The stimulation of low-affinity, nontolerized clones by heteroclitic antigen analogues causes the breaking of tolerance established to an immunodominant T cell epitope

H-2K mice injected, intravenously in saline or intraperitoneally in incompl ete Freund's adjuvant, with large quantities of the immunodominant I-E-k-re stricted epitope from moth cytochrome c (MCC) 88-103 fail to respond to sub sequent immunization with this epitope when administered in complete Freund 's adjuvant. This state of tolerance can be broken by immunization with cer tain MCC 88-103 analogues that are heteroclitic antigens as assessed on rep resentative MCC 88-103 specific T cell clones. In this paper, the mechanism of breaking tolerance by heteroclitic antigens was investigated. The follo wing observations were made: (a) T cell hybridomas derived from tolerance-b roken animals required higher concentrations of MCC 88-103 to be stimulated than hybridomas derived from normal immune animals, suggesting that they h ave T cell receptors (TCRs) of lower affinity; (b) in contrast to normal im mune animals whose MCC-specific TCRs are typically V beta 3(+)/V alpha 11(), none of the hybridomas derived from tolerance-broken animals expressed V beta 3, although they were all V alpha 11(+). Also, the V beta complementa rity determining region 3 (CDR3) regions from the tolerance-broken animals did not contain the canonical structure and length characteristics of the n ormal MCC 88-103 immune repertoire; and (c) adoptive transfer and tolerizat ion of MCC-specific V beta 3(+)/V alpha 11(+) transgenic T cells followed b y immunization with heteroclitic antigen failed to terminate the state of t olerance. Collectively, these data strongly suggest that the mechanism invo lved in breaking tolerance in this system is the stimulation of nontolerize d, low-affinity clones, rather than reversal of anergy. Further support for this mechanism was the finding that after activation, T cells apparently h ave a lowered threshold with respect to the affinity of interaction with an tigen required for stimulation.