Generation of T call-dependent immune responses involves a series of complex, and generally highly specific cellular interactions. Many of these interactions are controlled by the gene products of the Major Histocompatibility Complex (MHC). In the experiments described in this proposal, we will examine several aspects of MHC Class II restricted antigen presentation. The importance of MHC Class II intracellular trafficking for presentation of protein antigens will be examined through the analysis of sequence variants of these molecules. Wild type MHC molecules will be compared biochemically in order to identify the influence of polymorphism on immunologically relevant intracellular trafficking events. In addition to these studies, the role that conserved features in Class II-molecules have on activation of CD4 T calls will be analyzed, placing special emphasis on the region of the molecules that interact with accessory molecules such as CD4. The intracellular sorting events that control presentation of endogenously synthesized antigen will also be examined. A model antigen synthesized by APC and presented in the context Class II will be modified by recombinant DNA techniques, so that protein expression is redirected to particular subcellular sites. Transfectants bearing the Class II restriction element and the wild type or modified target antigen will be tested for their ability to stimulate a panel of specific T cells, so that the requirements for Class II restricted presentation of internally synthesized antigens can be defined. Finally, in order to study whether Class II positive cells of different lineages possess unique features of antigen processing or Class II trafficking, we propose a strategy to derive autonomously growing calls representing rare cell type in vivo, such as splenic dendritic cells and thymic epithelial calls. Transgenic mice will be constructed that express the SV40 T antigen under the control of the MHC Class II promoter. Class II positive tumors from such mice will be adapted to culture an used for biochemical and functional studies. Together, these studies will clarify the molecular events that control Class II restricted presentation of antigen in vivo. Such knowledge will offer possibilities for the development of rational approaches to modulating T call-dependent immune responses.
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