MICROBIAL SUPERANTIGENS INDUCE NF-KAPPA-B IN THE HUMAN MONOCYTIC CELL-LINE THP-1

Staphylococcal superantigens bind to MHC class II molecules and induce transcriptional activation of IL-1beta and TNF-alpha genes in human m onocytic cells. The understanding of the mechanisms by which superanti gens activate cytokine gene expression is incomplete. In this study, w e demonstrate that toxic shock syndrome toxin-1 (TSST-1) and staphyloc occal enterotoxins A and B induce the activation of NF-kappaB, a trans criptional enhancer that binds to sequences found in both the IL-1beta and TNF-alpha promoters. Electrophoretic mobility-shift assays showed a rapid induction of nuclear proteins that bound to the consensus kap paB motif. Furthermore, TSST-1 potently stimulated chloramphenicol ace tyltransferase (CAT) expression by THP-1 cells transfected with a cons ensus NF-kappaB-promoter CAT construct, indicative of induction of NF- kappaB enhancer function. Induction of both NF-kappaB DNA-binding prot eins and NF-kappaB enhancer function was down-regulated by inhibitors of protein kinase C and protein tyrosine kinase, indicating a role for these protein kinases in the induction of NF-kappaB by MHC class II l igands. Using neutralizing antibodies, we demonstrated that after the stimulation of cells with TSST-1, TNF-alpha, but not IL-1beta, acted t o up-regulate binding of NF-kappaB to DNA and the activation of the NF -kappaB-promoter CAT construct. These results indicate that induction of NF-kappaB by superantigens is up-regulated in part by an autocrine loop involving TNF-alpha.