Formation of supramolecular activation clusters on fresh ex vivo CD8(+) T cells after engagement of the T cell antigen receptor and CD8 by antigen-presenting cells

Upon productive interaction of CD4 T cells with antigen-presenting cells (A PCs), receptors and intracellular proteins translocate and form spatially s egregated supramolecular activation clusters (SMACs). It is not known wheth er SMACs are required for CD8 T cell activation. CD8 T cells, unlike CD4 T cells, can be activated by a single peptide-MHC molecule, or by purified mo novalent recombinant peptide-MHC molecules. We studied, by three-dimensiona l digital microscopy, cell conjugates of fresh ex vivo CD8 T cells (obtaine d from OT-1 mice, which are transgenic for T cell antigen receptor reactive with the complex of H-2K(b) and the ovalbumin octapeptide SIINFEKL) and pe ptide-pulsed APCs. Remarkably, even in T cell:APC conjugates that were form ed in the presence of the lowest concentration of peptide that was sufficie nt to elicit T cell proliferation and IFN-gamma production; the theta isofo rm of protein kinase C was clustered in a central SMAC, and lymphocyte func tion-associated antigen 1 and talin were clustered in the peripheral SMAC. Conjugation of T cells to APCs that were pulsed with concentrations of pept ide smaller than that required to activate T cells was greatly reduced, and SMACs were not formed at all. APCs expressing mutant H-2K(b) (Lys(227)) mo lecules that do not bind CD8 were unable to form stable conjugates with the se T cells, even at high peptide concentrations. Thus, although CD8 and CD4 T cells may display different sensitivity to the concentration and oligome rization of surface receptors, SMACs are formed and seem to be required fun ctionally in both cell types. However, unlike CD4 T cells, which can form S MACs without CD4, CD8 T cells from OT-1 transgenic mice depend on their cor eceptor, CD8, for the proper formation of SMACs.