Sp1 is a component of the cytokine-inducible enhancer in the promoter of vascular cell adhesion molecule-1

Abstract

Transcription of the vascular cell adhesion molecule-1 (VCAM-1) gene in endothelial cells is induced by the inflammatory cytokines interleukin-1β, tumor necrosis factor-α, and lipopolysaccharide. Previous studies demonstrated that the cytokine-response region in the VCAM1 promoter contains binding sites for the transcription factors nuclear factor-κB (NF-κB) and interferon regulatory factor-1. Using a saturation mutagenesis approach, we report that the cytokine-inducible enhancer consists of these previously characterized elements and a novel region located 3' of the NF-κB sites. Electrophoretic mobility shift assays and DNase I footprint studies with endothelial nuclear extracts and recombinant protein revealed that the transcriptional activator Sp1 interacts with this novel element in a specific manner. Transient transfection assays using vascular endothelial cells revealed that site-directed mutations in the Sp1 binding element decreased tumor necrosis factor-α-induced activity of the VCAM1 promoter. The cytokine-induced enhancer of the VCAM1 gene requires constitutively bound Sp1 and induced heterodimeric NF-κB for maximal promoter activity.