Janice White

Detection of Antigen-Specific T Cells with Multivalent Soluble Class II MHC Covalent Peptide Complexes

Multimeric soluble MHC class II molecules stably occupied with covalently attached peptides bind with appropriate specificity to T cell hybridomas and T cells from T cell receptor transgenic mice. There is a direct correlation between soluble T cell receptor affinity for monomeric MHC/peptide and level of binding of multimeric MHC/peptide to T cells. While binding of the multimeric MHC/peptide complex is proportional to T cell receptor affinity and expression level, there is little influence of T cell CD4.

A T cell receptor Vβ segment that imparts reactivity to a class II major histocompatibility complex product

We have identified in mice an allele of a new T cell receptor V beta gene, V beta 17a, whose product is bound by the monoclonal antibody KJ23a. Over 90% of T cell hybridomas prepared from V beta 17a+ T cells of SWR mice respond to allogeneic forms of the IE class II MHC protein, indicating that V beta 17a has an appreciable affinity for IE regardless of the other components of the T cell receptor. These results suggest a bias in the germ-line T cell receptor repertoire toward recognition of MHC proteins and indicate that the V beta portion of the receptor may form the most important contact points with MHC ligands.

How the T Cell Repertoire Becomes Peptide and MHC Specific

T cells bearing alphabeta T cell receptors (TCRs) recognize antigens in the form of peptides bound to class I or class II major histocompatibility proteins (MHC). TCRs on mature T cells are usually very specific for both peptide and MHC class and allele. They are picked out from a precursor population in the thymus by MHC-driven positive and negative selection. Here we show that the pool of T cells initially positively selected in the thymus contains many T cells that are very crossreactive for peptide and MHC and that subsequent negative selection establishes the MHC-restriction and peptide specificity of peripheral T cells. Our results also suggest that germline-encoded TCR variable elements have an inherent predisposition to react with features shared by all MHC proteins.

Immunological adjuvants promote activated T cell survival via induction of Bcl-3

Injection of soluble protein antigen into animals causes abortive proliferation of the responding T cells. Immunological adjuvants boost T cell responses at least in part by increasing the survival of activated T cells during and after the initial proliferative phase of their clonal expansion. To understand how adjuvants promote T cell survival, we used gene microarrays to analyze gene expression in T cells activated either with antigen alone or in the presence of two different adjuvants. Among the genes whose expression was increased by both adjuvants was the IκB family member Bcl-3. Retroviral infection experiments showed that expression of Bcl-3 increased survival of activated T cells in vitro and in vivo. Adjuvants may therefore improve survival of activated T cells via induction of Bcl-3.

The mouse T cell receptor: Comparison of MHC-restricted receptors on two T cell hybridomas

The receptors for antigen plus a major histocompatibility complex (MHC) gene product on a T cell hybridoma specific for ovalbumin plus a Class II MHC product were compared with those on another T cell hybridoma, specific for a Class I MHC product. In each case receptor material was identified by a clone-specific monoclonal antibody. The two receptors proved to have very similar gross structures, being 70-85 kd proteins, and reducing to an acidic alpha-chain and a slightly basic beta-chain, each 40-43 kd. The charge of both the acidic and basic polypeptides varied between the two receptors studied, showing that variable amino acid sequences occur in both chains.

The T cell receptor: the α and β chains define idiotype, and antigen and MHC specificity

Summary Three independent T cell hybridomas were isolated that have identical specificities for antigen and products of the major histocompatibility complex (MHC). All three react with the same clone-specific antireceptor antibody, and Southern blots show all three contain the same rearranged α and β genes. Variants of one of these hybridomas, DO-11.10, were isolated that had lost the ability to respond to antigen plus MHC. These proved to have lost the DO-11.10-specific α or β genes or both. Fusion of α-loss variants to β-loss variants restored reactivity. These results indicate that the specific recognition of antigen plus MHC is determined solely by the α/β-containing T cell receptor.

