Phagocyte-induced antigen-specific activation of unprimed CD8+ T cells in vitro.

The strict segregation of the major histocompatibility complex (MHC) class I and class II loading pathways has been challenged by recent reports indicating that MHC class I molecules can acquire antigen in the phagocytic pathway. We now show that this alternative peptide loading pathway can be used efficiently to generate macrophages able to activate unprimed antigen-specific cytotoxic T cells in vitro. Short peptides (8-11 residues), administered in the phagocytic pathway at nanomolar concentrations, were found to be effective in specifically activating naïve cytotoxic T lymphocytes (CTL) in vitro, but longer peptides or whole protein antigen were not. Whole protein antigen coated on beads did, however, render macrophages susceptible to lysis by an antigen-specific CTL clone. This indicates that proteolysis in the phagocytic pathway has limited capability for class I-restricted presentation. We propose a model for class I loading in the phagocytic pathway consisting of direct trafficking of nascent MHC class I from the trans-Golgi network to the phagosome, prior to appearance at the cell surface, and the use of the narrow cavity between bead and phagosomal membrane as a peptide exchange/loading compartment. Targeting immunogenic class I-binding peptide to the phagocytic pathway of macrophages facilitates presentation in association with class I. This is a useful tool for CTL response induction in vitro.