NF-κB activation induced by T cell receptor/CD28 costimulation is mediated by protein kinase C-θ

Protein kinase C-theta (PKC theta) is a Ca2+-independent member of the PKC family that is selectively expressed in skeletal muscle and T lymphocytes and plays an important role in T cell activation. However, the molecular basis for the important functions of PKC theta in T cells and the manner in which it becomes coupled to the T cell receptor-signaling machinery are unknown. We addressed the functional relationship between PKC theta and CD28 costimulation, which plays an essential role in T cell receptor-mediated IL-2 production. Here, we provide evidence that PKC theta is functionally coupled to CD28 costimulation by virtue of its selective ability to activate the CD28RE/activator protein-1 (AP-1) element in the IL-2 gene promoter. First, CD28 costimulation enhanced the membrane translocation and catalytic activation of PKC theta. Second, among several PKC isoforms, PKC theta was the only one capable of activating NF-kappa B or CD28RE/AP-1 reporters in T cells (but not in 293T cells). Third, wild-type PKC theta synergized with CD28/CD3 signals to activate CD28RE/AP-1. In addition, PKC theta selectively synergized with Tat to activate a CD28RE/AP-1 reporter. Fourth, CD3/CD28-induced CD28RE/AP-1 activation and NF-kappa B nuclear translocation were blocked by a selective PKC theta inhibitor. Last PKC theta-mediated activation of the same reporter was inhibited by the proteasome inhibitor MG132 (which blocks I kappa B degradation) and was found to involve I kappa B-kinase beta. These findings identify a unique PKC theta-mediated pathway for the costimulatory action of CD28, which involves activation of the I kappa B-kinase beta/I kappa B/NF-kappa B-signaling cascade.