Germline-encoded amino acids in the αβ T cell receptor control thymic selection

An αβ T-cell response depends on the recognition of antigen plus major histocompatibility complex (MHC) proteins by its antigen receptor (TCR). The ability of peripheral αβ T cells to recognize MHC is at least partly determined by MHC-dependent thymic selection, by which an immature T cell survives only if its TCR can recognize self MHC. This process may allow MHC-reactive TCRs to be selected from a repertoire with completely random and unbiased specificities. However, analysis of thymocytes before positive selection indicated that TCR proteins might have a predetermined ability to bind MHC. Here we show that specific germline-encoded amino acids in the TCR promote ‘generic’ MHC recognition and control thymic selection. In mice expressing single, rearranged TCR β-chains, individual mutation of amino acids in the complementarity-determining region (CDR) 2β to Ala reduced development of the entire TCR repertoire. Altogether, these results show that thymic selection is controlled by germline-encoded MHC contact points in the αβ TCR and indicate that the diversity of the peripheral T-cell repertoire is enhanced by this ‘built-in’ specificity.

Multiple binding sites for bacterial superantigens on soluble class II MHC molecules

We used surface plasmon resonance to study the binding of a set of soluble mouse I-E class II major histocompatibility molecules, each occupied by a different single peptide, to the staphylococcal enterotoxin superantigens, SEA and SEB. The rates of association and dissociation to SEA varied greatly depending on the I-E-bound peptide. By contrast, binding to SEB yielded fast association and dissociation rates, which were relatively peptide independent. The results also indicated nonoverlapping binding sites for SEB and SEA on class II and raised the possibility of enhanced SAg presentation to T cells by cross-linking of cell surface class II.

Bim mediates apoptosis of CD127lo effector T cells and limits T cell memory

Following an acute T cell response, most activated effector cells die, while some survive and become memory cells. The pro‐apoptotic Bcl‐2 family member, Bcl‐2 interacting mediator of death (Bim) is critical for eliminating most effector T cells, while expression of CD127 (IL‐7Rα) has been proposed to mark effector cells destined to become memory cells. Here, we examined the effects of Bim on the death of effector T cells in relationship to CD127 expression and on development of T cell memory following lymphocytic choriomeningitis virus (LCMV) infection. We found that large numbers of CD127lo LCMV‐specific CD4+ and CD8+ T cells were lost in wild‐type mice, but were spared in Bim–/– mice. Further, while the numbers of CD127hi T cells declined only slightly during contraction of the response in wild‐type mice, they increased significantly in Bim–/– mice due to re‐expression of CD127 on CD127lo T cells that had avoided apoptosis. Functional memory T cells were significantly increased in Bim–/– mice; however, they underwent a slow attrition due to decreased proliferative renewal. Taken together, these data suggest that the absence of Bim‐mediated death of LCMV‐specific CD4+ and CD8+ T cells in vivo can increase T cell memory, but other homeostatic mechanisms control the long‐term maintenance of memory cells.

Gene transfer of h-2 class ii genes: antigen presentation by mouse fibroblast and hamster b-cell lines.

We have transferred the mouse Ak alpha and Ak beta genes, which encode the class II I-Ak molecule, into mouse L-cell fibroblasts and hamster B cells. I-Ak molecules are expressed on the surface of both cell types. The L-cell and hamster B-cell I-Ak molecules appear normal by serological analyses and two-dimensional gel electrophoresis. Furthermore, the I-Ak molecules on L cells can act as targets for the allogenic T-cell killing of the transformed L cells. The I-Ak molecules in both mouse fibroblasts and hamster B cells can present certain antigens to T-cell helper hybridomas. Thus only class II molecules are required to convert the nonantigen-presenting cell. Accordingly, it will be possible to dissect the structure-function relationships existing between Ia molecules, foreign antigen, and T-cell receptor molecules by in vitro site-directed mutagenesis and gene transfer.