GABP FACTORS BIND TO A DISTAL INTERLEUKIN-2 (IL-2) ENHANCER AND CONTRIBUTE TO C-RAF-MEDIATED INCREASE IN IL-2 INDUCTION

Triggering of the T-cell receptor-CD3 complex activates two major sign al cascades in T lymphocytes, (i) Ca2+-dependent signal cascades and ( ii) protein kinase cascades, Both signal cascades contribute to the in duction of the interleukin 2 (IL-2) gene during T-cell activation, Pro minent protein kinase cascades are those that activate mitogen-activat ed protein (MAP) kinases, We show here that c-Raf, which is at the hel m of the classic MAP-Erk cascade, contributes to IL-2 induction throug h a distal enhancer element spanning the nucleotides from positions -5 02 to -413 in front of the transcriptional start site of the IL-2 gene , Induction of this distal IL-2 enhancer differs from induction of the proximal IL-2 promoter-enhancer, since it is induced by phorbol ester s alone and independent from Ca2+ signals, In DNA-protein binding stud ies, we detected the binding of transcription factors GABP alpha and - beta to a dyad symmetry element (DSE) of the distal enhancer, which is formed by palindromic binding sites of Ets-like factors, Introduction of point mutations suppressing GABP binding to the DSE interfered wit h the induction of the distal enhancer and the entire IL-2 promoter-en hancer, while overexpression of both GABP factors enhanced the IL-2 pr omoter-enhancer induction, Overexpression of BXB, a constitutive activ e version of c-Raf, and of further members of the Ras-Raf-Erk signal c ascade exerted an increase of GABP-mediated promoter-enhancer inductio n, In conjunction with previously published data on c-Raf-induced phos phorylation of GABP factors (E. Flory, A. Hoffmeyer, U. Smola, U. R. R app, and J. T. Bruder, J. Virol, 70:2260-2268, 1996), these results in dicate a contribution of GABP factors to the Raf-mediated enhancement of IL-2 induction during T-cell activation.