Lars Franksson

T Cell Tolerance Based on Avidity Thresholds Rather Than Complete Deletion Allows Maintenance of Maximal Repertoire Diversity

Given the flexible nature of TCR specificity, deletion or permanent disabling of all T cells with the capacity to recognize self peptides would severely limit the diversity of the repertoire and the capacity to recognize foreign Ags. To address this, we have investigated the patterns of CD8+ CTL reactivity to a naturally H-2Kb-presented self peptide derived from the elongation factor 1α (EF1α). EF1α occurs as two differentially expressed isoforms differing at one position of the relevant peptide. Low avidity CTLs could be raised against both variants of the EF1α peptide. These CTLs required 100-fold more peptide-H-2Kb complexes on the target cell compared with CTLs against a viral peptide, and did not recognize the naturally expressed levels of EF1α peptides. Thus, low avidity T cells specific for these self peptides escape tolerance by deletion, despite expression of both EF1α isoforms in dendritic cells known to mediate negative selection in the thymus. The low avidity in CTL recognition of these peptides correlated with low TCR affinity. However, self peptide-specific CTLs expressed elevated levels of CD8. Furthermore, CTLs generated against altered self peptide variants displayed intermediate avidity, indicating cross-reactivity in induction of tolerance. We interpret these data, together with results previously published by others, in an avidity pit model based on avidity thresholds for maintenance of both maximal diversity and optimal self tolerance in the CD8+ T cell repertoire.

PEPTIDE DEPENDENCY AND SELECTIVITY OF THE NK CELL INHIBITORY RECEPTOR LY-49C

MHC class I molecules can prevent NK cell‐mediated cytotoxicity by interacting with inhibitory receptors on the effector cells. Different conclusions have been reached regarding possible peptide selectivity of these receptors. To address whether peptide selectivity is an exclusive feature of human or immunoglobulin‐superfamily receptors, we have studied a system based on the murine NK receptor Ly‐49C in the lectin‐superfamily. Loading of TAP‐deficient RMA‐S cells with the H‐2Kb‐restricted, ovalbumin‐derived peptide OVA257 – 264 (pOVA) induced their ability to bind Ly‐49C‐transfected reporter cells, and also protected them from killing by Ly‐49C+ NK cells. Other peptides that bound and stabilized H‐2Kb equally well differed in their NK protective capacity. Comparison of the MHC class I peptide complexes (crystal structures and molecular models) revealed a conformational motif encompassing the C‐terminal parts of the α1 helix (73 – 77) and the bound peptide that was common for the protective complexes. Substitution analysis of pOVA suggested that position 7 in the peptide may be critical for optimal protection as well as for the conformational motif at position 73 – 77. In conclusion, protection mediated by the murine C‐type lectin receptor Ly‐49C is peptide dependent and selective.

β2 -Microglobulin-deficient NK cells show increased sensitivity to MHC class I-mediated inhibition, but self tolerance does not depend upon target cell expression of H-2Kb and Db heavy chains

Mice lacking β2 ‐microglobulin (β2 m− mice) express greatly reduced levels of MHC class I molecules, and cells from β2 m− mice are therefore highly sensitive NK cells. However, NK cells from β2 m− mice fail to kill β2 m− normal cells, showing that they are self tolerant. In a first attempt to understand better the basis of this tolerance, we have analyzed more extensively the target cell specificity of β2 m− NK cells. In a comparison between several MHC class I‐deficient and positive target cell pairs for sensitivity to β2 m− NK cells, we made the following observations: First, β2 m− NK cells displayed a close to normal ability to kill a panel of MHC class I‐deficient tumor cells, despite their nonresponsiveness to β2 m− concanavalin A (Con A)‐activated T cell blasts. Secondly, β2 m− NK cells were highly sensitive to MHC class I‐mediated inhibition, in fact more so than β2 m+ NK cells. Third β2 m− NK cells were not only tolerant to β2 m− Con A blasts but also to Con A blasts from H‐2Kb − /Db − double deficient mice in vitro. We conclude that NK cell tolerance against MHC class I‐deficient targets is restricted to nontransformed cells and independent of target cell expression of MHC class I free heavy chains. The enhanced ability of β2 m− NK cells to distinguish between MHC class I‐negative and ‐positive target cells may be explained by increased expression of Ly49 receptors, as described previously. However, the mechanisms for enhanced inhibition by MHC class I molecules appear to be unrelated to self tolerance in β2 m− mice, which may instead operate through mechanisms involving triggering pathways.