Establishment of an expression cloning system for CD4(+) T cell epitopes

We previously reported an epitope presenting vector, pCI, a derivative of a human invariant chain (Ii) expression vector, in which the class II associ ated invariant chain peptide (CLIP, Ii p89-101) could be substituted with a ntigenic peptides. In the current study, we used this vector to develop a n ew expression cloning system to identify CD4(+) T cell epitopes. We inserte d double-stranded oligo DNAs of randomized sequences into this vector and p repared an epitope-presenting library which loads randomized 13-mer peptide s onto HLA class II molecules coexpressed in COS-7 cells. Utilizing this li brary, we isolated a crossreactive epitope recognized by a glutamic acid de carboxylase (GAD) 65-autoreactive T cell clone established from a patient w ith insulin-dependent diabetes mellitus. Although the newly identified epit ope (PVQLSNQWHVVGATF) was far different from the original epitope, GAD65 p1 16-128 (NILLQYVVKSFDR), it did have the capacity to stimulate the T cell cl one comparable to that of the original GAD epitope. Our system may be appli cable not only for identifying of cross-reactive epitopes for CD4(+) T cell s of known specificity, but also for detection of epitopes stimulatory for CD4(+) T cells the epitopes of which are unknown. (C) 2001 Academic Press.