Identification of the T-cell and Ia contact residues of a T-cell antigenic epitope

The precise molecular structure of the antigenic determinant recognized by the T-cell receptor of the CD4-positive cell has not been completely resolved. A major advance in our understanding of this issue has been made by our demonstration of a direct association between an immunogenic peptide and a purified Ia molecule. The most likely and economical hypothesis is that antigen binds directly to an Ia molecule creating the antigenic determinant and that this antigen-Ia complex is recognized by the T-cell receptor. We examined in detail a determinant of hen egg-white lysozyme (HEL) contained in the tryptic fragment HEL(46-61), recognized by T cells in H-2k strains of mice. This peptide binds with a Kd of approximately 3 microM to I-Ak molecules. We have already ascertained that (1) the 10-mer HEL(52-61) is the shortest stimulatory peptide; (2) the Leu56 residue, the only residue different from mouse lysozyme, is responsible for the immunogenicity; (3) the Leu56 and Tyr53 residues are critical for recognition by the T-cell receptor and (4) HEL(46-61) generates multiple determinants when it associated with the I-Ak molecule. If antigen and Ia interact, the antigen must have two features: it must bind to an Ia molecule and also interact with the T-cell receptor. The two sites do not appear to be laterally separable in this peptide and are therefore probably composed of distinct but interspersed amino-acid residues. We have now identified the three residues of HEL(52-61) that contact the T-cell receptor and three other residues that contact the I-Ak molecule. From modelling studies we also propose that HEL(52-61) assumes an alpha-helical conformation as it is bound to I-Ak and recognized by the T-cell receptor.     

Pleiotropic loss of activation pathways in a T-cell receptor alpha-chain deletion variant of a cytolytic T-cell clone

Untransformed T-cell clones maintained in culture are dependent on signals transmitted through their antigen receptors (Ti; alpha and beta chains associated with the CD3 molecules) for growth and effector function. For cytolytic T cells (CTL), Ti stimulation also activates the killing machinery and induces synthesis of gamma interferon (IFN-gamma) messenger RNA and IFN-gamma secretion. The Thy-1 molecule, expressed on all murine cells of the T-cell lineage, has been suggested to function in transmembrane signalling, based on the ability of some anti-Thy-1 monoclonal antibodies (mAb) to activate T cells. Recently, it was suggested that Thy-1 could function as a signal-transduction molecule when expressed in B-cell lymphomas after transfection of the gene, leading to speculation that the molecule was part of an activation pathway independent of the Ti/CD3 structures. Here we report the immunoselection of a variant CTL clone which has lost expression of mRNA for the alpha-chain of the Ti. The Ti- variant was defective in lectin-mediated activation whether measured by increase in intracytoplasmic Ca2+, CTL effector function or IFN-gamma synthesis. The variant, which expressed normal levels of Thy-1, was also unresponsive to Thy-1 mAb activation as measured by IFN-gamma secretion, whereas it responded to calcium ionophore plus phorbol ester. These results indicate that in a non-transformed, functional mature T-cell, Thy-1 mediated signalling is not an alternative to, but might depend on elements associated with the Ti/CD3-mediated T-cell activation pathway.     

Identification of a putative second T-cell receptor

Framework monoclonal antibodies have identified a population of human lymphocytes that express the T3 glycoprotein but not the T-cell receptor (TCR) alpha- and beta-subunits. Chemical crosslinking experiments reveal that these lymphocytes express novel T3-associated polypeptides, one of which appears to be the product of the T gamma gene. The other polypeptide may represent a fourth TCR subunit, designated T delta.     

