Presentation of an influenza nucleoprotein epitope incorporated into the H-2D(b) signal sequence requires the transporter-associated with antigen presentation

In an effort to optimize the formation of peptide-specific CTL target structures in the context of plasmid DNA immunization, a strategy was developed to couple the biosynthesis of a class I heavy chain with its optimal binding epitope. Specifically, a cDNA expression vector was constructed with the influenza nucleoprotein epitope NP366-74 incorporated into the signal sequence of its restriction element H-2D(b), Transporter associated with Ag presentation-expressing murine cell lines P815 (H-2(d)) and BW5147 (H-2(k)) transfected with this modified heavy chain expressed normal levels of plasmid-encoded D-b at the cell surface, and were lysed by NP366-74-specific CTL, These results indicate that the modified signal sequence was successfully delivered to the endoplasmic reticulum, and the epitope within it processed for T cell recognition, In contrast, T2 cells, which lack the TAP transporter, when transfected with the same vector were not lysed by NP366-74 CTL, and exhibited an Ag-processing-defective phenotype. Thus, these data, which indicate TAP-dependent presentation of an optimal CTL epitope located in a signal sequence, challenge the effectiveness of Ag processing in the endoplasmic reticulum.