T cell receptor alpha/beta pairs expressed by CD4 T cells isolated from the lungs of chronic beryllium disease patients

The goal of the study was to discover dominant stimulatory epitopes for beryllium-specific T cells isolated from the bronchoalveolar lavage of HLA-DP2-expressing patients with chronic beryllium disease (CBD). CD4 T cells were single-cell sorted from seven CBD patients and then analyzed for TRA and TRB gene expression by PCR. Barcoding on the 5' and 3' ends of PCR products were used to enable pooling of wells for high-throughput sequencing. After TCR gene expression for each T cell was determined, selected pairs were expressed in hybridoma cell lines to determine if they were beryllium-specific. Selected hybridomas expressing an LKGGG amino acid CDR3beta motif found in multiple patients were beryllium-specific and were entered into our pathway of antigen discovery. Using decapeptide positional scanning libraries, biometrical analysis, and testing of individual synthesized peptides, we determined that T cells were specific to beryllium-modified self-peptides derived from C-C motif ligands 4 (CCL4) and 3 (CCL3). These findings have a profound impact on our understanding of the immunopathogenesis of CBD.