Reevaluating the biochemical valency of native TCR/CD3 complexes isolated from T cells (IRM7P.708)

Abstract

Abstract The αβ T cell antigen Receptor (TCR) is a multi-protein complex expressed together with CD3 subunits (γδεζ). The conventional model has it as monovalent with a stochiometry of (αβγδε2ζ2); however, some previously published work reported observation of bivalent complexes with two αβ heterodimers. To reevaluate the valency of the TCR, we have subjected T cell digitonin-solubilized complexes to immunoprecipitation detected by flow cytometry (IP-FCM), size exclusion chromatography, and other methods for detecting complex size and subunit composition. Our results indicate that precisely bivalent TCRs are found among surface-expressed complexes independent of cross-linking by strong antigenic ligands. Their detection can be masked in the presence of Coomassie reagent from Blue-native PAGE conditions, or if antibodies used in detection assays are themselves bivalent. Therefore, bivalent TCR/CD3 complexes can be found under the biochemical conditions that have classically defined the minimally expressed TCR/CD3 unit (digitonin solubilization). Further investigation is required to determine the functional relevance of these bivalent TCR complexes, although greater antigenic binding and signaling is predicted over that of monovalent TCRs.