The interaction with Sp1 and reduction in the activity of histone deacetylase 1 are critical for the constitutive gene expression of IL-1 alpha in human melanoma cells.
A375-6 human melanoma cells are sensitive to the antiproliferative effect of IL-1. After a long period of culturing, we have obtained cells resistant to IL-1. The resistant clone A375-R8 constitutively produced IL-1 alpha. In this study, we identified a sequence, CGCC, located at -48 to -45 upstream of the transcription start site, to be essential for the constitutive IL-1 alpha gene activation. Specificity protein 1 (Sp1) and Sp3 bound to the nucleotide containing the sequence. Although the binding level to the nucleotide and expression level of Sp1 and Sp3 are comparable in A375-R8 and A375-6 cells, transactivation activity of Sp1 is higher in A375-R8 cells as compared with A375-6 cells. Sp3 could not transactivate the IL-1 alpha promoter. These results suggest that Sp1 but not Sp3 is important for IL-1 alpha gene activation. Trichostatin A (TSA), an inhibitor of histone deacetylase (HDAC), greatly augmented the IL-1 alpha promoter activity in A375-6 cells to the level comparable with that in A375-R8 cells. TSA also induced IL-1 alpha mRNA expression in A375-6 cells. Sp1 and Sp3 bound to HDAC1 in A375-R8 and A375-6 cells. The chromatin immunoprecipitation assay revealed the binding of Sp1 and HDAC1 to the promoter region of the IL-1 alpha gene. The activities of HDAC bound to Sp1 and Sp3, and that of HDAC1 was lower in A375-R8 cells as compared with A375-6 cells. These results indicate that the reduction in the activity and interaction of HDAC1 with Sp1 are critical for the constitutive IL-1 alpha gene expression.
Enya
Kazuaki
K
Department of Molecular Health Sciences, Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabe-dori, Mizuho-ku, Nagoya, Aichi 467-8603, Japan.
Hayashi
Hidetoshi
H
Takii
Takemasa
T
Ohoka
Nobumichi
N
Kanata
Shinya
S
Okamoto
Takashi
T
Onozaki
Kikuo
K
D045744:Cell Line, Tumor; D004247:DNA / Q000276:immunology; D004789:Enzyme Activation / Q000276:immunology; D020869:Gene Expression Profiling; D056284:Histone Deacetylase 1; D006655:Histone Deacetylases / Q000378:metabolism*; D006801:Humans; D053582:Interleukin-1alpha / Q000096:biosynthesis* / Q000235:genetics*; D008545:Melanoma / Q000276:immunology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000096:biosynthesis / Q000378:metabolism*; D051705:Sp3 Transcription Factor / Q000096:biosynthesis / Q000378:metabolism; D014407:Tumor Cells, Cultured
D053582:Interleukin-1alpha; D016329:Sp1 Transcription Factor; D051705:Sp3 Transcription Factor; D004247:DNA; C110550:HDAC1 protein, human; D056284:Histone Deacetylase 1; D006655:Histone Deacetylases