Peritoneal macrophages from C57BL/6 mice process antigens from bacteria or coated on polystyrene beads for presentation by major histocompatibility complex (MHC) class I molecules. To investigate this antigen processing pathway, peritoneal macrophages from homozygous TAP1(-/-) mice, which lack the transporter associated with antigen processing (TAP) and are defective in presenting endogenous antigens on MHC class I, were used. TAP1(-/-) or C57BL/6 macrophages were co-incubated with either bacteria or polystyrene bends containing the 257-264 epitope from ovalbumin [OVA(257-264)], which binds the mouse class I molecule Kb. The, 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 fusion 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 coated onto polystyrene beads. The data reveal that peritoneal macrophages from C57BL/6 and TAP1(-/-) mice can process bacteria expressing Crl-OVA, MBP-Crl-OVA and rOVA as well as beads coated with native OVA, purified Crl-OVA, and purified MBP-Crl-OVA and present OVA(257-264) for recognition by OVA(257-264)/Kb-specific T hybridoma cells, albeit with different relative processing efficiencies. The processing efficiency of TAP1(-/-) macrophages co-incubated with bacteria or beads containing Crl-OVB or MBP-Crl-OVA was reduced approximately three to five times compared to C57BL/6 macrophages, but OVA(257-264) was presented 100 times less efficiently when the source of OVA(257-264) was full-length OVA. Chloroquine inhibition studies showed a differential requirement for acidic compartments in C57BL/6 versus TAP1(-/-) macrophages, which 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 cellular compartments and that the protein context of the OVA(257-264) epitope influences the extent of TAP-independent processing for MHC class I presentation.
- Antigen processing and presentation
- Major histocompatibility complex class I molecule
- Transporter associated with antigen processing (TAP)