PMA/ionomycin induces Igχ 3' enhancer activity which is in part mediated by a unique NFAT transcription complex

Abstract

The Igχ 3' enhancer is required for high levels of Igχ gene expression. We now show that χ3' enhancer function increases five- to eightfold after stimulation of primary murine B cells with phorbol 12-myristate 13-acetate (PMA) and the calcium ionophore ionomycin. In the presence of cyclosporin A this induction is almost halved, suggesting that transcription factors of the NFAT family contribute to χ3' enhancer induction. Indeed, we identify a novel NFAT binding site which is required for full enhancer function. We find that this site is transcriptionally active in stimulated B cells, T cells and fibroblasts and that both PMA and ionomycin are required for maximal induction. Time course analysis of the components of the protein-DNA complex in primary lymphocytes reveals that both NFATp and NFATc are present in the complex after 15 min, while only NFATc is detectable after 4 h. This suggests that NFATc plays the dominant role in controlling long-term responses of this transcription factor family. Furthermore, JunB, JunD, FosB and cFos form part of the DNA-protein complex in Bal-17 B cells. Complex formation as well as transcriptional activity can also be induced by crosslinking of surface Ig. We have, thus, identified a unique NFAT complex in B cells that contributes to Igχ gene expression.