Preferential expression of a defined T-cell receptor beta-chain gene in hapten-specific cytotoxic T-cell clones

A multitude of different antigens can be recognized by T cells through specific receptors. Both the alpha- and beta-chains of the T-cell receptor contribute to the antigen recognition portion. The repertoire of beta-chain variable region (V beta) gene segments is limited to some 20 elements which seem to be used randomly in different T cells. Diversity at the beta-chain level can be created in several ways: a multiplicity of germline gene segments; combinatorial diversity by rearranging different V, diversity (D), joining (J) and constant (C) region elements; junctional diversity by joining gene segments at different sites; N-region diversity, that is, insertion of random nucleotides at junctional sites; and somatic mutation. However, the major sources and the extent of diversity of the T-cell receptor are unclear. To address this issue, 42 H-2Kb-restricted, 2,4,6-trinitrophenyl (TNP)-specific cytotoxic T-cell (Tc) clones from C57BL/6 mice were characterized with respect to expression of different beta-chain gene segments in messenger RNA using specific oligonucleotide probes. We report here that nearly half of the Tc clones use identical elements for productive beta-chain gene rearrangement. Thus, there is a restriction in the use of beta-chain gene segments in this panel of Tc clones which favours a particular V beta--D beta--J beta--C beta combination with a defined D beta element.     

Protection of cottontop tamarins against Epstein-Barr virus-induced malignant lymphoma by a prototype subunit vaccine

Epstein-Barr (EB) virus is one of the five herpesviruses of man. Strong links between this agent and the chain of events causing two human cancers, endemic Burkitt's lymphoma and undifferentiated nasopharyngeal carcinoma, have long been evident (reviewed in ref. 1). Because of this, and because of the very high incidence of nasopharyngeal carcinoma in certain large populations, it was suggested in 1976 that a vaccine should be developed against EB virus to prevent infection and thereby reduce tumour incidence amongst those at risk. The virus-determined membrane antigen (MA) was proposed as immunogen because it was known to elicit naturally occurring virus-neutralizing antibodies in man and because analogous antigens had been shown to act as effective experimental vaccines for preventing the herpesvirus-induced lymphomas of Marek's disease in chickens. Progress has been achieved in defining, quantifying and preparing MA molecules, and in enhancing their immunogenicity; a sensitive assay for antibodies to MA has been elaborated. Here we report that isolated cell membranes expressing MA, or purified MA glycoprotein of relative molecular mass (Mr) 340,000 (gp340), have been used to vaccinate cottontop tamarins (Saguinus oedipus oedipus), and that animals receiving either preparation were protected against the effects of a 100% tumour-inducing challenge dose of EB virus.     

Selecting and maintaining a diverse T-cell repertoire

To provide a T-cell population that will respond promptly to foreign antigen, the immune system looks inward, using the variety of self-antigens to select and maintain a diverse repertoire of receptors. A protective immune system must include a T-lymphocyte population that is poised to respond to foreign antigenic peptides presented by self-major histocompatibility complex molecules. As the organism cannot predict the precise pathogen-derived antigens that will be encountered, the system uses the diverse array of self-peptides bound to self-major histocompatibility complex molecules, not only to select a receptor repertoire in the thymus, but also to keep na?ve T cells alive and 'ready for action' in the periphery.     

Structure of a human gammadelta T-cell antigen receptor.

T-cell antigen receptors composed of gamma and delta polypeptide chains (gammadelta TCRs) can directly recognize antigens in the form of intact proteins or non-peptide compounds, unlike alphabeta TCRs, which recognize antigens bound to major histocompatibility complex molecules (MHC). About 5% of peripheral blood T cells bear gammadelta TCRs, most of which recognize non-peptide phosphorylated antigens. Here we describe the 3.1 A resolution structure of a human gammadelta TCR from a T-cell clone that is phosphoantigen-reactive. The orientation of the variable (V) and constant (C) regions of the gammadelta TCR is unique when compared with alphabeta TCRs or antibodies, and results from an unusually small angle between the Vgamma and Cgamma domains. The complementarity-determining regions (CDRs) of the V domains exhibit a chemically reasonable binding site for phosphorylated antigens, providing a possible explanation for the canonical usage of the Vgamma9 and Vdelta2 gene segments by phosphoantigen-reactive receptors. Although the gammadelta TCR V domains are similar in overall structure to those of alphabeta TCRs, gammadelta TCR C domains are markedly different. Structural differences in Cgamma and Cdelta, and in the location of the disulphide bond between them, may enable gammadelta TCRs to form different recognition/signalling complexes than alphabeta TCRs.