Differing processing requirements of four recombinant antigens containing a single defined T-cell epitope for...

Area of Science:

• Immunology
• Molecular Biology
• Biochemistry

Background:

• Predictive rules for generating peptide-major histocompatibility complex (MHC) class II complexes from intact antigens are not fully understood.
• The T-cell epitope moth cytochrome c (MCC) (88-103) recognized by monoclonal antibody (mAb) D4 complexed with H-2Ek serves as a model system.

Purpose of the Study:

• To investigate how positional and structural constraints of an epitope influence its presentation within recombinant antigens.
• To determine the binding of recombinant antigens to H-2Ek and the formation of the D4 epitope by antigen-presenting cells.

Main Methods:

• Design of four recombinant antigens containing the MCC (88-103) epitope either peripherally or internally.
• Assessment of direct binding of antigens to soluble H-2Ek.
• Evaluation of epitope formation from antigens by antigen processing-competent and -incompetent cell lines.

Main Results:

• Three of four antigens yielded MCC(88-103)/H-2Ek complexes independently of conventional antigen processing pathways.
• Peripheral epitope placement in these three antigens facilitated direct binding (two cases) or processing by cell-surface proteases (one case).
• An internally placed epitope required conventional processing for MCC(88-103) presentation.

Conclusions:

• Epitope presentation requirements for MHC class II complex generation are context-dependent.
• Peripheral epitope location can enable antigen presentation independent of standard intracellular processing.
• The designed recombinant antigens provide a framework for defining predictive rules for antigen processing and presentation.