This project aims to identify and dissect important biochemical functions in antigen processing for presentation by class II MHC molecules (MHC-II). In particular, we are interested in the functions contributed by particular subcellular organelles, endosomes and lysosomes, which we can separate by subcellular fractionation. Preliminary evidence indicates that a lysosomal compartment appears to contribute to antigen processing. We hope to further define the characteristics of these organelles and to define the patterns of transport of MHC-II molecules to and from them. We will attempt to localize the compartment that mediates the formation of peptide-MHC-II complexes, using subcellular fractionation coupled with biochemical or immunological (T cell) assays to quantitate peptide binding to MHC-II. We hope to define the role of invariant chain and other possible accessory molecules in targeting MHC-II into a productive antigen processing pathway. These studies should provide important advances in our understanding of the basic mechanisms of antigen processing that determine antigenicity and immune recognition. These principles are important to aa wide range of problems in clinical immunology, including vaccination for infectious disease, therapy for autoimmunity or immunodeficiency, or enhancing immune responses to tumor cells to combat cancer.