GLU227-]LYS SUBSTITUTION IN THE ACIDIC LOOP OF MAJOR HISTOCOMPATIBILITY COMPLEX CLASS-I ALPHA-3 DOMAIN DISTINGUISHES LOW AVIDITY CD8 CORECEPTOR AND AVIDITY-ENHANCED CD8 ACCESSORY FUNCTIONS

Cytotoxic T lymphocyte (CTL) activation requires specific T cell recep tor (TCR)-class I major histocompatibility complex (MHC) antigen compl ex interactions as well as the participation of coreceptor or accessor y molecules on the surface of CTL. CD8 can serve as a coreceptor in th at it binds to the same MHC class I molecules as the TCR to facilitate efficient TCR signaling. In addition, CD8 can be ''activated'' by TCR stimulation to bind to class I molecules with high avidity, including class I not recognized by the TCR as antigenic complexes (non-antigen [Ag] class I), to augment CTL responses and thus serve an accessory m olecule function. A Glu/Asp227-->Lys substitution in the class I alpha 3 domain acidic loop abrogates lysis of target cells expressing these mutant molecules by alloreactive CD8-dependent CTL. Lack of response is attributed to the destruction of the CD8 binding site in the alpha 3 domain which is likely to disrupt CD8 coreceptor function. The relat ive importance of the class I alpha 3 domain acidic loop Glu227 in cor eceptor as opposed to accessory functions of CD8 is unclear. To addres s this issue, we examined CTL adhesion and degranulation in response t o immobilized class I-peptide complexes formed in vitro from antigenic peptides and purified class I molecules containing wild-type or Glu22 7-->Lys substituted alpha 3 domains. The alpha 3 domain mutant class I -peptide complexes. were bound by CTL and triggered degranulation, how ever to much lower levels than wild-type class I-peptide complexes. In further experiments, it is directly demonstrated that the alpha 3 dom ain mutant class I molecules, which lack the Glu227 CD8 binding site, still serve as TCR-activated, avidity-enhanced CD8 accessory ligands. However, mutant class I-peptide Ag complexes failed to effectively ser ve as CD8 coreceptor ligands to initiate TCR-dependent signals require d to induce avidity-enhanced CD8 binding to coimmobilized non-Ag class I molecules. Thus the Glu227-->Lys mutation effectively distinguishes CD8 coreceptor and avidity-enhanced CD8 accessory functions.