Activation of the Interleukin-6 promoter by a dominant negative mutant of c-Jun.
The human IL-6 promoter contains multiple regulatory elements such as those binding transcription factors belonging to the NF-kappaB (-75/-63), C/EBP (-158/-145 and -87/-76) and AP-1 (-283/-277) families. Herein, we report that ectopic expression of c-Jun, C/EBPdelta, and the p65 subunit of NF-kappaB synergistically activates an IL-6 promoter construct containing only a TATA box and a kappaB binding site. These results suggest that interactions among NF-kappaB, C/EBP, and AP-1, which are all activated by the most powerful physiological inducers of the IL-6 gene, namely TNF-alpha and IL-1, may be crucial for maximal activation of the IL-6 promoter in response to the two cytokines. Furthermore, we show that a mutated form of c-Jun lacking the transactivation domain (TAM-67) was a much stronger activator of the IL-6 promoter than c-Jun. In combination with p65 and/or C/EBPdelta, TAM-67 also synergistically activated the IL-6 promoter, while it inhibited TNF-alpha induced AP-1 activity directing an AP-1-responsive reporter plasmid. Lastly, electrophoretic mobility shift assay (EMSA) results strongly suggest the formation of complexes between p65, C/EBPdelta, and/or c-Jun or TAM-67 on the kappaB site, supporting the idea that the functional synergism is determined by a physical interaction. These data provide new insight into the molecular mechanisms regulating the formation of the transcription complex responsible for IL-6 promoter activation.
Faggioli
Laura
L
Department of Neurological and Vision Sciences, Biochemistry Section, University of Verona, Strada Le Grazie 8, 37134 Verona, Italy.
Costanzo
Chiara
C
Donadelli
Massimo
M
Palmieri
Marta
M
D001665:Binding Sites; D051743:CCAAT-Enhancer-Binding Protein-delta; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism; D002135:Calcium-Binding Proteins; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics*; D008562:Membrane Glycoproteins / Q000378:metabolism; D009154:Mutation; D016328:NF-kappa B / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D016755:Proto-Oncogene Proteins c-jun / Q000235:genetics* / Q000378:metabolism; D050863:Synaptotagmin I; D050857:Synaptotagmins; D018808:Transcription Factor AP-1 / Q000378:metabolism; D051996:Transcription Factor RelA; D014157:Transcription Factors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D022762:CCAAT-Enhancer-Binding Proteins; C497252:CEBPD protein, human; D002135:Calcium-Binding Proteins; D015850:Interleukin-6; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D016755:Proto-Oncogene Proteins c-jun; D050863:Synaptotagmin I; C439528:TAM67 peptide; D018808:Transcription Factor AP-1; D051996:Transcription Factor RelA; D014157:Transcription Factors; D050857:Synaptotagmins; D051743:CCAAT-Enhancer-Binding Protein-delta