Exogenous Peptides Presented by Transporter Associated with Antigen Processing (TAP)-Deficient and TAP-Competent Cells: Intracellular Loading and Kinetics of Presentation

• Published in: The Journal of immunology (1950) 2001-09, Vol.167 (5), p.2529-2537
• Main Authors: Luft, Thomas, Rizkalla, Mark, Tai, Tsin Yee, Chen, Qiyuan, MacFarlan, Roderick I, Davis, Ian D, Maraskovsky, Eugene, Cebon, Jonathan
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Antigen Presentation - physiology; Antigens, Neoplasm; ATP Binding Cassette Transporter, Subfamily B, Member 2; ATP-Binding Cassette Transporters - immunology; ATP-Binding Cassette Transporters - metabolism; Binding, Competitive; Cell Differentiation; Cell Line; Cell Membrane - immunology; Cell Membrane - metabolism; Dendritic Cells - cytology; Dendritic Cells - immunology; Dendritic Cells - metabolism; Histocompatibility Antigens Class I - metabolism; HLA-A2 Antigen - metabolism; Humans; Interferon-gamma - biosynthesis; Intracellular Fluid - immunology; Intracellular Fluid - metabolism; Kinetics; MART-1 Antigen; Neoplasm Proteins - genetics; Neoplasm Proteins - immunology; Neoplasm Proteins - metabolism; Peptides - immunology; Peptides - metabolism; T-Lymphocytes, Cytotoxic - immunology; TAP protein
• ISSN: 0022-1767; 1550-6606
• DOI: 10.4049/jimmunol.167.5.2529

This study investigates the differential capacity of TAP-deficient T2 cells, TAP-competent EBV cells, and immature and mature dendritic cells to present peptides to preformed CTL lines. It demonstrates that presentation of exogenous peptides involves peptide uptake and loading onto newly synthesized MHC class I molecules. This mechanism was best demonstrated for low affinity peptides in the presence of irrelevant peptides competing for HLA binding sites. Under these circumstances, inhibition of protein synthesis with cycloheximide or vesicular trafficking with brefeldin A significantly reduced the presentation of low affinity peptides. This was not restored by adding exogenous beta(2)-microglobulin to stabilize the MHC complex on the cell surface. In contrast, presentation of high affinity peptides was not sensitive to cycloheximide or brefeldin A, which suggests that different mechanisms may operate for presentation of high and low affinity peptides by TAP-competent cells. High affinity peptides can apparently compete with peptides in preloaded MHC class I molecules at the cell surface, whereas low affinity peptides require empty MHC molecules within cells. Accordingly, very high concentrations of exogenous low affinity peptides in conjunction with active MHC class I metabolism were required to allow successful presentation against a background of competing intracellular high affinity peptides in TAP-competent cells. These findings have implications for the design of peptide and protein-based vaccines.