Level of interleukin-8 expression by metastatic human melanoma cells directly correlates with constitutive NF-kappaB activity.
The purpose of this study was to determine whether constitutive NF-kappaB activity regulates the expression level of interleukin-8 (IL-8) in metastatic human melanoma cells. Cultures of metastatic human A375 melanoma cells expressed higher levels of IL-8 mRNA and protein than nonmetastatic A375 human melanoma cells. No discernible differences in IL-8 half-life were found between metastatic and nonmetastatic cells, but cells that overexpressed IL-8 had a higher transcription rate and increased IL-8 promoter activity. Analysis of the IL-8 promoter using deletion mutants revealed that the region within -133 was essential for constitutive IL-8 promoter activity and that mutation of NF-kappaB binding sites eliminated the constitutive IL-8 promoter activity. The activation of constitutive IL-8 transcription directly correlated with the level of constitutive NF-kappaB activity. Transfection of melanoma cells with a dominant-negative mutant IkappaBalpha expression vector (pLXSN-IkappaBalphaM) significantly decreased the level of constitutive NF-kappaB activity and expression of IL-8, demonstrating that constitutive NF-kappaB/relA activities contribute to overexpression of IL-8 in highly metastatic human melanoma cells.
Huang
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S
Department of Cancer Biology, The University of Texas MD Anderson Cancer Center, Houston 77030, USA.
DeGuzman
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A
Bucana
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CD
Fidler
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IJ
D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D004587:Electrophoresis, Agar Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D020672:I-kappa B Proteins; D016209:Interleukin-8 / Q000096:biosynthesis / Q000235:genetics*; D008156:Luciferases / Q000378:metabolism; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000556:secondary; D008545:Melanoma / Q000235:genetics / Q000378:metabolism* / Q000556:secondary; D009154:Mutation; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis*; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000378:metabolism
D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016209:Interleukin-8; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D012333:RNA, Messenger; D000072000:NF-KappaB Inhibitor alpha; D008156:Luciferases