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Naive CD4+ T-cell differentiation to T helper 1 (Th1) and Th2 cells is dependent on T-bet and GATA-3 factors, respectively. T-bet and GATA-3, indeed, through chromatin remodelling allow transcriptional activation of Ifngamma and Th2 cytokine (Il4, Il5, Il13) genes, respectively. We investigated the effects of the negative costimulatory receptor cytotoxic T-lymphocyte antigen-4 (CTLA-4) on GATA-3 and T-bet mRNA expression and Th cell differentiation in mouse naive CD4+ T cells. Our results show that CTLA-4 inhibits GATA-3 mRNA expression and Th2 cell differentiation. At variance, CTLA-4 does not affect T-bet mRNA expression and Th1 cell differentiation. GATA-3 mRNA expression is inhibited when CD4+ cells are stimulated under both neutral (i.e. absence of cytokines) and Th2-polarizing (i.e. presence of interleukin (IL)-4) conditions, the effect being larger under the latter condition. Hence CTLA-4 might affect the IL-4/signal transducer and activator of transcription-6 (STAT6) pathway leading to GATA-3 mRNA up-regulation. We found, indeed, that CTLA-4 engagement inhibits STAT6 activation leaving unaffected the STAT6 protein level. Moreover, CTLA-4 engagement drastically inhibits IL-4Ralpha mRNA and protein up-regulation under Th2-polarizing conditions. Thus, CTLA-4 exerts a tight control on Th2 cell differentiation by negatively regulating both the CD3/CD28 and the IL-4/STAT6 pathways.

The negative role exerted by cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) in the regulation of T-cell activity, as induced by T-cell receptor (TCR)/CD3 and CD28 costimulation, has been widely described. In the present work we investigated the role of CTLA-4 in the control of cell activation, as induced by costimulation of the adhesion molecule lymphocyte function-associated antigen-1 (LFA-1) in murine CD4+ T cells. Results show that CTLA-4 engagement inhibits interleukin-2 (IL-2) production, not only when induced by CD3/CD28 costimulation, but also when CD4+ T cells are costimulated by anti-CD3 and anti-LFA-1 monoclonal antibodies (mAbs). LFA-1 has been described to induce Ca2+ mobilization also in the absence of TCR engagement. Moreover, we found that CTLA-4 engagement negatively affects Ca2+ mobilization and NF-AT activation, as induced by LFA-1 engagement alone. PLCgamma1 phosphorylation was also dampened within minutes after CTLA-4 engagement. Altogether these data indicate that through the control of signals induced by different receptors, CTLA-4 could be a global attenuator of T-cell activation.