TCR alpha beta-independent CD28 signaling and costimulation require non-CD4-associated Lck

Whether the sequelae of signals generated through CD28 either directly or i n circumstances of costimulation require proximal events mediated by p56(lc k) remains contentious. We demonstrate that CD4(-), but not CD4(+) clonal v ariants respond to CD28-specific mAb with both early and late indicators of activation. Forced expression of A418/A420-mutated CD4 or wild-type CD4 in the CD4- variant recapitulated the CD28-mediated responses of the CD4(-) a nd CD4(+) variants, respectively. The implicated involvement of non-CD4-ass ociated Lck is formally demonstrated by overexpressing S20/S23 Lck or wild- type Lck in CD4(+) variants, The former, but not latter, rescues direct CD2 8 signaling, and supports costimulation, The results demonstrate that const itutive levels of non-CD4-associated Lck functionally limit CD28-mediated s ignaling.