Structure of the chicken CD3epsilon/delta/gamma heterodimer and its assembly with the alphabetaT cell receptor

In mammals, the alphabetaT cell receptor (TCR) signaling complex is composed of a TCRalphabeta heterodimer that is noncovalently coupled to three dimeric signaling molecules, CD3epsilondelta, CD3epsilongamma, and CD3zetazeta. The nature of the TCR signaling complex and subunit arrangement in different species remains unclear however. Here we present a structural and biochemical analysis of the more primitive ancestral form of the TCR signaling complex found in chickens. In contrast to mammals, chickens do not express separate CD3delta and CD3gamma chains but instead encode a single hybrid chain, termed CD3delta/gamma, that is capable of pairing with CD3epsilon. The NMR structure of the chicken CD3epsilondelta/gamma heterodimer revealed a unique dimer interface that results in a heterodimer with considerable deviation from the distinct side-by-side architecture found in human and murine CD3epsilondelta and CD3epsilongamma. The chicken CD3epsilondelta/gamma heterodimer also contains a unique molecular surface, with the vast majority of surface-exposed, nonconserved residues being clustered to a single face of the heterodimer. Using an in vitro biochemical assay, we demonstrate that CD3epsilondelta/gamma can assemble with both chicken TCRalpha and TCRbeta via conserved polar transmembrane sites. Moreover, analogous to the human TCR signaling complex, the presence of two copies of CD3epsilondelta/gamma is required for zetazeta assembly. These data provide insight into the evolution of this critical receptor signaling apparatus.