Overexpression of CD3η during thymic development does not alter the negative selection process

To elucidate the role of CD3η in thymic development and to determine whether CD3η is involved in the negative selection process, CD3η was overexpressed >100 fold in transgenic (tg) mice using a Thy-1 promoter and regulatory elements. CD3η was readily observed in the majority of cortical thymocytes and in a fraction of medullary thymocytes in tg mice by immunohistochemical staining with an anti-CD3η-specific mAb. In contrast, endogenous CD3η levels were too low to detect in normal littermates. Flow cytometric analysis demonstrated an increased level of TCR on thymocytes with intermediate TCR density in tg animals and parallel biochemical studies showed a marked increased in TCR-associated CD3ζ-η heterodimers and CD3η-η homodimers relative to controls. Despite this change in surface TCR phenotype, there was no significant alteration in the total numbers or proportion of CD4⁺CD8⁺ double-positive or CD4⁺CD8^- or CD4^-CD8⁺ single-positive thymocytes or peripheral T cells. Percentages of SP Vβ5, Vβ6, and Vβ8 thymocytes in tg animals were unaltered compared to normal littermates when backcrossed either to C57BL/6 (H-2^b) or DBA/2 (H-2^d) backgrounds. Furthermore, induction of DNA fragmentation with anti-CD3ε mAb treatment in vivo was not significantly different for tg and normal littermates. Collectively, these data imply that CD3η is not a limiting component of the negative selection process.