The Impact of Single Amino Acid Substitutions in CD3γ on the CD3ϵγ Interaction and T-Cell Receptor–CD3 Complex Formation

The human T-cell receptor–CD3 complex consists of at least eight polypeptide chains; CD3γϵ- and δϵ-dimers associate with the disulfide linked αβ- and ζζ-dimers to form a functional receptor complex. The exact structure of this complex is still unknown. We now have examined the interaction between CD3γ and CD3ϵ in human T-cells. For this purpose, we have generated site-directed mutants of CD3γ that were introduced in human T-cells defective in CD3γ expression. Cell-surface and intracellular expression of the introduced CD3γ chains was determined, as was the association with CD3δ, CD3ϵ, and the T-cell receptor. Although the introduction of wild type CD3γ and CD3γ (78Y-F) fully restored T-cell receptor assembly and expression, the introduction of CD3γ (82C-S), CD3γ (85C-S), and CD3γ (76Q-E) all resulted in an impaired association between CD3γ and CD3ϵ and a lack of cell-surface expressed CD3γ. Finally, the introduction of CD3γ (76Q-L) and CD3γ (78Y-A) restored the expression of TCR-CD3δϵγϵζ2 complexes, although the association between CD3γ and CD3ϵ was impaired. These results indicate that several amino acids in CD3γ are essential for an optimal association between CD3γ and CD3ϵ and the assembly of a cell-surface expressed TCR-CD3δϵγϵζ2 complex.