Corinne Moulon

T cell immune responses to haptens. Structural models for allergic and autoimmune reactions

Protein-reactive chemicals, metal salts and drugs, commonly classified as immunological haptens, are major environmental noxes targeted at the immune system of vertebrates. They may not only interfere with this defense system by toxicity alone, but more often by evoking hapten-specific immune responses resulting in allergic and eventually autoimmune responses. Here, we review recent developments in the analysis of the structural basis of hapten recognition, particularly by T lymphocytes, which represent central elements in cell-mediated, as well as in IgE dependent, allergies. A break-through in this field was the finding that T cells detect haptens as structural entities, attached covalently or by complexation to self-peptides anchored in binding grooves of major histocompatibility antigens (MHC-proteins). Synthetic hapten-peptide conjugates were shown to induce hapten-specific contact sensitivity in mice, opening new routes for studying hapten-induced immune disorders.

A New Type of Metal Recognition by Human T Cells: Contact Residues for Peptide-independent Bridging of T Cell Receptor and Major Histocompatibility Complex by Nickel

In spite of high frequencies of metal allergies, the structural basis for major histocompatibility complex (MHC)-restricted metal recognition is among the unanswered questions in the field of T cell activation. For the human T cell clone SE9, we have identified potential Ni contact sites in the T cell receptor (TCR) and the restricting human histocompatibility leukocyte antigen (HLA)-DR structure. The specificity of this HLA-DR–promiscuous VA22/VB17+ TCR is primarily harbored in its α chain. Ni reactivity is neither dependent on protein processing in antigen-presenting cells nor affected by the nature of HLA-DR–associated peptides. However, SE9 activation by Ni crucially depends on Tyr29 in CDR1α, an N-nucleotide–encoded Tyr94 in CDR3α, and a conserved His81 in the HLA-DR β chain. These data indicate that labile, nonactivating complexes between the SE9 TCR and most HLA-DR/peptide conjugates might supply sterically optimized coordination sites for Ni ions, three of which were identified in this study. In such complexes Ni may effectively bridge the TCR α chain to His81 of most DR molecules. Thus, in analogy to superantigens, Ni may directly link TCR and MHC in a peptide-independent manner. However, unlike superantigens, Ni requires idiotypic, i.e., CDR3α-determined TCR amino acids. This new type of TCR–MHC linkage might explain the high frequency of Ni-reactive T cells in the human population.

Components of the Ligand for a Ni++ Reactive Human T Cell Clone

The major histocompatibility complex (MHC) restriction element for a human Ni2+ reactive T cell, ANi-2.3, was identified as DR52c. A series of experiments established that the functional ligand for this T cell was a preformed complex of Ni2+ bound to the combination of DR52c and a specific peptide that was generated in human and mouse B cells, but not in fibroblasts nor other antigen processing–deficient cells. In addition, ANi-2.3 recognition of this complex was dependent on His81 of the MHC β chain, suggesting a role for this amino acid in Ni2+ binding to MHC. We propose a general model for Ni2+ recognition in which βHis81 and two amino acids from the NH2-terminal part of the MHC bound peptide coordinate Ni2+ which then interacts with some portion of the Vα CDR1 or CDR2 region.

Metal-Protein Complex-Mediated Transport and Delivery of Ni2+ to TCR/MHC Contact Sites in Nickel-Specific Human T Cell Activation

Nickel allergy clearly involves the activation of HLA-restricted, skin-homing, Ni-specific T cells by professional APCs. Nevertheless, knowledge concerning the molecular details of metal-protein interactions underlying the transport and delivery of metal ions to APC during the early sensitization phase and their interactions with HLA and TCRs is still fragmentary. This study investigates the role of human serum albumin (HSA), a known shuttling molecule for Ni2+ and an often-disregarded, major component of skin, in these processes. We show that Ni-saturated HSA complexes (HSA-Ni) induce and activate Ni-specific human T cells as potently as Ni salt solutions when present at equimolar concentrations classically used for in vitro T cell stimulation. However, neither HSA itself nor its Ni-binding N-terminal peptide are involved in determining the specificity of antigenic determinants. In fact, HSA could be replaced by xenogeneic albumins exhibiting sufficient affinity for Ni2+ as determined by surface plasmon resonance (Biacore technology) or atomic absorption spectroscopy. Moreover, despite rapid internalization of HSA-Ni by APC, it was not processed into HLA-associated epitopes recognizable by Ni-specific T cells. In contrast, the presence of HSA-Ni in the vicinity of transient contacts between TCR and APC-exposed HLA molecules appeared to facilitate a specific transfer of Ni2+ from HSA to high-affinity coordination sites created at the TCR/HLA-interface.

New Structural Model for HLA-Restricted Presentation of Nickel to T Cells

Nickel ions are potent inducers of T cell-mediated contact hypersensitivity in humans. Human leukocyte antigen (HLA)-restricted, nickel (Ni)-specific T cells have been repeatedly cloned from Ni-allergic patients.1 However, unlike the case for peptideor hapten-reactive T cells, the structural identity of Ni-containing allergenic epitopes remains to be defined. We analyzed the unique T cell receptor (TCR) of the human CD4+ T cell clone SE9, which specifically reacts to Ni in the context of most HLA-DR alleles. Furthermore, its Ni-specificity and DR-restriction are defined largely by the α-chain of its α,β-heterodimer.2 Ni recognition by this TCR is independent of the nature of HLA-DR-associated peptides, but crucially relies on a conserved, surface-exposed histidine in position 81 of HLA-DR β-chains. A variety of His81 mutations completely abolished nickel presentation, while a mutant replacing His by Tyr retained partial activity. Antigen contact residues within the TCR α-chain were studied by expressing wild-type and mutated TCR αand β-chains in the TCRdeficient murine hybridoma line 54ζ17,3 resulting in hybridoma T913. These studies revealed a lack of involvement of the α-chain CDR2 loop, but identified two tyrosines as absolutely essential contact residues of the TCR α-chain: one in position 29 of CDR1, the other in the N-nucleotide-encoded position 94 of CDR3. Exchange of either of the two tyrosines to Ala or Phe extinguished Ni specificity, whereas Tyr29 in CDR1 could be functionally replaced by His. Other mutations in CDR1 or CDR2 had either no or only quantitative effects on Ni recognition. Even an elongation of the CDR3 loop by insertion of Ala between positions 92 and 93 did not alter the hybridoma’s sensitivity for nickel. These findings reveal that Ni ions may activate TCRs by a mechanism differing not only from HLA-allele-specific peptide recognition, but also from peptide-independent activation by superantigens. In the case of hybridoma T913, peptide-independent bridging of the TCR and the major histocompatibility complex results from the cross-linking by Ni2+ of Tyr or His residues in hypervariable (and particularly in idiotypic) sequences of the TCR with conserved