Differing processing requirements of four recombinant antigens containing a single defined T-cell epitope for presentation by major histocompatibility complex class II

A set of predictive rules governing the likelihood of generating a particul ar peptide-major histocompatibility complex (MHC) class II complex from an intact antigen has not been fully elucidated. We investigated the influence of positional and structural constraints in the region of the epitope by d esigning a set of recombinant antigens that each contained the well-charact erized T-cell epitope moth cytochrome c (MCC) (88-103), which is specifical ly recognized by the monoclonal antibody (mAb) D4 when complexed with H-2E( k). Our model antigens contained MCC(88-103) either peripherally, at or tow ards the C-terminus, or internally. Their abilities to bind directly to sol uble H-2E(k), and the extent of D4 epitope formation from them by antigen p rocessing-competent and -incompetent cell lines, were determined. Here we r eport that three of these four antigens yielded MCC(88-103)/H-2E(k) complex es independently of the conventional MHC class II antigen-processing and pr esentation pathway, and in each case the epitope was carried peripherally; two bound directly as intact proteins, probably as a result of spatial sepa ration of the epitope from the major globular domain, and one was processed to peptide by a cell-surface protease. One protein, which carried the epit ope inserted into an internal loop, acted as a conventional processing-depe ndent MCC(88-103) delivery vehicle. Thus, this epitope has different presen tation requirements depending on its context. These antigens constitute a p anel whose framework could be modified to further define predictive rules f or antigen processing for presentation through the different MHC class II c omplex-generating pathways.