Mouse MHC class I tetramers that are unable to bind to CD8 reveal the need for CD8 engagement in order to activate naive CD8 T cells.

Although the role of CD8 as a supplier of lck is unchallenged, its role in contributing to the formation of a stable complex between class I molecules and the TCR, as well as its role as an adhesion molecule, is less clear. To address the role of CD8/MHC-I interactions, we generated tetramers composed of H2-K(b) molecules with mutations in the alpha 3 domain of H2-K(b) that abolish CD8 binding. We show that the ability of tetramers to stain and activate CD8 T cells is strongly dependent on binding of CD8 to the same class I molecule engaged by the TCR. We characterize a mutation in the alpha 3 domain that results in H2-K(b) molecules capable of staining specific CD8 T cells with little ensuing activation. Although CD8 to some extent serves an adhesive function, this contribution is modest and does not substitute for lack of binding of CD8 to the class I molecule engaged by the TCR. We show that CD8 and the TCR associate in a process independent of binding of CD8 to class I. Our data support the notion that CD8 is required to form a stable complex between class I and the TCR.