Dominant mechanisms of tolerance have obvious therapeutic implications for intervention in autoimmunity and transplant-rejection. Our knowledge of how "regulatory" T (Treg) cells are induced and their mode of action is largely derived from studies of polyclonal T cell responses. The exact nature of the cellular interactions that govern the generation and homeostasis of regulatory T cells remains elusive and is subject of the present proposal. We will make use of two double-transgenic models that share an MHC class II restricted T cell receptor (TCR) specific for influenza hemagglutinin (HA), while the model antigen HA is expressed rather ubiquitously in one system and in a "tissue-specific" fashion in the other system. In both models, we have observed "anergic" CD4 T cells that can suppress the response of naive CD4 T cells in vitro. The regulatory properties of TCR transgenic CD4 T cells segregate with the expression of CD25 only in one system, while in the other CD25 negative cells are likewise inhibitory. Intriguingly, thymic epithelium is involved in the generation of regulatory cells not only in the situation of widespread antigen expression, but likewise when antigen expression is driven by a "tissue-specific" promoter, the latter being reminiscent of "promiscuous" expression of tissue-antigens in thymic epithelial cells. Both transgenic systems will be employed and compared with respect to the role of hematopoietic versus thymic epithelial cells in the generation of Treg cells and to address questions concerning the antigen-dependency of thymus derived Treg cells after exit from the thymus. Furthermore, we will test the hypothesis that "promiscuous" expression of so-called tissue-antigens in thymic epithelium is involved in the generation of Treg cells. Ultimately, based on the observations in our transgenic systems, we will test different protocols to experimentally induce Treg cells in vivo.