Human T lymphocytes, bearing
T cell receptor (TCR) gammadelta, play an important role in anti-
tumor/microbe immune responses. However, few
tumor antigens recognized by TCRgammadelta have been defined so far. To investigate antigenic
epitopes/
proteins recognized by gammadeltaT cells, we have established a new immunobiochemical strategy that uses
complementarity-determining region 3 of TCR delta chain (CDR3delta)
peptide-mediated
epitope/protein-binding assays. CDR3delta
peptides synthesized using the CDR3 region in TCR Vdelta2 chain were validated for their binding specificity to target cells or tissues. These CDR3delta
peptides were then employed as probes to pan putative
epitopes in a 12-mer random
peptide phage-displayed library and to identify putative
protein ligands within
tumor protein extracts by affinity chromatography and liquid chromatography/electrospray ionization-tandem mass spectrometry analysis. As a result, we have identified nine
peptides and two
proteins for TCRgammadelta, including human
mutS homolog 2 (hMSH2) and
heat shock protein (HSP) 60. All nine tested
epitope peptides not only bind to gammadeltaT cells but also functionally activate gammadeltaT cells in vitro. Identification of HSP60 confirms the validity of this method as HSP60 is an identified
ligand for TCRgammadelta. We show that hMSH2 is expressed on the surface of SKOV3
tumor cells, and cytotoxicity of Vdelta2 gammadeltaT cells to SKOV3 cells was blocked by anti-hMSH2 antibody, suggesting that hMSH2 may be a new
ligand for TCRgammadelta. Taken together, our findings provide a novel immunobiochemical strategy to identify
epitopes/
proteins recognized by gammadeltaT cells.