efficiently, while DC pulsed with short peptides failed to do so. Long‐lasting presentation of the long peptides was, at least in part, due to a stored persistent pool of antigen, which was still available for loading on MHC class I molecules after several days of chase. These results show that the use of long synthetic peptides allows the efficient, long‐lasting, presentation of tumor antigens, suggesting that long peptides represent an interesting approach for active anti‐tumor vaccination.     

T cells recognize antigen alone and not MHC molecules

It is a central dogma of contemporary immunology that T cells engaged in immune responses to foreign antigens or cells recognize determinants on major histocompatibility complex (MHC) molecules. Here Ole Werdelin argues that this dogma is false. Taking the case of T-cell responses which are controlled by MHC class II molecules, he suggests that la molecules serve to bind antigen fragments and stabilize them in the membrane of presenting cells, shielding them from proteolytic degradation and permitting T cells to bind the epitopes so displayed.     

Coupling Toll-like receptor signaling with phagocytosis: potentiation of antigen presentation

Much attention has focused on the role of co-stimulation in dictating tolerance versus immunity to internalized antigens, with the assumption that the presentation of antigen-derived peptides by MHC molecules occurs constitutively. Here, I highlight our new appreciation for the regulated presentation of phagocytosed antigens by MHC class II molecules as a direct result of controlling phagosome maturation by Toll-like receptors. I discuss how the mode of antigen delivery into dendritic cells coupled with the right type of signal transduction pathway can impact greatly not only the co-stimulatory context in which the antigen is presented to naive T cells (signal 2) but MHC-II presentation of the antigen and formation of the T-cell receptor ligand (signal 1) itself.     

Major histocompatibility complex class I presentation of exogenously acquired minor alloantigens initiates skin allograft rejection

Major histocompatibility complex (MHC) class I molecules present peptides from endogenous proteins. However, in some cases class I-restricted peptides can also derive from exogenous antigens. This MHC class I exogenous presentation could be involved in minor histocompatibility antigen (mHAg)-disparate allograft rejection when donor alloantigens are not expressed in graft antigen-presenting cells (APC) that initiate the rejection mechanism. Here we addressed this question by using a skin graft experimental model where donors (H-2^b or H-2^d Tgβ-gal mice) expressed the mHAg like β-galactosidase (β-gal) in keratinocytes but not in Langerhans' cells (LC) which have an APC function. Rejection of Tgβ-gal skin by a β-gal-specific CD8 cytotoxic T lymphocyte (CTL) effector mechanism should require presentation by donor and/or recipient LC of MHC class I-restricted peptides of exogenous β-gal shed by keratinocytes. Indeed, our results showed that 1) H-2^b Tgβ-gal skin was rejected by H-2^bxs and H-2^bxd recipients; 2) rejection was mediated by β-gal-specific CD8⁺ CTL effectors; and 3) H-2^bxd mice having rejected H-2^b Tgβ-gal skin generated β-gal-specific CTL restricted by H-2^b and H-2^d class I molecules and rejected subsequently grafted H-2^d Tgβ-gal skin in an accelerated fashion, demonstrating that recipient LC have presented exogenous β-gal-derived MHC class I epitopes. These results lead to the conclusion that MHC class I exogenous presentation of donor mHAg can initiate allograft rejection.     

Different types of allospecific CTL clones identified by their ability to recognize peptide loading-defective target cells

Allospecific immune responses against the MHC of another individual are remarkably strong, due t a high number of responding T cell clones. Although it has been demonstrated that some allospecific cytotoxic T lymphocytes (CTL) recognize peptides presented by allogeneic MHC class I molecules, it has remained unclear whether MHC molecules can be recognized directly. We used the H-2b-derived murine lymphoma mutant RMA-S, which has a defect affecting peptide loading of class I molecules, to test whether recognition by allospecific CTL always requires the presence of peptides. Three types of anti-H-2Kb CTL clones can be distinguished by their ability to lyse RMA-S target cells. Type A CTL clones efficiently lyse these target cells, the lysis by type B CTL clones is inefficient, and type C clones fail to lyse RMA-S. Up-regulation of the levels of H-2Kb density improved lysis by type B clones, but did not lead to lysis by type C clones. Some type A and B CTL clones apparently can recognize class I molecules devoid of peptides, while others are likely to recognize peptides which are not affected by the presentation defect of RMA-S. We suggest that type C clones are specific for peptides which are not presented by the mutant cells. The results show that the majority of alloreactive CTL recognize peptide/MHC complexes, while some CTL behave as if they can recognize class I molecules in the absence of MHC-bound peptides.     

Processing of Listeria monocytogenes antigens and the in vivo T-cell response to bacterial infection

Summary: Presentation of antigens to T lymphocytes is a critical step in the clearance of pathogens from their hosts and in the establishment of protective immunity. Several animal models have been developed to study this process, but few have been as informative as the murine immune response to Listerio monocytogenes infection. Herein we review the presentation of L, monocytogenes proteins by the MHC class I antigen-processing pathway and the in vivo T-cell response to these bacterial antigens. These studies demonstrate the following: I) The size of a peptide-specific T-cell response does not correlate with the amount of epitope presented by infected cells; 2) T cells specific for dominant epitopes do not, in the case of L. monocytogenes infection, inhibit responses to subdominant epitopes; 3) T cells responding to different epitopes presented by MHC class la molecules expand, contract and enter the memory pool synchronously; 4) Repeated in vivo expansion of antigen-specific T-cell populations results in a narrowing of their T-cell receptor repertoire and in an increase in their affinity for antigen; and S) T cells restricted by H2-M3 MHC class Ib molecules constitute a major part of the primary response to bacterial infection, but appear to play a relatively smaller role in memory responses. These studies have provided a novel glimpse of the relationship between antigen processing and in vivo T-cell responses to infection, and provide a foundation for more detailed analyses of T-cell-mediated adaptive immunity.