CTLA-4 ligation suppresses CD28-induced NF-kappaB and AP-1 activity in mouse T cell blasts

The effects of cytotoxic lymphocyte antigen 4 (CTLA-4) on CD3/CD28 monoclonal antibody (mAb) activation of CD4(+)/CTLA-4(+) blastoid T cells were studied in an in vitro model system. As previously reported, coligation of CTLA-4 mAb results in suppression of T cell proliferation and cytokine production. The proliferation but not the interleukin 2 (IL-2) production could be restored by addition of exogenous IL-2, suggesting that the inhibitory effect occurred at the level of IL-2 production rather than at the regulation of the IL-2 receptor pathway. To study the effects on nuclear factors critical for T cell activation, we analyzed the levels of the transcription factors NF-kappaB and AP-1. These were potently induced in CD3/CD28 mAb-restimulated T cells. In contrast, CTLA-4 ligation strongly suppressed the induction of both transcription factors. The compositions of NF-kappaB and AP-1 family members were similar, irrespective of stimulation conditions. Analyses of the NF-kappaB regulator IkappaB-alpha revealed similar levels of IkappaB-alpha protein in the preparations. However, a reduced phosphorylation of IkappaB-alpha in CTLA-4 coengaged T cell blasts compared with T cells ligated with CD3/CD28 was found. Previous studies have concluded that CTLA-4 ligation regulates T cell activation by inhibiting the T cell receptor-mediated signals. However, the present findings propose that the major impact of CTLA-4 ligation is inhibition of signals mediated by CD28.