Degradation of ZAP-70 following antigenic stimulation in human T lymphocytes: Role of calpain proteolytic pathway

T cell activation by the specific Ag results in dramatic changes of the T c ell phenotype that include a rapid and profound down-regulation and degrada tion of triggered TCRs, In this work, we investigated the fate of the TCR-a ssociated ZAP-70 kinase in Ag-stimulated T cells. T cells stimulated by pep tide-pulsed APCs undergo an Ag dose-dependent decrease of the total cellula r content of ZAP-70, as detected by FACS analysis and confocal microscopy o n fixed and permeabilized T cell-APC conjugates and by Western blot on tota l cell lysates, The time course of ZAP-70 consumption overlaps with that of zeta-chain degradation, indicating that ZAP-70 is degraded in parallel wit h TCR internalization and degradation. Pharmacological activation of protei n kinase C (PKC) does not induce ZAP-70 degradation, which, on the contrary , requires activation of protein tyrosine kinases, Two lines of evidence in dicate that the Ca2+-dependent cysteine protease calpain plays a major role in initiating ZAP-70 degradation: 1) treatment of T cells with cell-permea ting inhibitors of calpain markedly reduces ZAP-70 degradation; 2) ZAP-70 i s cleaved in vitro by calpain, Our results show that, in the course of T ce ll-APC cognate interaction, ZAP-70 is rapidly degraded via a calpain-depend ent mechanism.