MAJOR HISTOCOMPATIBILITY COMPLEX CLASS-I PRESENTATION OF OVALBUMIN PEPTIDE-257-264 FROM EXOGENOUS SOURCES - PROTEIN CONTEXT INFLUENCES THE DEGREE OF TAP-INDEPENDENT PRESENTATION

Peritoneal macrophages from C57BL/6 mice process antigens from bacteri a or coated on polystyrene beads for presentation by major histocompat ibility complex (MHC) class I molecules. To investigate this antigen p rocessing pathway peritoneal macrophages from homozygous TAP1(-/-) mic e, which lack the transporter associated with antigen processing (TAP) and are defective in presenting endogenous antigens on MHC class I, w ere used. TAP1(-/-) or C57BL/6 macrophages were co-incubated with eith er bacteria or polystyrene beads containing the 257-264 epitope from o valbumin [OVA(257-264)], which binds the mouse class I molecule K-b. T he source of the OVA(257-264) epitope was either the Crl-OVA(257-264) (Crl-OVA) fusion protein, the maltose binding protein (MBP)-Crl-OVA fu sion protein, native OVA or bacterial recombinant OVA (rOVA); Crl-OVA, MBP-Crl-OVA and rOVA were each expressed in bacteria, and Crl-OVA and MBP-Crl-OVA purified from bacterial lysates and native egg OVA were c oated onto polystyrene beads. The data reveal that peritoneal macropha ges from C57BL/6 and TAP1(-/-) mice can process bacteria expressing Cr l-OVA, MBP-Crl-OVA and rOVA as well as beads coated with native OVA, p urified Crl-OVA, and purified MBP-Crl-OVA and present OVA(257-264) for recognition by OVA(257-264)K-b-specific T hybridoma cells, albeit wit h different relative processing efficiencies. The processing efficienc y of TAP1(-/-) macrophages co-incubated with bacteria or beads contain ing Crl-OVA or MBP-Crl-OVA was reduced approximately three to five tim es compared to C57BL/6 macrophages, but OVA(257-264) was presented 100 times less efficiently when the source of OVA(257-264) was full-lengt h OVA. Chloroquine inhibition studies showed a differential requiremen t for acidic compartments in C57BL/6 versus TAP1(-/-) macrophages, whi ch also depended upon the source of the OVA (257-264) epitope (Crl-OVA versus full-length OVA). These data suggest that TAP1(-/-) and C57BL/ 6 macrophages may process Crl-OVA and full-length OVA in different cel lular compartments and that the protein context of the OVA(257-264) ep itope influences the extent of TAP-independent processing for MHC clas s I presentation.