Antigen processing and MHC I-restricted presentation of influenza PA polymerase.

Abstract

This laboratory has developed and is currently studying the cold-attenuated (ca) live influenza vaccine. Little is known of the immunogenicity of PA; however, PA, and only this polymerase, is recognized by MHC I-restricted T cells from mice with the H-2$\sp{\rm b}$ haplotype. Furthermore, given that the cytotoxic T lymphocyte (CTL) response to the viral internal proteins is more important than that to the antigenically variable glycoproteins, and is crossreactive among influenza virus subtypes, then CTL recognition of PA is also likely to be directed against a conserved antigenic site(s). Therefore, study of CTL recognition of the ca PA may help to determine the antigen processing required for this functionally important protein to become immunogenic. The intracellular trafficking of PA, and its interaction with MHC I, was examined in cells which express complete MHC I molecules, and was compared to the behavior in cell variants unable to assemble MHC I. Effects of inhibitors of MHC I-restricted antigenic processing were revealed by immunofluorescence and immunoprecipitation. Immunological localization within cells was determined with monospecific sera to PA, antibodies to free and/or assembled MHC I, and antibodies to endosome/lysosome, Golgi and endoplasmic reticulum markers, in combination with subcellular fractionation. Direct examination of the binding of cell-associated MHC I with antigenic peptides, and their transport to the cell surface, was investigated using synthetic peptides of PA which contain putative CTL epitopes and MHC I-binding motifs. The results indicate that ca PA is antigenically processed and presented by a MHC I-restricted mechanism.