Enhanced surface TCR replenishment mediated by CD28 leads to greater TCR engagement during primary stimulation.

When T cells are stimulated with high concentrations of strong TCR agonist, engaged TCR are internalized and degraded, resulting in greatly reduced surface TCR levels for up to several days post-stimulation. It has been noted that surface TCR levels rise subsequently, even in the presence of continuing stimulation, but the role of CD28 co-stimulation in surface TCR replenishment has not been investigated. Here, we have examined the return of surface TCR following activation, the availability of these TCR for antigenic engagement and the role of CD28 in that process. We report that within 24 h of stimulation, the level of surface TCR expression becomes dependent on the degree of CD28 signaling provided during T cell activation. In addition, when cells are removed from stimulus after 24 h, surface TCR expression recovers to a stable level which exceeds that of unstimulated cells and is proportional to the degree of CD28 co-stimulation. TCR that replenish the plasma membrane during T cell activation can be down-regulated by receptor occupancy with the same efficiency as TCR on freshly stimulated cells. Thus, as a result of enhanced surface TCR replenishment, CD28-co-stimulated cells can engage and down-regulate more TCR than co-stimulation-deprived cells in the face of ongoing stimulation. Furthermore, engagement of newly expressed TCR on activated T cells re-induces CD69, suggesting participation of these replenishing TCR in continued T cell signaling. These data identify the augmentation of surface TCR replenishment during activation as a novel mechanism that likely contributes to the enhanced antigenic sensitivity of CD28-co-stimulated T cells.