Activation of T cells through the antigen-specific T-cell receptor in combi
nation with a costimulatory signal results in efficient cytokine gene trans
cription. The CD28-induced signal represents a major costimulatory signal f
or T cells. A CD28 response element, named CD28RE, was first identified in
the interleukin-2 (IL-2) promoter region, Here we demonstrate that the NF-k
appa B sequence in the IL-6 promoter functions as a CD28 response element.
Mutations in this sequence rendered the IL-6 promoter unresponsive to CD28
costimulation, Moreover, this element could replace the IL-2 CD28RE in conf
erring CD28 responsiveness to the IL-2 promoter. In analogy to the known CD
28 response elements IL-2 CD28RE, IL-8 CD28RE, and the human immunodeficien
cy virus-1 (HIV-1) NF-kappa B motif, the IL-6 NF-kappa B motif efficiently
bound c-Rel, c-Rel/NFKB1, and the recently identified inducible T-cell fact
or NF-MATp35, However, the IL-6 NF-kappa B sequence together with the IL-8
CD28RE and HIV-1 NF-kappa B sequence differed from the IL-2 CD28RE in the b
inding of NF-kappa B/Rel family proteins. Although the IL-2 CD28RE exerted
selective binding with c-Rel and c-Rel/NFKB1, the other CD28REs allowed eff
icient binding of a wide range of NF-kappa B/Rel family proteins. The diffe
rence in binding specificity correlated with the capacity of the distinct C
D28 response elements to function in the context of the IL-6 promoter in re
sponse to T-cell activation. Domain swapping experiments revealed that the
IL-8 CD28RE and HIV-1 NF-kappa B motif conferred similar responsiveness as
the genuine IL-6 NF-kappa B motif in the transcriptional activation of the
IL-6 promoter upon CD28 costimulation, In contrast, replacement of the IL-6
NF-kappa B sequence by the IL-2 CD28RE motif strongly reduced the responsi
veness of the IL-6 promoter. These data indicate that despite the sequence
similarity, two different classes of CD28 responsive elements exist that di
ffer in their NF-kappa B binding capacity and the ability to confer CD28 co
stimulatory responsiveness toward a heterologous promoter.