T cell activation by recombinant Fc epsilon RI gamma-chain immune receptors: an extracellular spacer domain impairs antigen-dependent T cell activation but not antigen recognition

T cells can be endowed with antigen specificity by grafting with a chimeric receptor consisting of an extracellular antigen binding moiety (scFv) deri ved from an antibody and an intracellular signaling domain. Conflicting dat a exist on the impact of an extracellular spacer domain between the antigen binding and the signaling domain with respect to cellular activation. Here , we recorded conjugate formation and antigen-driven cellular activation of T cells grafted with receptor molecules that contain the same antigen bind ing site (anti-CD30 HRS3-scFv) and signaling domain (Fc epsilon Rl gamma-ch ain), however, with and without an IgG1 CH2CH3 (Fc) spacer domain between t he scFv and transmembrane moiety. Receptors of both configurations mediate equally efficient conjugate formation between receptor grafted T cells and antigen-positive target cells. Specific signaling by the spacer containing receptor, however, is blocked by five- to 10-fold lower concentrations of s oluble antigen than by the spacerless receptor indicating a higher avidity of the spacer containing receptor to soluble antigen. In contrast, cellular activation upon binding to antigen-positive cells is mediated more efficie ntly by the spacer-less receptor. This demonstrates that the extracellular spacer domain impairs antigen-dependent cellular activation by the chimeric immune receptor, but not intercellular conjugate formation.