Transcriptional activation of the human TNF-alpha promoter by superantigen in human monocytic cells: role of NF-kappa B.
We have studied the transcriptional activation of the human TNF-alpha gene by the superantigen staphylococcal enterotoxin A (SEA) in the human premonocytic cell line THP-1. Nuclear proteins from SEA-stimulated THP-1 cells bound strongly to kappa 3, the most proximal of three putative NF-kappa B binding sites (kappa 1-kappa 3) found in the 5' regulatory region of the TNF-alpha gene, but only weakly to kappa 1, the most distal of the NF-kappa B binding sites, and showed no binding to kappa 2. The mobility of the kappa 3-nucleoprotein complex was identical to that of complexes formed between nuclear proteins and the consensus NF-kappa B seuqence. Moreover, both 5' and 3' mutants of kappa 3 were unable to displace kappa 3 binding, suggesting that the kappa 3 binding complex induced by SEA has the characteristics of NF-kappa B. Studies using Abs directed against the NF-kappa B subunits p50 and p65 suggested that both p50 and p65 bind to the kappa 3 sequence. Reporter gene assays showed that deletion of kappa 3 (-99 to -89 bp) and point mutation of the three 5' guanine bases in the kappa 3 sequence reduced the inducibility of the TNF-alpha promoter by SEA and LPS. These results indicate that superantigen induces NF-kappa B in human monocytic cells and suggest that binding of NF-kappa B to the kappa 3 site of the TNF-alpha promoter plays an important role in the transcriptional activation of the TNF-alpha gene by superantigen.
Trede
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NS
Division of Immunology, Children's Hospital, Boston, MA 02115, USA.
Tsytsykova
A V
AV
Chatila
T
T
Goldfeld
A E
AE
Geha
R S
RS
D001483:Base Sequence; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000032:analysis; D004768:Enterotoxins / Q000494:pharmacology*; D006801:Humans; D008969:Molecular Sequence Data; D009000:Monocytes / Q000276:immunology*; D016328:NF-kappa B / Q000502:physiology; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics; D018089:Superantigens / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000187:drug effects* / Q000235:genetics*
D004768:Enterotoxins; D016328:NF-kappa B; D018089:Superantigens; D014409:Tumor Necrosis Factor-alpha; C026066:enterotoxin A, Staphylococcal; D015500:Chloramphenicol O-Acetyltransferase